BackgroundThis study aimed to investigate the pathophysiology of hepatic microcirculatory dysfunction in non-alcoholic fatty liver disease (NAFLD).MethodsIn Wistar rats, NAFLD model was induced by 20 weeks of high-fat diet (HFD) feeding. Rolling and adhesion of leukocytes and tissue perfusion in hepatic microcirculation were examined using in vivo microscopic and laser speckle contrast imaging (LSCI), respectively. Oxidative stress and inflamatory parameters were analysed by TBARs, catalase enzyme activity, RT-PCR and ELISA. The participation of advanced glycation end-products (AGE) and its receptor RAGE was evaluated by the measurement of gene and protein expression of RAGE by RT-PCR and Western-blot, respectively and by liver and serum quantification of fluorescent AGEs.ResultsWistar rats fed high-fat diet (HFD) showed increase in epididymal and abdominal fat content, systolic arterial blood pressure, fasting blood glucose levels, hepatic triglycerides and cholesterol, and impairment of glucose and insulin metabolisms. Liver histology confirmed the presence of steatosis and ultrasound analysis revealed increased liver size and parenchymal echogenicity in HFD-fed rats. HFD causes significant increases in leukocyte rolling and adhesion on hepatic microcirculation and decrease in liver microvascular blood flow. Liver tissue presented increase in oxidative stress and inflammtion. At 20 weeks, there was a significantly increase in AGE content in the liver and serum of HFD-fed rats and an increase in RAGE gene expression in the liver.ConclusionThe increase in liver AGE levels and microcirculatory disturbances could play a role in the pathogenesis of liver injury and are key components of NAFLD.

ObjectiveWe investigated the protective effects of pyridoxamine against metabolic and microcirculatory complications in nonalcoholic fatty liver disease.MethodsNonalcoholic fatty liver disease was established by a high‐fat diet administration over 28 weeks. Pyridoxamine was administered between weeks 20 and 28. The recruitment of leukocytes and the number of vitamin A‐positive hepatic stellate cells were examined by in vivo microscopy. Laser speckle contrast imaging was used to evaluate microcirculatory hepatic perfusion. Thiobarbituric acid reactive substances measurement and RT‐PCR were used for oxidative stress and inflammatory parameters. advanced glycation end products were evaluated by fluorescence spectroscopy.ResultsThe increase in body, liver, and fat weights, together with steatosis and impairment in glucose metabolism observed in the nonalcoholic fatty liver disease group were attenuated by pyridoxamine treatment. Regarding the hepatic microcirculatory parameters, rats with high‐fat diet‐induced nonalcoholic fatty liver disease showed increased rolling and adhesion of leukocytes, increased hepatic stellate cells activation, and decreased tissue perfusion, which were reverted by pyridoxamine. Pyridoxamine protected against the increased hepatic lipid peroxidation observed in the nonalcoholic fatty liver disease group. Pyridoxamine treatment was associated with increased levels of tumor necrosis factor alpha (TNF‐α) mRNA transcripts in the liver.ConclusionPyridoxamine modulates oxidative stress, advanced glycation end products, TNF‐α transcripts levels, and metabolic disturbances, being a potential treatment for nonalcoholic fatty liver disease‐associated microcirculatory and metabolic complications.

Introduction: Combined antiretroviral therapy (cART) used to treat acquired immunodeficiency virus (HIV) induces a number of adverse effects, such as insulin resistance and dyslipidemia, which ultimately increases the cardiovascular risk. Advanced glycation end products (AGEs) have been implicated in the etiology of cardiovascular diseases, diabetes and other chronic diseases. It is known that physical exercise improves the lipid profile, insulin resistance and reduces the risk of cardiovascular diseases. However, the impact of physical exercise on AGE levels in HIV-infected patients has not been so far investigated. Therefore, this study compared AGEs levels in people with and without HIV and verified the effect of physical training on serum AGE levels.Methods: Participants were initially assigned into three groups: healthy control (CTL, n = 35), physically inactive HIV-infected (In-HIV, n = 33) and physically active HIV-infected (Ac-HIV, n = 19). The In-HIV group underwent physical training for 3 months, consisting of 60-min sessions of multimodal supervised exercise (aerobic, resistance and flexibility) with moderate intensity (50–80% heart rate reserve), performed 3 times/week. AGEs were measured in serum by fluorescence spectrometry.Results: At baseline, serum AGEs fluorescence level was significantly higher in inactive HIV-patients when compared to controls or active HIV-patients (In-HIV: 0.93 ± 0.08 vs. controls: 0.68 ± 0.13 and Ac-HIV: 0.59 ± 0.04 A.U.; P < 0.001). Triglycerides were also higher in In-HIV than CTL (182.8 ± 102 vs. 132.8 ± 52.3 mg/dL; P < 0.05). Waist circumference was lower in Ac-HIV, compared to In-HIV and controls (83.9 ± 10.4 vs. 92.9 ± 13.5 and 98.3 ± 12.4, respectively; P < 0.05). Body mass, fasting blood glucose, LDL, HDL, and total cholesterol were similar between groups. After training, AGE levels decreased (Baseline: 0.93 ± 0.08 vs. 3 months follow-up: 0.59 ± 0.04 AU; P < 0.001), no further difference being detected vs. CTL or Ac-HIV. Conclusion: HIV-infected patients under cART exhibited elevated AGEs levels compared to healthy individuals and physically active patients. Short-term aerobic training of moderate intensity counteracted this condition.

In this study we have explored the pathogenesis of the hepatic alterations which occur in diabetes and the modulation of these complications by the combination of metformin adjunct treatment and insulin monotherapy. For this purpose, diabetic rats were treated with insulin (DM + Ins) or metformin plus insulin (DM + Met + Ins). Biochemical and cardiometabolic parameters were analysed by spectrophotometry. Intravital microscopy was used to study the hepatic microcirculation. In the liver tissue, real-time PCR was used to analyse oxidative stress enzymes, inflammatory markers and receptors for advanced glycation end products (AGE) (RAGE) gene expression. Lipid peroxidation was assessed by thiobarbituric acid reactive species (TBARs) analyses. AGE deposition and RAGE protein expression were studied by fluorescence spectrophotometry and Western blot respectively. Body weight, %HbA1c , urea, total proteins and oxidative stress parameters were found to be similarly improved by insulin or Met + Ins treatments. On the other hand, Met + Ins treatment showed a more pronounced effect on fasting blood glucose level than insulin monotherapy. Fructosamine, uric acid, creatinine, albumin and amylase levels and daily insulin dose requirements were found to be only improved by the combined Met + Ins treatment. Liver, renal and pancreatic dysfunction markers were found to be more positively affected by metformin adjunct therapy when compared to insulin treatment. Liver microcirculation damage was found to be completely protected by Met + Ins treatment, while insulin monotherapy showed no effect. Our results suggest that oxidative stress, microcirculatory damage and glycated proteins could be involved in the aetiology of liver disease due to diabetes. Additionally, metformin adjunct treatment improved systemic and liver injury in induced diabetes and showed a more pronounced effect than insulin monotherapy.

This study aimed to investigate changes in renal function and the AGE-RAGE axis in the kidney of a non-genetic animal model of metabolic syndrome (MetS) induced by high-fat diet (HFD). Additionally, we evaluated the protective effect of pyridoxamine (PM), a vitamin B6 analog with anti-AGE effects, in the context of diet-related renal endothelial dysfunction. Methodology: In Wistar rats, the MetS animal model was induced by 20 or 28 weeks of HFD feeding. When indicated, a subgroup of animals was treated daily with PM (60 mg/kg) for 2 months. Tissue perfusion in renal microcirculation was examined by laser speckle contrast imaging. Oxidative stress was analyzed by thiobarbituric acid reactive species and the inflammatory markers by ELISA (TNF-α and IL-1β). Reverse transcription polymerase chain reaction was used to analyze eNOs, IL-6, vascular cell adhesion molecule (VCAM), NADPH oxidase subunit 47 (N47), catalase, and receptor for AGE (RAGE) gene expression. Results: Wistar rats fed a HFD showed negligible alteration in renal function, decrease in catalase mRNA transcripts and catalase enzyme activity compared to control (CTL) animals. Increased levels of IL-1β were observed in the kidney of MetS-induced rats. HFD-fed rats exhibited kidney endothelial dysfunction, with no significant differences in basal microvascular blood flow. PM significantly improved kidney vasorelaxation in HFD-fed rats. eNOS, VCAM, and RAGE gene expression and AGE content were not altered in kidneys of HFDinduced MetS rats in comparison to CTLs. Conclusions: Our findings suggest that HFD-induced microvascular dysfunction precedes the decline in renal function, and could be related to antioxidant machinery defects and inflammation activation in the kidney. PM showed a vasoprotective effect, and thus, could be an important contributory factor in ameliorating diet-induced renal damage.