Alceu Afonso Jordão

Effects of creatine supplementation on homocysteine levels and lipid peroxidation in rats.

Hyperhomocysteinaemia is an independent risk factor for CVD. Recent data show a relationship between homocysteine (Hcy) and free radical formation. Since creatine synthesis is responsible for most of the methyl group transfers that result in Hcy formation, creatine supplementation might inhibit Hcy production and reduce free radical formation. The present study investigated the effects of creatine supplementation on Hcy levels and lipid peroxidation biomarkers. Thirty rats were divided into three groups: control group; diet with creatine group (DCr; 2 % creatine in the diet for 28 d); creatine overload plus diet with creatine group (CrO + D; 5 g creatine/kg by oral administration for 5 d+2 % in the diet for 23 d). Plasma Hcy was significantly lower (P < 0.05) in DCr (7.5 (sd 1.2) micromol/l) and CrO + D (7.2 (sd 1.7) micromol/l) groups compared with the control group (12.4 (sd 2.2) micromol/l). Both plasma thiobarbituric acid-reactive species (TBARS) (control, 10 (sd 3.4); DCr, 4.9 (sd 0.7); CrO + D, 2.4 (sd 1) micromol/l) and plasma total glutathione (control, 4.3 (sd 1.9); DCr, 2.5 (sd 0.8); CrO + D, 1.8 (sd 0.5) micromol/l) were lower in the groups that received creatine (P < 0.05). In addition, Hcy showed significant negative correlation (P < 0.05) with plasma creatine (r - 0.61) and positive correlation with plasma TBARS (r 0.74). Plasma creatine was negatively correlated with plasma TBARS (r - 0.75) and total peroxide (r - 0.40). We conclude that creatine supplementation reduces plasma Hcy levels and lipid peroxidation biomarkers, suggesting a protective role against oxidative damage. Modulating Hcy formation may, however, influence glutathione synthesis and thereby affect the redox state of the cells.

Creatine Supplementation Prevents the Accumulation of Fat in the Livers of Rats Fed a High-Fat Diet

The aim of the present study was to examine the effects of creatine supplementation on liver fat accumulation induced by a high-fat diet in rats. Rats were fed 1 of 3 different diets for 3 wk: a control liquid diet (C), a high-fat liquid diet (HF), or a high-fat liquid diet supplemented with creatine (HFC). The C and HF diets contained, respectively, 35 and 71% of energy derived from fat. Creatine supplementation involved the addition of 1% (wt:v) of creatine monohydrate to the liquid diet. The HF diet increased total liver fat concentration, liver TG, and liver TBARS and decreased the hepatic S-adenosylmethionine (SAM) concentration. Creatine supplementation normalized all of these perturbations. Creatine supplementation significantly decreased the renal activity of l-arginine:glycine amidinotransferase and plasma guanidinoacetate and prevented the decrease in hepatic SAM concentration in rats fed the HF diet. However, there was no change in either the phosphatidylcholine:phosphatidylethanolamine (PE) ratio or PE N-methyltransferase activity. The HF diet decreased mRNA for PPARα as well as 2 of its targets, carnitine palmitoyltransferase and long-chain acylCoA dehydrogenase. Creatine supplementation normalized these mRNA levels. In conclusion, creatine supplementation prevented the fatty liver induced by feeding rats a HF diet, probably by normalization of the expression of key genes of β-oxidation.

Inhibitory Effects of Lutein and Lycopene on Placental Glutathione S-Transferase-Positive Preneoplastic Lesions and DNA Strand Breakage Induced in Wistar Rats by the Resistant Hepatocyte Model of Hepatocarcinogenesis

Inhibitory effects of lutein (LUT) and lycopene (LYC) on hepatic preneoplastic lesions (PNLs) and DNA strand breakage induced in Wistar rats by the resistant hepatocyte (RH) model of hepatocarcinogenesis were investigated. Animals received by gavage during 8 consecutive weeks on alternate days 70 mg/kg body weight of LUT or LYC. Rats treated with only corn oil and submitted to this model were used as controls. At the end of the experiment, treatment of the animals with LUT or LYC resulted in an increase in the respective liver carotenoid concentrations (P < 0.05). Moreover, it tended to reduce the incidence, total number, and multiplicity of hepatocyte nodules compared with the control group, although the differences did not reach statistical significance. Animals treated with LUT or LYC presented also a lower number of hepatic placental glutathione S-transferase-positive (GST-P) PNLs (P < 0.05), which were smaller (P < 0.05) and occupied a smaller area of the liver section (P < 0.05). Finally, hepatic DNA strand breakage evaluated by the comet assay was lower (P < 0.05) in carotenoid-treated animals when compared with the control group. Therefore, the results indicate that LUT and LYC represent promising chemopreventive agents during hepatocarcinogenesis and whose anticarcinogenic actions could be related to a protection against DNA instability.

Effects of creatine supplementation on oxidative stress and inflammatory markers after repeated-sprint exercise in humans.

OBJECTIVE The goal of this study was to evaluate the effects of creatine (Cr) supplementation on oxidative stress and inflammation markers after acute repeated-sprint exercise in humans. METHODS Twenty-five players under age 20 y were randomly assigned to two groups: Cr supplemented and placebo. Double-blind controlled supplementation was performed using Cr (0.3 g/kg) or placebo tablets for 7 d. Before and after 7 d of supplementation, the athletes performed two consecutive Running-based Anaerobic Sprint Tests (RAST). RAST consisted of six 35-m sprint runs at maximum speed with 10 sec rest between them. Blood samples were collected just prior to start of test (pre), just after the completion (0 h), and 1 h after completion. RESULTS Average, maximum, and minimum power values were greater in the Cr-supplemented group compared with placebo (P < 0.05). There were significant increases (P < 0.05) in plasma tumor necrosis factor alpha (TNF-α) and C-reactive protein (CRP) up to 1 h after acute sprint exercise in the placebo-supplemented group. Malondialdehyde, lactate dehydrogenase (LDH), catalase, and superoxide dismutase enzymes also were increased after exercise in both groups. Red blood cell glutathione was lower after exercise in both groups. Cr supplementation reversed the increase in TNF-α and CRP as well as LDH induced by acute exercise. Controversially, Cr supplementation did not inhibit the rise in oxidative stress markers. Also, antioxidant enzyme activity was not different between placebo and Cr-supplemented groups. CONCLUSION Cr supplementation inhibited the increase of inflammation markers TNF-α and CRP, but not oxidative stress markers, due to acute exercise.

Antioxidant Effect of Thiamine on Acutely Alcoholized Rats and Lack of Efficacy Using Thiamine or Glucose to Reduce Blood Alcohol Content

Although there is no consensus about the use of glucose and thiamine for the treatment of acute ethanol intoxication, this is a routine practice in many countries. Our objective was to determine the efficacy of this treatment and the changes it causes in the antioxidant status of the liver. Male Wistar rats were intoxicated with an ethanol dose of 5 g/kg and divided into three groups: ethanol (EtOH; untreated), EtOH+G (treated with glucose), and EtOH+B1 (treated with thiamine). Blood and urinary ethanol as well as hepatic malondialdehyde, reduced glutathione and vitamin E were determined in all animals. Blood alcohol levels did not differ between groups, although urinary excretion was about four times higher in the group treated with thiamine (EtOH+B1). The malondialdehyde, reduced glutathione and vitamin E values used here as parameters of the antioxidant system of the liver showed improvement for the thiamine-treated group (EtOH+B1). Treatment with glucose or thiamine was ineffective in reducing blood alcohol levels in rats with acute ethanol intoxication. However, the beneficial effect of thiamine as an antioxidant for ethanol metabolism was demonstrated. Further investigations are necessary to clarify the urinary excretion of ethanol reported here for the first time and the possibility of using thiamine as an antioxidant in situations of chronic alcohol use.

Protein Oxidative Stress and Dyslipidemia in Dialysis Patients

Our aim was to investigate and determine the associations between oxidative stress (OS), dyslipidemia and inflammation in patients treated with continuous ambulatory peritoneal dialysis (CAPD) and hemodialysis (HD) using observational cross‐sectional study. Twenty patients in CAPD and 48 in HD for at least 8 weeks and aged ≥18 years were included in the study. Individuals with malignant or acute inflammatory disease were excluded. A control group of 17 healthy individuals was also recruited. The biochemical parameter evaluations were analyzed using colorimetric kits for albumin, serum glucose, total cholesterol (TC) and lipid fractions. To determine the inflammatory status, CRP, IL‐6 and TNF‐α were analyzed by automated chemiluminescence kits. Plasma advanced oxidation protein products (AOPP) were determined by spectrophotometry. Mean AOPP levels were significantly higher for the HD group compared to the control, and there was no difference in AOPP concentrations between the control and CAPD groups. Dialysis patients had levels of inflammatory parameters higher than controls, and showed a high prevalence of patients with dyslipidemia, especially in CAPD. In the HD group, AOPP was positively correlated with triglycerides (TG) and inversely associated with HDL. Also the HD group was observed to have negative associations between TNF‐α and HDL, LDL and TC. In the CAPD group, CRP was inversely correlated with HDL. Hemodialysis patients had increased protein OS and associations of inflammation and dyslipidemia were also observed in these dialysis groups. A more detailed characterization of the relations between oxidative stress and other more traditional risk factors has therapeutic importance, since cardiovascular diseases are the leading cause of death among dialysis patients.

Effect of NFκB Inhibition by CAPE on Skeletal Muscle Ischemia-Reperfusion Injury

BACKGROUND/AIMS Nuclear factor kappa B (NFkappaB) plays important role in the pathogenesis of skeletal muscle ischemia/reperfusion (I/R) injury. Caffeic acid phenyl ester (CAPE), a potent NFkappaB inhibitor, exhibits protective effects on I/R injury in some tissues. In this report, the effect of CAPE on skeletal muscle I/R injury in rats was studied. METHODS Wistar rats were submitted to sham operation, 120-min hindlimb ischemia, or 120-min hindlimb ischemia plus saline or CAPE treatment followed by 4-h reperfusion. Gastrocnemius muscle injury was evaluated by serum aminotransferase levels, muscle edema, tissue glutathione and malondialdehyde measurement, and scoring of histological damage. Apoptotic nuclei were determined by a terminal uridine deoxynucleotidyl transferase dUTP nick end labeling assay. Muscle neutrophil and mast cell accumulation were also assessed. Lipoperoxidation products and NFkappaB were evaluated by 4-hydroxynonenal and NFkappaB p65 immunohistochemistry, respectively. RESULTS Animals submitted to ischemia showed a marked increase in aminotransferases after reperfusion, but with lower levels in the CAPE group. Tissue glutathione levels declined gradually during ischemia to reperfusion, and were partially recovered with CAPE treatment. The histological damage score, muscle edema percentage, tissue malondialdehyde content, apoptosis index, and neutrophil and mast cell infiltration, as well as 4-hydroxynonenal and NFkappaB p65 labeling, were higher in animals submitted to I/R compared with the ischemia group. However, the CAPE treatment significantly reduced all of these alterations. CONCLUSIONS CAPE was able to protect skeletal muscle against I/R injury in rats. This effect may be associated with the inhibition of the NFkappaB signaling pathway and decrease of the tissue inflammatory response following skeletal muscle I/R.

Lipid peroxidation and vitamin E in serum and follicular fluid of infertile women with peritoneal endometriosis submitted to controlled ovarian hyperstimulation: a pilot study.

OBJECTIVE To assess the level of lipid peroxidation (LP) and vitamin E in the follicular fluid and serum of infertile patients, with or without endometriosis, who were submitted to ovulation induction for assisted reproduction procedures. DESIGN Prospective study. SETTING Assisted conception unit, university hospital. PATIENT(S) Infertile patients 20 to 38 years of age were selected prospectively and consecutively and were divided into the endometriosis group (17 patients with pelvic endometriosis) and the control group (19 patients with previous tubal ligation or male factor and without endometriosis). INTERVENTION(S) Peripheral blood samples were collected on D1 (before the beginning of the use of gonadotropins), D2 (day of hCG administration), and D3 (day of oocyte retrieval). On D3, follicular-fluid samples free from blood contamination also were collected and stored. MAIN OUTCOME MEASURE(S) Lipid peroxidation was assessed by malondialdehyde quantification by spectrophotometry, and measurement of vitamin E was performed by HLPC. RESULT(S) On D1, no significant difference in LP was observed between groups. However, vitamin E levels were significantly higher in the control group. On D2, LP levels were significantly higher in the endometriosis group compared with in the control group, and vitamin E levels continued to be significantly higher in the control group. On D3, there was no significant difference in serum and follicular-fluid levels of LP and vitamin E between groups. However, on D3, vitamin E levels were found to be significantly higher in serum than in follicular fluid in both groups, whereas malondialdehyde levels were significantly lower in follicular fluid than in serum only in the control group. CONCLUSION(S) Before the beginning of ovulation induction, a significant decrease in vitamin E was observed in patients with endometriosis, perhaps because antioxidants are consumed during oxidation reactions. After ovulation induction with exogenous gonadotropins, the group of patients with endometriosis not only presented increased lipid peroxidation but also maintained lower vitamin E levels than the control group, a fact that hypothetically could compromise oocyte quality in endometriotic patients. However, on the day of oocyte retrieval, both serum LP potential and vitamin E levels were found to be similar in the two groups.

Blood and salivary oxidative stress biomarkers following an acute session of resistance exercise in humans.

The aim of the present study was to compare oxidative stress biomarkers determined in blood and saliva before and after acute resistance exercise. 1 week after 1 maximum repetition (1RM) test 11 healthy well-trained males completed a hypertrophy acute session of resistance training including 3 sets of 10 repetitions at 75% of the 1RM, with 90 s rest periods between sets. Venous blood and saliva samples were collected before (pre) and 10 min after (post) the resistance training session. A significant (p<0.05) rise in blood lactate accumulation (pre: 1.6+/-0.4 vs. post: 9.5+/-2.4) was found post-acute resistance training compared with baseline values. Significant increases (p<0.05) in TBARS (42%), AOPP (28%), uric acid (27%) and GSH (14%) were detected post-acute resistance training in relation to pre in blood samples. A significant increase (p<0.05) in uric acid (36%) was found in saliva post-acute resistance training as well as a significant correlation (p<0.05) between uric acid determined in blood and saliva. Statistical analysis did not reveal any other change in the salivary oxidative stress biomarkers. In conclusion, an acute session of resistance exercise induces oxidative stress in plasma of trained men after acute resistance training, which was not found in saliva samples except for uric acid.

Validation of a Manual Headspace Gas Chromatography Method for Determining Volatile Compounds in Biological Fluids

Background We report the validation of a method for the determination of acetaldehyde, acetone, methanol, and ethanol in biological fluids using manual headspace sample introduction and an acetonitrile internal standard. Methods This method uses a capillary column ( l = 30 m, I.D. = 0.25 mm, d F = 0.25 μm) installed in a gas chromatography–flame ionization detector (GC-FID) apparatus with a run time of 7.5 minutes. Results Analysis of the retention times and the resolution of the analyte peaks demonstrated excellent separation without widening of the peaks. Precision and accuracy were good (interassay precision <15% and recovery between 85% and 115%) in both blood and urine. Conclusion The method was linear (r >0.99) over the analytical measurement range (AMR) of each analyte.