Assessment of eritropoetin effect on stem cells in damaged tissue of diabetic rats

Objective: Diabetes mellitus is a metabolic disease characterized with chronic hyperglycemia and disorders of carbohydrate, protein and lipid metabolism. Incidence of the disease increases in recent years and the pathology has become a global public health problem. Clinical importance of the disease is related with its complications. Diabetes mellitus increases the tendency of atherosclerotic disorders and ischemic heart disease. Upto date effort was given to the effects of diabetes on endothelium and coronary arteries in many researches. In this study, we planned to investigate the effects of diabetes directly on cardiac and aortic tissues. Studies performed on stem cells have gained advance on cardiac surgery in the recent years. However, the diabetic patient group has not been taken into consideration in most of these studies. On the other hand, it is very well known that diabetes mellitus is also a risk factor for many of the postoperative morbidities leading to alterations on stem cell levels. In this study, the effects of diabetes mellitus on cardiovascular system were investigated by assessing the stem cells levels in serum and heart and compared with the normal population. Additionally, efficacy of erythropoietin, which is known to increase stem cells, was studied on diabetic rats. Material and Methods: Twenty-five male Spragie Dawley rats were divided into three groups as; control group (group 1), diabetic group (group 2) and glutamin induced diabetic group (group 3). Diabetes model was created by damaging the pancreatic beta-cells irreversibly with intraperitoneal 50mg/kg Streptozocin (STZ). Rats were survived one month in order to see the side effects of diabetes. In group 3, with the purpose to induce stem cells, rats received 3000U/kg erythropoietin two times in the fouth week. Euthanasia was performed at the end of one month and blood and adequate tissue samples were obtained from right atrium, right ventricle, left atrium, and left ventricle. Blood reticulocyte levels were determined automatically with automatic hemogram device and changes were assessed in 3 groups. Tissues were stained with routine H/E and Mason-Tricrom to determine the degree of fibrozis. Tissues staining for immunohistochemical assessment of a reliable stem cell marker, the CD34 and VEGF, a necessary predictor of tissue regeneration were analyzed. Histologic analysis scores were determined as; 0 point: CD34/VEGF negative; 1 point: 10% of cells being stained for CD34/VEGF; 2 points: 10-50% of cells being stained for CD34/VEGF; 3 points: more than 50% of cells being stained for CD34/VEGF. Results: The glucose levels were measured to be 72,80±11,516mg/dl among the rats in all groups before the procedure. They increased to 358,70±44,749mg/dl and 382,80±34,941mg/dl at the end of second day and first week, respectively, following STZ administration. This increase in the blood glucose levels resulted in significant decrease in reticulocyte levels in group 2. Additionally, the increase in blood glucose levels resulted in statistically significant increase in tissue level of fibrozis, CD34 and VEGF. When the rats in Group 1 and 2 were compared the fibrozis, CD34 and VEGF levels were found to increase statistically significant. When the group 3 was compared with the other groups, the tissue level of CD34 and VEGF were more significantly enhanced. When group 2 and group 3 were compared, even though the myocardial tissues expressed fibrozis, especially the amount of fibrozis was lower in group 3 tissues. In addition, the levels of CD34 and VEGF were significantly higher in group 3 than group 2. Conclusion: Fibrozis occurs and increases in the cardiovascular system in rats in case of diabetes mellitus. The amount of CD34 and VEGF which function in cellular protection and tissue regeneration may be enhanced with safely applicable erythropoietin. We investigated the levels of CD34 and VEGF in the cardiovascular system in diabetic and erythropoietin induced diabetic rats for the first time in the literature. We believe the originality of the study is raised by measurement of reticulocyte levels in serum, and CD34 and VEGF levels in right atrium, right ventricle, left atrium, and left ventricle.