PRIMA-1 synergizes olaparib-induced cell death in p53 mutant triple negative human breast cancer cell line via restoring p53 function, arresting cell cycle, and inducing apoptosis.

This study concerned with assessing the effect of restoring p53 using PRIMA-1 on the anti-cancer activity of olaparib against TP53-mutant triple negative breast cancer (TNBC) cells and exploring the optimum synergistic concentrations and the underlying mechanism. Human BC cell lines, MDA-MB-231 with mutated TP53 gene, and MCF-7 with wild-type TP53 gene were treated with olaparib and/or PRIMA-1. The IC50 value for olaparib was significantly decreased by PRIMA-1 in MDA-MB-231 cells compared to MCF-7 cells. Contrary to MCF-7 cells, co-treatment with olaparib and PRIMA-1 had a synergistic anti-proliferative effect in MDA-MB-231 at all tested concentrations with the best synergistic combination at 45 and 8.5 µM, respectively, and furthermore PRIMA-1 enhanced olaparib-induced apoptosis. This synergistic apoptotic effect was associated with a significant boost in mRNA expression of TP53 gene, cell cycle arrest at G2/M phase, modulation of BRCA-1, BAX and Bcl2 proteins expressions, and induction of active caspase-3. These results present a clue for the utility of combined olaparib and PRIMA-1 in treatment of TP53-mutant TNBC invitro. PRIMA-1 triggers olaparib-induced MDA-MB-231 cell death in a synergistic manner via restoring TP53, decreasing BRCA-1 expression, cell cycle arrest, and enhancement of apoptosis via p53/BAX/Bcl2/caspase 3 pathway.

The Potential Protective Effects of Diosmin on Streptozotocin-Induced Diabetic Cardiomyopathy in Rats.

BACKGROUND: Diabetic cardiomyopathy (DCM) is a nonischemic myocardial disorder characterized by metabolic disturbances and oxidative stress in diabetic patients. The present paper aims to determine the protective effect of the phlebotrophic drug, diosmin, on DCM in a model of high-fat diet-fed and streptozotocin-induced type 2 diabetes in the rat. MATERIALS AND METHODS: The animals were divided into 4 groups (8 rats/group) as follows: vehicle-treated nondiabetic control group, vehicle-treated diabetic group, diosmin (50 mg/kg)-treated diabetic group and diosmin (100 mg/kg)-treated diabetic group. Treatment was given once daily orally by gavage for 6 weeks. Oxidant and antioxidant stress markers, inflammatory markers and proapoptotic and antiapoptotic gene expression using quantified real-time polymerase chain reaction were investigated. RESULTS: Diosmin treatment in diabetic rats lowered elevated blood glucose levels, homeostatic model assessment for insulin resistance, cardiac creatine kinase and lactate dehydrogenase enzymes, cardiac malondialdehyde and nitric oxide. Moreover, diosmin increased plasma insulin and c-peptide levels, cardiac glutathione content, superoxide dismutase, catalase and glutathione S-transferase activities. Also, diosmin treatment significantly (P < 0.05) lowered the levels of interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α), down-regulated cardiac Bcl-2-associated X protein and caspase 3 and 9 and up-regulated B-cell lymphoma 2 mRNA expression levels. CONCLUSIONS: Diosmin may have a sizeable therapeutic potential in the treatment of DCM due to antidiabetic, antioxidative stress, anti-inflammatory and antiapoptotic effects. Detailed studies are needed to disclose the precise mechanisms motivating the protective effect of diosmin .

Curcumin ameliorates streptozotocin-induced heart injury in rats.

Heart failure (HF) is one of diabetic complications. This work was designed to investigate the possible modulatory effect of curcumin against streptozotocin-induced diabetes and consequently HF in rats. Rats were divided into control, vehicle-treated, curcumin-treated, diabetic-untreated, diabetic curcumin-treated, and diabetic glibenclamide-treated groups. Animal treatment was started 5 days after induction of diabetes and extended for 6 weeks. Diabetic rats showed significant increase in serum glucose, triglycerides, total cholesterol, low-density lipoprotein-cholesterol, very low density lipoprotein-cholesterol, nitric oxide, lactate dehydrogenase, cardiac malondialdehyde, plasma levels of interleukin-6, and tumor necrosis factor-alpha, and also showed marked decrease in serum high-density lipoprotein-cholesterol, cardiac reduced glutathione, and cardiac antioxidant enzymes (catalase, superoxide dismutase, and glutathione-S-transferase). However, curcumin or glibenclamide treatment significantly mitigated such changes. In conclusion, curcumin has a beneficial therapeutic effect in diabetes-induced HF, an effect that might be attributable to its antioxidant and suppressive activity on cytokines.

Erythrocyte nitric oxide synthase as a surrogate marker for mercury-induced vascular damage: the modulatory effects of naringin.

In this study, endothelial nitric oxide synthase activity and nitric oxide (NO) production by human erythrocytes in the presence and absence of mercuric chloride (HgCl2 ), L-arginine (L-ARG), N ω- nitro-L-arginine methyl ester (L-NAME), and naringin (NAR) were investigated. In addition, the levels of reduced glutathione (GSH) and related enzymes were estimated in erythrocytes hemolysate. The protein carbonyl content (PCC) and thiobarbituric acid-reactive substances (TBARS) levels were also determined. The results of this study revealed that the treatment of erythrocytes with either HgCl2 or L-NAME induced a significant decrease in NOS activity and nitrite levels compared with control cells. Furthermore, mercury exposure significantly increased the levels of PCC and TBARS but reduced the GSH level. The activities of glucose-6-phosphate dehydrogenase, glutathione reductase, glutathione peroxidase, and glutathione-S-transferase (GST) were inhibited. The exposure of erythrocytes to HgCl2 in combination with L-ARG, NAR, or both ameliorated the investigated parameters compared with erythrocytes incubated with HgCl2 alone. These results indicate that mercury exposure decreased both erythrocyte NOS activity and nitrite production, and that these parameters might be indicative of mercury exposure. The data also suggest that concomitant treatment with NAR can restore NO bioavailability through either its metal-chelating properties or its antioxidant activity.

Uroprotective effect of pentoxifylline in cyclophosphamide-induced hemorrhagic cystitis in rats.

The role of phosphodiesterase inhibitor, pentoxifylline, in the prevention of cyclophosphamide-induced hemorrhagic cystitis was evaluated in a rat model. Hemorrhagic cystitis was induced in rats by an intraperitoneal (i.p.) injection of a single dose of cyclophosphamide (150 mg/kg). Pentoxifylline (150 mg/kg/day/ip) was administered for 10 days followed by cyclophosphamide. Hemorrhagic cystitis was well characterized macroscopically, microscopically, and biochemically. Cyclophosphamide induced bladder injury including acute severe inflammation, vascular congestion, severe edema, hemorrhage, inflammatory cell infiltration in the lamina propria, and epithelial denudation; as well as it notably elevated serum inflammatory cytokines (tumor necrosis factor-α, interleukin-6, and interleukin-1ß), bladder content of malondialdehyde and total nitrate, accompanied with depletion of bladder antioxidant enzymes activities (glutathione peroxidase, superoxide dismutase, glutathione-S-transferase, and catalase). Prior administration of pentoxifylline improved all biochemical and histologic alterations induced by the cytotoxic drug cyclophosphamide. In conclusion, pentoxifylline has proven uroprotective efficacy in the cyclophosphamide-induced hemorrhagic cystitis model, possibly through modulating the release of inflammatory cytokines and nitric oxide and restoring the oxidant/antioxidant balance.

Effects of nutritional and excessive levels of selenium on red blood cells of rats fed a high cholesterol diet.

In this study, we investigated the effects of selenium (Se) on the properties of erythrocytes and atherogenic index in the presence and absence of high cholesterol diet (HCD). The effect of selected two different doses (1 µg and 50 µg Se/kg/body weight) on HCD-induced oxidative stress was investigated. The hemolysis of the erythrocytes of the HCD rats as well as by high levels of selenium or their combination was markedly increased. Likewise, atherogenic index and plasma glutathione peroxidase (GPx) activity were significantly increased in the same groups of rats compared to control ones. In contrast, paraoxonase activity, glutathione levels and protein thiol levels, catalase, GPx, and superoxide dismutase activities were significantly decreased in rats that received the HCD, high selenium dose, or their combination. Malondialdehyde and protein carbonyl levels in the plasma and red blood cells were significantly increased by HCD and high selenium dose administration. Co-administration of selenium at low dose with or without an HCD restored all of the investigated parameters to near-normal values. The results of this study suggest that excess selenium administration with HCD worsens the atherogenic index and enhances formation of oxidized red blood cells. At dosage levels in the nutritional range such as 1 µg Se/kg body weight, selenium ameliorates the atherogenic index and preserves the antioxidant capacity of the erythrocytes.

The protective effect of aminoguanidine on doxorubicin-induced nephropathy in rats.

Reactive oxygen species and cytokines have been implicated in the nephrotoxicity induced by doxorubicin. The goal of the present study was to determine protective effect of aminoguanidine on doxorubicin-induced nephrotoxicity in rats. Different groups of male Wistar rats received doxorubicin (67.75 mg/kg/i.p./2 days), aminoguanidine alone and aminoguanidine (200 and 400 mg/kg/i.p./30 days) prior to doxorubicin, respectively. Doxorubicin significantly increased serum creatinine (505%), blood urea nitrogen (333%), nitric oxide (406%), and plasma tumor necrosis factor-alpha (706%) as well as urinary albumin (452%) and N-acetyl-ß-D-glucosaminidase (415%) compared to control. Moreover, renal glutathione (334%), superoxide dismutase (283%), and catalase (513%) were significantly reduced accompanied with elevation in renal malondialdehyde compared to control. Pretreatment with aminoguanidine mitigated such changes in all mentioned parameters. Histopathological changes showed that doxorubicin-caused significant structural damages to kidneys that were reduced with aminoguanidine. Results indicate that reactive oxygen species and cytokines are involved in doxorubicin-induced nephrotoxicity, which can be reduced by aminoguanidine.

Hepatoprotective activity of quercetin against acrylonitrile-induced hepatotoxicity in rats.

Acrylonitrile is a potent hepatotoxic, mutagen, and carcinogen. A role for free radical-mediated lipid peroxidation in the toxicity of acrylonitrile has been suggested. The present study was designed to assess the hepatoprotective effect of quercetin against acrylonitrile-induced hepatotoxicity in rats. Liver damage was induced by oral administration of acrylonitrile (50 mg/kg/day/5 weeks). Acrylonitrile produced a significant elevation of malondialdehyde (138.9%) with a marked decrease in reduced glutathione (72.4%), and enzymatic antioxidants; superoxide dismutase (81%), and glutathione peroxidase (53.2%) in the liver. Serum aspartate aminotransferase, alanine aminotransferases, direct bilirubin, and total bilirubin showed a significant increase in acrylonitrile alone treated rats (115.5%, 110.8%, 1006.8%, and 1000.8%, respectively). Pretreatment with quercetin (70 mg/kg/day/6 weeks) and its coadministration with acrylonitrile prevented acrylonitrile-induced alterations in hepatic lipid peroxides and enzymatic antioxidants as well as serum aminotransferases and bilirubin. Histopathological findings supported the biochemical results. We suggest that querectin possess hepatoprotective effect against acrylonitrile-induced hepatotoxicity through its antioxidant activity.

Effect of lead toxicity on coenzyme Q levels in rat tissues.

Lead is a persistent and common environmental contaminant, which chiefly plays a significant role in modern industry. Coenzyme Q acts as electron and proton carrier in mitochondria and functions as an antioxidant in its reduced form (ubiquinol). To investigate the hazardous effects of lead on the coenzyme Q level, rats were injected i.p. with lead acetate (5 mg/kg b.wt. daily for 6 weeks). Our results showed that the levels of both oxidized (ubiquinone) and reduced (ubiquinol) forms of coenzyme Q(9) and Q(10) in serum, brain, liver and kidney of lead-treated rats are quite different depending on the organ tissue type.

Bone mineral density and vitamin D receptor polymorphism in beta-thalassemia major.

Osteoporosis is the most prevalent bone complication in beta-thalassemic patients despite regular transfusions and iron chelation therapy. Although its etiology is multi-factorial, genetic factors play an important role in pathogenesis. These factors have not yet been clearly defined, however, osteoporosis may be related to vitamin D receptor gene BsmI polymorphism. In this study, BsmI vitamin D receptor gene polymorphism was analyzed using polymerase chain reaction and BsmI restriction fragment length polymorphism in 42 regularly treated-beta-thalassemic patients of different ages. Bone mineral density was measured by peripheral quantitative ultrasound at the heel of the foot. Serum levels of alkaline phosphatase, calcium, phosphorus, ferritin and 25-hydroxyvitamin D3 were determined. Patients were divided into two groups according to pubertal signs: group I (22 children), and group II (20 adolescents and adults). The Z-scores of bone mineral density in both groups were -1.32 +/- -0.9 and -2.30 +/- -1.02 respectively, with a significant difference between the two groups. The height standard deviation and 25-hydroxyvitamin D3 were significantly decreased in group II compared to group I. Moreover, significantly lower bone mineral density and height standard deviation were detected among patients with BB vitamin D receptor genotype. Therefore, this genotype may be considered as a risk factor for osteoporosis in beta-thalassemic patients.

Potential testicular toxicity of sodium nitrate in adult rats.

Nitrate is a common contaminant in groundwater aquifers. Current study aimed at evaluating the potential testicular toxicity of sodium nitrate in rats. Sodium nitrate was given orally to rats at doses of 50, 100 or 200 mg/kg/day for 60 consecutive days. Sperm count and motility, daily sperm production and testis weight were significantly decreased specially at high doses. Testicular activity of lactate dehydrogenase-X, glucose-6-phosphate dehydrogenase, and acid phosphatase were inhibited in a dose-related manner. Lipid peroxides and hydrogen peroxide production were significantly increased in all treated animals. This was accompanied by inhibition of testicular activities of superoxide dismutase and glutathione peroxidase. Fifty mg/kg of sodium nitrate did not significantly alter catalase or glutathione reductase activity. Glutathione was significantly decreased by sodium nitrate in a dose-related manner. The decrease in sperm count and motility and daily sperm production was confirmed by histopathological studies which indicated chromatolysis, pyknosis and necrosis in spermatocytes. In conclusion, subchronic exposure of rats to sodium nitrate results in testicular toxicity as evidenced by decreased sperm count and motility, daily sperm production and testis weight, inhibited activity of enzyme markers of spermatogenesis and induction of histopathological changes. These effects are attributed, at least partly, to testicular oxidative stress.

L-arginine augments the antioxidant effect of garlic against acetic acid-induced ulcerative colitis in rats.

Garlic contains many sulfhydryl compounds that act as antioxidants. However, the role of nitric oxide (NO) in inflammation is controversial. The aim of the present study is to investigate the possible protective effect of garlic against acetic acid-induced ulcerative colitis in rats, as well as the probable modulatory effect of L-arginine (NO precursor) on garlic activity. Intra-rectal inoculation of rats with 4% acetic acid for 3 consecutive days caused a significant increase in the colon weight and marked decrease in the colon length. In addition, acetic acid induced a significant increase in serum levels of nitrate as well as colonic tissue content of malondialdehyde (MDA). Moreover, colonic tissue contents of glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT) were markedly reduced. On the other hand, pre-treatment of rats with garlic (0.25 g/kgbwt, orally) for 4 consecutive weeks and 3 days during induction of colitis significantly reduced the increase in the colon weight induced by acetic acid and ameliorated alterations in oxidant and antioxidant parameters. Interestingly, oral co-administration of garlic (0.25 g/kgbwt) and L-arginine (625 mg/kgbwt) for the same period of garlic administration mitigated the changes in both colon weight and length induced by acetic acid and increased garlic effect on colon tissue contents of MDA and GSH. In conclusion, L-arginine can augment the protective effect of garlic against ulcerative colitis; an effect that might be mainly attributed to its NO donating property resulting in enhancement of garlic antioxidant effect. Further studies will be needed to determine which one of the active ingredients of garlic has the main antioxidant effect to be used with L-arginine.

Potential antidiabetic and hypolipidemic effects of propolis extract in streptozotocin-induced diabetic rats.

Free radicals have been implicated in the pathogenesis of diabetes mellitus leading to various complications including atherosclerosis. Propolis was reported to have oxygen radical scavenging activity. The present study was designed to investigate the possible antidiabetic, hypolipidemic and antioxidant effects of ethanolic extract of propolis (EEP). Type capital I, Ukrainian diabetes was induced in rats by injection of streptozotocin (STZ) in a dose of 60 mg/kg bwt, i.p. for 3 consecutive days. After 5 weeks of STZ injection, there were an apparent reduction in the animal body weight amounting to 21% and significant increases in serum glucose (184%), triglycerides (63%), total cholesterol (43%) and low density lipoprotein-cholesterol (LDL-C) (148%) with a concomitant decrease in serum high density lipoprotein-cholesterol (HDL-C) (51%) as compared to the control normal group. In addition, there was significant elevation in pancreatic lipid peroxides measured as malondialdehyde (MDA) and serum nitric oxide (NO) amounting to 185% and 224%, respectively with marked reduction in serum reduced glutathione (GSH) and catalase (CAT) (66% and 31%, respectively) and pancreatic superoxide dismutase (SOD) (54%) in STZ-treated rats. On the other hand, oral daily treatment of animals with EEP in a dose of 200 mg/kg bwt for a period of 5 weeks ameliorated STZ-induced alterations in the animal body weight as well as in serum glucose, lipids, lipoproteins, NO, GSH & CAT and pancreatic MDA & SOD. In conclusion, propolis extract offers promising antidiabetic and hypolipidemic effects that may be mainly attributed to its potent antioxidant potential. Further studies will be needed in future in order to determine which one(or more) of its active constituents has the main antidiabetic and hypolipidemic effects.

Experimental diabetic nephropathy can be prevented by propolis: Effect on metabolic disturbances and renal oxidative parameters.

Oxidative stress may play a key role in the pathogenesis of diabetic nephropathy. Propolis and its extract have antioxidant properties. The effect of ethanolic extract of propolis against experimental diabetes mellitus-associated changes was examined. Diabetes was induced experimentally in rats by i.p. injection of streptozotocin (STZ) in a dose of 60 mg/kg bwt for 3 successive days. Blood urea nitrogen (BNU), creatinine, glucose, lipid profile, malondialdehyde (MDA) and urinary albumin were measured. Superoxide dimutase (SOD), glutathione (GSH), catalase (CAT) and MDA were measured in the renal tissue. The results showed decreased body weight and increased kidney weight in diabetic animals. Compared to the control normal rats, diabetic rats had higher blood glucose, BNU, creatinine, total cholesterol, triglycerides, low-density lipoprotein-cholesterol (LDL-C), MDA and urinary albumin and lower high-density lipoprotein-cholesterol (HDL-C) levels. Moreover, renal tissue MDA was markedly increased while SOD, GSH and CAT were significantly decreased. Oral administration of propolis extract in doses of 100,200 & 300 mg/kg bwt improved the body and kidney weights, serum glucose, lipid profile, MDA and renal function tests. Renal GSH, SOD and CAT were significantly increased while MDA was markedly reduced. These results may suggest a strong antioxidant effect of propolis which can ameliorate oxidative stress and delay the occurrence of diabetic nephropathy in diabetes mellitus.

Hesperidin, an antioxidant flavonoid, prevents acrylonitrile-induced oxidative stress in rat brain.

Acrylonitrile (ACN) is a volatile, toxic liquid used as a monomer in the manufacture of synthetic rubber, styrene plastics, acrylic fiber, and adhesives. ACN is a potent neurotoxin. A role for free radical mediated lipid peroxidation in the toxicity of ACN has been suggested. We examined the ability of hesperidin, an antioxidant flavonoid, to attenuate ACN-induced alterations in lipid peroxidation in rat brains. The daily oral administration of ACN to male albino rats in a dose of 50 mg/kg bwt for a period of 28 days produced a significant elevation in brain lipid peroxides measured as malondialdehyde (MDA) amounting to 107%, accompanied by a marked decrease in brain-reduced glutathione (GSH) content reaching 63%. In addition, ACN administration resulted in significant reductions in the enzymatic antioxidant parameters of brain; superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), and glutathione-S-transferase (GST) recording 43%, 64%, 52%, and 43%, respectively. On the other hand, pretreatment with hesperidin and its coadministration with ACN once daily in a dose of 200 mg/kg bwt i.p. for 28 days ameliorated ACN-induced alterations in brain lipid peroxidation. These results suggest that hesperidin may have a beneficial role against ACN-induced oxidative stress in the brain; an effect that is mainly attributed to the antioxidant property of hesperidin.

Adenosine receptor subtype-selective antagonists in inflammation and hyperalgesia.

In this study, we examined the effects of systemic and local administration of the subtype-selective adenosine receptor antagonists PSB-36, PSB-1115, MSX-3, and PSB-10 on inflammation and inflammatory hyperalgesia. Pharmacological blockade of adenosine receptor subtypes after systemic application of antagonists generally led to a decreased edema formation after formalin injection and, with the exception of A(3) receptor antagonism, also after the carrageenan injection. The selective A(2B) receptor antagonist PSB-1115 showed a biphasic, dose-dependent effect in the carrageenan test, increasing edema formation at lower doses and reducing it at a high dose. A(1) and A(2B) antagonists diminished pain-related behaviors in the first phase of the formalin test, while the second, inflammatory phase was attenuated by A(2B) and A(3) antagonists. The A(2B) antagonist was particularly potent in reducing inflammatory pain dose-dependently reaching the maximum effect at a low dose of 3 mg/kg. Inflammatory hyperalgesia was totally eliminated by the A(2A) antagonist MSX-3 at a dose of 10 mg/kg. In contrast to the A(1) antagonist, the selective antagonists of A(2A), A(2B), and A(3) receptors were also active upon local administration. Our results demonstrate that the blockade of adenosine receptor subtypes can decrease the magnitude of inflammatory responses. Selective A(2A) antagonists may be useful for the treatment of inflammatory hyperalgesia, while A(2B) antagonists have potential as analgesic drugs for the treatment of inflammatory pain.

Visceral, inflammatory and neuropathic pain in glycine receptor alpha 3-deficient mice.

The alpha3-subunit of strychnine-sensitive glycine receptors is an important modulator of the pain-sensitizing effects of spinal prostaglandin prostaglandin E(2). Mice deficient for alpha3-subunit of strychnine-sensitive glycine receptors lack the prostaglandin E(2)-induced inhibition of glycinergic neurotransmission and recover faster from inflammation-induced hyperalgesia. It, however, remains unclear whether alpha3-subunit of strychnine-sensitive glycine receptors plays a role in other pain models involving prostaglandin synthesis, such as chemically induced pain or neuropathic pain. In this paper, we show a reduction of acetic acid-induced writhing responses in the absence of alpha3-subunit of strychnine-sensitive glycine receptors, but no changes in formalin-induced pain. Furthermore, alpha3-subunit of strychnine-sensitive glycine receptors-deficient mice develop normal thermal hyperalgesia and tactile allodynia. Thus, alpha3-subunit of strychnine-sensitive glycine receptors is involved in the modulation of moderate inflammatory acetic acid-induced pain responses, but neither in formalin-induced pain nor in neuropathic pain.

GlyR alpha3: an essential target for spinal PGE2-mediated inflammatory pain sensitization.

Prostaglandin E2 (PGE2) is a crucial mediator of inflammatory pain sensitization. Here, we demonstrate that inhibition of a specific glycine receptor subtype (GlyR alpha3) by PGE2-induced receptor phosphorylation underlies central inflammatory pain sensitization. We show that GlyR alpha3 is distinctly expressed in superficial layers of the spinal cord dorsal horn. Mice deficient in GlyR alpha3 not only lack the inhibition of glycinergic neurotransmission by PGE2 seen in wild-type mice but also show a reduction in pain sensitization induced by spinal PGE2 injection or peripheral inflammation. Thus, GlyR alpha3 may provide a previously unrecognized molecular target in pain therapy.

Antinociceptive effects of novel A2B adenosine receptor antagonists.

Caffeine, an adenosine A1, A2A, and A2B receptor antagonist, is frequently used as an adjuvant analgesic in combination with nonsteroidal anti-inflammatory drugs or opioids. In this study, we have examined the effects of novel specific adenosine receptor antagonists in an acute animal model of nociception. Several A2B-selective compounds showed antinociceptive effects in the hot-plate test. In contrast, A1- and A2A-selective compounds did not alter pain thresholds, and an A3 adenosine receptor antagonist produced thermal hyperalgesia. Evaluation of psychostimulant effects of these compounds in the open field showed only small effects of some antagonists at high doses. Coadministration of low, subeffective doses of A2B-selective antagonists with a low dose of morphine enhanced the efficacy of morphine. Our results indicate that analgesic effects of caffeine are mediated, at least in part, by A2B adenosine receptors.