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Cancer Res. 2016 Apr 1;76(7):2000-12. doi: 10.1158/0008-5472.CAN-15-1360. Epub 2016 Feb 3.
Aspirin Suppresses the Acquisition of Chemoresistance in Breast Cancer by Disrupting an NFκB-IL6 Signaling Axis Responsible for the Generation of Cancer Stem Cells.
Saha S, Mukherjee S, Khan P, Kajal K, Mazumdar M, Manna A, Mukherjee S, De S, Jana D, Sarkar DK, Das T
Acquired chemoresistance has curtailed cancer survival since the dawn of chemotherapy. Accumulating evidence suggests a major role for cancer stem cells (CSC) in chemoresistance, although their involvement in acquired resistance is still unknown. The use of aspirin has been associated with reduced cancer risk and recurrence, suggesting that the anti-inflammatory drug may exert effects on CSCs. In this study, we investigated the contribution of CSCs to acquired chemoresistance of breast cancer and the avenues for reversing such effects with aspirin. We observed that the residual risk of recurrence was higher in breast cancer patients who had acquired chemoresistance. Treatment of preexisting CSCs with a genotoxic drug combination (5-fluorouracil, doxorubicin, and cyclophosphamide) generated an NFκB-IL6-dependent inflammatory environment that imparted stemness to nonstem cancer cells, induced multidrug resistance, and enhanced the migration potential of CSCs. Treatment with aspirin prior to chemotherapy suppressed the acquisition of chemoresistance by perturbing the nuclear translocation of NFκB in preexisting CSCs. Therefore, disruptions to the NFκB-IL6 feedback loop prevented CSC induction and sensitized preexisting CSCs to chemotherapy. Collectively, our findings suggest that combining aspirin and conventional chemotherapy may offer a new treatment strategy to improve recurrence-free survival of breast cancer patients. Cancer Res; 76(7); 2000-12. ©2016 AACR.

Lab Invest. 2015 Jul;95(7):702-17. doi: 10.1038/labinvest.2015.49. Epub 2015 Apr 13.
Aspirin blocks growth of breast tumor cells and tumor-initiating cells and induces reprogramming factors of mesenchymal to epithelial transition.
Maity G, De A, Das A, Banerjee S, Sarkar S, Banerjee SK.
Acetylsalicylic acid (ASA), also known as aspirin, a classic, nonsteroidal, anti-inflammatory drug (NSAID), is widely used to relieve minor aches and pains and to reduce fever. Epidemiological studies and other experimental studies suggest that ASA use reduces the risk of different cancers including breast cancer (BC) and may be used as a chemopreventive agent against BC and other cancers. These studies have raised the tempting possibility that ASA could serve as a preventive medicine for BC. However, lack of in-depth knowledge of the mechanism of action of ASA reshapes the debate of risk and benefit of using ASA in prevention of BC. Our studies, using in vitro and in vivo tumor xenograft models, show a strong beneficial effect of ASA in the prevention of breast carcinogenesis. We find that ASA not only prevents breast tumor cell growth in vitro and tumor growth in nude mice xenograft model through the induction of apoptosis, but also significantly reduces the self-renewal capacity and growth of breast tumor-initiating cells (BTICs)/breast cancer stem cells (BCSCs) and delays the formation of a palpable tumor. Moreover, ASA regulates other pathophysiological events in breast carcinogenesis, such as reprogramming the mesenchymal to epithelial transition (MET) and delaying in vitro migration in BC cells. The tumor growth-inhibitory and reprogramming roles of ASA could be mediated through inhibition of TGF-β/SMAD4 signaling pathway that is associated with growth, motility, invasion, and metastasis in advanced BCs. Collectively, ASA has a therapeutic or preventive potential by attacking possible target such as TGF-β in breast carcinogenesis.

J Pharmacol Exp Ther. 1981 Aug;218(2):464-9.
Protective effects of aspirin in endotoxic shock.
Halushka PV, Wise WC, Cook JA.
Endotoxic shock is associated with increased metabolism of arachidonic acid to thromboxanes (TX) and prostaglandins (PG). This study assessed the effects of varied doses of aspirin, an inhibitor of arachidonic acid metabolism, on Salmonella enteriditis endotoxin (20 mg/kg)-induced mortality, plasma levels of arachidonate metabolites and other pathophysiological sequelae in Long-Evans rats. Aspirin, in doses of 3.75, 15 an 30 mg/kg, given 30 min before endotoxin significantly (P less than .01) improved 24-hr survival from 11% to 60 to 70%, but 100 mg/kg afforded no protection. Pretreatment with aspirin (15 or 100 mg/kg) 30 min before endotoxin significantly (P less than .001) decreased the endotoxin-induced elevations in plasma levels of immunoreactive (i) TXB2, a stable metabolite of TXA2, i6-keto PGF1 alpha, a stable metabolite of PGI2 and significantly (P less than .05) inhibited thrombin-induced in vitro platelet iTXB2 synthesis. Endotoxin-induced hypoglycemia and elevations in serum acid phosphatase and beta-glucuronidase activities, lysosomal enzymes, were all significantly (P less than .01) attenuated by pretreatment with aspirin (15 mg/kg) 30 min before endotoxin. Aspirin (15 or 100 mg/kg) given 24 h before challenge with endotoxin significantly improved 24-hr survival to 42 (P less than .01) and 44% (P less than .005), respectively. Although 24 hr pretreatment with aspirin (15 or 100 mg/kg) significantly (P less than .001) reduced endotoxin-induced elevations in iTXB2, only the 100 mg/kg dose significantly lowered plasma levels of i6-keto PGF1 alpha. These observations are consistent with the notion that the beneficial effects of aspirin seen in experimental endotoxic shock may be mediated, in part, via reduction of platelet TXA2 synthesis.

J Clin Invest. 2002 May;109(10):1321-6.
Mechanism by which high-dose aspirin improves glucose metabolism in type 2 diabetes.
Hundal RS1, Petersen KF, Mayerson AB, Randhawa PS, Inzucchi S, Shoelson SE, Shulman GI.
Recent studies have implicated fatty acid-dependent activation of the serine kinase IKKbeta, which plays a key role in tissue inflammation, in the pathogenesis of insulin resistance. High doses of salicylates have recently been shown to inhibit IKKbeta activity and might therefore ameliorate insulin resistance and improve glucose tolerance in patients with type 2 diabetes. To test this hypothesis, we studied nine type 2 diabetic subjects before and after 2 weeks of treatment with aspirin ( approximately 7 g/d). Subjects underwent mixed-meal tolerance tests and hyperinsulinemic-euglycemic clamps with [6,6-(2)H2]glucose to assess glucose turnover before and after treatment. High-dose aspirin treatment resulted in a approximately 25% reduction in fasting plasma glucose, associated with a approximately 15% reduction in total cholesterol and C-reactive protein, a approximately 50% reduction in triglycerides, and a approximately 30% reduction in insulin clearance, despite no change in body weight. During a mixed-meal tolerance test, the areas under the curve for plasma glucose and fatty acid levels decreased by approximately 20% and approximately 50%, respectively. Aspirin treatment also resulted in a approximately 20% reduction in basal rates of hepatic glucose production and a approximately 20% improvement in insulin-stimulated peripheral glucose uptake under matched plasma insulin concentrations during the clamp. In conclusion, these data support the hypothesis that IKKbeta represents a new target for treating type 2 diabetes mellitus.

Am J Med. 1983 Jun 14;74(6A):91-6.
Studies on the beneficial effects of aspirin in endotoxic shock. Relationship to inhibition of arachidonic acid metabolism.
Halushka PV, Wise WC, Cook JA.
Endotoxic shock is associated with increased metabolism of arachidonic acid into thromboxanes and prostaglandins. This study assessed the effects of varied doses of aspirin, an inhibitor of arachidonic acid metabolism, on Salmonella enteritidis endotoxin-induced mortality, plasma levels of arachidonate metabolites, and other pathophysiologic sequelae in Long-Evans rats. Aspirin in doses of 3.75, 15, and 30 mg/kg given 30 minutes prior to endotoxin challenge significantly (p less than 0.01) improved 24-hour survival rates from 11 percent to approximately 65 percent, but 100 mg/kg afforded no protection. Pretreatment with aspirin (15 or 100 mg/kg) 30 minutes prior to endotoxin also significantly (p less than 0.001) decreased the endotoxin-induced elevations in plasma levels of immunoreactive thromboxane B2, a stable metabolite of thromboxane A2, and immunoreactive 6-keto prostaglandin F1 alpha, a stable metabolite of prostacyclin. Aspirin doses of 15 or 100 mg/kg given 24 hours prior to challenge with endotoxin significantly improved 24-hour survival rates from 17 percent to 42 percent (p less than 0.01) and 44 percent (p less than 0.005), respectively. Pretreatment with an aspirin dose of 15 mg/kg 24 hours prior to challenge with endotoxin significantly (p less than 0.05) inhibited thrombin-induced immunoreactive thromboxane B2 synthesis in platelet-rich plasma (in vitro) and endotoxin-induced immunoreactive 6-keto-prostaglandin F1 alpha and immunoreactive thromboxane B2 synthesis by rat peritoneal macrophages. Although 24-hour pretreatment with aspirin (15 or 100 mg/kg) significantly (p less than 0.001) reduced endotoxin-induced elevations in immunoreactive thromboxane B2, only the 100 mg/kg dose significantly lowered plasma levels of immunoreactive 6-keto prostaglandin F1 alpha. These observations are consistent with the notion that the beneficial effects of aspirin seen in experimental endotoxic shock may be mediated, in part, via inhibition of arachidonic acid metabolism.

J Physiol Paris. 2001 Jan-Dec;95(1-6):51-7.
Protection by aspirin of indomethacin-induced small intestinal damage in rats: mediation by salicylic acid.
Takeuchi K, Hase S, Mizoguchi H, Komoike Y, Tanaka A.
Most of non-steroidal anti-inflammatory drugs (NSAIDs) except aspirin (ASA) produce intestinal damage in rats. In the present study, we re-examined the intestinal toxic effect of ASA in rats, in comparison with various NSAIDs, and investigated why ASA does not cause damage in the small intestine, in relation to its metabolite salicylic acid (SA). Various NSAIDs (indomethacin; 10 mg/kg; flurbiprofen; 20 mg/kg; naproxen; 40 mg/kg; dicrofenac; 40 mg/kg; ASA; 20-200 mg/kg) were administered s.c., and the small intestinal mucosa was examined macroscopically 24 h later. All NSAIDs tested, except ASA, caused hemorrhagic lesions in the small intestine, with a decrease of mucosal PGE(2) contents. ASA did not provoke any damage, despite inhibiting (prostaglandin) PG production, and prevented the occurrence of intestinal lesions induced by indomethacin, in a dose-related manner. This protective action of ASA was mimicked by the equimolar doses of SA (17.8-178 mg/kg). Indomethacin caused intestinal hypermotility, in preceding to the occurrence of lesion, and this event was followed by increases of enterobacterial translocation in the mucosa. Both ASA and SA prevented both the intestinal hypermotility and the bacterial translocation seen after indomethacin treatment. In addition, the protective effect of SA was not significantly influenced by either the adenosine deaminase or the adenosine receptor antagonists. Following administration of ASA, the blood SA levels reached a peak within 30 min and remained elevated for more than 7 h. These results suggest that SA has a cytoprotective action against indomethacin-induced small intestinal lesions, and this action may be associated with inhibition of the intestinal hypermotility and the bacterial translocation, but not mediated by endogenous adenosine. Failure of ASA to induce intestinal damage may be explained, at least partly, by a protective action of SA, the metabolite of ASA.

Proc Soc Exp Biol Med. 1985 Feb;178(2):250-3.
Salicylic acid blocks indomethacin-induced cyclooxygenase inhibition and lesion formation in rat gastric mucosa.
Ligumsky M, Guth PH, Elashoff J, Kauffman GL Jr, Hansen D, Paulsen G.
Salicylic acid has been shown to decrease gastric mucosal lesions induced by indomethacin in the rat. In vitro, it has also been shown to counteract the inhibitory effect of indomethacin and aspirin on the cyclooxygenase enzyme system in seminal vesicle microsomes and in platelets and vascular tissue. The hypothesis that the mechanism of salicylic acid “protection” against indomethacin-induced gastric lesions involves interference with indomethacin-induced mucosal cyclooxygenase inhibition was tested. Male, fasted rats were treated with intragastric salicylic acid in doses of 50, 100, 200, 300, or 400 mg/kg concomitantly with a sc injection of 20 mg/kg of indomethacin. Gastric mucosal lesions and mucosal cyclooxygenase activity (as measured by ex vivo prostaglandin F2 alpha synthesis) were examined 3 hr later. Intragastric salicylic acid, 200-400 mg/kg, significantly reduced indomethacin-induced lesion formation, while counteracting significantly indomethacin inhibition of prostaglandin synthesis. Salicylic acid alone did not significantly change cyclooxygenase activity. It is concluded that topical salicylic acid can decrease indomethacin-induced gastric mucosal lesion in the rat, in part, by counteracting the inhibitory effect of indomethacin at the cyclooxygenase level.

Lancet. 2011 Jan 1;377(9759):31-41. Epub 2010 Dec 6.
Effect of daily aspirin on long-term risk of death due to cancer: analysis of individual patient data from randomised trials.
Rothwell PM, Fowkes FG, Belch JF, Ogawa H, Warlow CP, Meade TW.
BACKGROUND:
Treatment with daily aspirin for 5 years or longer reduces subsequent risk of colorectal cancer. Several lines of evidence suggest that aspirin might also reduce risk of other cancers, particularly of the gastrointestinal tract, but proof in man is lacking. We studied deaths due to cancer during and after randomised trials of daily aspirin versus control done originally for prevention of vascular events.
METHODS:
We used individual patient data from all randomised trials of daily aspirin versus no aspirin with mean duration of scheduled trial treatment of 4 years or longer to determine the effect of allocation to aspirin on risk of cancer death in relation to scheduled duration of trial treatment for gastrointestinal and non-gastrointestinal cancers. In three large UK trials, long-term post-trial follow-up of individual patients was obtained from death certificates and cancer registries.
RESULTS:
In eight eligible trials (25 570 patients, 674 cancer deaths), allocation to aspirin reduced death due to cancer (pooled odds ratio [OR] 0·79, 95% CI 0·68-0·92, p=0·003). On analysis of individual patient data, which were available from seven trials (23 535 patients, 657 cancer deaths), benefit was apparent only after 5 years’ follow-up (all cancers, hazard ratio [HR] 0·66, 0·50-0·87; gastrointestinal cancers, 0·46, 0·27-0·77; both p=0·003). The 20-year risk of cancer death (1634 deaths in 12 659 patients in three trials) remained lower in the aspirin groups than in the control groups (all solid cancers, HR 0·80, 0·72-0·88, p<0·0001; gastrointestinal cancers, 0·65, 0·54-0·78, p<0·0001), and benefit increased (interaction p=0·01) with scheduled duration of trial treatment (≥7·5 years: all solid cancers, 0·69, 0·54-0·88, p=0·003; gastrointestinal cancers, 0·41, 0·26-0·66, p=0·0001). The latent period before an effect on deaths was about 5 years for oesophageal, pancreatic, brain, and lung cancer, but was more delayed for stomach, colorectal, and prostate cancer. For lung and oesophageal cancer, benefit was confined to adenocarcinomas, and the overall effect on 20-year risk of cancer death was greatest for adenocarcinomas (HR 0·66, 0·56-0·77, p<0·0001). Benefit was unrelated to aspirin dose (75 mg upwards), sex, or smoking, but increased with age-the absolute reduction in 20-year risk of cancer death reaching 7·08% (2·42-11·74) at age 65 years and older.
INTERPRETATION:
Daily aspirin reduced deaths due to several common cancers during and after the trials. Benefit increased with duration of treatment and was consistent across the different study populations. These findings have implications for guidelines on use of aspirin and for understanding of carcinogenesis and its susceptibility to drug intervention.

Cancer Epidemiol Biomarkers Prev. 2009 Oct;18(10):2726-33. Epub 2009 Sep 15.
Cyclooxygenase-2 polymorphisms, aspirin treatment, and risk for colorectal adenoma recurrence–data from a randomized clinical trial.
Barry EL, Sansbury LB, Grau MV, Ali IU, Tsang S, Munroe DJ, Ahnen DJ, Sandler RS, Saibil F, Gui J, Bresalier RS, McKeown-Eyssen GE, Burke C, Baron JA.
Cyclooxygenase-2 (COX-2) catalyzes the rate-limiting step in the production of prostaglandins, potent mediators of inflammation. Chronic inflammation plays an important role in the development and progression of colorectal cancer. Aspirin inhibits COX-2 activity and lowers the risk for colorectal adenomas and cancer. We investigated whether common genetic variation in COX-2 influenced risk for colorectal adenoma recurrence among 979 participants in the Aspirin/Folate Polyp Prevention Study who were randomly assigned to placebo or aspirin and followed for 3 years for the occurrence of new adenomas. Of these participants, 44.2% developed at least one new adenoma during follow-up. Adjusted relative risks and 95% confidence intervals (95% CI) were calculated to test the association between genetic variation at six COX-2 single-nucleotide polymorphisms and adenoma occurrence and interaction with aspirin treatment. Two single-nucleotide polymorphisms were significantly associated with increased adenoma recurrence: for rs5277, homozygous carriers of the minor C allele had a 51% increased risk compared with GG homozygotes (relative risk, 1.51; 95% CI, 1.01-2.25), and for rs4648310, heterozygous carriers of the minor G allele had a 37% increased risk compared with AA homozygotes (relative risk, 1.37; 95% CI, 1.05-1.79). (There were no minor allele homozygotes.) In stratified analyses, there was suggestive evidence that rs4648319 modified the effect of aspirin. These results support the hypothesis that COX-2 plays a role in the etiology of colon cancer and may be a target for aspirin chemoprevention and warrant further investigation in other colorectal adenoma and cancer populations.

Aspirin, which stimulates bones formation, has other thyroid-like actions, including activation of mitochondrial respiration and energy production, with an increase of cytochrome c oxidase (Cai, et al., 1996)., and it lower serotonin (Shen, et al., 2011). It also apparently protects against calcification of soft tissues, (Vasudev, et al., 2000), though there has been surprisingly little investigation of that. -Ray Peat, PhD

Arch Biochem Biophys. 1996 Jan 1;325(1):107-12.
Thyromimetic action of the peroxisome proliferators clofibrate, perfluorooctanoic acid, and acetylsalicylic acid includes changes in mRNA levels for certain genes involved in mitochondrial biogenesis.
Cai Y, Nelson BD, Li R, Luciakova K, dePierre JW.
The effects of three peroxisome proliferators on the mRNA levels for some mitochondrial inner-membrane proteins in rat liver were investigated. Clofibrate, perfluorooctanoic acid, and acetylsalicylic acid all increased the mRNA levels for the mitochondrial-encoded respiratory-chain components cytochrome c oxidase subunit I and NADH dehydrogenase subunit I. Mitochondrial 16S rRNA was also induced by clofibrate. The mRNA levels for the nuclear-encoded mitochondrial inner-membrane proteins adenine nucleotide translocator and cytochrome c1 were selectively induced by the different peroxisome proliferators. Malic enzyme, which is induced by thyroid hormone, was also induced by the three peroxisome proliferators tested. These effects are in some ways similar to those obtained with thyroid hormone.

Cell Biochem Biophys. 2011 Sep;61(1):23-31.
Aspirin attenuates pulmonary arterial hypertension in rats by reducing plasma 5-hydroxytryptamine levels.
Shen L, Shen J, Pu J, He B.
Pulmonary arterial hypertension (PAH) is characterized by increasing pulmonary pressure, right ventricular failure, and death. The typical pathological changes include medial hypertrophy, intimal fibrosis and in situ thrombosis. Serotonin (5-HT) and other factors contribute to the development of pathologic lesions. Aspirin (ASA), a platelet aggregation inhibitor, inhibits 5-HT release from platelets. The aim of this study was to determine the efficacy of ASA in preventing or attenuating PAH. Sprague-Dawley rats injected with monocrotaline (MCT) developed severe PAH within 31 days. One hundred forty rats were randomized to receive either vehicle or ASA (0.5, 1, 2, or 4 mg/kg/day). The pre-ASA group was treated with ASA (1 mg/kg/day) for 30 days before the MCT injection. Thirty-one days after the injection (day 61 for the pre-ASA group), pulmonary arterial pressure (PAP), right ventricular hypertrophy and pulmonary arteriole thickness were measured. Plasma 5-HT was measured by high-performance liquid chromatography. Aspirin suppressed PAH and increased the survival rate compared with the control group (84 vs. 60%, P < 0.05). Aspirin treatment also reduced right ventricular hypertrophy and pulmonary arteriole proliferation in ASA-treated PAH model. In addition, plasma 5-HT was decreased in our ASA-treated PAH model. The degree of 5-HT reduction was associated with systolic PAP, right ventricular hypertrophy and wall thickness of pulmonary arterioles in rats. These results showed that ASA treatment effectively attenuated MCT-induced pulmonary hypertension, right ventricular hypertrophy, and occlusion of the pulmonary arteries. The effects of ASA was associated with a reduction of 5-HT.

Artif Organs. 2000 Feb;24(2):129-36.
Synergistic effect of released aspirin/heparin for preventing bovine pericardial calcification.
Vasudev SC, Chandy T, Sharma CP, Mohanty M, Umasankar PR.
Calcification is a frequent cause of the clinical failure of bioprosthetic heart valves fabricated from glutaraldehyde pretreated bovine pericardium (GATBP). Aspirin, a potent antiplatelet drug, and heparin, an anticoagulant, are commonly used for postimplant complications such as thrombosis and thromboembolism. Aspirin and heparin were embedded in chitosan/polyethylene vinylacetate co-matrix to develop a prolonged release form. The effect of these drugs towards the bioprosthetic calcification was investigated by in vitro and in vivo models. In vitro and in vivo evaluation suggest that the released aspirin/heparin from the co-matrix had a synergistic effect in inhibiting GATBP calcification. In vivo subcutaneous co-implantation was performed with PEG-20,000 grafted bovine pericardium (PEG-GABP), aspirin, and heparin. Biochemical, histological, and scanning electron microscopic evaluation of retrieved samples demonstrated a significant reduction in calcium deposition and alkaline phosphatase activity on PEG-GABP compared to GATBP. It seems that the aspirin/heparin combination synergistically inhibits the pericardial calcification in addition to their antithrombotic function.

Zhonghua Yi Xue Za Zhi. 2011 Apr 5;91(13):925-9.
[Effect of aspirin administration for the treatment of osteoporosis in ovariectomized rat model].
[Article in Chinese]
Chen ZW, Wu ZX, Sang HX, Qin GL, Wang LS, Feng J, Wang J, Li XJ, Wang JC, Zhang D.
OBJECTIVE:
To explore the therapeutic effects of aspirin on postmenopausal osteoporosis and understand its action mechanism.
METHODS:
Forty three-month-old female SD rats were randomly divided into 5 groups (n = 8 each): sham group, OVX group and aspirin groups (A1, A2 & A3). The OVX and aspirin groups were ovariectomized (OVX). All rats underwent BMD (bone mineral density) scan at the time of OVX and 3 months after OVX. After the animal model of osteoporosis was established, aspirin groups were intragastrically administered at a dose of 8.93 mg×kg(-1)×d(-1) (A1), 26.79 mg×kg(-1)×d(-1) (A2) and 80.36 mg×kg(-1)×d(-1) (A3) daily in OVX rats. Three months later, the BMD and micro-architecture of vertebrae were measured by dual-energy X-ray absorptiometry and microtomography. Alkaline phosphatase and osteocalcin were measured in peripheral blood. The trabecular architecture changes were observed by histomorphology. Axial compression tests were used to evaluate the mechanical properties of vertebral specimens and three-point bending tests used for femur shaft.
RESULTS:
Three months after ovariectomy, BMD was significantly lower than preoperative. BMD in aspirin treated groups was significantly higher than that in OVX group. Alkaline phosphatase in peripheral blood decreased significantly in aspirin groups than those in OVX, but osteocalcin had no significant difference between aspirin and OVX groups. The microtomography reconstruction analysis also showed that the trabecular thickness, trabecular number and BMD increased significantly in aspirin groups than those in OVX. And there was no significant difference between A3 and sham groups. The results of biomechanical test showed that the maximum compression load of lumbar spine and three-point bending load of femur shaft were significantly higher in aspirin groups than those in OVX group.
CONCLUSION:
Aspirin can promote trabecular bone remodeling, improve three-dimensional structure of trabecular bone and increase bone density of cancellous in osteoporotic rats by stimulating bone formation. It may become a new drug for the treatment of osteoporosis.

Neuropharmacology. 2000 Apr 27;39(7):1309-18.
Mechanisms of the neuroprotective effect of aspirin after oxygen and glucose deprivation in rat forebrain slices.
Moro MA, De Alba J, Cárdenas A, De Cristóbal J, Leza JC, Lizasoain I, Díaz-Guerra MJ, Boscá L, Lorenzo P.
Acetylsalicylic acid (ASA, Aspirin) is an anti-inflammatory drug with a wide spectrum of pharmacological activities and multiple sites of action. Apart from its preventive actions against stroke due to its antithrombotic properties, recent data in the literature suggest that high concentrations of ASA also exert direct neuroprotective effects. We have used an in vitro model of brain ischaemia using rat forebrain slices deprived of oxygen and glucose to test ASA neuroprotective properties. We have found that ASA inhibits neuronal damage at concentrations lower than those previously reported (0.1-0.5 mM), and that these effects correlate with the inhibition of excitatory amino acid release, of NF-kappaB translocation to the nucleus and iNOS expression caused by ASA. All of these three mechanisms may mediate the neuroprotective effects of this drug. Our results also show that the effects of ASA are independent of COX inhibition. Taken together, our present findings show that ASA is neuroprotective in an in vitro model of brain ischaemia at doses close to those recommended for its antithrombotic effects.

J Neurochem. 2001 Oct;79(2):456-9.
Neuroprotective effect of aspirin by inhibition of glutamate release after permanent focal cerebral ischaemia in rats.
De Cristóbal J, Moro MA, Dávalos A, Castillo J, Leza JC, Camarero J, Colado MI, Lorenzo P, Lizasoain I.
Aspirin reduces the size of infarcts after ischaemic stroke. Although this fact has been attributed to its anti-platelet actions, direct neuroprotective effects have also been reported. We have recently demonstrated that aspirin is neuroprotective by inhibiting glutamate release in ‘in vitro’ models of brain ischaemia, via an increase in ATP production. The present study was designed to determine whether the inhibition of glutamate release induced by aspirin might be protective in a whole-animal model of permanent focal brain ischaemia. Focal brain ischaemia was produced in male adult Fischer rats by occluding both the common carotid and middle cerebral arteries. Central and serum glutamate levels were determined at fixed intervals after occlusion. The animals were then killed and infarct volume was measured. Aspirin (30 mg/kg i.p. administered 2 h before the occlusion) produced a significant reduction in infarct volume, an effect that correlated with the inhibition caused by aspirin on ischaemia-induced increase in brain and serum glutamate concentrations after the onset of the ischaemia. Aspirin also inhibited ischaemia-induced decrease in brain ATP levels. Our present findings show a novel mechanism for the neuroprotective effects of aspirin, which takes place at concentrations in the anti-aggregant-analgesic range, useful in the management of patients with risk of ischaemic events.

Stroke. 2002 Jan;33(1):261-7.
Inhibition of glutamate release via recovery of ATP levels accounts for a neuroprotective effect of aspirin in rat cortical neurons exposed to oxygen-glucose deprivation.
De Cristóbal J, Cárdenas A, Lizasoain I, Leza JC, Fernández-Tomé P, Lorenzo P, Moro MA.
BACKGROUND AND PURPOSE:
Aspirin is preventive against stroke not only because of its antithrombotic properties but also by other direct effects. The aim of this study was to elucidate its direct neuroprotective effects.
METHODS:
Viability parameters, glutamate release and uptake, and ATP levels were measured in cultured cortical neurons exposed to oxygen-glucose deprivation (OGD). In addition, ATP levels and oxygen consumption were studied in isolated brain mitochondria or submitochondrial particles.
RESULTS:
Aspirin inhibited OGD-induced neuronal damage at concentrations lower (0.3 mmol/L) than those reported to act via inhibition of the transcription factor nuclear factor-kappaB (which are >1 mmol/L), an effect that correlated with the inhibition caused by aspirin on glutamate release. This effect was shared by sodium salicylate but not by indomethacin, thus excluding the involvement of cyclooxygenase. A pharmacological dissection of the components involved indicated that aspirin selectively inhibits the increase in extracellular glutamate concentration that results from reversal of the glutamate transporter, a component of release that is due to ATP depletion. Moreover, aspirin-afforded neuroprotection occurred in parallel with a lesser decrease in ATP levels after OGD. Aspirin elevated ATP levels not only in intact cortical neurons but also in isolated brain mitochondria, an effect concomitant with an increase in NADH-dependent respiration by brain submitochondrial particles.
CONCLUSIONS:
Taken together, our present findings show a novel mechanism for the neuroprotective effects of aspirin, which takes place at concentrations in the antithrombotic-analgesic range, useful in the management of patients with high risk of ischemic events.

An R Acad Nac Med (Madr). 2002;119(2):311-20; discussion 320-6.
[Neuroprotection by aspirin in cerebrovascular pathology].
[Article in Spanish]
Lorenzo Fernández P.
Aspirin reduces the size of infarcts after ischaemic stroke. Although this fact has been attributed to its antiplatelet actions, direct neuroprotective effects have been also reported. We have demonstrated that aspirin is neuroprotective by inhibiting glutamate release in “in vitro” models of brain ischaemia. A pharmacological dissection of the components involved, using cell cortical culture exposed to oxygen-glucose deprivation, indicated that aspirin selectively inhibits the increase in extracellular glutamate concentration which results from reversal of the glutamate transporter, a component of release which is due to ATP depletion. Moreover, the neuroprotection afforded by aspirin occurred in parallel to a lesser decay in ATP levels after OGD. Aspirin not only elevated ATP levels in intact cortical neurones, but also in isolated brain mitochondria. On the other hand, using a whole-animal model of permanent focal brain ischaemia (MCAO; middle cerebral artery occlusion), wed have also demonstrated that aspirin (30 mg/Kg i.p. administered 2 h before the occlusion) produced a significant reduction in infarct volume, effect that correlated with the inhibition caused by aspirin on ischaemia-induced increase in brain and serum glutamate concentrations after the onset of the ischaemia. Aspirin also inhibited ischaemia-induced decrease in brain ATP levels. Finally, clinical studies showed that prior treatment with aspirin was associated with a lower risk of neurological deterioration after acute ischemic stroke which was related to a reduction of glutamate release. Our present findings show a novel mechanism for the neuroprotective effects of aspirin, that takes place at concentrations in the antiaggregant-analgesic range, useful in the management of patients with risk of ischaemic events.

Science 31 August 2001: Vol. 293 no. 5535 pp. 1673-1677
Reversal of Obesity- and Diet-Induced Insulin Resistance with Salicylates or Targeted Disruption of Ikkβ
Minsheng Yuan, Nicky Konstantopoulos1, Jongsoon Lee1, Lone Hansen, Zhi-Wei Li, Michael Karin and Steven E. Shoelson
We show that high doses of salicylates reverse hyperglycemia, hyperinsulinemia, and dyslipidemia in obese rodents by sensitizing insulin signaling. Activation or overexpression of the IκB kinase β (IKKβ) attenuated insulin signaling in cultured cells, whereas IKKβ inhibition reversed insulin resistance. Thus, IKKβ, rather than the cyclooxygenases, appears to be the relevant molecular target. Heterozygous deletion (Ikkβ +/−) protected against the development of insulin resistance during high-fat feeding and in obese Lepob/ob mice. These findings implicate an inflammatory process in the pathogenesis of insulin resistance in obesity and type 2 diabetes mellitus and identify the IKKβ pathway as a target for insulin sensitization.

Toxicology. 1982;24(1):33-43.
Inhibition of hepatic lipogenesis by salicylate.
Beynen AC, Buechler KF, van der Molen AJ, Geelen MJ
Salicylate has been found to be an inhibitor of fatty acid synthesis in isolated rat hepatocytes. Half-maximal inhibition of fatty acid synthesis occurs at approximately 2 mM. The inhibitory effect of salicylate on fatty acid synthesis is not relieve by the addition of acetate, suggesting that salicylate inhibits the conversion of acetate into fatty acids. Acetyl-CoA carboxylase activity in homogenates of hepatocytes is not influenced by previous exposure of the intact cells to salicylate. Partially purified acetyl-CoA carboxylase, isolated and assayed in the absence of citrate, is markedly inhibited by salicylate. However, in the presence of 0.5 mM citrate, which is the concentration of this metabolite in the cytosol of the liver cell, salicylate activates the enzyme. Upon treatment of acetyl-CoA carboxylase with salicylate (in the absence or presence of citrate), followed by separation of enzyme and effector on a Sephadex G-25 column, the enzyme activity is enhanced as compared to the salicylate-free control, demonstrating that the inhibitory effect of salicylate (in the absence of citrate) is reversible, but not the stimulatory effect (in the presence of citrate). Salicylate inhibition of fatty acid synthesis by hepatocytes is not rapidly reversible; hepatocytes preincubated with salicylate followed by a wash procedure (centrifugation and resuspension) still show depressed rates of fatty acid synthesis from acetate upon further incubation. Salicylate was found to prevent pyruvate accumulation in hepatocyte suspensions observed in the absence of this compound; salicylate even induces the disappearance of pyruvate and lactate initially present in the cell suspension. This suggests that salicylate activates pyruvate and lactate consumption, which is most likely related to the well-known fact that salicylate uncouples oxidative phosphorylation. The latter action of the drug will stimulate citric acid-cycle activity. This causes an inhibition of fatty acid and cholesterol synthesis since acetyl units will be specifically channelled into the citric acid cycle and not into the lipogenic pathway. It is concluded that part of the inhibitory effect of salicylate on fatty acid biosynthesis is exerted at (a) step(s) in the conversion of acetate into fatty acids, acetyl-CoA carboxylase not being a target of this compound. In addition, salicylate prevents that pyruvate, generated by glycolysis, enters the lipogenic pathway. The latter effect of salicylate would also explain the observed inhibition of cholesterol synthesis by this compound.

Fertil Steril. 1999 May;71(5):825-9.
Low-dose aspirin treatment improves ovarian responsiveness, uterine and ovarian blood flow velocity, implantation, and pregnancy rates in patients undergoing in vitro fertilization: a prospective, randomized, double-blind placebo-controlled assay.
Rubinstein M, Marazzi A, Polak de Fried E.
OBJECTIVE:
To determine the effects of low-dose aspirin on ovarian response, uterine and ovarian blood flow velocity, and implantation and pregnancy rates in patients undergoing IVF.
DESIGN:
Prospective, randomized, double-blind placebo-controlled assay.
SETTING:
Department of Reproductive Medicine, CER Medical Institute, Buenos Aires, Argentina.
PATIENT(S):
Two hundred ninety-eight infertile patients (mean [+/- SDI age, 35.6+/-4.09 years) undergoing IVF cycles.
INTERVENTION(S):
In the treatment group, 149 patients underwent controlled ovarian hyperstimulation and received a daily dose of 100 mg of aspirin. In the control group, 149 patients underwent controlled ovarian hyperstimulation in association with placebo.
MAIN OUTCOME MEASURE(S):
Number of follicles, number of oocytes retrieved, serum E2 levels, uterine and ovarian pulsatility index, cancellation rate, number of embryos transferred, and implantation and pregnancy rates.
RESULT(S):
There were statistically significant differences between the treatment group and the control group, respectively, in the number of follicles (19.8+/-7.2 versus 10.2+/-5.3), number of oocytes retrieved (16.2+/-6.7 versus 8.6+/-4.6), serum E2 levels (2,923.8+/-1,023.4 versus 1,614.3+/-791.7 pg/mL), uterine pulsatility index (1.22+/-0.34 versus 1.96+/-0.58), ovarian pulsatility index (1.18+/-0.31 versus 1.99+/-0.56), pregnancy rate (45% versus 28%), and implantation rate (17.8% versus 9.2%).
CONCLUSION(S):
Low-dose aspirin treatment significantly improves ovarian responsiveness, uterine and ovarian blood flow velocity, and implantation and pregnancy rates in IVF patients.

J Formos Med Assoc. 1997 Apr;96(4):253-7.
Aspirin improves uterine blood flow in the peri-implantation period.
Kuo HC, Hsu CC, Wang ST, Huang KE.
A prospective clinical study at the Infertility Clinic of National Cheng Kung University Hospital investigated the effect of aspirin on infertile women with impaired uterine perfusion. A total of 127 women with unexplained infertility or repeated failure with various assisted-conception techniques were enrolled. Uterine perfusion was assessed by Doppler ultrasound and classified as normal or impaired (pulsatility index < 3.0 or > or = 3.0, respectively). One-third (43/127) of the women were found to have impaired uterine perfusion during their menstrual cycles. Those with impaired uterine blood flow were given aspirin (100 mg/day) starting on day 3 of the next ovulatory cycle. Only 36 women completed both the screening and the aspirin-treated cycles. The pulsatility index was measured in the natural and aspirin-treated cycles in the same group of women and compared using repeated measures analyses of variance. A significant improvement in the uterine blood perfusion (p < 0.05) was detected on the day leutinizing hormone peaked and in the midluteal phase (peri-implantation period) of aspirin-treated cycles. Thus, the use of low-dose aspirin may improve uterine perfusion in women with unexplained infertility and impaired uterine blood flow.

Hum Reprod. 1994 Oct;9(10):1954-7.
The benefits of low-dose aspirin therapy in women with impaired uterine perfusion during assisted conception.
Wada I, Hsu CC, Williams G, Macnamee MC, Brinsden PR.
The objective of this long-running study was to determine whether the addition of low-dose aspirin to a standard hormone replacement therapy (HRT) protocol improved uterine perfusion during assisted conception. A total of 99 women scheduled for frozen embryo replacement were studied. Endometrial preparation was with a standard buserelin/HRT protocol. Uterine perfusion was assessed by Doppler ultrasound and classified as impaired or normal. In their first attempts, those with impaired perfusion (group I, n = 37) received low doses of aspirin [150 mg (n = 26) or 300 mg daily (n = 11)], starting from day 13 of HRT. Women with normal perfusion (group II) did not receive aspirin. In subsequent attempts, those from group I were arbitrarily allocated to start aspirin on day 1 or day 13 of HRT, and 10 women from group II were arbitrarily selected to receive aspirin from day 1 of HRT. In group I, the cancellation (46 versus 36%) and pregnancy rates (15 versus 25%) in those who received 150 or 300 mg aspirin daily were similar. In those with cancelled first attempts, good perfusion was achieved in 82 versus 20% (P < 0.02) of subsequent attempts using aspirin from day 1 versus day 13 of HRT. Higher pregnancy rates (47 versus 17%) were achieved in those taking aspirin from day 1 of HRT. In group II, pregnancy rates were not statistically different in those who did or did not receive aspirin during their subsequent attempts (10 versus 35%). The addition of low-dose aspirin to a standard HRT protocol in women with impaired uterine perfusion is associated with improved blood flow and satisfactory pregnancy rates.

Presse Med. 1996 Jan 6-13;25(1):31-6.
[Aspirin during pregnancy. Indications and modalities of prescription after the publication of the later trials].
[Article in French]
Uzan S, Merviel P, Beaufils M, Bréart G, Salat-Baroux J.
Aspirin, an inhibitor of cyclo-oxygenase, is prescribed in a number of conditions related to abnormal production of prostaglandins including gravidic hypertension. Results of the most recent trials demonstrate that in patients with a past history of pre-eclampsia or intra-uterine growth retardation, a pathological Doppler examination of the uterus, a pathological angiotensin test or an antiphospholipid syndrome, prescription of aspirin at the dose of 100 mg/day can prevent recurrence or development of pre-eclampsia or intra-uterine growth retardation. Treatment should begin as soon as possible during pregnancy, certainly before development of clinical manifestations. After history taking and identification of possible contraindications, bleeding time (Ivy method) is recorded before and after prescription and should be lower than 8 minutes. In case bleeding time exceeds 10 minutes 10 to 15 days after initiating aspirin, doses may be reduced to 50 mg per day or even 50 mg every two or three days to reach the target level. Treatment should generally be continued up to 36 weeks gestation.

Croat Med J. 2005 Oct;46(5):826-31.
Usefulness of aspirin therapy in high-risk pregnant women with abnormal uterine artery Doppler ultrasound at 14-16 weeks pregnancy: randomized controlled clinical trial.
Ebrashy A, Ibrahim M, Marzook A, Yousef D.
AIM:
To assess the effectiveness of low-dose aspirin in the prevention of preeclampsia and intrauterine growth restriction (IUGR) in high-risk pregnant women with abnormal findings at uterine artery Doppler velocimetry performed at 14-16 weeks.
DESIGN:
Randomized controlled clinical trial.
SETTING:
Department of Obstetrics and Gynecology, Faculty of Medicine, Cairo University, Egypt.
METHODS:
The trial enrolled 139 women at risk of preeclampsia or IUGR, with abnormal uterine artery Doppler findings that included the presence of unilateral or bilateral diastolic notch, high resistance index (RI), or high pulsatility index (PI) at 14-16 weeks of gestation. The women were randomly allocated into two groups, one receiving aspirin since admission to hospital (n=74) and the other serving as control (n=65). All women were followed up until delivery to assess maternal and perinatal outcomes. T-test was used for comparison of quantitative variables, and categorical variables were compared by chi2 test.
OUTCOME CRITERIA: Development of mild or severe preeclampsia, time of onset of preeclampsia, preterm delivery, and the development of IUGR.
RESULTS:
Preeclampsia developed in 35% of women receiving aspirin and 62% of women in the control group (P=0.003), with severe preeclampsia developing in 8% and 23% of women (P=0.215), respectively. Preeclampsia before 37 weeks of gestation was recorded in only 4% of women receiving aspirin as opposed to 83% of controls (P<0.001). In the group of women receiving aspirin, 19% of newborns suffered from IUGR as opposed to 32%of newborns in the control group (P=0.106). There was no significant difference between the two groups in the rate of preterm delivery (P=0.080), mode of delivery (P=0.971), Apgar score <5 after one minute (P=0.273) and after 5 minutes (P=0.941), maternal or neonatal bleeding (P=0.948), and neonatal birth weight (P=0.399).
CONCLUSION:
Low-dose aspirin administered as early as 14-16 weeks of gestation to pregnant women at high risk of preeclampsia with abnormal uterine Doppler findings may reduce or modify the course of severe preeclampsia. Its effects on the prevention of IUGR need further evaluation.

J Indian Med Assoc. 1997 Feb;95(2):43-4, 47.
Role of low dose aspirin in prevention of pregnancy induced hypertension.
Tewari S, Kaushish R, Sharma S, Gulati N.
In a prospective randomised clinical study 50 primi or multigravidae with history of essential hypertension or pregnancy induced hypertension (PIH) in previous pregnancy were selected from antenatal clinics, on the basis of positive roll over test (ROT) carried out between 28 to 30 weeks of pregnancy. The study group comprised 25 women who received 50 mg aspirin daily from day of positive ROT till 37 weeks of pregnancy. Other 25 women served as control. The incidence of PIH in study and control group was 4% versus 28% and that of pre-term birth was 4% versus 24%. The mean birth weight of newborns in the two groups was 3.04 +/- 0.38 kg and 2.71 +/- 0.48 kg respectively. All these differences were statistically significant. No adverse maternal or neonatal complication due to aspirin was observed.

Teratology. 1991 Nov;44(5):521-9.
Aspirin dose-dependently reduces alcohol-induced birth defects and prostaglandin E levels in mice.
Randall CL, Anton RF, Becker HC, Hale RL, Ekblad U.
The purpose of the present study was threefold. The first purpose was to determine if aspirin (ASA) decreases alcohol-induced birth defects in mice in a dose-dependent fashion. The second purpose was to see if the antagonism of alcohol-induced birth defects afforded by ASA pretreatment was related to dose-dependent decreases in prostaglandin E (PGE) levels in uterine/embryo tissue. The third purpose was to determine if ASA pretreatment altered maternal blood alcohol level. In experiments 1 and 2, pregnant C57BL/6J mice were administered ASA (0, 18.75, 37.5, 75, 150, or 300 mg/kg) on gestation day 10. One hour following the subcutaneous injection of ASA, mice received alcohol (5.8 g/kg) or an isocaloric sucrose solution intragastrically. In experiment 1 the incidence of birth defects was assessed in fetuses delivered by caesarean section on gestation day 19. In experiment 2 uterine/embryo tissue samples were collected on gestation day 10 1 hr following alcohol intubation for subsequent PGE analysis. In experiment 3 blood samples were taken at five time points following alcohol intubation from separate groups of alcohol-treated pregnant mice pretreated with 150 mg/kg ASA or vehicle. The results from the three experiments indicated that 1) ASA dose-dependently reduced the frequency of alcohol-induced birth defects in fetuses examined at gestation day 19, (2) ASA decreased the levels of PGE in gestation day 10 uterine/embryo tissue in a similar dose-dependent fashion, and 3) ASA pretreatment did not significantly influence maternal blood alcohol levels. These results provide additional support for the hypothesis that PGs may play an important role in mediating the teratogenic actions of alcohol.

Lancet. 1991 Jun 15;337(8755):1427-31.
Prevention of fetal growth retardation with low-dose aspirin: findings of the EPREDA trial.
Uzan S, Beaufils M, Breart G, Bazin B, Capitant C, Paris J.
The efficacy of low-dose aspirin in preventing fetal growth retardation was tested in a randomised, placebo-controlled, double-blind trial. A secondary aim was to find out whether dipyridamole improves the efficacy of aspirin. 323 women at 15-18 weeks’ amenorrhoea were selected at twenty-five participating centres on the basis of fetal growth retardation and/or fetal death or abruptio placentae in at least one previous pregnancy. They were randomly allocated to groups receiving placebo, 150 mg/day aspirin, or 150 mg/day aspirin plus 225 mg/day dipyridamole, for the remainder of the pregnancy. In the first phase of the trial all actively treated patients (n = 156) were compared with the placebo group (n = 73). Mean birthweight was significantly higher in the treated than in the placebo group (2751 [SD 670] vs 2526 [848] g; difference 225 g [95% CI 129-321 g], p = 0.029) and the frequency of fetal growth retardation in the placebo group was twice that in the treated group (19 [26%] vs 20 [13%]; p less than 0.02). The frequencies of stillbirth (4 [5%] vs 2 [1%]) and abruptio placentae (6 [8%] vs 7 [5%]) were also higher in the placebo than in the treated group. The benefits of aspirin treatment were greater in patients with two or more previous poor outcomes than in those with only one. In the second analysis, of aspirin only (n = 127) vs aspirin plus dipyridamole (n = 119), no significant differences were found. There was no excess of maternal or neonatal side-effects in the aspirin-treated patients.

Clin Perinatol. 1992 Jun;19(2):305-17.
An aspirin a day to prevent prematurity.
Sibai BM.
Intrauterine fetal growth retardation and preeclampsia remain a substantial cause of preterm birth world wide. There is evidence to suggest that a functional imbalance between vascular prostacyclin and platelet-derived thromboxane A2 production plays a central role in the pathogenesis of these disorders. Low-dose aspirin appears to reverse the above functional balance resulting in increased prostacyclin to thromboxane ratio. The efficacy and safety of low-dose aspirin in preventing preeclampsia and fetal growth retardation were tested in several randomized and uncontrolled trials. The data in the literature suggest that low-dose aspirin is effective in reducing preterm birth due to the above complications in selected high-risk pregnant women.

J Endocrinol. 1989 Jun;121(3):513-9.
Indomethacin inhibits the effects of oestrogen in the anterior pituitary gland of the rat.
Rosental DG, Machiavelli GA, Cherñavsky AC, Speziale NS, Burdman JA.
Two inhibitors of prostaglandin synthesis, indomethacin and aspirin, blocked the increase of oestrogen-binding sites in the nuclear subcellular fraction, an increase which occurs after the administration of oestradiol. Consequently the biological effects of oestrogens in the anterior pituitary gland of the rat (prolactin synthesis, concentration of progesterone-binding sites and cell proliferation) are diminished. The anterior pituitary gland synthesized prostaglandin F2 alpha (PGF2 alpha), PGE2 and PGD2 from arachidonic acid. This synthesis was blocked when indomethacin was added to the culture media. Oestrogen increased the concentration of PGE2: an increase that was partially prevented by indomethacin. Prostaglandins may have an important role on the effects of oestrogen in the anterior pituitary gland of the rat.

Free Radic Biol Med. 2000 Dec;29(11):1135-42.
Synergistic inhibition of cyclooxygenase-2 expression by vitamin E and aspirin.
Abate A, Yang G, Dennery PA, Oberle S, Schröder H.
The use of aspirin in rheumatoid arthritis is limited since inhibition of the pro-inflammatory enzyme cyclooxygenase-2 occurs only at higher aspirin doses that are often associated with side effects such as gastric toxicity. Using a macrophage cell line (J774. 1A), the present study explores possible synergistic effects of aspirin and vitamin E on the expression and activity of cyclooxygenase-2. Lipopolysaccharide-induced prostaglandin E(2) formation was significantly reduced by aspirin (1-100 microM) or vitamin E (100-300 microM). When combined with vitamin E, aspirin-dependent inhibition of prostaglandin E(2) formation was increased from 59% to 95% of control. Likewise, lipopolysaccharide-induced cyclooxygenase-2 protein and mRNA expression were virtually abolished by the combined treatment of aspirin and vitamin E, whereas the two agents alone were only modestly effective. Vitamin C did not mimic the actions of vitamin E under these conditions, suggesting that redox-independent mechanisms underlie the action of vitamin E. In agreement with this, vitamin E and aspirin were without effect on lipopolysaccharide-induced translocation of the redox-sensitive transcription factor NF-kappa B. Our results show that co-administration of vitamin E renders cyclooxygenase-2 more sensitive to inhibition by aspirin by as yet unknown mechanisms. Thus, anti-inflammatory therapy might be successful with lower aspirin doses when combined with vitamin E, thereby possibly avoiding the side effects of the usually required high dose aspirin treatment.

Int J Biochem. 1982;14(9):783-6.
Inhibition of anaerobic glycolysis in bovine retina extracts by salicylate and acetylsalicylate.
Rinaudo MT, Curto M, Bruno R, Ponzetto C.
1. Na salicylate 31 mM inhibits anaerobic glycolysis from glucose in bovine retina extracts. The formation rate of DAP and GAP increases while that of FDP, G6P, F6P and lactate decreases. All the above modifications are almost completely removed by 1.4 mM NAD+. 2. Bovine retina extracts, preincubated for 1 hr at 0 degrees C with 31 mM Na salicylate show a strongly reduced glycolytic activity. In this system G6P and F6P do accumulate, FDP, DAP, GAP and lactate decrease. These effects are not altered adding 3.5 mM NAD+ to the preincubation mixture. 3. Acetylsalicylate 31 mM inhibits anaerobic glycolysis in crude retina extracts. As the rate of lactate formation decreases, G6P and F6P do accumulate, while FDP, DAP and GAP diminish. 4. Identical modifications are observed adding the inhibitor directly to the incubation mixture, or preincubating it with the extracts at 0 degrees C for 4 hr. 3.5 mM NAD+ does not remove the effects of acetylsalicylate.

J Pharm Pharmacol. 2002 Apr;54(4):577-82.
Non-steroidal anti-inflammatory drugs inhibit epinephrine- and cAMP-mediated lipolysis in isolated rat adipocytes.
de Zentella PM, Vázquez-Meza H, Piña-Zentella G, Pimentel L, Piña E.
Acute ethanol intoxication increased triacylglycerides (TAG) and thiobarbituric acid reactive substances (TBARS) in liver and promoted the liberation of epinephrine. Four non-steroidal anti-inflammatory drugs (NSAIDs)–aspirin, naproxen, nimesulide and piroxicam–prevented this increase in TAG and TBARS. Because fatty acids provided by adipose tissue contribute substantially to elevated hepatic TAG in ethanol-intoxicated rats, it was thought that the NSAIDs might reduce epinephrine-stimulated lipolysis in these rats. Isolated rat adipocytes were activated with epinephrine in the presence or absence of the NSAIDs. The NSAIDs inhibited epinephrine-stimulated lipolysis. These drugs did not modify the binding of dihydroalprenolol (beta-adrenergic agonist) to their receptors in isolated guinea-pig liver membranes. The NSAIDs, at concentrations 3,000-fold lower than that of cAMP, inhibited stimulated lipolysis by this messenger. In conclusion, aspirin, naproxen, nimesulide and piroxicam reduce the release of fatty acids from adipose tissue to the liver by inhibiting the epinephrine-stimulated lipolysis, and this, in part, explains the protective action of these NSAIDs against hepatic signs of acute ethanol intoxication.

Physiol Behav. 1996 Jan;59(1):133-9.
Nonsteroidal anti-inflammatory drugs alter body temperature and suppress melatonin in humans.
Murphy PJ, Myers BL, Badia P.
Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit prostaglandin synthesis in humans. Prostaglandins are involved in thermoregulation, melatonin synthesis, and sleep. To determine effects of NSAIDs on body temperature (BT) and melatonin synthesis (MT) in humans, and to elucidate mechanisms by which NSAIDs may alter sleep patterns, a series of experiments using the NSAIDs aspirin and ibuprofen was conducted. Seventy-five subjects were tested under several experimental protocols. BT after NSAID or placebo was assessed in both between- and within-subjects designs at night and during the day. MT levels were assessed after NSAID or placebo at night in a within-subjects design. The normal nocturnal BT decrease was attenuated and MT was suppressed after NSAID relative to after placebo administration. Lower MT levels were associated with a relative flattening of BT. Daytime BT was not affected by NSAIDs. These results are compatible with the hypothesis that some of the behavioral changes associated with NSAIDs, including changes in sleep, are due to changes in BT and MT. We speculate that NSAID effects on sleep and BT are related to prostaglandin synthesis inhibition and/or suppression of MT.

Regul Pept. 2003 Jul 15;114(2-3):101-7.
Inhibition of cytosolic phospholipase A2 mRNA expression: a novel mechanism for acetylsalicylic acid-mediated growth inhibition and apoptosis in colon cancer cells.
Yu HG, Huang JA, Yang YN, Luo HS, Yu JP, Meier JJ, Schrader H, Bastian A, Schmidt WE, Schmitz F.
Acetylsalicylic acid (ASA) has been confirmed to inhibit proliferation and to induce apoptosis in human colorectal cancer cells in vitro. However, the mechanism by which ASA exhibits antiproliferative and proapoptotic effects in cyclooxygenase 2 (COX-2)-negative cells remains to be further elucidated. In the present study, SW480, a COX-2-negative colon cancer cell line, was treated with various concentrations of ASA (0, 2.5, 5, and 10 mM). The antiproliferative and proapoptotic effects of ASA were confirmed by MTT assay, flow cytometry of propidium iodide (PI)-stained cells, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assay. After treatment with ASA, intracellular cyclic AMP (cAMP) levels were increased and the production of prostaglandin E2 (PGE2) was decreased. RT-PCR analysis revealed that treatment of ASA induced a concentration-dependent downregulation of cytosolic phospholipase A2 (cPLA2) mRNA expression in SW480 cells and also in two other colorectal cancer cell lines, Colo320 and HT-29 cells. Intracellular calcium levels were unaffected by ASA treatment. Our results indicate that the ASA-induced downregulation of cytosolic phospholipase A2 mRNA expression might be a novel mechanism for ASA-mediated growth inhibition and apoptosis in colon cancer cells.

Cancer. 2013 Mar 11. doi: 10.1002/cncr.27817. [Epub ahead of print]
Aspirin is associated with lower melanoma risk among postmenopausal Caucasian women: The Women’s Health Initiative.
Gamba CA, Swetter SM, Stefanick ML, Kubo J, Desai M, Spaunhurst KM, Sinha AA, Asgari MM, Sturgeon S, Tang JY.
BACKGROUND:
Nonsteroidal anti-inflammatory drugs (NSAIDs) have been associated with decreased risk of gastric, colorectal, and breast cancer. However, the impact of NSAIDs on the risk of melanoma has been inconsistent. The authors evaluated the association between NSAID use and cutaneous melanoma risk in the Women’s Health Initiative (WHI) Observational Study (OS).
METHODS:
At study entry, use of aspirin (acetylsalicylic acid [ASA]) and nonaspirin NSAIDs was assessed among 59,806 postmenopausal Caucasian women ages 50 to 79 years. Cox proportional hazards models were constructed after adjusting for participant skin type, sun exposure history, and medical indications for NSAID use among other confounders.
RESULTS:
During a median follow-up of 12 years, 548 incident melanomas were confirmed by medical review. Women who used ASA had a 21% lower risk of melanoma (hazard ratio, 0.79; 95% confidence interval, 0.63-0.98) relative to nonusers. Increased duration of ASA use (<1 year, 1-4 years, and ≥5 years) was associated with an 11% lower risk of melanoma for each categorical increase (Ptrend = .01), and women with ≥5 years of use had a 30% lower melanoma risk (hazard ratio, 0.70; 95% confidence interval, 0.55-0.94). In contrast, use of non-ASA NSAIDs and acetaminophen were not associated with melanoma risk.
CONCLUSIONS:
Postmenopausal women who used ASA had a significantly lower risk of melanoma, and longer duration of ASA use was associated with greater protection. Although this study was limited by the observational design and self-report of NSAID use, the findings suggest that ASA may have a chemopreventive effect against the development of melanoma and warrant further clinical investigation. Cancer 2012. © 2012 American Cancer Society.

Curr Atheroscler Rep. 2007 Nov;9(5):409-16.
Aspirin in the treatment and prevention of cardiovascular disease: current perspectives and future directions.
Hennekens CH.
In secondary prevention among male and female survivors of prior myocardial infarction (MI), occlusive stroke, transient ischemic attack, and other high-risk conditions, long-term use of aspirin confers very similar statistically significant and clinically important reductions in MI, stroke, and cardiovascular death. In men and women suffering acute MI or acute occlusive stroke, aspirin confers similar benefits. In primary prevention, aspirin confers a statistically significant and clinically important reduction in risk of a first MI, but the data on stroke and cardiovascular disease death remain inconclusive, so aspirin should be prescribed on an individual basis by the healthcare provider who weighs this clear benefit against long-term side effects. Worldwide, aspirin used more widely and appropriately would avoid many premature deaths in secondary prevention, during acute MI, and during acute stroke, as well as many first MI in primary prevention.

Circulation. 1990 Sep;82(3):897-902.
Circadian variation of acute myocardial infarction and the effect of low-dose aspirin in a randomized trial of physicians.
Ridker PM, Manson JE, Buring JE, Muller JE, Hennekens CH.
Increased platelet aggregation in the morning and upon assuming an upright posture may account at least in part for the observed circadian variation in onset of acute myocardial infarction. The Physicians’ Health Study, a randomized, double-blind, placebo-controlled trial of alternate-day aspirin intake (325 mg) among 22,071 US male physicians, afforded the opportunity to assess this circadian pattern and examine whether it is altered by aspirin therapy. During a 5-year period of follow-up, 342 cases of nonfatal myocardial infarction were confirmed, of which the time of onset was available in 211 (62%). The placebo group showed a bimodal circadian variation in onset of myocardial infarction with a primary peak between 4:00 AM and 10:00 AM (p less than 0.001). In the aspirin group, however, this circadian variation was minimal (p = 0.16), due primarily to a marked reduction in the morning peak of infarction. Specifically, aspirin was associated with a 59.3% reduction in the incidence of infarction during the morning waking hours, compared with a 34.1% reduction for the remaining hours of the day. The greater reduction was observed during the 3-hour interval immediately after awakening, a period with a risk of infarction twice that of any other comparable time interval (p less than 0.001). Aspirin intake was associated with a mean reduction in the incidence of infarction of 44.8% over the entire 24-hour cycle. These data support the hypothesis that increased platelet aggregability in the morning and upon arising contributes to the occurrence of myocardial infarction and that aspirin reduces the risk of infarction by inhibiting platelet aggregation during these critical periods.

Br J Cardiol. 2010;17(4):185-189.
‘Time is Muscle’: Aspirin Taken During Acute Coronary Thrombosis
Peter C Elwood, Gareth Morgan, Malcolm Woollard, Andrew D Beswick
“Randomised trials have shown that the earlier aspirin is taken by patients with myocardial infarction, the greater the reduction in deaths. We suggest, therefore, that patients known to be at risk of an AMI, including older people, should be advised to carry a few tablets of soluble aspirin at all times, and chew and swallow a tablet immediately, if they experience severe chest pain.”
“It is accepted practice for paramedics to give aspirin to patients for whom an emergency call has been received because of chest pain
It is suggested that patients at increased vascular risk, including older people, should carry tablets of soluble aspirin at all times, and chew and swallow a tablet immediately they experience severe chest pain”

Published OnlineFirst June 26, 2014; doi: 10.1158/1055-9965.EPI-13-1284
Case–Control Study of Aspirin Use and Risk of Pancreatic Cancer
Samantha A. Streicher, Herbert Yu, Lingeng Lu, Mark S. Kidd, and Harvey A. Risch
Background: Pancreas-cancer prognosis is dismal, with 5-year survival less than 5%. Significant relationships between aspirin use and decreased pancreas-cancer incidence and mortality have been shown in four of 13 studies.
Methods: To evaluate further a possible association between aspirin use and risk of pancreatic cancer, we used data from a population-based Connecticut study conducted from January 2005 to August 2009, of 362 pancreas-cancer cases frequency matched to 690 randomly sampled controls.
Results: Overall, regular use of aspirin was associated with reduced risk of pancreatic cancer [odds ratio (OR), 0.52; 95% confidence interval (CI), 0.39–0.69]. Increments of decreasing risk of pancreatic cancer were observed for each year of low-dose or regular-dose aspirin use (OR, 0.94; 95% CI, 0.91–0.98 and OR, 0.98; 95% CI, 0.96–1.01, respectively) and for increasing years in the past that low-dose or regular-dose aspirin use had started (OR, 0.95; 95% CI, 0.92–0.99 and OR, 0.98; 95% CI, 0.96–1.00, respectively). Reduced risk of pancreatic cancer was seen in most categories of calendar time period of aspirin use, for both low-dose aspirin and regular-dose aspirin use. Relative to continuing use at the time of interview, termination of aspirin use within 2 years of interview was associated with increased risk of pancreatic cancer (OR, 3.24; 95% CI, 1.58–6.65).
Conclusions: Our results provide some support that a daily aspirin regimen may reduce risk of developing pancreatic cancer.
Impact: Long-term aspirin use has benefits for both cardiovascular disease and cancer, but appreciable bleeding complications that necessitate risk–benefit analysis for individual applications. Cancer Epidemiol Biomarkers Prev; 23(7); 1–10. ©2014 AACR.

Lab Invest. 2015 Jul;95(7):702-17. doi: 10.1038/labinvest.2015.49. Epub 2015 Apr 13.
Aspirin blocks growth of breast tumor cells and tumor-initiating cells and induces reprogramming factors of mesenchymal to epithelial transition.
Maity G, De A, Das A, Banerjee S, Sarkar S, Banerjee SK.
Acetylsalicylic acid (ASA), also known as aspirin, a classic, nonsteroidal, anti-inflammatory drug (NSAID), is widely used to relieve minor aches and pains and to reduce fever. Epidemiological studies and other experimental studies suggest that ASA use reduces the risk of different cancers including breast cancer (BC) and may be used as a chemopreventive agent against BC and other cancers. These studies have raised the tempting possibility that ASA could serve as a preventive medicine for BC. However, lack of in-depth knowledge of the mechanism of action of ASA reshapes the debate of risk and benefit of using ASA in prevention of BC. Our studies, using in vitro and in vivo tumor xenograft models, show a strong beneficial effect of ASA in the prevention of breast carcinogenesis. We find that ASA not only prevents breast tumor cell growth in vitro and tumor growth in nude mice xenograft model through the induction of apoptosis, but also significantly reduces the self-renewal capacity and growth of breast tumor-initiating cells (BTICs)/breast cancer stem cells (BCSCs) and delays the formation of a palpable tumor. Moreover, ASA regulates other pathophysiological events in breast carcinogenesis, such as reprogramming the mesenchymal to epithelial transition (MET) and delaying in vitro migration in BC cells. The tumor growth-inhibitory and reprogramming roles of ASA could be mediated through inhibition of TGF-β/SMAD4 signaling pathway that is associated with growth, motility, invasion, and metastasis in advanced BCs. Collectively, ASA has a therapeutic or preventive potential by attacking possible target such as TGF-β in breast carcinogenesis.

Cell Death Dis. 2015 Jun 11;6:e1786. doi: 10.1038/cddis.2015.153.
Aspirin delays mesothelioma growth by inhibiting HMGB1-mediated tumor progression.
Yang H, Pellegrini L, Napolitano A, Giorgi C, Jube S, Preti A, Jennings CJ, De Marchis F, Flores EG, Larson D, Pagano I, Tanji M, Powers A, Kanodia S, Gaudino G, Pastorino S, Pass HI, Pinton P, Bianchi ME, Carbone M.
High-mobility group box 1 (HMGB1) is an inflammatory molecule that has a critical role in the initiation and progression of malignant mesothelioma (MM). Aspirin (acetylsalicylic acid, ASA) is the most widely used nonsteroidal anti-inflammatory drug that reduces the incidence, metastatic potential and mortality of many inflammation-induced cancers. We hypothesized that ASA may exert anticancer properties in MM by abrogating the carcinogenic effects of HMGB1. Using HMGB1-secreting and -non-secreting human MM cell lines, we determined whether aspirin inhibited the hallmarks of HMGB1-induced MM cell growth in vitro and in vivo. Our data demonstrated that ASA and its metabolite, salicylic acid (SA), inhibit motility, migration, invasion and anchorage-independent colony formation of MM cells via a novel HMGB1-mediated mechanism. ASA/SA, at serum concentrations comparable to those achieved in humans taking therapeutic doses of aspirin, and BoxA, a specific inhibitor of HMGB1, markedly reduced MM growth in xenograft mice and significantly improved survival of treated animals. The effects of ASA and BoxA were cyclooxygenase-2 independent and were not additive, consistent with both acting via inhibition of HMGB1 activity. Our findings provide a rationale for the well documented, yet poorly understood antitumorigenic activity of aspirin, which we show proceeds via HMGB1 inhibition. Moreover, the use of BoxA appears to allow a more efficient HMGB1 targeting while eluding the known gastrointestinal side effects of ASA. Our findings are directly relevant to MM. Given the emerging importance of HMGB1 and its tumor-promoting functions in many cancer types, and of aspirin in cancer prevention and therapy, our investigation is poised to provide broadly applicable information.