Effects of Human Type P Insulin Mediator on Protein Expression in Developing Mouse Cerebral Cortex Cells

Type 2 diabetes, the more prevalent form of diabetes mellitus, is caused by insulin resistance in peripheral tissues due to a post-receptor defect in the insulin-signaling pathway. Binding of insulin to its receptor in the cell membrane generates second messengers or mediators. These insulin mediators are inositol phosphoglycan in structure. The type P insulin mediator is defective in type 2 diabetics and likely plays a crucial role in insulin resistance. Previous work in this lab has shown that human type P insulin mediator (hPIM) increases cell proliferation and differentiation and inhibits apoptosis in chicken fibroblasts and rat hippocampal cells. This study looked at specific protein expression changes induced by hPIM in embryonic mouse cerebral cortex cells. Sephadex size exclusion chromatography columns were used to isolate hPIM from human plasma. Cortices were dissected from E15 mouse embryos. After 4 days of growth, cortical cells were treated with hPIM for 24 hours. Protein was extracted from cells using FASP method, and proteomic analysis was performed using LC-MS/MS. The MS data showed up regulation of many proteins that promote mitogenesis, cell differentiation, protein expression, and overall cell health. Less than 10 proteins were down regulated by hPIM. Down regulation of these proteins also promotes cell proliferation and protein expression. The results from this study support previous findings that hPIM promotes cell viability, proliferation, and differentiation and provide further insight into hPIM's direct action in the cell. Optimizing protein extraction and LC-MS/MS results from cell culture is also discussed.