José Antonio González-Correa

Neuroprotective effect of hydroxytyrosol and hydroxytyrosol acetate in rat brain slices subjected to hypoxia-reoxygenation.

Hydroxytyrosol (HT) and hydroxytyrosol acetate (HT-AC) are two well-known phenolic compounds with antioxidant properties that are present in virgin olive oil (VOO). Because VOO has shown neuroprotective effects in rats, the purpose of the present study was to investigate the possible neuroprotective effect of HT and HT-AC in a model of hypoxia-reoxygenation in rat brain slices after in vitro incubation of these compounds or after 7 days of oral treatment with 5 or 10 mg/kg per day. Lactate dehydrogenase (LDH) efflux to the incubation medium was measured as a marker of brain cell death. HT and HT-AC inhibited LDH efflux in a concentration-dependent manner, with 50% inhibitory concentrations of 77.78 and 28.18 microM, respectively. Other well-known antioxidants such as vitamin E and N-acetyl-cysteine had no neuroprotective effect in this experimental model. After 1 week of treatment, HT (5 and 10 mg/kg per day p.o.) reduced LDH efflux by 37.8% and 52.7%, respectively, and HT-AC reduced LDH efflux by 45.4% and 67.8%. These data are additional evidence of the cytoprotective effect of VOO administration, and provide a preliminary basis for further study of these polyphenols as potential neuroprotective compounds.

Effects of Hydroxytyrosol and Hydroxytyrosol Acetate Administration to Rats on Platelet Function Compared to Acetylsalicylic Acid

Virgin olive oil (VOO) contains the polyphenols hydroxytyrosol (HT) and hydroxytyrosol acetate (HT-AC). This study investigated the antiplatelet effect of HT and HT-AC in healthy rats and compared their effects to acetylsalicylic acid (ASA). All compounds were administered orally for 7 days. HT and HT-AC inhibited platelet aggregation in whole blood, with a 50% inhibitory dose (ID50) of 48.25 mg/kg per day for HT, 16.05 mg/kg per day for HT-AC, and 2.42 mg/kg per day for ASA. Platelet synthesis of thromboxane B2 was inhibited by up to 30% by HT and 37% by HT-AC; the ID50 of this effect for ASA was 1.09 mg/kg per day. Vascular prostacyclin production was inhibited by up to 27.5% by HT and 32% by HT-AC; the ID50 of this effect for ASA was 6.75 mg/kg per day. Vascular nitric oxide production was increased by up to 34.2% by HT, 66% by HT-AC, and 64% by ASA. We conclude that HT and HT-AC administered orally inhibited platelet aggregation in rats and that a decrease in thromboxane synthesis along with an increase in nitric oxide production contributed to this effect.

Antioxidant effect of acetylsalicylic and salicylic acid in rat brain slices subjected to hypoxia

Acetylsalicylic acid (ASA) reduces the incidence of ischemic stroke mainly through its antithrombotic action; however, it also has a direct neuroprotective effect. The present study was designed to evaluate the effect of ASA on oxidative stress and the activity of nitric oxide synthase (NOS) in an in vitro model of hypoxia in rat brain slices. Rat brain slices were perfused with nitrogen (hypoxia) for a maximum of 120 min, after which we measured lipid peroxidation, glutathione levels, glutathione‐related enzyme activities, and constitutive nitric oxide synthase (cNOS) and inducible nitric oxide synthase (iNOS) activities. In brain tissue subjected to hypoxia, ASA reduced oxidative stress and iNOS activity (all increased by hypoxia), but only when used at higher concentrations. The effects of salicylic acid (SA) were similar but more intense than were those of ASA. After oral administration, the effect of SA was much greater than that of ASA, and the decrease in cell death with SA was seen much more clearly. In view of the greater effect of SA compared to ASA on changes in oxidative stress parameters in a model of hypoxia, and higher brain concentrations of SA when it is administered alone than when ASA is given (undetectable levels), we conclude that SA plays an important role in the cytoprotective effect in brain tissue after ASA administration.

Role of the inhibition of oxidative stress and inflammatory mediators in the neuroprotective effects of hydroxytyrosol in rat brain slices subjected to hypoxia reoxygenation

The aim of this study was to analyze the mechanism of the neuroprotective effect of hydroxytyrosol (HT) in an experimental model of hypoxia-reoxygenation in rat brain slices. After reoxygenation the increase in lactate dehydrogenase efflux was inhibited by HT in a concentration-dependent manner and dose-dependent inhibition after oral administration to rats for 7 days (1, 5 and 10 mg/kg per day). Maximum inhibition was 57.4% in vitro and 38.7% ex vivo. Hydroxytyrosol reduced oxidative stress parameters: it inhibited lipid peroxidation and increased enzymatic activities related with the glutathione system both in vitro and after oral administration to rats. The increase in prostaglandin E2 and interleukin 1β after reoxygenation were inhibited after incubation of brain slices with HT and after oral administration. The accumulation of nitric oxide in brain slices was reduced in a concentration-dependent manner. In conclusion, HT exerts a neuroprotective effect in a model of hypoxia-reoxygenation in rat brain slices, both in vitro and after 7 days of oral administration to rats. HT exerts an antioxidant activity and lowered some inflammatory markers in this model.

Antioxidant effects of a single dose of acetylsalicylic acid and salicylic acid in rat brain slices subjected to oxygen-glucose deprivation in relation with its antiplatelet effect.

The aim of the present study was to analyze the relative participation of the antiplatelet and the antioxidant effects of acetylsalicylic acid (ASA) and salicylic acid (SA) after a single dose (1 or 10 mg/kg i.p.) in an in vitro model of anoxia in slices of rat brain. After 20 min of drug administration, blood and brain were obtained (n=6 rats per group). We measured: lipid peroxidation, glutathione levels and lactate dehydrogenase efflux (LDH), ASA and SA concentrations and platelet aggregation in whole blood. An increase in lipid peroxidation (80%) and in LDH efflux (520%) and a decrease in glutathione levels (35%) were observed after 120 min anoxia in saline-treated rats. SA reduced this oxidative stress and LDH efflux, but it did not modify platelet aggregation. ASA strongly inhibited platelet aggregation but exerted a poor antioxidant effect. ASA was not detectable in brain tissue. We conclude that repeated doses of ASA are necessary to obtain a tissular antioxidant effect, probably when liver generates enough SA.

Aminoglycoside-associated nephrotoxicity in extrahepatic obstructive jaundice

Experimental data demonstrate that biliary obstruction increases renal sensitivity to gentamicin. In the present study the incidence of and risk factors for aminoglycoside nephrotoxicity were prospectively studied in patients with extrahepatic obstructive jaundice. Two hundred and thirty-seven hospitalized adult patients were classified into three groups. Group I consisted of 84 patients with extrahepatic obstructive jaundice, who received aminoglycoside (gentamicin or tobramycin). Group II consisted of 81 patients with extrahepatic obstructive jaundice, who received either antibiotics other than aminoglycoside or no antimicrobial therapy. Group III consisted of 72 noncholestatic patients receiving aminoglycosides for different disorders. Nephrotoxicity developed in 27 patients (32%) in group I vs 9 patients (11%) in group II and 4 patients (5.6%) in group III (p < 0.00001). In group I, a comparison of patients with and without nephrotoxicity revealed significantly higher values in the former for mean serum bilirubin concentration, initial steady-state trough aminoglycoside concentration and estimated half-life. Stepwise multivariate analysis with nephrotoxicity status as the dependent variable determined that the most significant variable for predicting nephrotoxicity was serum total bilirubin level. In extrahepatic cholestasis a high serum bilirubin level is a distinct factor predisposing to aminoglycoside nephrotoxicity.

Effects of S-adenosyl-l-methionine on lipid peroxidation and glutathione levels in rat brain slices exposed to reoxygenation after oxygen-glucose deprivation

We analyzed the effects of S-adenosyl-L-methionine (AdoMet) on tissue oxidative stress in rat brain slices exposed to reoxygenation after oxygen-glucose deprivation. The thiobarbituric acid reactive substances (TBARS), total and oxidized glutathione, and lactate-dehydrogenase efflux (LDH) from tissue to the incubation medium, were measured. Brain slices were incubated without glucose and with N2, then glucose was added and O2 was perfused. After the anoxic-reoxygenation period, increase in TBARS, oxidized glutathione and LDH efflux, and decrease in total glutathione levels, were observed. The incubation with AdoMet before the anoxic period reduced TBARS (31-1000 micromol/l), glutathione production was increased (31-1000 micromol/l), LDH efflux decreased 6.41% with 15 micromol/l and 61.5% with 500 micromol/l). In the ex vivo experiments, we administered 50 mg/kg per day p.o., AdoMet for 3 days, then brain slices were collected and the anoxia-reoxygenation experiment was carried out. AdoMet led to the inhibition of brain lipid peroxidation and increased total glutathione production, after 3 h-reoxygenation. The increase of LDH efflux in non-treated rats was reduced by 77%. We conclude that AdoMet exerts citoprotective effects in an experimental model of brain slices reoxygenation after oxygen-glucose deprivation.

Effect of virgin olive oil plus acetylsalicylic acid on brain slices damage after hypoxia-reoxygenation in rats with type 1-like diabetes mellitus.

Aspirin is the most widely used drug for the secondary prevention of ischemic stroke in patients suffering from diabetes mellitus. Moreover virgin olive oil (VOO) administration exerts a neuroprotective effect in healthy rat brain slices. The aim of the present study was to determine the possible influence of VOO administration to streptozotocin-diabetic rats (DR) on the neuroprotective effect of aspirin in rat brain. DR were treated during 3 months with saline, aspirin (2mg/kg/day p.o.), VOO (0.5 mL/kg/day p.o.) or its association; a control normoglycemic group was treated with saline. Brain slices were subjected to oxygen-glucose deprivation before a reoxygenation period. All the treatments significantly reduced lactate dehydrogenase LDH efflux after reoxygenation (-54.1% for aspirin, -51.3% for VOO and -72.9% for aspirin plus VOO). Lipid peroxides in brain slices were also reduced after the treatment with aspirin (-17.90%), VOO (-37.3%) and aspirin plus VOO (-49.2%). Production of nitric oxide after reoxygenation was inhibited by all the treatments (-46.5% for ASA, -48.2% for VOO and -75.8% for ASA plus VOO). The activity of the inducible isoform (iNOS) was inhibited by the three types of treatment (-31.8% for ASA, -29.1% for VOO and -56.0% for ASA plus VOO). The main conclusion of our study is that daily oral administration of VOO to diabetic rats may be a natural way to increase the neuroprotective effect of aspirin in diabetic animals.

Effects of dexibuprofen on platelet function in humans: comparison with low-dose aspirin.

Background:The aim of the current study is to evaluate the antiplatelet effect of dexibuprofen in healthy volunteers in comparison with low-dose aspirin. Methods:Healthy volunteers (n = 12) were treated in a crossover manner with 100 mg daily aspirin or with 800 mg daily dexibuprofen. Blood samples were obtained within 24 h; 3, 7, and 14 days after repeated doses; and 24 h after the last dose. In each sample, the authors measured platelet aggregation, thromboxane B2, 6-keto-prostaglandin F1α, and nitric oxide. Results:The antiplatelet effect of dexibuprofen (maximal inhibition of aggregation was 48–55% for adenosine diphosphate and 90–95% for collagen and arachidonic acid) was equal to the effect of aspirin. The main difference between the two drugs was in the degree of recovery of platelet function. The effect of aspirin persisted for 24 h after the last dose (remaining inhibition 50%, respect to the pretreatment value), whereas platelet aggregation had returned to baseline pretreatment values within 24 h after dexibuprofen was stopped. Conclusions:Both aspirin and dexibuprofen inhibited platelet function with a similar intensity, but dexibuprofen exerted a reversible effect for 24 h after the last dose.

Influence of vitamin E on the antiplatelet effect of acetylsalicylic acid in human blood

We analysed the in vitro interaction between acetylsalicylic acid and vitamin E on the principal antiplatelet sites of action of acetylsalicylic acid, i.e., platelet aggregation, prostanoid production in platelets and leukocytes, and nitric oxide synthesis. Aggregation was measured in whole blood and in platelet-rich plasma (PRP) with ADP, collagen or arachidonic acid as platelet inducers, and we measured the production of thromboxane B2, prostacyclin and nitric oxide. Vitamin E potentiated the antiplatelet effect of acetylsalicylic acid in both whole blood and PRP. In PRP induced with collagen the IC50 for acetylsalicylic acid alone was 339 ± 11.26, and that of acetylsalicylic acid + vitamin E was 0.89 ± 0.09 (P < 0.05). Vitamin E did not enhance inhibition of platelet thromboxane production by acetylsalicylic acid. Vitamin E spared or even increased prostacyclin levels, and acetylsalicylic acid + vitamin E diminished the inhibition of prostacyclin synthesis by acetylsalicylic acid (IC50 acetylsalicylic acid alone = 1.81 ± 0.15 µM; IC50 acetylsalicylic acid + vitamin E = 12.92 ± 1.10 µM, P < 0.05). Vitamin E increased the effect of acetylsalicylic acid on neutrophil nitric oxide production 42-fold (P < 0.05). We conclude that vitamin E potentiates the antiplatelet effect of acetylsalicylic acid in vitro, and thus merits further research in ex vivo studies.