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UBC Theses and Dissertations

Investigation of insulin secretory abnormalities in pancreatic islets from diabetic and non diabetic BB rats Curtis, Susan B.


Functional studies of islets isolated from diabetic, nondiabetic and prediabetic BB rats were performed to investigate the hypothesis that a primary alteration exists in the pancreatic β-cells of BB rats which results in abnormal insulin secretion and precedes cell mediated autoimmune mechanisms. Islets from diabetic and nondiabetic BB rats were isolated and compared with similar islets from Wistar Furth (WF) rats after short-term culture to determine if β-cells demonstrate altered insulin secretion in response to in vitro stimuli. A marked attenuation in the insulin response to glucose and GIP was present in newly diagnosed diabetic BB rats, as well as in nondiabetic BB rats (120 days of age), which would never develop diabetes. Coincident with the marked reduction in glucose-stimulated insulin secretion, was an exaggerated arginine-stimulated insulin secretion. The appropriate response to the inhibitory action of somatostatin was retained. The insulin content of islets isolated from recent onset diabetic rats was similar to control rats, indicating that insulin stores were not limiting insulin secretion The amount of insulin released from a given β-cell mass was profoundly reduced compared to control animals, not just from diabetic but also from nondiabetic BB rats. Preproinsulin mRNA was detected in islets from diabetic rats using nonisotopicin situ hybridization (ISH) techniques. These results indicated that insulin biosynthesis was still present, and that the islets from these rats were not simply releasing stored insulin. A marked variation in the intensity of staining was observed both between and within islets of the same animal. In many instances, a stronger intensity of the ISH signal was observed in diabetic islets compared with normal islets. Immunocytochemical data from this study demonstrated a pronounced reduction in GLUT-2 immunoreactivity in remaining β-cells at the onset of diabetes, which may have contributed to the loss of glucose-stimulated insulin secretion at the onset of diabetes. The glucose response of cultured islets from diabetic animals was profoundly reduced, yet levels of glucose transporters in unaffected islets of diabetic animals were in many instances normal. In addition, a minor reduction in GLUT-2 immunoreactivity was observed in nondiabetic rats which also had an attenuated response to glucose. Assuming normal glucose transporter function of the islets in this study, then factors other than glucose transport must be considered to explain the reduced insulin response to glucose. Due to possible insulitis, isolation of islets from diabetic and nondiabetic BB rats does not entirely remove them from influences of the immune system. These experiments, therefore, would not distinguish between a defect intrinsic to the β-cell from one that is imposed on it by the immune system. Consequently similar studies were performed using islets isolated from young BB rats (40, 50, and 60 days of age) prior to the cell mediated immune response was examined. Islets from BB rats have normal glucose-induced insulin secretion, but an unusually high arginine response is present in the prediabetic (D60) period. BB rats demonstrated circulating autoantibodies directed to an unidentified cytoplasmic target (or targets) in enteroendocrine cells, nerve cells, and pancreatic islet cells. Islet cytoplasmic autoantibodies occurred at an early stage in both diabetic and nondiabetic animals and, therefore, could not be used as a predictive test to differentiate the two groups of animals. It was concluded that because the incidence of diabetes in litters from the Ottawa colony of BB rats varies from 50-70%, the presence of young diabetes resistant rats could obscure a possible effect in true prediabetic rats. To date, there is no predictive test to differentiate between the two groups of animals. In an attempt to overcome this problem rats from the Worcester colony (BB/W) which have an incidence of diabetes that is > 90% were obtained. Static incubation experiments were repeated using BB/W rat islets, to determine if glucose recognition was impaired in truly ‘prediabetic’ rats. Diabetes resistant (DR) BB rats were also included as controls. Two significant changes to β-cell function were observed in the BB/W rat islets. Glucose-induced insulin secretion was found to be reduced prior to the onset of diabetes (by D60), in DP and DR rats relative to BB and WF rat islets, supporting the existence of an intrinsic defect to the β-cell of BB/W rats. The presence of the defect in the DR rats led to the conclusion that the defect was not linked to the autoimmune response. Hyper responsiveness to arginine was identified as a second abnormality of insulin secretion found in BB/W as well as BB rats. A decline in GLUT-2 immunoreactivity was not found to be an early marker of the prediabetic period in this study, and cannot account for the decrease in glucose-stimulated insulin secretion.

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