Insulin-dependent diabetes is an autoimmune disease characterized by T cell-mediated invasion and destruction of pancreatic beta cells. Studies have shown that the insulin receptor located in the membrane of T cells is a chemotactic receptor capable of directing cell movement to the islet in response to Recently, increased insulin receptor expression on T cells of non-obese diabetic mouse has been associated with the aggressively diabetogenic mechanism for delivering autoreactive T cells to the pancreatic islet. Based on these observations, the investigators have designed a study to evaluate the role of the insulin receptor expression in the pathogenesis of type I diabetes. The main objective of the proposal is to quantify the expression of insulin receptor of a transfected well-known immortal T cell line (molt4) in response to changes in insulin and glucose concentration, using Enhanced Green Fluorescent Protein (EGFP) as a marker for gene expression. The PI will test the hypothesis that increased insulin receptor expression on T cells due to alterations in glucose levels, facilitates pancreatic T cell infiltration and consequently, P cell death The following specific aims have been identified: (a) Characterize the EGFP calibration standards to be used to quantitate the expression of the insulin receptor; (b) Generate the EGFP/insulin receptor construct and transfect Molt4 T cells. (c) Quantify the expression of the insulin receptor as a function of glucose concentration on transfected Molt4 T cells. (d) Examine the effect of glucose concentration on insulin binding and insulin receptor internalization, as well as (e) Assess the effect of selected cytokines on the expression of the insulin receptor in transfected Molt4 T cells.
| Gerena-Lopez, Y; Nolan, J; Wang, L et al. (2004) Quantification of EGFP expression on Molt-4 T cells using calibration standards. Cytometry A 60:21-8 |
