Categories:

Role of Serotonin in Preeclampsia

Also see:
Estrogen Increases Serotonin
Anti Serotonin, Pro Libido
Gelatin > Whey
Thyroid peroxidase activity is inhibited by amino acids
Whey, Tryptophan, & Serotonin
Tryptophan, Fatigue, Training, and Performance
Carbohydrate Lowers Free Tryptophan
Protective Glycine
Intestinal Serotonin and Bone Loss
Hypothyroidism and Serotonin
Estrogen Increases Serotonin
Gelatin, Glycine, and Metabolism
Whey, Tryptophan, & Serotonin
Tryptophan, Sleep, and Depression
Menstrual Cycle Related Epilepsy (Catamenial Epilepsy)
Estrogen’s Role in Seizures
The Brain – Estrogen’s Harm and Progesterone’s Protection
Estrogen, Glutamate, & Free Fatty Acids
Women, Estrogen, and Circulating DHA
Epilepsy and Progesterone by Ray Peat, PhD

“In women with preeclampsia, there are abnormally high levels of serotonin, nitric oxide, and lipid peroxidation. In a study of more than 3000 women (Clausen, et al., 2001), the consumption of sugar and polyunsaturated fat was strongly associated with the development of preeclampsia. Women who don’t eat enough protein are likely to substitute sugar and fat for the absent protein, so this study is consistent with Brewer’s work, but it’s very important to see that it was polyunsaturated fats, not saturated or monounsaturated fats, that caused the problem. Eclampsia (pregnancy-related seizures) and preeclampsia are caused by oxidative stress, produced by the excessive unstable fats. The increased serotonin and nitric oxide are exactly what would be expected to result from the high consumption of polyunsaturated fats, especially with a deficiency of protein in the diet.” -Ray Peat, PhD

Hypertension. 2007 Oct;50(4):773-9. Epub 2007 Jul 23.
Pregnant rats treated with a serotonin precursor have reduced fetal weight and lower plasma volume and kallikrein levels.
Salas SP, Giacaman A, Romero W, Downey P, Aranda E, Mezzano D, Vío CP.
Pregnant women with preeclampsia have increased serotonin levels, suggesting a possible role of this amine in abnormal pregnancy. With the hypothesis that an increase in serotonin would reduce volume expansion and cause fetal growth restriction, we evaluated the maternal and fetal effects of the administration of the serotonin precursor 5-hidroxytryptophan (5-HTP) to Sprague-Dawley rats. At pregnancy day 13 (n=19) or in random cycle nonpregnant rats (n=10), animals were assigned to a single injection of 5-HTP (100 mg/kg IP) or to a control group. Animals were studied at day 21, after overnight urinary collection. Additional pregnant rats received ketanserin (1 mg/kg), a 5-HT(2) receptor antagonist, 1 hour before 5-HTP injection. In pregnant rats, 5-HTP lowered plasma volume (control: 22+/-1.1; 5-HTP: 17+/-0.7 mL; P<0.001) and creatinine clearance, whereas serum creatinine and urinary protein excretion were increased; no changes were observed in nonpregnant rats. Systolic blood pressure did not change significantly. Urinary kallikrein activity and plasma aldosterone levels decreased only in pregnant animals. Fetal (control: 5.5+/-0.1; 5-HTP: 4.2+/-0.2 g; P<0.001) and placental weights were reduced. In nonpregnant and pregnant animals, 5-HTP caused profound renal morphological alterations and decreased kallikrein immunostaining. Preadministration of ketanserin abolished all of the changes associated with the use of 5-HTP. These data indicate that the administration of a serotonin precursor to pregnant rats limits plasma volume expansion and fetal growth via 5-HT(2) receptors, suggesting a possible role for serotonin in abnormal pregnancy. We postulate that an increased vascular resistance, both at the placental and renal levels, mediates these effects.

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Aldosterone, Sodium Deficiency, and Insulin Resistance

Also see:
The Randle Cycle
Free Fatty Acids Suppress Cellular Respiration
Aldosterone as an endogenous cardiovascular toxin
Aldosterone and Thrombosis
Sodium Deficiency and Stress
Low Sodium Diet: High FFA, Insulin Resistance, Atherosclerosis

Dietary salt restriction, which increases aldosterone levels, is also associated with an increase in insulin resistance. -Garg and Adler, 2012

Curr Opin Endocrinol Diabetes Obes. 2012 Jun;19(3):168-75.
Role of mineralocorticoid receptor in insulin resistance.
Garg R, Adler GK.
PURPOSE OF REVIEW:
Recent data suggest that mineralocorticoid receptor activation can affect insulin resistance independent of its effects on blood pressure. This review discusses new evidence linking mineralocorticoid receptor to insulin resistance and the underlying mechanisms of these effects.
RECENT FINDINGS:
Observational studies have shown mineralocorticoid activity to be associated with insulin resistance irrespective of race, blood pressure or body weight. Increased mineralocorticoid activity may be the common link between obesity, hypertension, dyslipidemia and insulin resistance, features that make up the metabolic syndrome. Treatment of primary aldosteronism is associated with a decrease in insulin resistance and provides one of the most convincing evidences in favor of the contribution of mineralocorticoid receptor to insulin resistance. Dietary salt restriction, which increases aldosterone levels, is also associated with an increase in insulin resistance. Potential mechanisms by which mineralocorticoid receptor may contribute to insulin resistance include a decreased transcription of the insulin receptor gene, increased degradation of insulin receptor substrates, interference with insulin signaling mechanisms, decreased adiponectin production and increased oxidative stress and inflammation. Advantages of mineralocorticoid receptor antagonists on insulin resistance have been demonstrated in animal models.
SUMMARY:
There may be a benefit of mineralocorticoid receptor antagonists in human insulin resistance states, but more clinical research is needed to explore these possibilities.

Prog Cardiovasc Dis. 2010 Mar-Apr;52(5):401-9.
Aldosterone: role in the cardiometabolic syndrome and resistant hypertension.
Whaley-Connell A, Johnson MS, Sowers JR.
The prevalence of diabetes, hypertension, and cardiovascular disease (CVD) and chronic kidney disease (CKD) is increasing in concert with obesity. Insulin resistance, metabolic dyslipidemia, central obesity, albuminuria. and hypertension commonly cluster to comprise the cardiometabolic syndrome (CMS). Emerging evidence supports a shift in our understanding of the crucial role of elevated serum aldosterone in promoting insulin resistance and resistant hypertension. Aldosterone enhances tissue generation of oxygen free radicals and systemic inflammation. This increase in oxidative stress and inflammation, in turn, contributes to impaired insulin metabolic signaling, reduced endothelial-mediated vasorelaxation, and associated cardiovascular and renal structural and functional abnormalities. In this context, recent investigation indicates that hyperaldosteronism, which is often associated with obesity, contributes to impaired pancreatic beta-cell function as well as diminished skeletal muscle insulin metabolic signaling. Accumulating evidence indicates that the cardiovascular and renal abnormalities associated with insulin resistance are mediated, in part, by aldosterone’s nongenomic as well as genomic signaling through the mineralocorticoid receptor (MR). In the CMS, there are increased circulating levels of glucocorticoids, which can also activate MR signaling in cardiovascular, adipose, skeletal muscle, neuronal, and liver tissue. Furthermore, there is increasing evidence that fat tissue produces a lipid soluble factor that stimulates aldosterone production from the adrenal zona glomerulosa. Recently, we have learned that MR blockade improves pancreatic insulin release, insulin-mediated glucose utilization, and endothelium-dependent vasorelaxation as well as reduces the progression of CVD and CKD. In summary, aldosterone excess exerts detrimental metabolic effects that contribute to the development of the CMS and resistant hypertension as well as CVD and CKD.

Ann Intern Med. 2009 Jun 2;150(11):776-83.
Narrative review: the emerging clinical implications of the role of aldosterone in the metabolic syndrome and resistant hypertension.
Sowers JR, Whaley-Connell A, Epstein M.
The prevalence of obesity, diabetes, hypertension, and cardiovascular and chronic kidney disease is increasing in developed countries. Obesity, insulin resistance, and hypertension commonly cluster with other risk factors for cardiovascular and chronic kidney disease to form the metabolic syndrome. Emerging evidence supports a paradigm shift in our understanding of the renin-angiotensin-aldosterone system and in aldosterone’s ability to promote insulin resistance and participate in the pathogenesis of the metabolic syndrome and resistant hypertension. Recent data suggest that excess circulating aldosterone promotes the development of both disorders by impairing insulin metabolic signaling and endothelial function, which in turn leads to insulin resistance and cardiovascular and renal structural and functional abnormalities. Indeed, hyperaldosteronism is associated with impaired pancreatic beta-cell function, skeletal muscle insulin sensitivity, and elevated production of proinflammatory adipokines from adipose tissue, which results in systemic inflammation and impaired glucose tolerance. Accumulating evidence indicates that the cardiovascular and renal abnormalities associated with insulin resistance are mediated in part by aldosterone acting on the mineralocorticoid receptor. Although we have known that mineralocorticoid receptor blockade attenuates cardiovascular and renal injury, only recently have we learned that mineralocorticoid receptor blockade improves pancreatic insulin release, insulin-mediated glucose utilization, and endothelium-dependent vasorelaxation. In summary, aldosterone excess has detrimental metabolic effects that contribute to the metabolic syndrome and endothelial dysfunction, which in turn contribute to the development of resistant hypertension as well as cardiovascular disease and chronic kidney disease.

Curr Hypertens Rep. 2011 Apr;13(2):163-72.
The role of aldosterone in the metabolic syndrome.
Briet M, Schiffrin EL.
The metabolic syndrome associates metabolic abnormalities such as insulin resistance and dyslipidemia with increased waist circumference and hypertension. It is a major public health concern, as its prevalence could soon reach 30% to 50% in developed countries. Aldosterone, a mineralocorticoid hormone classically involved in sodium balance regulation, is increased in patients with metabolic syndrome. Besides its classic actions, aldosterone and mineralocorticoid receptor (MR) activation affect glucose metabolism, inducing insulin resistance through various mechanisms that involve oxidative stress, inflammation, and downregulation of proteins involved in insulin signaling pathways. Aldosterone and MR signaling exert deleterious effects on the cardiovascular system and the kidney that influence the cardiovascular risk associated with metabolic syndrome. Salt load plays a major role in cardiovascular injury induced by aldosterone and MR signaling. Large multicenter, randomized clinical trials testing the beneficial effects of MR antagonists on cardiovascular events and mortality in patients with metabolic syndrome are needed.

Curr Hypertens Rep. 2010 Aug;12(4):252-7.
Mineralocorticoid receptor antagonists and the metabolic syndrome.
Tirosh A, Garg R, Adler GK.
Key components of the metabolic syndrome (MetS), ie, obesity and insulin resistance, are associated with increased aldosterone production and mineralocorticoid receptor (MR) activation. Both MetS and hyperaldosteronism are proinflammatory and pro-oxidative states associated with cardiovascular disease. This review discusses emerging data that MR activation may contribute to abnormalities seen in MetS. In view of these data, MR antagonists may be beneficial in MetS, not only by controlling hypertension but also by reversing inflammation, oxidative stress, and defective insulin signaling at the cellular-molecular level. Clinical trials have demonstrated benefits of MR antagonists in heart failure, hypertension, and diabetic nephropathy, but additional trials are needed to demonstrate the clinical significance of MR blockade in MetS.

Klin Wochenschr. 1991;69 Suppl 25:51-7.
Short-term dietary sodium restriction increases serum lipids and insulin in salt-sensitive and salt-resistant normotensive adults.
Ruppert M, Diehl J, Kolloch R, Overlack A, Kraft K, Göbel B, Hittel N, Stumpe KO.
Evidence suggests that dietary salt reduction similar to diuretic therapy may adversely affect lipid and glucose metabolism. We studied 147 non-obese normotensive subjects (60 females and 87 males) aged 19-78 years who entered a single-blind crossover trial and were randomly assigned to a low salt diet of 20 mmol or a high salt diet of 300 mmol sodium per day, for 7 days each. Sodium restriction lowered mean arterial blood pressure (MAP) by a mean of 7.5 mmHg in 17% (salt-sensitive), had no hemodynamic effect in 67% (salt-resistant) and raised MAP by a mean of 6 mmHg in 16% of the subjects (reverse reactors). With dietary salt restriction serum total- and LDL-cholesterol as well as serum insulin and uric acid concentrations increased significantly in all three groups. The largest increases in total (10%) and LDL- (12%) cholesterol occurred in the reverse reactors. Salt-sensitives had significant higher lipoprotein(a) values than the other two groups. Salt-restriction had no significant effect on this parameter. Plasma renin activity, as well as plasma aldosterone and noradrenaline concentrations rose in all three groups during the low salt diet, the largest increases being observed in the reverse reactors. Short-term sodium restriction in normotensive adults has unfavourable effects on lipid and glucose metabolism, especially in subjects who do not derive hemodynamic benefit. Further studies are necessary to examine the effects of more moderate salt reduction for longer periods on the risk factor profile for cardiovascular disease before a low salt diet can be regarded as a safe public health measure for the general population.

Aust J Exp Biol Med Sci. 1976 Feb;54(1):71-8.
Regulation of aldosterone in the guinea-pig–effect of oestrus cycle, pregnancy and sodium status.
Whipp GT, Wintour EM, Coghlan JP, Scoggins BA.
The blood concentrations of aldosterone, corticosterone and cortisol were measured in conscious, non-stressed guinea-pigs using a double isotope dilution derivative assay procedure. Aldosterone levels in the guinea-pig were high when compared with those of other species. The concentration of aldosterone, 37-7 +/- 15-9 ng/100 ml (x +/- SD), and cortisol, 31-8 +/- 10-1 mug/100 ml, found in non-pregnant females on a moderate sodium intake was significantly greater than in males (aldosterone 22-2 +/- 2-4 ng/100 ml and cortisol 19-3 +/- 5-7 mug/100 ml). There was no sex difference in corticosterone concentrations; females, 0-25 +/- 0-06 mug/100 ml and males, 0-23 +/- 0-10 mug/100 ml. The oestrus cycle had no effect on levels of the three steroids measured. Two thirds of the way through the 68-day gestation period aldosterone levels were significantly elevated compared with non-pregnant values (68-7 +/- 50-9 ng/100 ml, p less than 0-05). Values at day 20 (33-2 +/- 11-7 ng/100 ml) and day 60 of gestation (51-9 +/- 21-7 ng/100 ml) were similar to those of non-pregnant animals. Cortisol and corticosterone levels were significantly elevated at 20 days gestation and they continued to rise until, at day 60, cortisol was 9 times and corticosterone 4 times higher than the non-pregnant values. Compared with a moderate Na intake, salt loading suppressed aldosterone levels and Na restriction raised them.

J Clin Invest. 1972 October; 51(10): 2645–2652.
Studies of the control of plasma aldosterone concentration in normal man III
Robert G. Dluhy, Lloyd Axelrod, Richard H. Underwood, and Gordon H. Williams
The peripheral plasma levels of aldosterone, renin activity, potassium, sodium, corticosterone, and cortisol were measured in six normal subjects four times daily—10 a.m., 2 p.m., 5 p.m., 11 p.m.—on 3 consecutive days. A constant daytime activity program was maintained throughout the study. After 5 days on a 10 mEq sodium/100 mEq potassium isocaloric intake, the mean upright 10 a.m. plasma renin activity was 1773±186 ng/100 ml per 3 hr and the mean plasma aldosterone, 81±14 ng/100 ml. These two parameters fell continuously throughout the day parallel to the fall in plasma cortisol and corticosterone. In response to 2 liters of normal saline infused from 10 a.m. to 2 p.m. on 2 consecutive days, plasma aldosterone levels fell significantly to 13±5 ng/100 ml at 2 p.m. after the 1st day’s infusion and to 6±1 ng/100 ml at 2 p.m. after the 2nd. Plasma renin activity demonstrated a parallel fall to 368±63 ng/100 ml per 3 hr and 189±27 ng/100 ml per 3 hr at 2 p.m. on the 1st and 2nd days, respectively. There was no significant alteration in plasma levels of cortisol, corticosterone, potassium, or sodium on the 2 days of sodium loading in comparison with the control day. In an additional study, five normal supine subjects received 500 ml saline/hr for 6 hr. As in the 2 day study, plasma aldosterone and renin activity had parallel decrements at 1, 2, 4, and 6 hr after the start of the saline infusion. From these studies, it is concluded that plasma renin activity is the dominant factor controlling plasma aldosterone when sodium-depleted normal subjects are acutely repleted.

A low salt diet increases the free fatty acids, leading to insulin resistance, and contributing to atherosclerosis (Prada, et al., 2000; Mrnka, et al., 2005; Catanozi, et al., 2003; Garg, et al., 2011). -Ray Peat, PhD

J Endocrinol. 2005 Jun;185(3):429-37.
Low salt intake modulates insulin signaling, JNK activity and IRS-1ser307 phosphorylation in rat tissues.
Prada PO, Coelho MS, Zecchin HG, Dolnikoff MS, Gasparetti AL, Furukawa LN, Saad MJ, Heimann JC.
A severe restriction of sodium chloride intake has been associated with insulin resistance and obesity. The molecular mechanisms by which the low salt diet (LS) can induce insulin resistance have not yet been established. The c-jun N-terminal kinase (JNK) activity has been involved in the pathophysiology of obesity and induces insulin resistance by increasing inhibitory IRS-1(ser307) phosphorylation. In this study we have evaluated the regulation of insulin signaling, JNK activation and IRS-1(ser307) phophorylation in liver, muscle and adipose tissue by immunoprecipitation and immunoblotting in rats fed with LS or normal salt diet (NS) during 9 weeks. LS increased body weight, visceral adiposity, blood glucose and plasma insulin levels, induced insulin resistance and did not change blood pressure. In LS rats a decrease in PI3-K/Akt was observed in liver and muscle and an increase in this pathway was seen in adipose tissue. JNK activity and IRS-1(ser307) phosphorylation were higher in insulin-resistant tissues. In summary, the insulin resistance, induced by LS, is tissue-specific and is accompanied by activation of JNK and IRS-1(ser307) phosphorylation. The impairment of the insulin signaling in these tissues, but not in adipose tissue, may lead to increased adiposity and insulin resistance in LS rats.

Physiol Res. 2000;49(2):197-205.
Low-salt diet alters the phospholipid composition of rat colonocytes.
Mrnka L, Nováková O, Novák F, Tvrzická E, Pácha J.
he effect of low-salt diet on phospholipid composition and remodeling was examined in rat colon which represents a mineralocorticoid target tissue. To elucidate this question, male Wistar rats were fed a low-salt diet and drank distilled water (LS, low-salt group) or saline instead of water (HS, high-salt group) for 12 days before the phospholipid concentration and fatty acid composition of isolated colonocytes were examined. The dietary regimens significantly influenced the plasma concentration of aldosterone which was high in LS group and almost zero in HS group. Plasma concentration of corticosterone was unchanged. When expressed in terms of cellular protein content, a significantly higher concentration of phospholipids was found in LS group, with the exception of sphingomyelin (SM) and phosphatidylserine (PS). Phosphatidylcholine (PC) and phosphatidylethanolamine (PE) accounted for more than 70% of total phospholipids in both groups. A comparison of phospholipid distribution in LS and HS groups demonstrated a higher percentage of PE and a small, but significant, decrease of PC and SM in LS group. The percentage of phosphatidylinositol (PI), PS and cardiolipin (CL) were not affected by mineralocorticoid treatment. With respect to the major phospholipids (PE, PC), a higher level of n-6 polyunsaturated fatty acids (PUFA) and lower levels of monounsaturated fatty acids were detected in PC of LS group. The increase of PUFA predominantly reflected an increase in arachidonic acid by 53%. In comparison to the HS group, oleic acid content was decreased in PC and PE isolated from colonocytes of the LS group. Our data indicate that alterations in phospholipid concentration and metabolism can be detected in rats with secondary hyperaldosteronism. The changes in phospholipid concentration and their fatty acid composition during fully developed effect of low dietary Na+ intake may reflect a physiologically important phenomenon with long-term consequences for membrane structure and function.

J Lipid Res. 2003 Apr;44(4):727-32. Epub 2003 Jan 16.
Dietary sodium chloride restriction enhances aortic wall lipid storage and raises plasma lipid concentration in LDL receptor knockout mice.
Catanozi S, Rocha JC, Passarelli M, Guzzo ML, Alves C, Furukawa LN, Nunes VS, Nakandakare ER, Heimann JC, Quintão EC.
This study aimed at measuring the influence of a low salt diet on the development of experimental atherosclerosis in moderately hyperlipidemic mice. Experiments were carried out on LDL receptor (LDLR) knockout (KO) mice, or apolipoprotein E (apoE) KO mice on a low sodium chloride diet (LSD) as compared with a normal salt diet (NSD). On LSD, the rise of the plasma concentrations of TG and nonesterified fatty acid (NEFA) was, respectively, 19% and 34% in LDLR KO mice, and 21% and 35% in apoE KO mice, and that of plasma cholesterol was limited to the LDLR KO group alone (15%). Probably due to the apoE KO severe hypercholesterolemia, the arterial inner-wall fat storage was not influenced by the diet salt content and was far more abundant in the apoE KO than in the LDLR KO mice. However, in the less severe hypercholesterolemia of the LDLR KO mice, lipid deposits on the LSD were greater than on the NSD. Arterial fat storage correlated with NEFA concentrations in the LDLR KO mice alone (n = 14, P = 0.0065). Thus, dietary sodium chloride restriction enhances aortic wall lipid storage in moderately hyperlipidemic mice.

Metabolism. 2011 Jul;60(7):965-8. Epub 2010 Oct 30.
Low-salt diet increases insulin resistance in healthy subjects.
Garg R, Williams GH, Hurwitz S, Brown NJ, Hopkins PN, Adler GK. Low-salt (LS) diet activates the renin-angiotensin-aldosterone and sympathetic nervous systems, both of which can increase insulin resistance (IR). We investigated the hypothesis that LS diet is associated with an increase in IR in healthy subjects. Healthy individuals were studied after 7 days of LS diet (urine sodium <20 mmol/d) and 7 days of high-salt (HS) diet (urine sodium >150 mmol/d) in a random order. Insulin resistance was measured after each diet and compared statistically, unadjusted and adjusted for important covariates. One hundred fifty-two healthy men and women, aged 39.1 ± 12.5 years (range, 18-65) and with body mass index of 25.3 ± 4.0 kg/m(2), were included in this study. Mean (SD) homeostasis model assessment index was significantly higher on LS compared with HS diet (2.8 ± 1.6 vs 2.4 ± 1.7, P < .01). Serum aldosterone (21.0 ± 14.3 vs 3.4 ± 1.5 ng/dL, P < .001), 24-hour urine aldosterone (63.0 ± 34.0 vs 9.5 ± 6.5 μg/d, P < .001), and 24-hour urine norepinephrine excretion (78.0 ± 36.7 vs 67.9 ± 39.8 μg/d, P < .05) were higher on LS diet compared with HS diet. Low-salt diet was significantly associated with higher homeostasis model assessment index independent of age, sex, blood pressure, body mass index, serum sodium and potassium, serum angiotensin II, plasma renin activity, serum and urine aldosterone, and urine epinephrine and norepinephrine. Low-salt diet is associated with an increase in IR. The impact of our findings on the pathogenesis of diabetes and cardiovascular disease needs further investigation.

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Stress and Aging: The Glucocorticoid Cascade Hypothesis

Also see:
The Streaming Organism
Stress, Portrait of a Killer – Full Documentary (2008)
Belly Fat, Cortisol, and Stress

Sci. Aging Knowl. Environ., 25 September 2002
Vol. 2002, Issue 38, p. cp21
The Neuroendocrinology of Stress and Aging: The Glucocorticoid Cascade Hypothesis
Robert M. Sapolsky, Lewis C. Krey, and Bruce S. McEwen
Abstract: Over the past 5 yr, we have examined some of the sharpest edges of the pathology of aging. We have studied the capacity of aged organisms to respond appropriately to stress and the capacity of stress to cumulatively damage aging tissue. The idea of a relationship between stress and aging has permeated the gerontology literature in two forms. First, senescence has been thought of as a time of decreased adaptiveness to stress. This idea has been supported frequently, as many aged physiological systems function normally under basal conditions, yet do not adequately respond to a challenge. For example, aged and young humans have similar basal body temperatures, but the former are relatively impaired in thermoregulatory capacities when heat- or cold-challenged. A second theme in gerontology concerning stress is that chronic stress can accelerate the aging process. Selye and Tuchweber for example, postulated a finite “adaptational energy” in an organism, with prolonged stress prematurely depleting such reserves, thus accelerating the onset of senescence. This idea was derivative of earlier idea that the “rate of living” could be a pacemaker of aging. Experimentally, varied approaches have supported the notion that at least some biomarkers of age can be accelerated by stress.

The above hypotheses led us to examine the adrenocortical axis, the endocrine axis which is among the most central to the stress response. Our findings support both of these concepts. We find that the aged male rat is impaired in terminating the secretion of adrenocortical stress hormones, glucocorticoids, at the end of stress. This hormonal excess may be due to degenerative changes in a region of the brain which normally inhibits glucocorticoid release; the degeneration, in turn, is caused by cumulative exposure to glucocorticoids. Together, these effects form a feed-forward cascade with potentially serious pathophysiological consequences in the aged subject.

In the adrenal glands, renewing cells stream from the capsule on the surface of the gland toward the center of the gland. The first cells to be produced in a regenerating gland are those that produce aldosterone, the next in the stream are the cortisol producing cells, and the last to be formed are the cells that produce the sex hormones, the androgens, including DHEA, and progesterone. In aging, after the age of thirty, the renewal slows, but the dissolution of the sex hormone zone continues, so the proportion shifts, increasing the ration of aldosterone and cortisol producing cells to the layer that produces the protective androgens and progesterone (Parker, et al., 1997). -Ray Peat, PhD

J Clin Endocrinol Metab. 1997 Nov;82(11):3898-901.
Aging alters zonation in the adrenal cortex of men.
Parker CR Jr, Mixon RL, Brissie RM, Grizzle WE.
Whereas aging has been shown to be associated with striking reductions in circulating levels of adrenal androgens in humans, the alteration in adrenal function that occurs in aging has not been identified. We sought to determine if there are changes in the zonation of the adrenal in aging men by performing histomorphologic analyses of adrenal specimens that had been obtained at autopsy following sudden death due to trauma. We evaluated adrenals from 21 young men (20-29 yrs) and 12 older men (54-90 yrs); inclusion criteria required the presence of medullary tissue in the specimen and fixation within the first 24 hrs postmortem. Sections stained with H/E were examined microscopically and areas of the cortex that included adjacent medullary tissue were chosen for quantitative evaluation by use of a computerized image analysis system. The average width (arbitrary units, pixels) of the zona reticularis and that of the combined zonae fasciculata/glomerulosa were determined from sections stained for reticulum fibers. The zona reticularis represented 37.1 +/- 1.9% of the total cortical width in the young men, which was significantly greater than that of the older men (27.1 +/- 3.3%, P = 0.0082). The zona fasciculata/glomerulosa to zona reticularis ratio in the young men (1.84 +/- 0.15) was significantly less that that of the older men (3.29 +/- 0.47, P = 0.0011). There was no significant difference in the total width of the cortex in young compared to older men. These data suggest that aging results in alterations within the cortex of the adrenals in men such that there is a reduction in the size of the zona reticularis and a relative increase in the outer cortical zones. A reduced mass of the zona reticularis could be responsible for the diminished production of dehydroepiandrosterone and dehydroepiandrosterone sulfate that occurs during aging.

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Estrogen, Glutamate, & Free Fatty Acids

Also see:
Menstrual Cycle Related Epilepsy (Catamenial Epilepsy)
Phospholipases, PUFA, and Inflammation
Brain Swelling Induced by Polyunsaturated Fats (PUFA)
Women, Estrogen, and Circulating DHA
Arachidonic Acid’s Role in Stress and Shock
Estrogen’s Role in Seizures
The Brain – Estrogen’s Harm and Progesterone’s Protection
PUFA Kill Thymocytes
PUFA Breakdown Products Depress Mitochondrial Respiration
PUFA, Fish Oil, and Alzheimers
Fish Oil Toxicity
Benefits of Aspirin

“Estrogen increases the activity of the excitatory transmitter glutamate (Weiland, 1992), and glutamate increases the release of free fatty acids (Kolko, et al., 1996). DHA (more strongly even the arachidonic acid) inhibits the uptake of the excitotoxic amino acid aspartate, and in some situations glutamate, prolonging their actions.” -Ray Peat, PhD

“Albumin, besides maintaining blood volume and preventing edema, serves to protect respiration, by binding free fatty acids. Estrogen blocks the liver’s ability to produce albumin, and increases the level of circulating free fatty acids. Free fatty acids cause brain edema. This is probably another aspect of estrogen’s contribution to seizure susceptibility.” -Ray Peat, PhD

Endocrinology. 1992 Dec;131(6):2697-702.
Glutamic acid decarboxylase messenger ribonucleic acid is regulated by estradiol and progesterone in the hippocampus.
Weiland NG.
Ovarian steroids modulate learning, memory, and epileptic seizure activity, functions that are mediated in part by the hippocampus. Normal function depends on precise interactions between the inhibitory gamma-aminobutyric acid (GABA)ergic and excitatory glutamatergic neurons of the hippocampus. To determine whether estradiol and progesterone interact with GABAergic neurons, the levels of mRNA for glutamic acid decarboxylase (GAD), the rate-limiting enzyme for GABA synthesis, were measured by in situ hybridization histochemistry with 35S-labeled riboprobes complimentary to the feline GAD cDNA. The levels of mRNA for GAD were analyzed in selected region of the dorsal hippocampus and medial basal hypothalamus in ovariectomized, ovariectomized estradiol-treated, and ovariectomized estradiol- and progesterone-treated rats. In estradiol-treated rats, GAD mRNA levels increased in GABAergic neurons associated with the CA1 pyramidal cell layer, but not in the stratum oriens of CA1 or any other region of the hippocampus. Estradiol plus progesterone treatment reversed the estradiol-induced increase in GAD mRNA in CA1 and induced a small decrease in the hilus. No effect of estradiol or progesterone was observed in the dorsomedial, ventromedial, or arcuate nuclei of the hypothalamus. Estradiol or progesterone may alter cognitive performance and seizure activity by increasing or decreasing, respectively, the activity of GABAergic neurons in the hippocampus.

J Biol Chem. 1996 Dec 20;271(51):32722-8.
Synergy by secretory phospholipase A2 and glutamate on inducing cell death and sustained arachidonic acid metabolic changes in primary cortical neuronal cultures.
Kolko M, DeCoster MA, de Turco EB, Bazan NG.
Secretory and cytosolic phospholipases A2 (sPLA2 and cPLA2) may contribute to the release of arachidonic acid and other bioactive lipids, which are modulators of synaptic function. In primary cortical neuron cultures, neurotoxic cell death and [3H]arachidonate metabolism was studied after adding glutamate and sPLA2 from bee venom. sPLA2, at concentrations eliciting low neurotoxicity (Synergy in neurotoxicity and [3H]arachidonate release was observed when low, nontoxic (10 ng/ml, 0.71 nM), or mildly toxic (25 ng/ml, 1. 78 nM) concentrations of sPLA2 were added together with glutamate (80 microM). A similar synergy was observed with the sPLA2 OS2, from Taipan snake venom. The NMDA receptor antagonist MK-801 blocked glutamate effects and partially inhibited sPLA2 OS2 but not sPLA2 from bee venom-induced arachidonic acid release. Thus, the synergy with glutamate and very low concentrations of exogenously added sPLA2 suggests a potential role for this enzyme in the modulation of glutamatergic synaptic function and of excitotoxicity.

Increased intracellular calcium activates lipolysis (by phospholipases), producing more free fatty acids, as well as excitation and protein breakdown, and in the brain neurodegenerative diseases, calcium excess contributes to clumping synuclein (Wojda, et al., 2008), an important regulator of cytoskeleton proteins. -Ray Peat, PhD

IUBMB Life. 2008 Sep;60(9):575-90.
Calcium ions in neuronal degeneration.
Wojda U, Salinska E, Kuznicki J.
Neuronal Ca(2+) homeostasis and Ca(2+) signaling regulate multiple neuronal functions, including synaptic transmission, plasticity, and cell survival. Therefore disturbances in Ca(2+) homeostasis can affect the well-being of the neuron in different ways and to various degrees. Ca(2+) homeostasis undergoes subtle dysregulation in the physiological ageing. Products of energy metabolism accumulating with age together with oxidative stress gradually impair Ca(2+) homeostasis, making neurons more vulnerable to additional stress which, in turn, can lead to neuronal degeneration. Neurodegenerative diseases related to aging, such as Alzheimer’s disease, Parkinson’s disease, or Huntington’s disease, develop slowly and are characterized by the positive feedback between Ca(2+) dyshomeostasis and the aggregation of disease-related proteins such as amyloid beta, alfa-synuclein, or huntingtin. Ca(2+) dyshomeostasis escalates with time eventually leading to neuronal loss. Ca(2+) dyshomeostasis in these chronic pathologies comprises mitochondrial and endoplasmic reticulum dysfunction, Ca(2+) buffering impairment, glutamate excitotoxicity and alterations in Ca(2+) entry routes into neurons. Similar changes have been described in a group of multifactorial diseases not related to ageing, such as epilepsy, schizophrenia, amyotrophic lateral sclerosis, or glaucoma. Dysregulation of Ca(2+) homeostasis caused by HIV infection or by sudden accidents, such as brain stroke or traumatic brain injury, leads to rapid neuronal death. The differences between the distinct types of Ca(2+) dyshomeostasis underlying neuronal degeneration in various types of pathologies are not clear. Questions that should be addressed concern the sequence of pathogenic events in an affected neuron and the pattern of progressive degeneration in the brain itself. Moreover, elucidation of the selective vulnerability of various types of neurons affected in the diseases described here will require identification of differences in the types of Ca(2+) homeostasis and signaling among these neurons. This information will be required for improved targeting of Ca(2+) homeostasis and signaling components in future therapeutic strategies, since no effective treatment is currently available to prevent neuronal degeneration in any of the pathologies described here.

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Women, Estrogen, and Circulating DHA

Also see:
Fish Oil Toxicity
PUFA, Fish Oil, and Alzheimers
The Randle Cycle
Estrogen, Glutamate, & Free Fatty Acids
Menstrual Cycle Related Epilepsy (Catamenial Epilepsy
Estrogen’s Role in Seizures
Phospholipases, PUFA, and Inflammation

Quotes by Ray Peat, PhD:
“Women are known to have a greater susceptibility than men to lipolysis, with higher levels of free fatty acids in the serum and liver, because of the effects of estrogen and related hormones.

Women on average have more DHA circulating in the serum than men (Giltay, et al., 2004; McNamara, et al., 2008; Childs, et al., 2008). This high unsaturated fatty acid is the first to be released during stress, and biologically, the meaning of estrogen is to mimic stress. Estrogen and polyunsaturated fatty acids have similar actions on cells, increasing their water content and calcium uptake. Long before the Women’s Health Initiative reported in 2002 that the use of estrogen increased the risk of dementia, it was known that the incidence of Alzheimer‘s was 2 to 3 times higher in women than in men.”

“Under the influence of estrogen, or unsaturated fats, brain cells swell, and their shape and interactions are altered. Memory is impaired by an excess of estrogen. Estrogen and unsaturated fat and excess iron kill cells by lipid peroxidation, and this process is promoted by oxygen deficiency. The fetus and the very old have high levels of iron in the cells. Estrogen increases iron uptake. Estrogen treatment produces elevation of free fatty acids in the blood, and lipid peroxidation in tissues. This tends to accelerate the accumulation of lipofuscin, age-pigment. Lactic acid, the production of which is promoted by estrogen, lowers the availability of carbon dioxide, leading to impairment of blood supply to the brain.”

“One of estrogen’s effects is to chronically increase the circulation of free fatty acids, and to favor the long chain polyunsaturated fatty acids, such as EPA and DHA.”

“Some types of dementia, such as Alzheimer’s disease, involve a life-long process of degeneration of the brain, with an inflammatory component, that probably makes them comparable to osteoporosis and muscle-wasting. (In the brain, the microglia, which are similar to macrophages, and the astrocytes, can produce TNF.) The importance of the inflammatory process in Alzheimer’s disease was appreciated when it was noticed that people who used aspirin regularly had a low incidence of that dementia. Aspirin inhibits the formation of TNF, and aspirin has been found to retard bone loss. In the case of osteoporosis (A. Murrillo-Uribe, 1999), as in Alzheimer’s disease, the incidence is two or three times as high in women as in men. In both Alzheimer’s disease and osteoporosis, the estrogen industry is arguing that the problems are caused by a suddenly developing estrogen deficiency, rather than by prolonged exposure to estrogen.”

“Estrogen, which is promoted intensively as prevention or treatment for Alzheimer’s disease was finally shown to contribute to its development.”

“Our innate immune system is perfectly competent for handling our normal stress induced exposures to bacterial endotoxin, but as we accumulate the unstable fats, each exposure to endotoxin creates additional inflammatory stress by liberating stored fats. The brain has a very high concentration of complex fats, and is highly susceptible to the effects of lipid peroxidative stress, which become progressively worse as the unstable fats accumulate during aging.”

“Estrogen causes elevation of free fatty acids, and there are many interactions between the unsaturated fatty acids and estrogen, including their metabolism to prostaglandins, and their peroxidation.”

“Estrogen increases the free fatty acids circulating in the blood, and this shifts metabolism away from the oxidation of glucose to oxidation of fat, and it reduces oxidative metabolism for example by lowering thyroid function (Vandorpe and Kuhn, 1989)”

“In the 1970s, after reading Szent-Gyorgyi’s description of the antagonistic effect of progesterone and estrogen on the heart, I reviewed the studies that showed that progesterone protects against estrogen’s clotting effect. I experimented with progesterone, showing that it increases the muscle tone in the walls of veins, which is very closely related to the effects Szent-Gyorgyi described in the heart. And progesterone opposes estrogen’s ability to increase the amount of free fatty acids circulating in the blood.

“For example, the brain toxic effects of estrogen were usually neglected, and the much higher incidence of Alzheimer’s disease in women was usually interpreted as evidence that the disease is caused by a deficiency of estrogen. The neurotoxic effects of lipid peroxides and prostaglandins were ignored, while fish oil was advocated to prevent and treat dementia. The toxic effects of serotonin and nitric oxide were seldom considered, whle drugs to increase those were advocated to treat Alzheimer’s.”

“Acrolein’s self-stimulating production from DHA is another factor that could account for the predominance of Alzheimer’s disease in women, since, under the influence of estrogen, women accumulate significantly more DHA than men (Giltay, et al., 2004), and similar effects can be seen in animal studies (McNamara, et al., 2008).”

Am J Clin Nutr. 2004 Nov;80(5):1167-74.
Docosahexaenoic acid concentrations are higher in women than in men because of estrogenic effects.
Giltay EJ, Gooren LJ, Toorians AW, Katan MB, Zock PL.
BACKGROUND:
During pregnancy there is a high demand for docosahexaenoic acid (DHA), which is needed for formation of the fetal brain. Women who do not consume marine foods must synthesize DHA from fatty acid precursors in vegetable foods.
OBJECTIVE:
We studied sex differences in DHA status and the role of sex hormones.
DESIGN:
First, DHA status was compared between 72 male and 103 female healthy volunteers who ate the same rigidly controlled diets. Second, the effects of sex hormones were studied in 56 male-to-female transsexual subjects, who were treated with cyproterone acetate alone or randomly assigned to receive oral ethinyl estradiol or transdermal 17beta-estradiol combined with cyproterone acetate, and in 61 female-to-male transsexual subjects, who were treated with testosterone esters or randomly assigned for treatment with the aromatase inhibitor anastrozole or placebo in addition to the testosterone regimen.
RESULTS:
The proportion of DHA was 15 +/- 4% (x +/- SEM; P < 0.0005) higher in the women than in the men. Among the women, those taking oral contraceptives had 10 +/- 4% (P = 0.08) higher DHA concentrations than did those not taking oral contraceptives. Administration of oral ethinyl estradiol, but not transdermal 17beta-estradiol, increased DHA by 42 +/- 8% (P < 0.0005), whereas the antiandrogen cyproterone acetate did not affect DHA. Parenteral testosterone decreased DHA by 22 +/- 4% (P < 0.0005) in female-to-male transsexual subjects. Anastrozole decreased estradiol concentrations significantly and DHA concentrations nonsignificantly (9 +/- 6%; P = 0.09).
CONCLUSION:
Estrogens cause higher DHA concentrations in women than in men, probably by upregulating synthesis of DHA from vegetable precursors.

Psychoneuroendocrinology. 2009 May;34(4):532-9. Epub 2008 Nov 28.
Gender differences in rat erythrocyte and brain docosahexaenoic acid composition: role of ovarian hormones and dietary omega-3 fatty acid composition.
McNamara RK, Able J, Jandacek R, Rider T, Tso P.
The two-fold higher prevalence rate of major depression in females may involve vulnerability to omega-3 fatty acid deficiency secondary to a dysregulation in ovarian hormones. However, the role of ovarian hormones in the regulation of brain omega-3 fatty acid composition has not been directly evaluated. Here we determined erythrocyte and regional brain docosahexaenoic acid (DHA, 22:6n-3) composition in intact male and female rats, and in chronically ovariectomized (OVX) rats with or without cyclic estradiol treatment (2 microg/4d). All groups were maintained on diets with or without the DHA precursor alpha-linolenic acid (ALA, 18:3n-3). We report that both male (-21%) and OVX (-19%) rats on ALA+ diet exhibited significantly lower erythrocyte DHA composition relative to female controls. Females on ALA+ diet exhibited lower DHA composition in the prefrontal cortex (PFC) relative males (-5%). OVX rats on ALA+ diet exhibited significantly lower DHA composition in the hippocampus (-6%), but not in the PFC, hypothalamus, or midbrain. Lower erythrocyte and hippocampus DHA composition in OVX rats was not prevented by estrogen replacement. All groups maintained on ALA- diet exhibited significantly lower erythrocyte and regional brain DHA composition relative to groups on ALA+ diet, and these reductions were greater in males but not in OVX rats. These preclinical data corroborate clinical evidence for gender differences in peripheral DHA composition (female>male), demonstrate gender differences in PFC DHA composition (male>female), and support a link between ovarian hormones and erythrocyte and region-specific brain DHA composition.

Proc Nutr Soc. 2008 Feb;67(1):19-27.
Gender differences in the n-3 fatty acid content of tissues.
Childs CE, Romeu-Nadal M, Burdge GC, Calder PC.
Dietary n-3 PUFA have many beneficial effects on cell and tissue function and on human health. In mammals the n-3 essential fatty acid alpha-linolenic acid (ALNA) can be converted into longer-chain (LC) n-3 PUFA such as EPA and DHA via a series of desaturase and elongase enzymes that are mainly active in the liver. Human studies have identified that males and females appear to differ in their ability to synthesise EPA and DHA from ALNA, with associated differences in circulating concentrations. Based on studies of women using the contraceptive pill or hormone-replacement therapy and of trans-sexual subjects it is suggested that sex hormones play a role in these differences. The rat has been used to investigate gender differences in n-3 PUFA status since this model allows greater dietary control than is possible in human subjects. Like human subjects, female rats have higher plasma DHA concentrations than males. Rats also respond to increased dietary ALNA in a way that is comparable with available human data. The concentrations of LC n-3 PUFA in rat plasma and tissues are positively associated with circulating concentrations of oestradiol and progesterone and negatively associated with circulating concentrations of testosterone. These findings suggest that sex hormones act to modify plasma and tissue n-3 PUFA content, possibly by altering the expression of desaturase and elongase enzymes in the liver, which is currently under investigation.

Am J Clin Nutr. 2006 Dec;84(6):1330-9.
Dietary fish intake and plasma phospholipid n-3 polyunsaturated fatty acid concentrations in men and women in the European Prospective Investigation into Cancer-Norfolk United Kingdom cohort.
Welch AA, Bingham SA, Ive J, Friesen MD, Wareham NJ, Riboli E, Khaw KT.
BACKGROUND:
The n-3 polyunsaturated fatty acids (n-3 PUFAs) docosahexaenoic acid and eicosapentaenoic acid, found in fish and fish-oil supplements and also formed by conversion of alpha-linolenic acid in soy and rapeseed (canola) oils, are thought to have cardioprotective effects.
OBJECTIVE:
Because the relative feasibility and measurement error of dietary methods varies, this study compared fish and fish-oil intakes obtained from 4 dietary methods with plasma n-3 PUFAs in men and women in a general population.
DESIGN:
The study participants were 4949 men and women aged 40-79 y from the European Prospective Investigation into Cancer-Norfolk United Kingdom cohort. Measurements of plasma phospholipid n-3 PUFA concentrations and fish intakes were made with the use of 4 dietary methods (food-frequency questionnaire, health and lifestyle questionnaire, 7-d diary, and first-day recall from the 7-d diary).
RESULTS:
Amounts of fish consumed and relations with plasma phospholipid n-3 PUFAs were not substantially different between the 4 dietary methods. Plasma n-3 PUFA concentrations were significantly higher in women than in men, were 20% higher in fish-oil consumers than in non-fish-oil consumers, and were twice as high in fatty fish consumers as in total fish consumers. Only approximately 25% of the variation in plasma n-3 PUFA was explained by fish and fish-oil consumption.
CONCLUSIONS:
This large study found no substantial differences between dietary methods and observed clear sex differences in plasma n-3 PUFAs. Because variation in n-3 PUFA was only partially determined by fish and fish-oil consumption, this could explain the inconsistent results of observational and intervention studies on coronary artery disease protection.

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Ray Peat, PhD on the Menstrual Cycle

Also see:
Light is Right
Hot flashes, energy, and aging
SOS for PMS
Collection of Ray Peat Quote Blogs by FPS
Progesterone Decreases Aromatase Activity
Quotes: Thyroid, Estrogen, Menstrual Symptoms, PMS, and Infertility
Estrogen Stimulates Insulin, Promotes Weight Gain
Possible Indicators of Excess Estrogen
Progesterone: Essential to Your Well-Being
Plasma Estrogen Does Not Reflect Tissue Concentration of Estrogen
Estrogen and PCOS
Shock Increases Estrogen
Autoimmune Disease and Estrogen Connection
Hormonal profiles in women with breast cancer
PUFA Increases Estrogen
PUFA Inhibit Glucuronidation
PUFA Promote Cancer
Maternal PUFA Intake Increases Breast Cancer Risk in Female Offspring
Vitamin A: Anti-Cancer and Anti-Estrogen
Toxic Plant Estrogens
The Dire Effects of Estrogen Pollution
Alcohol Consumption – Estrogen and Progesterone In Women
Estrogen, Endotoxin, and Alcohol-Induced Liver Injury
Estrogen Levels Increase with Age
Fat Tissue and Aging – Increased Estrogen
Estrogen Related to Loss of Fat Free Mass with Aging
Bisphenol A (BPA), Estrogen, and Diabetes
PUFA, Estrogen, Obesity and Early Onset of Puberty
Estrogen Related to Loss of Fat Free Mass with Aging
Nutrition and Brain Growth in Chick Embryos
The Brain: Estrogen’s Harm and Progesterone’s Protection
Estrogen Increases Serotonin
Estrogen, Serotonin, and Aggression
Women, Estrogen, and Circulating DHA
PUFA, Fish Oil, and Alzheimers
Estrogen, Glutamate, & Free Fatty Acids
Phospholipases, PUFA, and Inflammation
Estrogen’s Role in Seizures
Benefits of Aspirin
Estrogen’s Role in Asthma
Menstrual Cycle Related Epilepsy (Catamenial Epilepsy)

Picture 4

“Women’s monthly cycles, in which a brief estrogen dominance is followed by sustained exposure to progesterone, are probably an important factor in the renewal of the cells of the brain and other organs, as well as those of the reproductive organs. The daily rhythms of hormones and metabolism are known to be involved in the regulation of cell renewal.”

“Cells respond to stimulation by estrogen by producing a variety of molecules, including the “progesterone receptor” protein. When progesterone enters the cell, binding to these proteins, the estrogenic stimulation is halted, by a series of reactions in which the estrogen receptors disintegrate, and in which estrogen is made water soluble by the activation of enzymes that attach sulfate or a sugar acid, causing it leave the cell and move into the bloodstream, and by reactions that prevent its reentry into the cell by inactivating another type of enzyme, and that suppress its de novo formation in the cell, and that oxidize it into a less active form. Progesterone terminates estrogen’s cellular functions with extreme thoroughness.”

“In polycystic ovaries, menopausal symptoms, arthritis, angina pectoris, multiple sclerosis, some kinds of dementia, migraine, and emphysema, the relief achieved with a simple improvement of cellular energy can be rapid and complete.”

“For example, the follicular phase is a time of low steroid production by the ovary, until near the end of the phase, just before ovulation, when estrogen rises. The luteal phase is a time of high estrogen and high progesterone synthesis. Many publications describe the follicular phase as a time of high estrogen, and the luteal phase as a time of low estrogen, roughly the opposite of the actual situation. And an even larger number of studies get the results they want by using a short exposure to estrogen to study something which takes a long time to develop.”

“The normal cyclic function of the ovaries is a model for the potentially creative role of an inflammation-like stress. Every month (in a rhythm influenced by many cues), a productive crisis comes to a focus in the ovary, with the formation of estrogen, prostaglandins, carbon monoxide, and other signal substances, causing rapid changes both locally and systemically, with water, hormones, and nutrients gathering around the ovum (as well as in other parts of the body, such as the feet). Then as the follicle ruptures with the release of an ovum, the excitatory, inflammation-like state is resolved, with a massive increase in the production of antiinflammatoy, antistress substance, progesterone, leading to the suppression of the excitatory substances. These monthly processes are developmental, they are part of the epigenetic development of the organ.

Most, if not all, of the substances involved in ovarian physiology are involved in the disease of stress and degeneration, which progress in proportion to the inability to produce the resolution of inflammation and restoration of the stable condition. The ovary is a major source of estrogen which can produce the excited, activated, inflammatory, and proliferative state in any tissue of the body, though it acts mainly on the uterus, breasts, and pituitary. But the ovary is also, in response, able to produce massive amounts of the protective progesterone, which interrupts the inflammatory effects of estrogen on the various tissues and organs, largely by suppressing the proteins that hold estrogen within cells (especially the “estrogen receptor”), but also by changing the activities of many enzymes away from the estrogen-controlled, inefficient pattern. The developmental actions of the ovary cause continuing epigenetic process in other organs, causing noticeable changes in their structure every month.”

“The estrogen dominance which is needed to start the reproductive cycle, with cell proliferation in the endometrium, breast, and pituitary, is not otherwise useful to the organism, and is controlled and opposed during pregnancy by a constantly rising production of progesterone. The state of estrogen dominance is essentially unstable.”

“Minks that have mated in the winter don’t implant the fertilized ovum in the uterus until the spring equinox, when progesterone rises suddenly, and allows both the uterus and the embryo to begin the process of pregnancy and gestation. Progesterone provides the oxygen needed for the successful implantation of the embryo, while estrogen and serotonin lower the intrauterine oxygen.

Under good conditions, the (premenstrual) luteal phase of the monthly cycle resembles pregnancy, as a period of progesterone dominance, in which the abundance of progesterone causes cells to decrease their estrogen content. The luteal phase is actually the first stage of pregnancy, and if there is implantation of an embryo all of the processes that begin at ovulation progress continuously until childbirth occurs. When there is no implantation, the luteal phase progesterone dominance is terminated, allowing estrogen to enter tissues and producing menstruation. The sudden decrease of progesterone production before menstruation is similar to the decrease of hormone production just before childbirth. The same conditions that produce the premenstrual syndrome, if they aren’t corrected by the placenta’s massive production of progesterone, will produce preeclampsia, toxemia of pregnancy, eclampsia, and postpartum depression. They are also related to the problems that become so common at menopause. Whenever the production of progesterone falls, tissues are susceptible to estrogen.

There are several common causes of a progesterone deficiency. Deficiencies of thyroid, vitamin A, and cholesterol are often responsible for a progesterone deficiency. Inadequate light exposure can cause it. Excess polyunsaturated fats, interfering with gonads and thyroid, can cause it. And excess serotonin can cause it.”

“In the ovary and uterus, the healthy alternation of excitation and quiescence usually continues for many years, and in rodents it often ends in a state of “persistent estrus,” in which the excitatory state can’t be terminated in the usual way, by the production of progesterone. In humans, menopause is analogous, because the excitatory FSH hormone from the pituitary becomes excessive, with the ovary continuing to produce estrogen but failing to produce progesterone, sometimes with the pituitary failing to shift from FSH to LH. In rodents, it’s recognized that persistent estrus is caused by chronically elevated estrogen, but in humans there has been tremendous resistance to the recognition of estrogen’s central role in menopause and senescence. An excess of the basic promoter of inflammation, serotonin, which is closely associated with estrogen’s influence, can have similar effects on the reproductive cycle (Cooper, et al., 1986). The industry has devoted the necessary funding to making the easily manipulated medical culture, and the public, believe the opposite, i.e., that reproductive aging is mainly caused by estrogen deficiency.”

“Stress uses progesterone and can cause menstrual periods to stop. Girls who begin regular exercise (such as dancing) before puberty have later sexual development.”

“Stress, trauma, and shock start an inflammatory process, that can cause progressive damage to the organs, including the liver. Giving progesterone following the injury protects against the increase of TNF, IL-6, and leakage of liver enzymes (Kuebler, et al., 2003). It has similar protective effects in the brain, lungs, and other organs. During the normal menstrual cycle, IL-6 is inversely related to the level of progesterone (Angstwurm, et aI., 1997).”

Dr. Peat’s (Progesterone) Dosage Recommendations
For topical treatment of sun damaged skin, or acne, wrinkles, dark marks, the oil can be applied directly to the affected areas.

Since progesterone has none of the harmful side effects of other hormones (except the alteration of the menstrual cycle if taken at the wrong time of the month), the basic procedure should be to use it in sufficient quantity to make the symptoms disappear, and to time its use so that menstrual cycles are not disrupted. This normally means using it only between ovulation and menstruation unless symptoms are sufficiently serious that a missed period is not important to you.

If a person has an enlarged thyroid gland, progesterone encourages secretion and unloading of the stored colloid and can bring on a temporary hyperthyroid state which is a corrective process. A thyroid supplement, like Thyodine may be used to shrink the goiter before progesterone is given.

Normal amounts of progesterone promote thyroid secretion, while a deficiency, especially when too much estrogen is present, causes the thyroid to enlarge. Some mention euphoria as a “side-effect” of progesterone usage, but euphoria is simply an indication of good physiological state. To avoid unexpected anesthesia, the correct dose should be determined by taking about 10 mg (3-4 drops) at a time, allowing it to spread into the membranes of the mouth, and repeating the dose after 10 minutes until the symptoms are controlled.

An excessive estrogen/progesterone ratio (should be 1 part to 10 parts, estrogen to progesterone), is involved in producing aggravating symptoms such thin, bluish skin. Low thyroid is one cause of excess estrogen, and when high estrogen is combined with low thyroid function, the skin can look relatively bloodless.

Symptoms in cycling women are most common around ovulation and in the premenstrual week, when the estrogen/progesterone ratio is normally highest. At puberty, in the early twenties and in the late thirties and menopause are the ages when the ratio is most often disturbed – and these are also the ages when thyroid disorders are most common.

The individual who suffers from one aspect of progesterone (and/or thyroid) deficiency will tend to develop other problems at varying times. With cyclic depression or migraine headaches at 22, there will possibly be breast disease after, and often there will be problems with pregnancy. These people with a history of severe symptoms are the ones most likely to have problems around menopause. Prenatal exposure to poorly balanced hormones seems to predispose children to later hormone imbalances.

Excess stress (which can block progesterone synthesis and elevate estrogen) may being on symptoms in someone who never had them. As well as darker days without sun, toxins and nutritional deficiencies. A very common cause of an estrogen excess is a dietary protein deficiency because the liver cannot detoxify estrogen when it is under nourished.

With a diet high in protein and vitamin A, progesterone can usually be reduced each month. Using thyroid supplementation will also usually reduce the amount of progesterone needed. Occasionally, a women won’t feel any effect even from 100 mg of progesterone and this may indicate they need to use thyroid supplementation and diet changes to normalize their estrogen, prolactin and cortisol levels.

Progesterone stimulates the ovaries and adrenals to produce progesterone, and it also activates the thyroid. It shouldn’t be necessary to use progesterone indefinitely unless the ovaries have been removed. In slender post-menopausal women, 10 mg per day is usually enough to prevent progesterone deficiency.

Ina 10% solution of progesterone in vitamin E, one drop contains about three milligrams of progesterone. Normally the body produces 10 to 20 milligrams per day. A dose of 3 or 4 drops usually brings the blood levels up to normal range, but this dose can be repeated throughout the day to control symptoms.

For general purposes, it is most economical and effective to take progesterone dissolved in vitamin E orally, for example taking a few drops on the lips and tongue, or rubbing it into the gums. (It is good for the general health of the gums, too.) These membranes are very thin and hence the progesterone can easily and quickly enter the blood. when swallowed, the vitamin E allows it to be absorbed through the walls of the stomach and intestine.

As was mentioned previously, topical application of this oil aids sun damaged skin. For topical treatment of arthritis, osteoporosis, tendinitis, bursitis, or varicose veins, to speed absorption it is best to apply a few drops of olive oil to the area, and then to rub the progesterone -vitamin E solution into and around the affected area.”

PROGESTERONE SUPPLEMENTATION
SYMPTOMATIC: For tendonitis, bursitis, arthritis, sunburn, etc., progesterone in vitamin E can be applied locally after a little olive oil has been put on the skin to make it easier to spread the progesterone solution. For migraines, it has been taken orally just as the symptoms begin.

FOR PMS: The normal pattern of progesterone secretion during the month is for the ovaries to produce a large amount in the 2nd two weeks of the menstrual cycle, (i.e., day 14 through day 28) beginning at ovulation and ending around the beginning of menstruation, and then to produce little for the following two weeks. An average person produces about 30 milligrams daily during the 2nd two weeks. The solution I have used contains approximately 3 or 4 milligrams of progesterone per small drop. Three to four drops, or about 10 to 15 milligrams of progesterone, is often enough to bring the progesterone level up to normal. That amount can be taken days 14 through 28 of the menstrual cycle; this amount may be repeated once or twice during the day as needed to alleviate symptoms. Since an essential mechanism of progesterone’s action involves its opposition to estrogen, smaller amounts are effective when estrogen production is low, and if estrogen is extremely high, even large supplements of progesterone will have no clear effect; in that case, it is essential to regulate estrogen metabolism, by improving the diet, correcting a thyroid deficiency, etc. (Unsaturated fat is antithyroid and synergizes with estrogen.)

PERIMENOPAUSAL: The symptoms and body changes leading up to menopause are associated with decreasing production of progesterone, at a time when estrogen may be at a lifetime high. The cyclic use of progesterone, two weeks on, two weeks off, will often keep the normal menstrual cycle going. Three to our drops, providing ten or twelve milligrams of progesterone, is typical for a day, but some women prefer to repeat that amount. Progesterone is always more effective when the diet contains adequate protein, and when thee isn’t an excessive amount of unsaturated fat in the diet.

POSTMENOPAUSAL: Some women continue the cyclic use of progesterone ater menopause, because the pituitary gland and brain may continue to cycle long after menstruation has stopped, and progesterone is an important regulator of pituitary and brain function. The cycling pituitary affects the adrenal glands and other organs, and progesterone tends to protect against the unopposed actions of prolactin, cortisol, and adrenal androgenic hormones. Progesterone’s effects on the pituitary apparently contribute to its protective effect against osteoporosis, hypertension, hirsutism, etc. But some women prefer to use progesterone without interruption after the menopause, for its protective antistress effects. Slender people usually find that two or three drops are enough, but this amount may be repeated once or twice as needed to relieve symptoms. Adequate protein in the diet and good thyroid function help the body to produce its own progesterone; even if the ovaries have been removed, the adrenal glands and brain continue to produce progesterone.

DOSAGE OF PROGESTERONE
Since progesterone has none of the harmful side effects of other hormones (except for alteration of the menstrual cycle if it is taken at the wrong time of month), the basic procedure should be to use it in sufficient quantity to make the symptoms disappear, and to time its use so that menstrual cycles are not disrupted. This normally means using it only between ovulation and menstruation unless symptoms are sufficiently serious that a missed period is not important. The basic idea of giving enough to stop the symptoms can be refined by some information on a few of the factors that condition the need for progesterone.

An excessive estrogen/progesterone ratio is more generally involved in producing or aggravating symptoms than either a simple excess of estrogen or a deficiency of progesterone, but even this ratio is conditioned by other factors, including age, diet, other steroids, thyroid, and other hormones. The relative estrogen excess seems to act by producing tissue hypoxia (as reported in my dissertation, University of Oregon, 1972), and this is the result of changes induced by estrogen in alveolar diffusion, peripheral vascular changes, and intracellular oxygen wastage.

Hypoxia in turn produces edema (as can be observed in the cornea when it is deprived of oxygen, as by a contact lens) and hypoglycemia (e.g., diminished ATP acts like insulin), because glycolysis must increase greatly for even a small deficiency of oxygen. Elevated blood lactic acid is one sign of tissue hypoxia. Edema, hypoglycemia, and lactic academia can also be produced by other “respiratory” defects, including hypothyroidism, in which the tissue does not use enough oxygen. In hypoxia, the skin will be bluer (in thin places, such as around the eyes), than when low oxygen consumption is the main problem. Low thyroid is one cause of excess estrogen, and when high estrogen is combined with low thyroid, the skin looks relatively bloodless.

Symptoms in cycling women are most common around ovulation and in the premenstrual week, when the estrogen/progesterone ratio is normally highest. At puberty, in the early twenties and in the late thirties and menopause are the ages when the ratio is most often disturbed–and these are also the ages when thyroid disorders are commonest in women.

The individual who suffers from one aspect of the progesterone (and/or thyroid) deficiency will tend to develop other problems at different times. With cyclic depressions or migraine headaches at age 22, there will possibly be breast disease later, and often there will be problems with pregnancy. These people with a history of sever symptoms are the ones most likely to have severe problems around menopause. Prenatal exposure to poorly balanced hormones seems to predispose the child to later hormone problems.

Excess stress (which can block progesterone synthesis and elevate estrogen) may bring on symptoms in someone who never had them. Spending a summer in Alaska, with an unusually long day, may relieve the symptoms of a chronic sufferer. Dark cloudy winters in England or the Pacific Northwest are powerful stressors, and cause lower production of progesterone in women, and testosterone in men. Toxins can produce similar symptoms, as can nutritional deficiencies. A very common cause of an estrogen excess is a dietary protein deficiency–the liver simply cannot detoxify estrogen when it is under-nourished.

With a diet high in protein (e.g., at least 70-100 grams per day, including eggs) and vitamin A (not carotene), I have found that the dose of progesterone can be reduced each month. Using thyroid will usually reduce the amount of progesterone needed. Occasionally, a woman won’t feel any effect even from 100 mg. of progesterone; I think this indicates that they need to use thyroid and diet, to normalize their estrogen, prolactin, and cortisol.

Progesterone stimulates the ovaries and adrenals to produce progesterone, and it also activates the thyroid, so one dose can sometimes have prolonged effects. It shouldn’t be necessary to keep using progesterone indefinitely, unless the ovaries have been removed. In slender post-menopausal women, 10 mg. per day is usually enough to prevent progesterone deficiency symptoms.

In a 10% solution of progesterone in vitamin E, one drop contains about three milligrams of progesterone. Normally, the body produces 10 to 20 milligrams per day. A dose of 3 or 4 drops usually brings the blood levels up to the normal range, but this dose can be repeated several times during the day if it is needed to control symptoms.

For general purposes, it is most economical and effective to take progesterone dissolved in vitamin E orally, for example taking a few drops on the lips and tongue, or rubbing it into the gums. (It is good for the general health of the gums.) These membranes are very thin, and the progesterone quickly enters the blood. When it is swallowed, the vitamin E allows it to be absorbed through the walls of the stomach and intestine, and it can be assimilated along with food, in the chylomicrons, permitting it to circulate in the blood to all of the organs before being processed by the liver. These droplets are smaller than red blood cells, and some physicians seem to forget that red blood cells pass freely through the liver.

For the topical treatment of sun damaged skin, or acne, wrinkles, etc. the oil can be applied directly to the affected area.”

“I have observed the use of progesterone transdermally (applied to the skin in about 400 women suffering from the full range of peri-menstrual symptoms, including migraine, acne, depression, mastalgia (breast pain), edema (water retention and swelling), and lethargy. I found that nearly all the women who apply the lotion themselves, are able to find the appropriate dosage for controlling their symptoms. Occasionally, thyroid therapy, weight reduction: or change in some aspect of lifestyle is necessary for complete relief from symptoms. When some women said the progesterone had no effect, it turned out that they were applying it as sparingly as they would a rare perfume-just touching it to their wrists.

Anesthesia (or drunkenness) from a very large overdose is the only negative side effective reported in the literature. Experimenting with very large doses of natural progesterone on myself, for migraine, I found that there is a temporary antagonism to testosterone, causing small symptoms, including a slight sense of gregariousness, an urge to socialize. However, synthetic progestins do have harmful side effects including breast tumors. Unfortunately, most physicians do not yet know the difference between natural progesterone and synthetic progestins.”

“Estrogen’s name, derived from the gadfly, accurately suggests its role as an excitant, getting things started. Progesterone’s name, relating to pregnancy, is compatible with thinking of it as an agent of calming and fulfillment. But these properties show up in every aspect of physiology, and the special cases of female estrus and pregnancy can be properly understood only in the larger context, in which, for example, progesterone is a brain hormone in both sexes and at all ages, and estrogen is an essential male hormone involved in the sperm cell’s function and male libido.”

“Sometimes progesterone can cause an underactive enlarged thyroid gland to begin secreting, temporarily producing mild hypothyroidism while the gland returns to a normal size. Supplemental progesterone can reduce excessive cortisol production.

Since progesterone helps the thyroid to secrete, and helps the liver to regulate glucose and convert T4 to T3, women who are low in progesterone usually have hypothyroid symptoms (because of insufficient T3), including high cortisol, which promotes the synthesis of estrogen (in several ways, but never from progesterone). Cortisol is made from progesterone, but increasing the supply of progesterone reliably lowers cortisol synthesis, acting on the brain, pituitary, and adrenal glands. Progesterone, by many mechanisms, including its antagonism to cortisol, lowers the amount of estrogen in cells (causing the estrogen-binding proteins to be degraded, inhibiting the enzymes that release estrogen from the sulfates and glucuronides, and activating the enzymes that detoxify estrogen). So I think the symptoms of increased estrogen and cortisol are the result of either extraneous ingredients in the creams, or from using it at the wrong time, for example, too early, triggering premature ovulation. Supplementing a small amount of T3, Cytomel or Cynomel, usually stops symptoms such as breast pain, irritability, and restless energy, in less than an hour.”

“(progesterone) Protection against hypoglycemia is probably the main mechanism. Diabetic mothers often have precocious children, if they aren’t damaged by drugs and irrational diets. Ten to 15 grams per pregnancy, at increasing dosage, seems to be the optimal amount, when there is some sign of excessive estrogen or unexpected toxemia.”

“In good health, an animal’s systems are designed so that certain tissues will be intensely but briefly stimulated by estrogen.”

“It [progest-e] can help with cramps, but it would probably take a lot; I think it’s better to use thyroid (including T3) to solve the basic problem, since it will let you regulate the balance between estrogen and progesterone, while allowing your cells to balance the minerals, retaining the magnesium needed to prevent cramping. Increasing your intake of all the main minerals, calcium, sodium, potassium, and magnesium usually helps in the short term, but the balance isn’t stable if your thyroid is low. Milk, orange juice, coffee (even decaffeinated coffee is a good source of magnesium), and well salted foods, support thyroid functions. Aspirin helps with thyroid function and mineral balance, even helps to prevent excessive estrogen production.”

“More recently, it has been discovered that progesterone inhibits the expression of the enzyme nitric oxide synthase while estrogen stimulates its expression. At the time of ovulation, when estrogen is high, a woman breathes out 50% more nitric oxide (“NO”) than men do, but at other times, under the influence of increased progesterone and thyroid, and reduced estrogen, women exhale much less NO than men do. (Nitric oxide is a free radical, and it decomposes’ into other toxic compounds, including the free radical peroxyrutnle} which damages cells, including the blood vessels, brain, and heart. Carbon dioxide tends to inhibit the production of peioxynitrile.)”

“When estrogen is given to an animal, it almost instantly causes capillaries to become leaky, allowing water to move out of the blood stream, and at the same time, estrogen causes cells to take up water. Both of these processes are the same as the early effects of oxygen deprivation. In the normal reproductive cycle, the surge of estrogen lasts only a few hours, and normal permeability is quickly restored by increasing progesterone. During those intermittent short exposures to estrogen, there isn’t a massive leakage of serum proteins into the tissues. During the time of estrogenic influence, all kinds of cells are influenced, with the excitatory equilibrium of nerve cells, glandular cells, and immune system cells being shifted, lowering the threshold of excitation, or prolonging the excited state.”

“Our body controls estrogen’s effects by decreasing excitation while increasing efficient energy production.”

“Estrogen’s survival value exists only in the context of a whole organism with multiple ways for limiting estrogen’s destabilizing actions. Estrogen’s harmful effects occur when our systems for opposing it fail.”

“Women and other mammals that are deficient in progesterone, and/or that have an excess of estrogen, have a higher than average incidence of cancer. Animal experiments have shown that administering progesterone could prevent cancer. Cells in the most cancer-susceptible tissues proliferate in proportion to the ratio of estrogen to progesterone. When the estrogen dominance persists for a long time without interruption, there are progressive distortions in the structure of the responsive organs–the uterus, breast, pituitary, lung, liver, kidney, brain, and other organs–and those structural distortions tend to progress gradually from fibroses to cancer.

As a result of the early studies in both humans and animals, progesterone was used by many physicians to treat the types of cancer that were clearly caused by estrogen, especially uterine, breast, and kidney cancers. But by the 1950s, the drug industry had created the myth that their patented synthetic analogs of progesterone were medically more effective than progesterone itself, and the result has been that medroxyprogesterone acetate and other synthetics have been widely used to treat women’s cancers, including breast cancer.

Unfortunately, those synthetic compounds have a variety of functions unlike progesterone, including some estrogenic and/or androgenic and/or glucocorticoid and/or antiprogesterone functions, besides other special, idiosyncratic side effects. The rationale for their use was that they were “like progesterone, only better.” The unpleasant and unwanted truth is that, as a group, they are seriously carcinogenic, besides being toxic in a variety of other ways. Thousands of researchers have drawn conclusions about the effects of progesterone on the basis of their experiments with a synthetic progestin.”

“Each type of cell and organ is subject to injury, and in some cases these injuries are cumulative. In the healthy liver, which stores glycogen, toxins can be inactivated, for example by combining with glucuronic acid, derived from the stored glucose. With injury, such as alcoholism combined with a diet containing polyunsaturated fats, the liver’s detoxifying ability is reduced. Even at an early stage, before there is a significant amount of fibrosis, the reduced activity of the liver causes estrogen to accumulate in the body. Estrogen’s valuable actions are, in health, exerted briefly, and then the synthesis of estrogen is stopped, and its excretion reduces its activity, but when the liver’s function is impaired, estrogen’s activity continues, causing further deterioration of liver function, as well as injury of nerves such as Desjardins described, and the systemic energy shifts and stress activations mentioned above.

Besides lowering the liver’s detoxifying ability, stress, hypoglycemia, malnutrition, hypothyroidism, and aging can cause estrogen to be synthesized inappropriately and continuously. With aging, estrogen begins to be produced throughout the body–in fat, muscles, skin, bones, brain, liver, breast, uterus, etc. Polyunsaturated fats are a major factor in the induction and activation of the aromatase enzyme, which synthesizes estrogen.

Increased synthesis of estrogen, with aromatase, and decreased excretion of it, by the liver and kidneys, are only two of the processes that affect the influence of estrogen during aging. Cellular stress (chemical, mechanical, hypoxemic, hypoglycemic [Clere, et al., 2012; Aguirre, et al., 2007, Zaman, et al., 2006, Saxon, et al., 2007; Tamir, et al., 2002; Briski, et al., 2010]) increases estrogen receptors (which activate CRH and the stress response). The presence of estrogen receptors means that estrogen will be bound inside cells, where it acts to modify those cells. Before estrogen can reach the liver to be inactivated, it must be released from cells. Ordinarily, the cyclic production of progesterone has that function, by destroying the estrogen-binding proteins. Progesterone also inhibits the aromatase which synthesizes estrogen, and shifts the activities of other enzymes, including sulfatases and dehydrogenates, in a comprehensive process of eliminating the presence and activity of estrogen. At menopause, when the ovary fails to produce the cyclic progesterone, all of these processes of estrogen inactivation fail. In the absence of progesterone, cortisol becomes more active, increasing aromatase activity, which now becomes chronic and progressive. The decrease of progesterone causes many other changes, including the increased conversion of polyunsaturated fatty acids to prostaglandins, and the formation of nitric oxide, all of which contribute to the tendency to flush.”

“During the reproductive years, women’s cyclic production of large amounts of progesterone probably retards their aging enough to account for their greater longevity. Childbearing also has a residual antiestrogenic effect and is associated with increased longevity.”

“Estrogen blocks the release of hormone from the thyroid gland, and progesterone facilitates the release. Estrogen excess or progesterone deficiency tends to cause enlargement of the thyroid gland, in association with a hypothyroid state. Estrogen can activate the adrenals to produce cortisol, leading to various harmful effects, including brain aging and bone loss. Progesterone stimulates the adrenals and the ovaries to produce more progesterone, but since progesterone protects against the catabolic effects of cortisol, its effects are the opposite of estrogen’s. Progesterone has antiinflammatory and protective effects, similar to cortisol, but it doesn’t have the harmful effects. In hypothyroidism, there is a tendency to have too much estrogen and cortisol, and too little progesterone.

The blood tests can be useful to demonstrate to physicians what the problem is, but I don’t think they are necessary. There is evidence that having 50 or 100 times as much progesterone as estrogen is desirable, but I don’t advocate “progesterone replacement therapy” in the way it’s often understood. Progesterone can instantly activate the thyroid and the ovaries, so it shouldn’t be necessary to keep using it month after month. If progesterone is used consistently, it can postpone menopause for many years.

Cholesterol is converted to pregnenolone and progesterone by the ovaries, the adrenals, and the brain, if there is enough thyroid hormone and vitamin A, and if there are no interfering factors, such as too much carotene or unsaturated fatty acids. Progesterone deficiency is an indicator that something is wrong, and using a supplement of progesterone without investigating the nature of the problem isn’t a good approach. The normal time to use a progesterone supplement is during the “latter half” of the cycle, the two weeks from ovulation until menstruation. If it is being used to treat epilepsy, cancer, emphysema, migraine or arthritis, or something else so serious that menstrual regularity isn’t a concern, then it can be used at any time. If progesterone is used consistently, it can postpone menopause for many years.”

“Most people are aware of some of the variations of bleeding and clotting that occur commonly. Bleeding gums, nose-bleeds, menstruation and its variations, and the spontaneous bruising (especially on the thighs) that many women have premenstrually, are familiar events that don’t seem to mean much to the medical world. Sometimes nose-bleeds are clearly stress-related, but the usual “explanation” for that association is that high blood pressure simply blows out weak blood vessels. Bleeding gums are sometimes stress related, but high blood pressure is seldom invoked to explain that problem.”

“Menstrual bleeding is a good place to start the investigation of bleeding problems, since its relatively harmless abnormalities are physiologically related to some very serious health problems, such as pregnancy bleeding, abruptio placentae, and eclampsia. Women who die from eclampsia have been found to have massively clotted blood vessels in their brains, but the variety of names for the pregnancy disorders have prevented most people from thinking of pregnancy as a time when there is a high risk of the “thrombohemorrhagic disorders,” a time when the clotting system is under stress. (For about fifteen years after Selye coined the term, only he and some Russians were publishing research on it, and Americans still don’t show much interest in the subject.)

Women with a chronic menstrual problem resulting from progesterone deficiency often continue to bleed each month even when they are pregnant, and these women tend to develop toxemia, and to have a high incidence of pregnancy complications, and to deliver premature, poorly developed babies.

In 1933 James Shute was recommending the use of vitamin E for preventing the clotting problems associated with pregnancy, that often lead to miscarriage. He based his work on animal studies, that led to vitamin E’s being known as the “fertility vitamin.” Later, his sons Wilfred and Evan reported that vitamin E could prevent heart attacks, birth defects, complications of diabetes, phlebitis, hypertension, and some neurological problems.”

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Fish Oils Increase Intestinal Permeability

Also see:
PUFA and Liver Toxicity; Protection by Saturated Fats
Endotoxin: Poisoning from the Inside Out
Ray Peat, PhD on the Benefits of the Raw Carrot
Protection from Endotoxin
Endotoxin-lipoprotein Hypothesis
Protective Bamboo Shoots
The effect of raw carrot on serum lipids and colon function
How does estrogen enhance endotoxin toxicity? Let me count the ways.
Bowel Toxins Accelerate Aging
Liver saving saturated fats

“Besides causing a general slowing of metabolism, aging and toxic PUFA have specific actions on the detoxifying system. The enzymes that help to detoxify PUFA and estrogen and serotonin are inhibited by both PUFA and estrogen. All systems, including blood vessels and the intestine, are made leaky by estrogen and the PUFA and their products.” -Ray Peat, PhD

“In the bowel, the capillary malfunction increases the absorption of endotoxin, which intensifies the systemic energy problem. (Polyunsaturated oils, especially fish oil, damage the bowel capillaries, allowing more endotoxin to be absorbed.)” -Ray Peat, PhD

J Nutr. 2011 Sep;141(9):1635-42. Epub 2011 Jul 20.
Ingestion of (n-3) fatty acids augments basal and platelet activating factor-induced permeability to dextran in the rat mesenteric vascular bed.
Dombrowsky H, Lautenschläger I, Zehethofer N, Lindner B, Schultz H, Uhlig S, Frerichs I, Weiler N.
Loss of intestinal barrier function and subsequent edema formation remains a serious clinical problem leading to hypoperfusion, anastomotic leakage, bacterial translocation, and inflammatory mediator liberation. The inflammatory mediator platelet activating factor (PAF) promotes eicosanoid-mediated edema formation and vasoconstriction. Fish oil-derived (n-3) fatty acids (FA) favor the production of less injurious eicosanoids but may also increase intestinal paracellular permeability. We hypothesized that dietary (n-3) FA would ameliorate PAF-induced vasoconstriction and enhance vascular leakage of dextran tracers. Rats were fed either an (n-3) FA-rich diet (EPA-rich diet; 4.0 g/kg EPA, 2.8 g/kg DHA) or a control diet (CON diet; 0.0 g/kg EPA and DHA) for 3 wk. Subsequently, isolated and perfused small intestines were stimulated with PAF and arterial pressure and the translocation of fluid and macromolecules from the vasculature to lumen and lymphatics were analyzed. In intestines of rats fed the EPA-rich diet, intestinal phospholipids contained up to 470% more EPA and DHA at the expense of arachidonic acid (AA). The PAF-induced increase in arterial pressure was not affected by the EPA-rich diet. However, PAF-induced fluid loss from the vascular perfusate was higher in intestines of rats fed the EPA-rich diet. This was accompanied by a greater basal loss of dextran from the vascular perfusate and a higher PAF-induced transfer of dextran from the vasculature to the lumen (P = 0.058) and lymphatics. Our data suggest that augmented intestinal barrier permeability to fluid and macromolecules is a possible side effect of (n-3) FA-rich diet supplementation.

DHA increases the leakiness of the bowel, allowing more endotoxin to enter the circulation (RoigPerez, et al., 2004). -Ray Peat, PhD

J Lipid Res. 2004 Aug;45(8):1418-28. Epub 2004 Jun 1.
Lipid peroxidation induced by DHA enrichment modifies paracellular permeability in Caco-2 cells: protective role of taurine.
Roig-Pérez S, Guardiola F, Moretó M, Ferrer R.
Dietary enrichment with docosahexaenoic acid (DHA) has numerous beneficial effects on health. However, the intake of high doses of polyunsaturated fatty acids can promote lipid peroxidation and the subsequent propagation of oxygen radicals. The purpose of this study was to evaluate the effect of DHA on lipid peroxidation and tight junction structure and permeability in Caco-2 cell cultures. Moreover, the effects of taurine, a functional ingredient with antioxidant properties, were also tested. Differentiated Caco-2 cell monolayers were maintained in DHA-supplemented conditions with or without added taurine. Incubation with 100 microM DHA increased lipid peroxidation and paracellular permeability, in parallel with a redistribution of the tight junction proteins occludin and ZO-1. Taurine partially prevented all of these effects. The participation of reactive oxygen and nitrogen species in increased paracellular permeability was also examined using various agents that modify the formation of superoxide radical, hydrogen peroxide, nitric oxide, and peroxynitrite. We conclude that hydrogen peroxide and peroxynitrite may be involved in the DHA-induced increase in paracellular permeability and that the protective role of taurine may be in part related to its capacity to counteract the effects of hydrogen peroxide.

Implications:
J Clin Gastroenterol. 2002 Apr;34(4):385-96.
Intestinal permeation and gastrointestinal disease.
DeMeo MT, Mutlu EA, Keshavarzian A, Tobin MC.
The gastrointestinal tract constitutes one of the largest sites of exposure to the outside environment. The function of the gastrointestinal tract in monitoring and sealing the host interior from intruders is called the gut barrier. A variety of specific and nonspecific mechanisms are in operation to establish the host barrier; these include luminal mechanisms and digestive enzymes, the epithelial cells together with tight junctions in between them, and the gut immune system. Disruptions in the gut barrier follow injury from various causes including nonsteroidal anti-inflammatory drugs and oxidant stress, and involve mechanisms such as adenosine triphosphate depletion and damage to epithelial cell cytoskeletons that regulate tight junctions. Ample evidence links gut barrier dysfunction to multiorgan system failure in sepsis and immune dysregulation. Additionally, contribution of gut barrier dysfunction to gastrointestinal disease is an evolving concept and is the focus of this review. An overview of the evidence for the role of gut barrier dysfunction in disorders such as Crohn’s disease, celiac disease, food allergy, acute pancreatitis, non-alcoholic fatty liver disease, and alcoholic liver disease is provided, together with critical insight into the implications of this evidence as a primary disease mechanism.

“Essential” polyunsaturated fat linoleic acid also increase intestinal permeability. Saturated fats had no such effect.
Alcohol Clin Exp Res. 2012 May;36(5):835-46. doi: 10.1111/j.1530-0277.2011.01673.x. Epub 2011 Dec 7.
The type of dietary fat modulates intestinal tight junction integrity, gut permeability, and hepatic toll-like receptor expression in a mouse model of alcoholic liver disease.
Kirpich IA, Feng W, Wang Y, Liu Y, Barker DF, Barve SS, McClain CJ.
BACKGROUND:
Interactions between the gut, immune system, and the liver, as well as the type of fat in the diet, are critical components of alcoholic liver disease (ALD). The goal of the present study was to determine the effects of saturated fat (SF) and unsaturated fat (USF) on ethanol (EtOH)-induced gut-liver interactions in a mouse model of ALD.
METHODS:
C57BL/6N mice were fed Lieber-DeCarli liquid diets containing EtOH and enriched in USF (corn oil) or SF (medium chain triglycerides:beef tallow). Control mice were pair-fed on an isocaloric basis. Liver injury and steatosis, blood endotoxin levels, intestinal permeability, and tight junction (TJ) integrity, as well as hepatic Toll-like receptor (TLR) gene expression, were evaluated.
RESULTS:
After 8 weeks of EtOH feeding, liver injury and steatosis were observed in USF + EtOH group compared with control and SF + EtOH. Significantly increased intestinal permeability in conjunction with elevated blood endotoxin levels were observed in the ileal segments of the mice fed USF + EtOH. USF diet alone resulted in down-regulation of intestinal TJ protein mRNA expression compared with SF. Importantly, alcohol further suppressed TJ proteins in USF + EtOH, but did not affect intestinal TJ in SF + EtOH group. The type of fat in the diet alone did not affect hepatic TLR expression. Compared with control animals, hepatic TLR (TLR 1, 2, 3, 4, 7, 8, 9) mRNA expression was significantly (p < 0.05) increased in USF + EtOH, but not in SF + EtOH group. Notably, TLR5 was the only up-regulated TLR in both SF + EtOH and USF + EtOH groups. CONCLUSIONS: Dietary fat is an important cofactor in alcohol-associated liver injury. We demonstrate that USF (corn oil/linoleic acid) by itself results in dysregulation of intestinal TJ integrity leading to increased gut permeability, and alcohol further exacerbates these alterations. We postulate that elevated blood endotoxin levels in response to USF and alcohol in conjunction with up-regulation of hepatic TLRs combine to cause hepatic injury in ALD.

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PUFA Kill Thymocytes

“Polyunsaturated fatty acids, derived from foods, have a special role in the immune system, intensifying the effects of stress (cholesterol newsletter, September, 2005) in killing lymphocytes, and blocking the proliferative response of thymic cells (Rotondo, et aI., 1994). They tend to shift immune functions from cellular immunity to humoral (antibody) immunity, and this pattern predisposes to autoimmunity. They are probably directly toxic to the liver (Ritskes-Hoitinga, 1998). DHA increases the leakiness of the bowel, allowing more endotoxin to enter the circulation (RoigPerez, et al., 2004).” -Ray Peat, PhD

Biochim Biophys Acta. 1994 Sep 8;1223(2):185-94.
Inhibition of cytokine-stimulated thymic lymphocyte proliferation by fatty acids: the role of eicosanoids.
Rotondo D, Earl CR, Laing KJ, Kaimakamis D.
The effect of individual fatty acids on the proliferation of thymic lymphocytes in response to interleukin-1 (IL-1) was investigated. Proliferation was estimated by measuring [3H]thymidine incorporation into the acid insoluble fraction of the thymocytes. A concentration-dependent inhibition (in the range 1-100 microM) in the IL-1-stimulated proliferation was observed with the C20 fatty acids dihomo-gamma-linolenic acid (DGLA), arachidonic acid and eicosapentaenoic acid (EPA). A less pronounced concentration-dependent inhibitory response was observed with the C18 fatty acids linoleic acid, alpha-linolenic acid and gamma-linolenic acid. Palmitic acid and oleic did not have any effect on either basal or IL-1-stimulated proliferation at concentrations up to 100 microM. The potencies of each fatty acid for this effect at a concentration of 100 microM were: arachidonic acid > EPA > or = DGLA > linoleic acid. DGLA, arachidonic acid and EPA also attenuated IL-2-stimulated proliferation. The inhibitory action of these fatty acids was not mediated by conversion to prostaglandins or other eicosanoids as the cyclooxygenase inhibitor, ketoprofen and NDGA did not alter their action. Incubation of thymocytes with radiolabelled DGLA and EPA followed by reverse-phase HPLC analysis revealed that DGLA is predominantly converted to a more polar metabolite which is not PGE1 whereas EPA does not appear to be converted to any other detectable metabolite. The data indicate that the inhibitory actions of fatty acids on cell proliferation do not occur as a result of conversion to other metabolites but may be direct effects. The inhibition of cytokine-stimulated lymphocyte proliferation by unsaturated fatty acids would imply that they may attenuate cell-mediated immune reactions.

Thymocytes are much more easily killed by stress than nerve cells, and they are easy to study. The PUFA kill them by increasing their intracellular calcium. The toxicity of DHA is greater than that of EPA, whose toxicity is greater than the alpha-linoleic acid, and linoleic acid was the most potent (Prasad, et al., 2010)…The process by which excitotoxicity kills cells is probably a foreshadowed version of the aging process. -Ray Peat, PhD

J Cell Physiol. 2010 Nov;225(3):829-36.
Role of calcium and ROS in cell death induced by polyunsaturated fatty acids in murine thymocytes.
Prasad A, Bloom MS, Carpenter DO.
We investigated the mechanisms whereby omega-3 and -6 polyunsaturated fatty acids (PUFAs) cause cell death of mouse thymocytes using flow cytometry, focusing on the respective roles of intracellular calcium concentration, [Ca(2+)](i) and reactive oxygen species (ROS). We applied the C-22, 20, and 18 carbon omega-3 (DHA, EPA, ALA) and omega-6 (DTA, ARA, and LNA) fatty acids to isolated thymocytes and monitored cell death using the DNA-binding dye, propidium iodide. When applied at 20 µM concentration, omega-3 fatty acids killed thymocytes over a period of 1 h with a potency of DHA > EPA > ALA. The omega-6 PUFAs were more potent. The C18 omega-6 fatty acid, LNA, was the most potent, followed by DHA and ARA. Cell death was always accompanied by an increase in the levels of [Ca(2+)](i) and ROS. Both increases were in proportion to the potency of the PUFAs in inducing cell death. Removing extracellular calcium did not prevent the elevation in [Ca(2+)](i) nor cell death. However, the intracellular calcium chelator, BAPTA, almost totally reduced both the elevation in [Ca(2+)](i) and cell death, while vitamin E reduced the elevation in ROS and cell death. BAPTA also prevented the elevation in ROS, but vitamin E did not prevent the elevation in [Ca(2+)](i). Thapsigargin, which depletes endoplasmic reticulum calcium, blocked the elevation in [Ca(2+)](i), but CCCP, a mitochondrial calcium uptake inhibitor, did not. These results suggest that the six PUFAs we studied kill thymocytes by causing release of calcium from endoplasmic reticulum, which causes release of ROS from mitochondria which leads to cell death.

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Cardiolipin, Cytochrome Oxidase, Metabolism, & Aging

Also see:
Protect the Mitochondria
Mitochondrial Medicine
PUFA Accumulation & Aging
“Curing” a High Metabolic Rate with Unsaturated Fats
Fat Deficient Animals – Activity of Cytochrome Oxidase
Dietary PUFA Reflect in Human Subcutaneous Fat Tissue
Toxicity of Stored PUFA
PUFA, Development, and Allergy Incidence
Estrogen Levels Increase with Age
Fat Tissue and Aging – Increased Estrogen
Unsaturated Fats and Age Pigment
Ray Peat, PhD Quotes on Coconut Oil
Medium Chain Fats from Saturated Fat – Weight Management Friendly
Unsaturated Fats and Longevity
Endotoxin: Poisoning from the Inside Out
Ray Peat, PhD on Endotoxin
PUFA Decrease Cellular Energy Production
Fish Oil Toxicity
Protective “Essential Fatty Acid Deficiency”
Glucocorticoids, Cytochrome Oxidase, and Metabolism
Copper: The Forgotten Essential Nutrient

Quotes Ray Peat, PhD:
“A crucial enzyme in the mitochondrion is cytochrome oxidase, which reacts directly with oxygen, completing (or beginning) the process of chemical respiration. It is this enzyme (which is most sensitive to cyanide) which appears to be a “choke point” for energy production in various situations. Learning how to preserve and promote the activity of this enzyme is an important issue for everything having to do with biological energy…Kunkel and Williams (J. Biol. Chem., 1951) found that the very high respiratory rate of animals fed a diet lacking polyunsaturated fats was caused primarily by a great increase in the activity of cytochrome oxidase, and that adding an “essential fatty acid” strongly inhibited this enzyme’s activity.”

“The absorption and retention of magnesium, sodium, and copper, and the synthesis of proteins, are usually poor in hypothyroidism. Salt craving is common in hypothyroidism, and eating additional sodium tends to raise the body temperature, and by decreasing the produc- tion of aldosterone, it helps to minimize the loss of magnesium, which in turn allows cells to respond better to the thyroid hormone. This is probably why a low sodium diet increases adrenalin production, and why eating enough sodium lowers adrenalin and improves sleep. The lowered adrenalin is also likely to improve intestinal motility.”

“Thyroid hormone is essential for forming carbon dioxide. In the early 1940s, experimental rabbits were fed their standard diet, with the addition of 1% desiccated thyroid gland, which would be equivalent to about 150 grains of Armour thyroid for a person. They became extremely hypermetabolic, and couldn’t eat enough to meet their nutritional needs for growth and tissue maintenance. When they died, all of their tissues weighed much less than those of animals that hadn’t received the toxic dose of thyroid, except for their bones, which were larger than normal. Experiments with the thin skull bones of mice have shown that the active thyroid hormone, T3, increases the formation of bone. To increase cellular respiration and carbon dioxide production, T3 increases the activity of the enzyme cytochrome oxidase, which uses copper as a co-factor. Increased thyroid activity increases the absorption of copper from foods.”

“Just a diet can affect it [cytochrome c oxidase], but stress tends to intensify [the loss of] it. Not getting enough light, anything that stresses your energy production system such as high estrogen or nitric oxide, or low thyroid, or too much darkness will tend to make you lose copper from your respiratory system. And as copper gets lost, iron just tends to fill in for it, binding to some of the same enzymes.”

“Iron and copper are best known for their roles in oxygen transport and utilization… Copper is a component or cofactor for mitochondrial cytochrome oxidases, for dopamine B-oxidase (required for the biosynthesis of norepinephrine and epinephrine, of tyrosinase (involved in the biosynthesis of melanin pigments), and lysyl oxidase (which participates in the formation of collage and elastin).”

“I think the issue is just one of propaganda analysis, because scientifically, no one ever refuted the refutation of essentiality which occurred when the “EFA deficiency syndrome” was cured with vitamin B6. The German demonstration that spontaneous cancer was eliminated on a fat free diet preceded the really awful, incompetent study that supposedly demonstrated the essentiality of polyunsaturated fatty acids, and in the 75 years since the German study a tremendous amount of information has accumulated showing both the toxicity and the non-essentiality of the polyunsaturated fatty acids. But there has been no financial support for publicizing the protective effect of not eating vegetable oils or fish oils. To the contrary, vast amounts of money are being spent in the promotion of the various polyunsaturated fats as foods.”

“The animals that don’t eat them do have increased nutritional needs for vitamins and minerals, because their metabolic rate is so much greater than the PUFA-replete animals whose cardiolipin has degenerated. The recent Stanford study that shows a much greater longevity for old people who have a very high oxygen consumption capacity is consistent with the historical animal studies. PUFA-deprived animals have a very high oxygen consumption, and are resistant to practically all causes of death and disease, including trauma and poisoning.”

“The editorial boards of many of the journals are packed with industry flacks who are apparently willing to publish any junk that helps to sell soy oil, canola, waste fish oil, or algae oils. And researchers have to get grants to stay in business.”

“As early as 1951, it was known (Kunkel and Williams, J. BioI. Chern.) that the polyunsaturated fatty acids strongly inhibit the crucial respiratory enzyme, cytochrome oxidase, and that inhibition of this enzyme has a very important effect on the whole animal suppressing its metabolic rate, reducing the number of calories it can burn. It is now known that polyunsaturated fats interfere with thyroid hornone in just about every conceivable way.”

“The mitochondria are responsible for the efficient production of energy needed for the functioning of complex organisms, and especially for nerves. The enzyme in the mitochondria that reacts directly with oxygen, and that is often rate limiting, is cytochrome oxidase.

“This enzyme is dependent upon the thyroid hormone and is inhibited by nitric oxide, carbon monoxide, estrogen, polyunsaturated fatty acids, serotonin, excess or free iron, ionizing radiation, and many toxins, including bacterial endotoxin. Red light, which passes easily through the tissues, reactivates the enzyme, which slowly loses its function during darkness.”

“Estrogen impairs the mitochondria in multiple ways, including blocking the function of cytochrome oxidase, decreasing the activity of ATP synthase, increasing heme oxygenase which produces carbon monoxide and free iron, damaging mitochondrial DNA, and shifting metabolism from glucose oxidation to fat oxidation, especially by inhibiting pyruvate dehyrogenase complex. These changes including the loss of cytochrome oxidase, are seen in the Alzheimer’s brain. The fact that this kind of energy impairment can be produced by estrogen doesn’t imply that estrogen is the cause, since many other things can cause similar effects–radiation, aluminum, endotoxin, for example.”

“The choice of foods which have less unsaturated fat tends to reinforce the achievements of evolution.”

“The fetus produces saturated fats such as palmitic acid, and the monounsaturated fat, oleic acid, which can be turned into the Mead acid, ETrA (5,8,11-eicosatrienoic acid), and its derivatives, which are antiinflammatory, and some of which act on the “bliss receptor,” or the cannibinoid receptor.

“At birth, the baby’s mitochondria contain a phospholipid, cardiolipin, containing palmitic acid, but as the baby eats foods containing polyunsaturated fatty acids, the palmitic acid in cardiolipin is replaced by the unsaturated fats. As the cardiolipin becomes more unsaturated, it becomes less stable, and less able to support the activity of the crucial respiratory enzyme, cytochrome oxidase.”

“The respiratory activity of the mitochondria declines as the polyunsaturated oils replace palmitic acid, and this change corresponds to the life-long decline of the person’s metabolic rate.”

“Cytochrome oxidase is one of the enzymes damaged by stress and by blue light, and activated or restored by red light, thyroid, and progesterone. It’s a copper enzyme, so it’s likely to be damaged by excess iron. It is most active when it is associated with a mitochondrial lipid, cardiolipin, that contains saturated palmitic acid; the substitution of polyunsaturated fats lowers its activity. Mitochonrial function in general is poisoned by the unsaturated fats, especially arachidonic acid and DHA.”

“The suppressive effects of unsaturated fats on mitochondrial energy production have been widely investigated, since it is that effect that makes animal fattening with PUFA so economical. Rather than interpreting that as a toxic effect, using the innate structure and function of the mitochondrion as a point of reference from which to evaluate dietary components, the consumption of “good” oils is being used as the reference point from which to evaluate the meaning of metabolism (“efficiency is good,” “low oxygen consumption is good”). Building on the idea that the oils are health-promoters which increase metabolic efficiency, the never-viable “rate of aging” theory was resuscitated: The anti-respiratory effect of PUFA is used (illogically) to return to the idea that aging occurs in proportion to the amount of oxygen consumed, because animals which lack the supposedly essential nutrients (“defective animals”) consume oxygen rapidly–burning calories rapidly, they are supposed to be like a candle that won’t last as long if it burns intensely. The old theory is simply resuscitated to explain why the anti-respiratory action of PUFA might be beneficial, justifying further promotion of their use as food and drugs.”

“When mitochondria are functioning fully, either glucose or saturated fats can safely provide energy. Some glucose or saturated fat can be converted to polyunsaturated fats, that can be used as regulators or signals, for example to activate the formation of stem cells. But those PUFA don’t create disruptive cascades of increasing excitation or inflammation or excessive growth, and, from the evidence of animals that are fed fat free diets, or diets lacking omega -3 and omega -6 fatty acids, they aren’t toxic to mitochondria.”

“Increased estrogen exposure, decreased thyroid hormone, an increased ratio of iron to copper, and lack of light, are other factors that impair the cytochrome oxidase enzyme.”

“The crucial mitochondrial respiratory enzyme, cytochrome c oxidase, declines with aging (Paradies, et al., 1997), as the lipid cardiolipin declines, and the enzyme’s activity can be restored to the level of young animals by adding cardiolipin. The composition of cardiolipin changes with aging, “specifically an increase in highly unsaturated fatty acids” (Lee, et al., 2006). Other lipids, such as a phosphatidylcholine containing two myristic acid groups, can support the enzyme’s activity (Hoch, 1992). Even supplementing old animals with hydrogenated peanut oil restores mitochondrial respiration to about 80% of normal (Bronnikov, et al., 2010).”

“Supplementing thyroid hormone increases mitochondrial cardiolipin (Paradies and Ruggiero, 1988). Eliminating the polyunsaturated fats from the diet increases mitochondrial respiration (Rafael, et al., 1984).”

“On a typical diet, tissues progressively accumulate linoleic acid, and this alters the structure of mitochondrial cardiolipin, which governs the response of the mitochondrial enzymes to the thyroid hormone. This process is especially evident in the female liver. In the “autoimmune” diseases, such as lupus, there are typically antibodies to cardiolipin, as if the body were trying to reject its own tissues, which have been altered by the storage of linoleic acid.

The altered mitochondrial function, which is involved in so many symptoms, can become part of a vicious circle, with endotoxin and estrogen having central roles, once the stage has been set by the combination of diet, stress, and toxins.”

J Biol Chem. 1951 Apr;189(2):755-61.
The effects of fat deficiency upon enzyme activity in the rat.
KUNKEL HO, WILLIAMS JN Jr.
The activity of the cytochrome oxidase, however, is markedly increased in fat deficiency…In each case the activity of livers from rats fed the basal diet was 38 per cent greater than from the linoleate-supplemented animals or from the animals receiving corn oil. This is particularly interesting in view of the observation of Burr and Beeber (8) and Wesson and Burr (9) that fat-deficient rats had a markedly increased metabolic rate. The latter authors reported that the basal and assimilatory metabolic rates of fat-deficient animals were 25 per cent greater than the rates of the control animals. Thus the liver cytochrome oxidase activity appears to parallel the met,abolic rate in fat deficiency. This increased cytochrome oxidase activity in liver and perhaps other tissues may account in a large part for the increased metabolic rate.

Summary
A fat deficiency in the rat causes a marked increase in liver cytochrome oxidase activity, a slight increase in choline oxidase activity, and a marked decrease in endogenous respiration. The activity of the succinic oxidase system is not altered by this deficiency condition. Supplementation with 100 mg. of methyl linoleate per rat per day reduced the cytochrome oxidase to the level of that produced by a 5 percent corn oil diet.

FEBS Lett. 1997 Apr 7;406(1-2):136-8.
Age-dependent decline in the cytochrome c oxidase activity in rat heart mitochondria: role of cardiolipin.
Paradies G, Ruggiero FM, Petrosillo G, Quagliariello E.
Cardiolipin is a major mitochondrial membrane lipid and plays a pivotal role in mitochondrial function. We have recently suggested a possible involvement of this phospholipid in the age-linked decline of cytochrome c oxidase activity in rat heart mitochondria [G. Paradies et al. (1993) Arch. Gerontol. Geriatr. 16, 263-272]. The aim of this work was to test our earlier proposal. We have investigated whether addition of exogenous cardiolipin to mitochondria is able to reverse, in situ, the age-linked decrease in the cytochrome oxidase activity. The method of fusion of liposomes with mitochondria developed by Hackenbrock [Hackenbrock and Chazotte (1986) Methods Enzymol. 125, 35-45] was employed in order to enrich the mitochondria cardiolipin content. We demonstrate that the lower cytochrome c oxidase activity in heart mitochondria from aged rats can be fully restored to the level of young control rats by exogenously added cardiolipin. No restoration was obtained with other phospholipids or with peroxidized cardiolipin. Our data support a key role for cardiolipin in the age-linked decline of rat heart mitochondrial cytochrome c oxidase activity.

FEBS Lett. 1999 Jul 9;454(3):207-9.
The effect of aging and acetyl-L-carnitine on the pyruvate transport and oxidation in rat heart mitochondria.
Paradies G, Petrosillo G, Gadaleta MN, Ruggiero FM.
The effect of aging and acute treatment with acetyl-L-carnitine on the pyruvate transport and oxidation in rat heart mitochondria was studied. The activity of the pyruvate carrier as well as the rates of pyruvate-supported respiration were both depressed (around 40%) in heart mitochondria from aged rats, the major decrease occurring during the second year of life. Administration of acetyl-L-carnitine to aged rats almost completely restored the rates of these metabolic functions to the level of young control rats. This effect of acetyl-L-carnitine was not due to changes in the content of pyruvate carrier molecules. The heart mitochondrial content of cardiolipin, a key phospholipid necessary for mitochondrial substrate transport, was markedly reduced (approximately 40%) in aged rats. Treatment of aged rats with acetyl-L-carnitine reversed the age-associated decline in cardiolipin content. As the changes in cardiolipin content were correlated with changes in rates of pyruvate transport and oxidation, it is suggested that acetyl-L-carnitine reverses the age-related decrement in the mitochondrial pyruvate metabolism by restoring the normal cardiolipin content.

Lipids Health Dis. 2006 Jan 23;5:2.
Selective remodeling of cardiolipin fatty acids in the aged rat heart.
Lee HJ, Mayette J, Rapoport SI, Bazinet RP.
BACKGROUND:
The heart is rich in cardiolipin, a phospholipid acylated in four sites, predominately with linoleic acid. Whether or not aging alters the composition of cardiolipin acyl chains is controversial. We therefore measured the fatty acid concentration of cardiolipin in hearts of 4, 12 and 24 month old rats that consumed one diet, adequate in fatty acids for the duration of their life.
RESULTS:
The concentration (nmol/g) of linoleic acid was decreased in 24 month old rats (3965 +/- 617, mean +/- SD) vs 4 month old rats (5525 +/- 656), while the concentrations of arachidonic and docosahexaenoic acid were increased in 24 month old rats (79 +/- 9 vs 178 +/- 27 and 104 +/- 16 vs 307 +/- 68 for arachidonic and docosahexaenoic acids, 4 months vs 24 months, respectively). Similar changes were not observed in ethanolamine glycerophospholipids or plasma unesterified fatty acids, suggesting specificity of these effects to cardiolipin.
CONCLUSION:
These results demonstrate that cardiolipin remodeling occurs with aging, specifically an increase in highly unsaturated fatty acids.

Biochemistry (Mosc). 2010 Dec;75(12):1491-7.
Dietary supplementation of old rats with hydrogenated peanut oil restores activities of mitochondrial respiratory complexes in skeletal muscles.
Bronnikov GE, Kulagina TP, Aripovsky AV.
The effect of dietary supplementation of old rats (26-33 months) with hydrogenated peanut oil on the activity of mitochondrial enzymes in skeletal muscles has been studied. The activities of NADH-coenzyme Q1 oxidoreductase, cytochrome c oxidase, and citrate synthase were determined spectrophotometrically in muscle homogenates. The activities of respiratory complexes I and IV were shown to significantly decrease with the age compared to the activity of the same enzymes in young animals, while the activity of citrate synthase was virtually unchanged. The fatty acid composition of muscle homogenates of old rats differed from that of young animals by a reduced content of myristic, oleic, linoleic, and α-linolenic acids and enhanced content of dihomo-γ-linolenic, arachidonic, and docosahexaenoic acids. Per oral supplementation of the old rats with hydrogenated peanut oil completely restored the activity of complex IV and increased the activity of complex I to 80% of the value observed in muscles of young animals, reducing the content of stearic, dihomo-γ-linolenic, arachidonic, eicosapentaenoic, docosapentaenoic, and docosahexaenoic acids relative to that in the groups of old and young rats. The content of oleic and linoleic acids increased relatively to that in the group of the old rats, as well as young animals. The possible mechanisms of the restoration of the activity of the respiratory enzymes under the administration of hydrogenated peanut oil are discussed.

Biochim Biophys Acta. 1988 Aug 17;935(1):79-86.
Effect of hyperthyroidism on the transport of pyruvate in rat-heart mitochondria.
Paradies G, Ruggiero FM.
A comparative study of the transport of pyruvate in heart mitochondria from normal and triiodothyronine-treated rats has been carried out. It has been found that the rate of carrier-mediated (alpha-cyanocinnamate-sensitive) pyruvate uptake is significantly enhanced in mitochondria from triiodothyronine-treated rats as compared with mitochondria from control rats. The kinetic parameters of the pyruvate uptake indicate that only the Vmax of this process is enhanced whilst there is no change in the Km value. The enhanced rate of pyruvate uptake is not dependent on the increase of the transmembrane delta pH value (both mitochondria from normal and triiodothyronine-treated rats exhibit the same delta pH value) neither does it depend on the increase of the pyruvate carrier molecules (titration of these last with alpha-cyanocinnamate gives the same total number of binding sites). the pyruvate-dependent oxygen uptake is stimulated by 35-40% in mitochondria from hyperthyroid rats when compared with mitochondria from control rats. There is, however, no difference in either the respiratory control ratios or in the ADP/O ratios between these two types of mitochondria. The heart mitochondrial phospholipid composition is altered significantly in hyperthyroid rats; in particular, negatively charged phospholipid such as cardiolipin and phosphatidylserine were found to increase by more than 50%. Minor alterations were found in the pattern of fatty acids with an increase of 20:4/18:2 ratio. It is suggested that the changes in the kinetic parameters of pyruvate transport in mitochondria from hyperthyroid rats involve hormone-mediated changes in the lipid composition of the mitochondrial membranes which in turn modulate the activity of the pyruvate carrier.

J Nutr. 1984 Feb;114(2):255-62.
The effect of essential fatty acid deficiency on basal respiration and function of liver mitochondria in rats.
Rafael J, Patzelt J, Schäfer H, Elmadfa I.
Rats were fed a diet poor (0.05%) in essential fatty acids (EFA) with hydrogenated coconut oil as fat component, or a control diet containing 4% of the total energy intake in the form of linoleic acid. Effects of dietary EFA deficiency were investigated during a period of 2-30 weeks. Growth retardation becomes significant after 4 weeks of deficiency and attains about 25% when the deficiency is maintained for longer than 12 weeks. Respiration, body weight and age of EFA-deficient rats and controls are in a nonlinear relationship. Basal respiration in relation to the body weight is significantly increased by EFA deficiency; it is unchanged when related to total animals under the employed experimental conditions. Oxidative phosphorylation in isolated liver mitochondria is unaffected by EFA deficiency, i.e., the increased metabolic rate of EFA-deficient rats, related to the body weight, cannot be explained from impaired functional integrity of the inner mitochondrial membrane. Respiratory chain enzyme activities in mitochondria from heart and skeletal muscle and specific amounts of mitochondria in these tissues are unchanged by EFA deficiency.

Arch Gerontol Geriatr. 1993 May-Jun;16(3):263-72.
Age-dependent decrease in the cytochrome c oxidase activity and changes in phospholipids in rat-heart mitochondria.
Paradies G, Ruggiero FM, Petrosillo G, Quagliariello E.
The effect of aging on the kinetic characteristics of cytochrome c oxidase in rat heart mitochondria was investigated. Mitochondria from young and aged rats had equivalent Km values for cytochrome c, while the maximal activity of the oxidase was significantly reduced in mitochondria from aged rats. The cytochrome aa3 content was the same in both these two types of mitochondria. The Arrhenius plot characteristics differ for cytochrome oxidase activity in mitochondria from aged rats as compared with young rats in that the breakpoint of the biphasic plot was shifted to a higher temperature. Cardiolipin content was markedly decreased in mitochondrial membrane from aged rats. No alterations were found in the patterns of the phospholipid fatty acid distribution of mitochondrial membranes from young and aged rats. The results support the conclusion that the reduced mitochondrial cytochrome c oxidase activity in aged animals is due to a specific decrease in the cardiolipin content.

Biochim Biophys Acta. 1995 May 24;1271(1):165-70.
Mitochondrial decay in aging.
Ames BN, Shigenaga MK, Hagen TM.
Several mitochondrial functions decline with age. The contributing factors include, the intrinsic rate of proton leakage across the inner mitochondrial membrane (a correlate of oxidant formation), decreased membrane fluidity, and decreased levels and function of cardiolipin, which supports the function of many of the proteins of the inner mitochondrial membrane. Oxidants generated by mitochondria appear to be the major source of the oxidative lesions that accumulate with age. Evidence supports the suggestion that age-associated accumulation of mitochondrial deficits due to oxidative damage is likely to be a major contributor to cellular, tissue, and organismal aging.

“With aging, cells have less ability to produce energy, and are often more easily stimulated. The accumulation of polyunsaturated fats is one of the factors that reduce the ability of mitochondria to produce energy (Zhang, et al., 2006, 2009; Yazbeck, et al., 1989). Increased estrogen exposure, decreased thyroid hormone, an increased ratio of iron to copper, and lack of light, are other factors that impair the cytochrome oxidase enzyme.” -Ray Peat, PhD

Comp Biochem Physiol A Comp Physiol. 1989;94(2):273-6.
The effects of essential fatty acid deficiency on brown adipose tissue activity in rats maintained at thermal neutrality.
Yazbeck J, Goubern M, Senault C, Chapey MF, Portet R.
1. The consequences of essential fatty acid (EFA) deficiency on the resting metabolism, food efficiency and brown adipose tissue (BAT) thermogenic activity were examined in rats maintained at thermal neutrality (28 C). 2. Weanling male Long-Evans rats were fed a hypolipidic semi-purified diet (control diet: 2% sunflower oil; EFA-deficient diet: 2% hydrogenated coconut oil) for 9 weeks. 3. They were kept at 28 C for the last 5 weeks. Compared to controls, in EFA-deficient rats the growth shortfall reached 21% at killing. 4. As food intake was the same in EFA-deficient and control rats, food efficiency was thus decreased by 40%. 5. Resting metabolism expressed per surface unit was 15% increased. 6. Non-renal water loss was increased by 88%. 7. BAT weight was 28% decreased but total and mitochondrial proteins were not modified. 8. Heat production capacity, tested by GDP binding per BAT was 69% increased in BAT of deficient rats. 9. The stimulation of BAT was established by two other tests: GDP inhibition of mitochondrial O2 consumption and swelling of mitochondria. 10. It is suggested that the observed enhancement of resting metabolism in EFA-deficient rats is, in part, due to an activation of heat production in BAT.

Am J Physiol Cell Physiol. 2006 May;290(5):C1321-33.
Polyunsaturated fatty acids mobilize intracellular Ca2+ in NT2 human teratocarcinoma cells by causing release of Ca2+ from mitochondria.
Zhang BX, Ma X, Zhang W, Yeh CK, Lin A, Luo J, Sprague EA, Swerdlow RH, Katz MS.
In a variety of disorders, overaccumulation of lipid in nonadipose tissues, including the heart, skeletal muscle, kidney, and liver, is associated with deterioration of normal organ function, and is accompanied by excessive plasma and cellular levels of free fatty acids (FA). Increased concentrations of FA may lead to defects in mitochondrial function found in diverse diseases. One of the most important regulators of mitochondrial function is mitochondrial Ca(2+) ([Ca(2+)](m)), which fluctuates in coordination with intracellular Ca(2+) ([Ca(2+)](i)). Polyunsaturated FA (PUFA) have been shown to cause [Ca(2+)](i) mobilization albeit by unknown mechanisms. We have found that PUFA but not monounsaturated or saturated FA cause [Ca(2+)](i) mobilization in NT2 human teratocarcinoma cells. Unlike the [Ca(2+)](i) response to the muscarinic G protein-coupled receptor agonist carbachol, PUFA-mediated [Ca(2+)](i) mobilization in NT2 cells is independent of phospholipase C and inositol-1,4,5-trisphospate (IP(3)) receptor activation, as well as IP(3)-sensitive internal Ca(2+) stores. Furthermore, PUFA-mediated [Ca(2+)](i) mobilization is inhibited by the mitochondria uncoupler carboxyl cyanide m-chlorophenylhydrozone. Direct measurements of [Ca(2+)](m) with X-rhod-1 and (45)Ca(2+) indicate that PUFA induce Ca(2+) efflux from mitochondria. Further studies show that ruthenium red, an inhibitor of the mitochondrial Ca(2+) uniporter, blocks PUFA-induced Ca(2+) efflux from mitochondria, whereas inhibitors of the mitochondrial permeability transition pore cyclosporin A and bongkrekic acid have no effect. Thus PUFA-gated Ca(2+) release from mitochondria, possibly via the Ca(2+) uniporter, appears to be the underlying mechanism for PUFA-induced [Ca(2+)](i) mobilization in NT2 cells.

PLoS ONE 4(6): e6048. doi:10.1371/journal.pone.0006048
Linoleic Acid-Induced Mitochondrial Ca2+ Efflux Causes Peroxynitrite Generation and Protein Nitrotyrosylation
Hong-Mei Zhang1, Howard Dang2, Chih-Ko Yeh3,4, Bin-Xian Zhang1,4*
It is well known that excessive non-esterified fatty acids in diabetes contribute to the pathogenesis of renal complications although the mechanism remains elusive. Enhanced oxidative stress has been hypothesized as a unified factor contributing to diabetic complications and increased protein nitrotyrosylation has been reported in the kidneys of diabetic patients. In the current manuscript we described that linoleic acid (LA) caused mitochondrial Ca2+ efflux and peroxynitrite production, along with increased nitrotyrosine levels of cellular proteins in primary human mesangial cells. The peroxynitrite production by LA was found to depend on mitochondrial Ca2+ efflux. Downregulation of hsp90β1, which has been previously shown to be essential for polyunsaturated fatty acid-induced mitochondrial Ca2+ efflux, significantly diminished LA-responsive mitochondrial Ca2+ efflux and the coupled peroxynitrite generation, implicating a critical role of hsp90β1 in the LA responses. Our results further demonstrated that mitochondrial complexes I and III were directly involved in the LA-induced peroxynitrite generation. Using the well established type 2 diabetic animal model db/db mice, we observed a dramatically enhanced LA responsive mitochondrial Ca2+ efflux and protein nitrotyrosylation in the kidney. Our study thus demonstrates a cause-effect relationship between LA and peroxynitrite or protein nitrotyrosylation and provides a novel mechanism for lipid-induced nephropathy in diabetes.

Endocrine. 2011 Apr;39(2):128-38. Epub 2010 Dec 15.
Long-term exposure of INS-1 rat insulinoma cells to linoleic acid and glucose in vitro affects cell viability and function through mitochondrial-mediated pathways.
Tuo Y, Wang D, Li S, Chen C.
Obesity with excessive levels of circulating free fatty acids (FFAs) is tightly linked to the incidence of type 2 diabetes. Insulin resistance of peripheral tissues and pancreatic β-cell dysfunction are two major pathological changes in diabetes and both are facilitated by excessive levels of FFAs and/or glucose. To gain insight into the mitochondrial-mediated mechanisms by which long-term exposure of INS-1 cells to excess FFAs causes β-cell dysfunction, the effects of the unsaturated FFA linoleic acid (C 18:2, n-6) on rat insulinoma INS-1 β cells was investigated. INS-1 cells were incubated with 0, 50, 250 or 500 μM linoleic acid/0.5% (w/v) BSA for 48 h under culture conditions of normal (11.1 mM) or high (25 mM) glucose in serum-free RPMI-1640 medium. Cell viability, apoptosis, glucose-stimulated insulin secretion, Bcl-2, and Bax gene expression levels, mitochondrial membrane potential and cytochrome c release were examined. Linoleic acid 500 μM significantly suppressed cell viability and induced apoptosis when administered in 11.1 and 25 mM glucose culture medium. Compared with control, linoleic acid 500 μM significantly increased Bax expression in 25 mM glucose culture medium but not in 11.1 mM glucose culture medium. Linoleic acid also dose-dependently reduced mitochondrial membrane potential (ΔΨm) and significantly promoted cytochrome c release from mitochondria in both 11.1 mM glucose and 25 mM glucose culture medium, further reducing glucose-stimulated insulin secretion, which is dependent on normal mitochondrial function. With the increase in glucose levels in culture medium, INS-1 β-cell insulin secretion function was deteriorated further. The results of this study indicate that chronic exposure to linoleic acid-induced β-cell dysfunction and apoptosis, which involved a mitochondrial-mediated signal pathway, and increased glucose levels enhanced linoleic acid-induced β-cell dysfunction.

FEBS Lett. 1996 Nov 18;397(2-3):260-2.
Stimulation of carnitine acylcarnitine translocase activity in heart mitochondria from hyperthyroid rats.
Paradies G, Ruggiero FM, Petrosillo G, Quagliariello E.
The effect of hyperthyroidism on fatty acid oxidation and on carnitine-acylcarnitine translocase activity in rat heart mitochondria has been studied. The rates of palmitoylcarnitine supported respiration as well as the carnitine-palmitoylcarnitine exchange reaction were both stimulated (approx. 36%) in heart mitochondria from hyperthyroid rats. Kinetic analysis of the carnitine-carnitine exchange reaction showed that thyroid hormone affects the Vmax of this process, while having no effect on the Km values. The level of cardiolipin was significantly higher (approx. 40%) in heart mitoplasts from hyperthyroid rats than from the control rats. It can be concluded that thyroid hormones produce a stimulation of heart mitochondrial carnitine translocase activity and that the basis of this effect is likely an increase in the cardiolipin content.

FEBS Lett. 1998 Mar 13;424(3):155-8.
Peroxidative damage to cardiac mitochondria: cytochrome oxidase and cardiolipin alterations.
Paradies G, Ruggiero FM, Petrosillo G, Quagliariello E.
Rat heart mitochondrial membranes exposed to the free radicals generating system tert-butylhydroperoxide/Cu2+ undergo lipid peroxidation as evidenced by the accumulation of thyobarbituric acid reactive substances. Mitochondrial lipid peroxidation resulted in a marked loss of both cytochrome c oxidase activity and cardiolipin content. The alterations in the properties of cytochrome c oxidase were confined to a decrease in the maximal activity (Vmax) with no change in the affinity (Km) with respect to the substrate cytochrome c. Various lipid soluble antioxidants could prevent the lipid peroxidation reaction and the associated loss of cytochrome c oxidase activity. External added cardiolipin but no other phospholipids, nor peroxidized cardiolipin was able to prevent the loss of cytochrome oxidase activity induced by lipid peroxidation. These results establish a close correlation between oxidative damage to cardiolipin and alterations in the cytochrome oxidase activity and may prove useful in probing molecular mechanism of free radicals induced peroxidative damage of mitochondria which has been proposed to contribute to aging and to chronic degenerative diseases.

Biochim Biophys Acta. 1994 Jan 11;1225(2):165-70.
Enhanced cytochrome oxidase activity and modification of lipids in heart mitochondria from hyperthyroid rats.
Paradies G, Ruggiero FM, Petrosillo G, Quagliariello E.
In order to further investigate the mechanism regulating the control of mitochondrial respiration by thyroid hormones, the effect of the hyperthyroidism on the kinetic characteristics of cytochrome c oxidase in rat heart mitochondria was studied. Mitochondrial preparations from both control and hyperthyroid rats had equivalent Km values for cytochrome c, while the maximal activity of cytochrome oxidase was significantly increased (by around 30%) in mitochondrial preparation from hyperthyroid rats. This enhanced activity of cytochrome oxidase was associated to a parallel increase in mitochondrial State 3 respiration. The hormone treatment resulted in a decrease in the flux control coefficient of the oxidase. The enhanced activity of cytochrome oxidase in hyperthyroid rats does not appear to be dependent on an increase in the mass of this enzyme complex in that the heme aa3 content was equivalent in both hyperthyroid and control preparations. The Arrhenius plot characteristics differ for cytochrome oxidase activity in mitochondria from hyperthyroid rats as compared with control rats in that the breakpoint of the biphasic plot is shifted to a lower temperature. Cardiolipin content was significantly increased in mitochondrial preparations from hyperthyroid rats, while there were no significant alterations in the fatty acid composition of cardiolipin of control and hyperthyroid preparations. The results support the conclusion that the enhanced cytochrome oxidase activity in heart mitochondrial preparations from hyperthyroid rats is due to a specific increase in the content of cardiolipin.

Biochim Biophys Acta. 1997 Dec 31;1362(2-3):193-200.
Alterations in carnitine-acylcarnitine translocase activity and in phospholipid composition in heart mitochondria from hypothyroid rats.
Paradies G, Ruggiero FM, Petrosillo G, Quagliariello E.
Changes in mitochondrial fatty acid metabolism may underlie the decline in cardiac function in the hypothyroid animals. The effect of hypothyroidism on fatty acid oxidation, carnitine-acylcarnitine translocase activity and lipid composition in rat heart mitochondria has been examined. Rates of mitochondrial fatty acid oxidation as well as carnitine-carnitine and carnitine-palmitoylcarnitine exchange reactions were all depressed in heart mitochondria isolated from hypothyroid rats. Kinetic analysis of the carnitine-carnitine exchange reaction showed that the hypothyroid state affects the Vmax of this process, while having no effect on the K(m) value. Heart mitochondrial inner membrane lipid composition was significantly altered in hypothyroid rats. Cardiolipin, particularly, was found to decrease (by around 36%). Alterations in fatty acid pattern of mitochondrial inner membrane preparations from hypothyroid rats were also found. The effects of the hypothyroid state on fatty acids oxidation, carnitine translocase activity and phospholipid composition were completely reversed by following treatment of hypothyroid rats with thyroid hormone. A lower cardiolipin content in the mitochondrial inner membrane offers a plausible mechanism to explain the decline in the translocase activity in hypothyroidism.

Arch Biochem Biophys. 1993 Nov 15;307(1):91-5.
Decreased cytochrome oxidase activity and changes in phospholipids in heart mitochondria from hypothyroid rats.
Paradies G, Ruggiero FM, Dinoi P, Petrosillo G, Quagliariello E.
The effect of hypothyroidism on kinetic characteristics of cytochrome oxidase in rat heart mitochondria was studied. Mitochondrial preparations from control and hypothyroid rats had equivalent Km values for cytochrome c, while the maximal activity of the oxidase was significantly decreased (more than 30%) in mitochondrial preparations from hypothyroid rats. This decrease is associated to a parallel decrease in state 3 respiration. The cytochrome aa3 content was slightly decreased (by around 15%) in mitochondria from hypothyroid rats. The Arrhenius plot characteristics differ for cytochrome oxidase activity in mitochondria from hypothyroid rats as compared with control rats in that the breakpoint of the biphasic plot is shifted to a higher temperature. Cardiolipin content was markedly decreased in the mitochondrial membrane from hypothyroid rats. No alterations were found in the pattern of cardiolipin fatty acid distribution of mitochondrial membrane from control and hypothyroid rats. The effects of the hypothyroid state on the activity of cytochrome oxidase, on cytochrome aa3 levels, and on cardiolipin contents were completely reversed by following the treatment of hypothyroid rats with thyroid hormone. The results support the conclusion that the depressed mitochondrial cytochrome oxidase activity in the hypothyroid state is due, at least in part, to a decrease in the cardiolipin content of the mitochondrial inner membrane.

Biochim Biophys Acta. 1997 Mar 28;1319(1):5-8.
Cardiolipin-dependent decrease of cytochrome c oxidase activity in heart mitochondria from hypothyroid rats.
Paradies G, Petrosillo G, Ruggiero FM.
Cardiolipin plays an important role in mitochondrial membrane structure and function. We have recently reported a decrease in the cytochrome c oxidase activity in heart mitochondria from hypothyroid rats (G. Paradies et al. (1993) Arch. Biochem Biophys. 307, 91-95). A possible involvement of cardiolipin in such a decrease has been proposed. The aim of this work was to test our earlier proposal. We have investigated whether addition of exogenous cardiolipin to hypothyroid mitochondria is able to reverse, in situ, their decreased cytochrome oxidase activity. The method of fusion of liposomes with mitochondria developed by Hackenbrock (Hackenbrock and Chazotte (1986) Methods Enzymol. 125, 35-45) was employed in order to enrich the mitochondrial cardiolipin content. We demonstrate that the decreased activity of this enzyme complex in heart mitochondria from hypothyroid rats can be completely restored to the level of control rats by exogenously added cardiolipin but not by other phospholipids. These data provide strong evidence for the involvement of cardiolipin in the thyroid hormone induced changes of mitochondrial cytochrome oxidase activity.

Mitochondrion. 2001 Aug;1(2):151-9.
Reactive oxygen species generated by the mitochondrial respiratory chain affect the complex III activity via cardiolipin peroxidation in beef-heart submitochondrial particles.
Paradies G, Petrosillo G, Pistolese M, Ruggiero FM.
The aim of this study was to investigate the effect of reactive oxygen species (ROS), produced by the mitochondrial respiratory chain, on the activity of complex III and on the cardiolipin content in bovine-heart submitochondrial particles (SMP). ROS were produced by treatment of nicotinamide adenine dinucleotide (NADH) respiring SMP with rotenone. This treatment resulted in a production of superoxide anion, detected by the epinephrine method, which was blocked by superoxide dismutase (SOD). Exposure of SMP to mitochondrial-mediated ROS generation resulted in a marked loss of complex III activity and in a parallel loss of mitochondrial cardiolipin content. Both these effects were completely abolished by SOD + catalase. Exogenous added cardiolipin was able to almost completely prevent the ROS-mediated loss of complex III activity. No effect was obtained with other major phospholipid components of the mitochondrial membrane such as phosphatidylcholine and phosphatidylethanolamine, or with peroxidized cardiolipin. The results demonstrate that mitochondrial-mediated ROS generation affects the activity of complex III via peroxidation of cardiolipin, which is required for the functioning of this multisubunit enzyme complex. These results may prove useful in probing molecular mechanisms of ROS-induced peroxidative damage to mitochondria, which have been proposed to contribute to those physiopathological conditions characterized by an increase in the basal production of ROS such as aging, ischemia/reperfusion and chronic degenerative diseases.

Free Radic Biol Med. 1999 Jul;27(1-2):42-50.
Lipid peroxidation and alterations to oxidative metabolism in mitochondria isolated from rat heart subjected to ischemia and reperfusion.
Paradies G, Petrosillo G, Pistolese M, Di Venosa N, Serena D, Ruggiero FM.
Ischemia-reperfusion injury to cardiac myocytes involves membrane damage mediated by oxygen free radicals. Lipid peroxidation is considered a major mechanism of oxygen free radical toxicity in reperfused heart. Mitochondrial respiration is an important source of these reactive oxygen species and hence a potential contributor to reperfusion injury. We have examined the effects of ischemia (30 min) and ischemia followed by reperfusion (15 min) of rat hearts, on the kinetic parameters of cytochrome c oxidase, on the respiratory activities and on the phospholipid composition in isolated mitochondria. Mitochondrial content of malonyldialdheyde (MDA), an index of lipid peroxidation, was also measured. Reperfusion was accompanied by a significant increase in MDA production. Mitochondrial preparations from control, ischemic and reperfused rat heart had equivalent Km values for cytochrome c, although the maximal activity of the oxidase was 25 and 51% less in ischemic and reperfused mitochondria than that of controls. These changes in the cytochrome c oxidase activity were associated to parallel changes in state 3 mitochondrial respiration. The cytochrome aa3 content was practically the same in these three types of mitochondria. Alterations were found in the mitochondrial content of the major phospholipid classes, the most pronounced change occurring in the cardiolipin, the level that decreased by 28 and by 50% as function of ischemia and reperfusion, respectively. The lower cytochrome c oxidase activity in mitochondria from reperfused rat hearts could be almost completely restored to the level of control hearts by exogenously added cardiolipin, but not by other phospholipids nor by peroxidized cardiolipin. It is proposed that the reperfusion-induced decline in the mitochondrial cytochrome c oxidase activity can be ascribed, at least in part, to a loss of cardiolipin content, due to peroxidative attack of its unsaturated fatty acids by oxygen free radicals. These findings may provide an explanation for some of the factors that lead to myocardial reperfusion injury.

Gene. 2002 Mar 6;286(1):135-41.
Reactive oxygen species affect mitochondrial electron transport complex I activity through oxidative cardiolipin damage.
Paradies G, Petrosillo G, Pistolese M, Ruggiero FM.
The aim of this study was to investigate the influence of reactive oxygen species (ROS) on the activity of complex I and on the cardiolipin content in bovine heart submitochondrial particles (SMP). ROS were generated through the use of xanthine/xanthine oxidase (X/XO) system. Treatment of SMP with X/XO resulted in a large production of superoxide anion, detected by acetylated cytochrome c method, which was blocked by superoxide dismutase (SOD). Exposure of SMP to ROS generation resulted in a marked loss of complex I activity and to parallel loss of mitochondrial cardiolipin content. Both these effects were completely abolished by SOD+catalase. Exogenous added cardiolipin was able to almost completely restore the ROS-induced loss of complex I activity. No restoration was obtained with other major phospholipid components of the mitochondrial membrane such as phosphatidylcholine and phosphatidylethanolamine, nor with peroxidized cardiolipin. These results demonstrate that ROS affect the mitochondrial complex I activity via oxidative damage of cardiolipin which is required for the functioning of this multisubunit enzyme complex. These results may prove useful in probing molecular mechanisms of ROS-induced peroxidative damage to mitochondria, which have been proposed to contribute to those pathophysiological conditions characterized by an increase in the basal production of reactive oxygen species such as aging, ischemia/reperfusion and chronic degenerative diseases.

FEBS Lett. 2000 Jan 28;466(2-3):323-6.
The effect of reactive oxygen species generated from the mitochondrial electron transport chain on the cytochrome c oxidase activity and on the cardiolipin content in bovine heart submitochondrial particles.
Paradies G, Petrosillo G, Pistolese M, Ruggiero FM.
The effect of reactive oxygen species (ROS), produced by the mitochondrial respiratory chain, on the activity of cytochrome c oxidase and on the cardiolipin content in bovine heart submitochondrial particles (SMP) was studied. ROS were produced by treatment of succinate-respiring SMP with antimycin A. This treatment resulted in a large production of superoxide anion, measured by epinephrine method, which was blocked by superoxide dismutase (SOD). Exposure of SMP to mitochondrial mediated ROS generation, led to a marked loss of cytochrome c oxidase activity and to a parallel loss of cardiolipin content. Both these effects were completely abolished by SOD+catalase. Added cardiolipin was able to almost completely restore the ROS-induced loss of cytochrome c oxidase activity. No restoration was obtained with peroxidized cardiolipin. These results demonstrate that mitochondrial mediated ROS generation affects the activity of cytochrome c oxidase via peroxidation of cardiolipin which is needed for the optimal functioning of this enzyme complex. These results may prove useful in probing molecular mechanism of ROS-induced peroxidative damage to mitochondria which have been proposed to contribute to aging, ischemia/reperfusion and chronic degenerative diseases.

Circ Res. 2004 Jan 9;94(1):53-9. Epub 2003 Dec 1.
Decrease in mitochondrial complex I activity in ischemic/reperfused rat heart: involvement of reactive oxygen species and cardiolipin.
Paradies G, Petrosillo G, Pistolese M, Di Venosa N, Federici A, Ruggiero FM.
Reactive oxygen species (ROS) are considered an important factor in ischemia/reperfusion injury to cardiac myocytes. Mitochondrial respiration is an important source of ROS production and hence a potential contributor to cardiac reperfusion injury. In this study, we have examined the effect of ischemia and ischemia followed by reperfusion of rat hearts on various parameters related to mitochondrial function, such as complex I activity, oxygen consumption, ROS production, and cardiolipin content. The activity of complex I was reduced by 25% and 48% in mitochondria isolated from ischemic and reperfused rat heart, respectively, compared with the controls. These changes in complex I activity were associated with parallel changes in state 3 respiration. The capacity of mitochondria to produce H2O2 increased on reperfusion. The mitochondrial content of cardiolipin, which is required for optimal activity of complex I, decreased by 28% and 50% as function of ischemia and reperfusion, respectively. The lower complex I activity in mitochondria from reperfused rat heart could be completely restored to the level of normal heart by exogenous added cardiolipin. This effect of cardiolipin could not be replaced by other phospholipids nor by peroxidized cardiolipin. It is proposed that the defect in complex I activity in ischemic/reperfused rat heart could be ascribed to a ROS-induced cardiolipin damage. These findings may provide an explanation for some of the factors responsible for myocardial reperfusion injury.

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PUFA Inhibit Glucuronidation

Also see:
PUFA Increases Estrogen
PUFA and Liver Toxicity; Protection by Saturated Fats
Protective “Essential Fatty Acid Deficiency”

Quotes by Ray Peat, PhD:
“One of the essential protective functions that decline with aging is the liver’s ability to detoxify chemicals, by combining them with glucuronic acid, making them water soluble so that they can be excreted in the urine. The liver (and also the intestines and stomach) efficiently process DHA by glucuronidation.”

“Two ubiquitous carcinogenic factors that can be manipulated without toxins are the polyunsaturated fatty acids (PUFA) and estrogen. These closely interact with each other, and there are many ways in which they can be modulated.”

“Besides causing a general slowing of metabolism, aging and toxic PUFA have specific actions on the detoxifying system. The enzymes that help to detoxify PUFA and estrogen and serotonin are inhibited by both PUFA and estrogen. All systems, including blood vessels and the intestine, are made leaky by estrogen and the PUFA and their products.”

“But when the endogenous omega-9 fatty acids have been thoroughly displaced by dietary omega-6 and omega -3 fatty acids, the systemic release of fatty acids becomes an amplifier of the stress state initiated by injury or other stress. The liver, for example, decreases its detoxification of estrogen in the presence of polyunsaturated fatty acids.”

Drug Metab Dispos. 2002 May;30(5):531-3.
Glucuronidation of the dietary fatty acids, phytanic acid and docosahexaenoic acid, by human UDP-glucuronosyltransferases.
Little JM, Williams L, Xu J, Radominska-Pandya A.
Linoleic acid has recently been shown to be glucuronidated in vitro by human liver and intestinal microsomes and recombinant UGT2B7. In the present study, the dietary fatty acids (FA), phytanic acid (PA), and docosahexaenoic acid (DHA) have been used as substrates for human UDP-glucuronosyltransferases (UGTs). Both compounds were effectively glucuronidated by human liver microsomes (HLM; 1.25 +/- 0.36 and 1.12 +/- 0.32 nmol/mg x min for PA and DHA, respectively) and UGT2B7 (0.71 and 0.53 nmol/mg x min). Kinetic analysis produced relatively low K(m) values for PA with both HLM and UGT2B7 (149 and 108 microM, respectively). The K(m) for DHA glucuronidation by HLM (460 microM) was considerably higher than that for UGT2B7 (168 microM), suggesting the involvement in microsomes of other UGT isoforms in addition to UGT2B7. Glucuronidation of PA and DHA by gastrointestinal microsomes from 16 human subjects was determined. In general, both PA and DHA were glucuronidated by gastric and intestinal microsomes, and activity toward both substrates was lowest in the stomach, increased in the small intestine, and lower in the colon. However, there were large interindividual variations in UGT activity toward both substrates in all segments of the intestine, as has been seen with other substrates. Thus, PA and DHA are effective in vitro substrates for human liver, gastric and intestinal microsomes, and glucuronidation may play a role in modulating the availability of these FA as ligands for nuclear receptors.

Glucuronidation is inhibited by PUFA increasing accumulation of these fats with aging.

Biochem Pharmacol. 2004 Jan 1;67(1):191-9.
Evidence that unsaturated fatty acids are potent inhibitors of renal UDP-glucuronosyltransferases (UGT): kinetic studies using human kidney cortical microsomes and recombinant UGT1A9 and UGT2B7.
Tsoutsikos P, Miners JO, Stapleton A, Thomas A, Sallustio BC, Knights KM.
Renal ischaemia is associated with accumulation of fatty acids (FA) and mobilisation of arachidonic acid (AA). Given the capacity of UDP-glucuronosyltransferase (UGT) isoforms to metabolise both drugs and FA, we hypothesised that FA would inhibit renal drug glucuronidation. The effect of FA (C2:0-C20:5) on 4-methylumbelliferone (4-MU) glucuronidation was investigated using human kidney cortical microsomes (HKCM) and recombinant UGT1A9 and UGT2B7 as the enzyme sources. 4-MU glucuronidation exhibited Michaelis-Menten kinetics with HKCM (apparent K(m) (K(m)(app)) 20.3 microM), weak substrate inhibition with UGT1A9 (K(m)(app) 10.2 microM, K(si) 289.6 microM), and sigmoid kinetics with UGT2B7 (S(50)(app)440.6 microM) Similarly, biphasic UDP-glucuronic acid (UDPGA) kinetics were observed with HKCM (S(50) 354.3 microM) and UGT1A9 (S(50) 88.2 microM). In contrast, the Michaelis-Menten kinetics for UDPGA observed with UGT2B7 (K(m)(app) 493.2 microM) suggested that kinetic interactions with UGTs were specific to the xenobiotic substrate and the co-substrate (UDPGA). FA (C16:1-C20:5) significantly inhibited (25-93%) HKCM, UGT1A9 or UGT2B7 catalysed 4-MU glucuronidation. Although linoleic acid (LA) and AA were both competitive inhibitors of 4-MU glucuronidation by HKCM (K(i)(app) 6.34 and 0.15 microM, respectively), only LA was a competitive inhibitor of UGT1A9 (K(i)(app) 4.06 microM). In contrast, inhibition of UGT1A9 by AA exhibited atypical kinetics. These data indicate that LA and AA are potent inhibitors of 4-MU glucuronidation catalysed by human kidney UGTs and recombinant UGT1A9 and UGT2B7. It is conceivable therefore that during periods of renal ischaemia FA may impair renal drug glucuronidation thus compromising the protective capacity of the kidney against drug-induced nephrotoxicity.

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