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Insulin Stimulates Primary β-Cell Proliferation via Raf-1 Kinase

Laboratory of Molecular Signalling in Diabetes, Diabetes Research Group, Department of Cellular and Physiological Science and Department of Surgery, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3

Address all correspondence and requests for reprints to: James D. Johnson, Ph.D., Assistant Professor. Diabetes Research Group, Department of Cellular and Physiological Sciences, and Department of Surgery, University of British Columbia, 5358 Life Sciences Building, 2350 Health Sciences Mall, Vancouver, British Columbia, Canada V6T 1Z3. E-mail: jimjohn{at}interchange.ubc.ca.

A relative decrease in β-cell mass is key in the pathogenesis of type 1 diabetes, type 2 diabetes, and in the failure of transplanted islet grafts. It is now clear that β-cell duplication plays a dominant role in the regulation of adult β-cell mass. Therefore, knowledge of the endogenous regulators of β-cell replication is critical for understanding the physiological control of β-cell mass and for harnessing this process therapeutically. We have shown that concentrations of insulin known to exist in vivo act directly on β-cells to promote survival. Whether insulin stimulates adult β-cell proliferation remains unclear. We tested this hypothesis using dispersed primary mouse islet cells double labeled with 5-bromo-2-deoxyuridine and insulin antisera. Treating cells with 200-pM insulin significantly increased proliferation from a baseline rate of 0.15% per day. Elevating glucose from 5–15 mM did not significantly increase β-cell replication. β-Cell proliferation was inhibited by somatostatin as well as inhibitors of insulin signaling. Interestingly, inhibiting Raf-1 kinase blocked proliferation stimulated by low, but not high (superphysiological), insulin doses. Insulin-stimulated mouse insulinoma cell proliferation was dependent on both phosphatidylinositol 3-kinase/Akt and Raf-1/MAPK kinase pathways. Overexpression of Raf-1 was sufficient to increase proliferation in the absence of insulin, whereas a dominant-negative Raf-1 reduced proliferation in the presence of 200-pM insulin. Together, these results demonstrate for the first time that insulin, at levels that have been measured in vivo, can directly stimulate β-cell proliferation and that Raf-1 kinase is involved in this process. These findings have significant implications for the understanding of the regulation of β-cell mass in both the hyperinsulinemic and insulin-deficient states that occur in the various forms of diabetes.