Jelizaveta Sokolovska

Influence of metformin on GLUT1 gene and protein expression in rat streptozotocin diabetes mellitus model.

Context: Metformin improves hyperglycaemia via mechanisms which include activation of AMP-activated protein kinase (AMPK). Recent findings indicate that some metabolic actions of metformin occur also by AMPK-independent mechanisms. Objective: To study the action of metformin on expression of GLUT1 glucose transporter in rat streptozotocin model of diabetes mellitus. Materials and methods: Streptozotocin-induced rats were treated with metformin while monitoring parameters of carbohydrate and lipid metabolism. GLUT1 mRNA and protein expression in kidneys, heart, liver and muscles were studied by means of real time quantitative RT-PCR and immunohistochemistry correspondingly. Results: Metformin treatment decreased glucose concentration, glycated haemoglobin % and improved glucose tolerance. Streptozotocin diabetes provoked increase of both GLUT1 gene and protein expression in kidneys, metformin treatment produced normalization of the GLUT1 expression levels. In the liver, diabetes triggered an increase in GLUT1 protein expression, which was normalized by metformin. Conclusion: Metformin is prospective for treatment of diabetic nephropathy.

Correction of glycaemia and GLUT1 level by mildronate in rat streptozotocin diabetes mellitus model.

Anti‐ischaemic drug mildronate suppresses fatty acid metabolism and increases glucose utilization in myocardium. It was proposed that it could produce a favourable effect on metabolic parameters and glucose transport in diabetic animals. Rats with streptozotocin diabetes mellitus were treated with mildronate (100 mg/kg daily, per os, 6 weeks). Therapeutic effect of mildronate was monitored by measuring animal weight, concentrations of blood glucose, insulin, blood triglycerides, free fatty acids, blood ketone bodies and cholesterol, glycated haemoglobin per cent (HbA1c%) and glucose tolerance. GLUT1 mRNA and protein expression in kidneys, heart, liver and muscles were studied by means of real time RT‐PCR and immunohistochemistry correspondingly. In the streptozotocin + mildronate group, mildronate treatment caused a significant decrease in mean blood glucose, cholesterol, free fatty acid and HbA1c concentrations and improved glucose tolerance. Induction of streptozotocin diabetes mellitus provoked increase of both GLUT1 gene and protein expression in kidneys, heart and muscle, mildronate treatment produced normalization of the GLUT1 expression levels. In the liver a similar effect was observed for GLUT1 protein expression, while GLUT1 gene expression was increased by mildronate. Mildronate produces therapeutic effect in streptozotocin diabetes model. Mildronate normalizes the GLUT1 expression up‐regulated by streptozotocin diabetes mellitus in kidneys, heart, muscle and liver. Copyright

1,4-Dihydropyridine derivatives without Ca2+-antagonist activity up-regulate Psma6 mRNA expression in kidneys of intact and diabetic rats

Impaired degradation of proteins by the ubiquitin–proteasome system (UPS) is observed in numerous pathologies including diabetes mellitus (DM) and its complications. Dysregulation of proteasomal degradation might be because of altered expression of genes and proteins involved in the UPS. The search for novel compounds able to normalize expression of the UPS appears to be a topical problem. A novel group of 1,4‐dihydropyridine (1,4‐DHP) derivatives lacking Ca2+‐antagonists activities, but capable to produce antidiabetic, antioxidant and DNA repair enhancing effects, were tested for ability to modify Psma6 mRNA expression levels in rat kidneys and blood in healthy animals and in rats with streptozotocin (STZ) induced DM. Psma6 gene was chosen for the study, as polymorphisms of its human analogue are associated with DM and cardiovascular diseases. 1,4‐DHP derivatives (metcarbatone, etcarbatone, glutapyrone, J‐9‐125 and AV‐153‐Na) were administered per os for three days (0.05 mg/kg and/or 0.5 mg/kg). Psma6 gene expression levels were evaluated by quantitative PCR.

New 1,4-Dihydropyridines Down-regulate Nitric Oxide in Animals with Streptozotocin-induced Diabetes Mellitus and Protect Deoxyribonucleic Acid against Peroxynitrite Action.

Diabetes mellitus (DM) and its complications cause numerous health and social problems throughout the world. Pathogenic actions of nitric oxide (NO) are responsible to a large extent for development of complications of DM. Search for compounds regulating NO production in patients with DM is thus important for the development of pharmacological drugs. Dihydropyridines (1,4‐DHPs) are prospective compounds from this point of view. The goals of this study were to study the in vivo effects of new DHPs on NO and reactive nitrogen and oxygen species production in a streptozotocin (STZ)‐induced model of DM in rats and to study their ability to protect DNA against nocive action of peroxynitrite. STZ‐induced diabetes caused an increase in NO production in the liver, kidneys, blood and muscles, but a decrease in NO in adipose tissue of STZ‐treated animals. Cerebrocrast treatment was followed by normalization of NO production in the liver, kidneys and blood. Two other DHPs, etaftorone and fenoftorone, were effective in decreasing NO production in kidneys, blood and muscles of diabetic animals. Furthermore, inhibitors of nitric oxide synthase (NOS) and an inhibitor of xanthine oxidoreductase (XOR) decreased NO production in kidneys of diabetic animals. Treatment with etaftorone decreased expression of inducible NOS and XOR in kidneys, whereas it increased the expression of endothelial NOS. In vitro, the studied DHPs did not significantly inhibit the activities of NOS and XOR but affected the reactivity of peroxynitrite with DNA. These new DHPs thus appear of strong interest for treatment of DM complications.

Effects of an Antimutagenic 1,4‐Dihydropyridine AV‐153 on Expression of Nitric Oxide Synthases and DNA Repair‐related Enzymes and Genes in Kidneys of Rats with a Streptozotocin Model of Diabetes Mellitus

Development of complications of diabetes mellitus (DM), including diabetic nephropathy, is a complex multi‐stage process, dependent on many factors including the modification of nitric oxide (NO) production and an impaired DNA repair. The goal of this work was to study in vivo effects of 1,4‐dihydropyridine AV‐153, known as antimutagen and DNA binder, on the expression of several genes and proteins involved in NO metabolism and DNA repair in the kidneys of rats with a streptozotocin (STZ)‐induced model of DM. Transcription intensity was monitored by means of real‐time RT‐PCR and the expression of proteins by immunohistochemistry. Development of DM significantly induced PARP1 protein expression, while AV‐153 (0.5 mg/kg) administration decreased it. AV‐153 increased the expression of Parp1 gene in the kidneys of both intact and diabetic animals. Expression of H2afx mRNA and γH2AX histone protein, a marker of DNA breakage, was not changed in diabetic animals, but AV‐153 up‐regulated the expression of the gene without any impact on the protein expression. Development of DM was followed by a significant increase in iNOS enzyme expression, while AV‐153 down‐regulated the enzyme expression up to normal levels. iNos gene expression was also found to be increased in diabetic animals, but unlike the protein, the expression of mRNA was found to be enhanced by AV‐153 administration. Expression of both eNOS protein and eNos gene in the kidneys was down‐regulated, and the administration of AV‐153 normalized the expression level. The effects of the compound in the kidneys of diabetic animals appear to be beneficial, as a trend for the normalization of expression of NO synthases is observed.

Poly-ADP ribose polymerase, xanthine oxidase and nitric oxide synthase expression in kidney tissue of experimental diabetes

Background: The spatial distribution of inflammatory and DNA reparation markers in the kidney tissue in diabetes is poorly understood. Aim: The present study investigated the role of endothelial and inducible nitric oxide synthase (eNOS and iNOS), Poly ADP ribose polymerase (PARP) xanthine oxidase (XO) in the pathogenesis of streptozotocin-induced diabetic changes in the kidney tissue. Methods: Diabetes mellitus was induced in rats by a single injection of streptozotocin (STZ) at a dose of 50 mg/kg. The XO, PARP, eNOS and iNOS protein expression in the kidney was studied by immunohistochemistry. Results: Obtained results showed that STZ administration incresed the numbers of PARP and XO-positive cells in kidney tissue compared to control group (respectively, 42 ± 8 vs 10 ± 2, p = 0.01; and 27 ± 7 vs 8 ± 2 cells/mm2, p = 0.002). At contrast, the eNOS expression was also observed in medullary collecting tubules, and it was decreased in STZ-treated rats. Conclusion: The present study suggests that increased PARP, XO in proximal tubules and decreased eNOS expression in medullary collection tubules, with concomitant increased eNOS and iNOS expression in the microvasculature plays a pivotal role in the kidney damage in diabetes mellitus. Supported by LCS grant Number 278/2012.