| First Author | Morran MP | Year | 2018 |
| Journal | J Biol Methods | Volume | 5 |
| Issue | 1 | PubMed ID | 29862308 |
| Mgi Jnum | J:341545 | Mgi Id | MGI:7541152 |
| Doi | 10.14440/jbm.2018.209 | Citation | Morran MP, et al. (2018) Insulin receptor based lymphocyte trafficking in the progression of type 1 diabetes. J Biol Methods 5(1):e85 |
| abstractText | The insulin receptor (IR) is a transmembrane receptor which recognizes and binds the hormone insulin. We describe two models that were devised to explore the role of IR over-expression on T-lymphocytes and their chemotactic motility in the progression of type 1 diabetes. FVB/NJ-CD3-3xFLAG-mIR/MFM mice were generated to selectively over-express 3xFLAG tagged murine IR in T-lymphocytes via an engineered CD3 enhancer and promoter construct. Insertion of the 3xFLAG-mIR transgene into FVB/NJ mice, a known non-autoimmune prone strain, lead to a minor population of detectable 3xFLAG-mIR tagged T-lymphocytes in peripheral blood and the presence of a few lymphocytes in the pancreas of the Tg+/- compared to age matched Tg-/- control mice. In order to induce stronger murine IR over-expression then what was observed with the CD3 enhancer promoter construct, a second system utilizing the strong CAG viral promoter was generated. This system induces cell specific IR over-expression upon Cre-Lox recombination to afford functional 3xFLAG tagged murine IR with an internal eGFP reporter. The pPNTlox2-3xFLAG-mIR plasmid was constructed and validated in HEK-Cre-RFP cells to ensure selective Cre recombinase based 3xFLAG-mIR expression, receptor ligand affinity towards insulin, and functional initiation of signal transduction upon insulin stimulation. |
