Early diabetes mellitus is characterized by glomerular hyperfiltration, hyperperfusion and increase in kidney size in both human and experimental animals (1,2). These alterations may play a role in the subsequent nephropathy (sclerosis) associated with diabetes (3). We have been able to establish and characterize mesangial cell lines derived from normal and streptozotocin (STZ)-treated diabetic Sprague-Dawley rats and demonstrate that cultured mesangial cells derived from STZ-treated diabetic Sprague- Dawley rats and demonstrate that cultured mesangial cells derived from STZ- treated rats achieved higher cell saturation densities than the normal counterparts (4). This cell growth was associated with a higher proportion of """"""""diabetic"""""""" cells in the S+G2-M stages of the cell cycle during both the exponential cell growth and the plateau phase. In addition, we observed that there was a diminished intracellular calcium response to angiotensin II (AII) in """"""""diabetic"""""""" compared to normal mesangial cells (50. An impaired response to AII is consistent with other reported findings and can partially explain the hemodynamic changes seen in early diabetes mellitus. There appears to be no defect in calcium mobilization, however, since these """"""""diabetic"""""""" mesangial cells responded strongly to arginine vasopressin (AVP). Based on these observations, we proposed to further evaluate mesangial cell function in early diabetes, using this unique in vitro """"""""diabetic"""""""" mesangial cell model. The overall objective of the study is to assess the role of mesangial cells in diabetes mellitus.
Specific aims are: 1. To evaluate whether the enhanced proliferative response of """"""""diabetic"""""""" mesangial cells is associated with an impaired response to stimulatory and/or inhibitory mitogenic factors. 2. To determine the expression of surface receptors ot insulin-like growth factor in normal and 'diabetic"""""""" mesangial cells during normal and high glucose conditions, and the regulation of the IGF-1 receptor expression at different stages of cell growth. 3. To examine whether athe alterations in intracellular calcium concentration in normal and 'diabetic"""""""" mesangial cells in response to AII and AVP are associated with changes in the contractile response to these agents and/or the receptor expression for AII and AVP. We will test the hypothesis that changes in mesangial cell function mediate the alterations in glomerular hemodynamics characteristic of the onset of diabetes mellitus. To achieve this purpose, in vitro mesangial cell lines derived from normal and STZ-treated (STZ; 65 mg/kg body weight) diabetic male Sprague-Dawley rats will be used. Mesangial cell proliferation and cell cycle distribution in response to a variety of mitogenic factors will be determined by thymidine incorporation and flow cytometric analysis. The contractile response of mesangial cells to vasoactive hormones will be measured by image analysis, while binding studies will be performed to assess receptor expression. This research project will provide a unique training experience for both graduate and undergraduate students interested in learning and applying biomedical research techniques such as experimental animal handling, cell culture methodology , flow cytometry analysis, image analysis, radiobinding techniques, as well as statistical data analysis.
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