In addition to IGF 1 and insulin receptors, mammary epithelial cells may also express insulin IGF 1 hybrid receptors. Hybrid receptors have already been detected in most tissues that express each insulin receptor and IGF 1 receptor. An IGF 1 concentration of two. six nM won’t activate the insulin receptor, but could potentially cause the activation from the insulin IGF 1 hybrid recep tors. Data presented in Figure 3C supports this hypoth esis and suggests that IGF 1 signaling has led to the formation of insulin IGF 1 hybrid receptors. Functional studies with hybrid receptors demonstrate that they behave far more like IGF 1 receptors in lieu of insulin receptors simply because they bind IGF 1 with a a lot greater affinity than insulin. As anticipated, we did not observe activation from the hybrid receptor with ten nM insulin.
Although the significance from the hybrid receptors in mammary epithelial cells in unclear, we hypothesize that the insulin IGF 1 hybrids could be much more abundant in MCF10A cells selleck chemical than otherwise anticipated and this hypothesis is supported by reports that insulin and hybrid insulin IGF 1 receptors are important regulators of breast cancer cells. Throughout this study, we are going to refer to the IGF 1R mediated induction in LIP for simplicity, however the reader really should realize that hybrid receptors could also be involved in regulation of LIP LAP. Due to the fact LIP expression is analyzed 16 hr soon after addi tion of ligand, we also checked p EGFR expression at this later time point. EGFR was not phosphorylated in MCF10A cells or MCF 7 cells 16 hr just after addition of IGF 1 To confirm that IGF 1 was indeed activating the IGF 1R signaling cascade, we analyzed p IGF 1R and p Akt expression at 20 min and 16 hr.
To further assess the possibility that EGFR activity may perhaps play a function inside the IGF 1R stimulated boost in LIP expression, we tested the sensitivity of IGF 1 treated MCF10A cells to the selective EGFR kinase inhibitor, MAPK activation AG1478. Pretreatment of cells for 30 minutes with 0. 1, 1 or five uM AG1478 before addition of 2. six nM IGF 1 for 16 hr didn’t inhibit or decrease the IGF 1 mediated increases in LIP expression and did not inhibit the raise in the LIP LAP ratio. As a manage, five uM AG1478 did lead to the expected reduce in p EGFR, decreases in EGF mediated LIP expression and the LIP LAP ratio, and lesser reductions with 0. 1 and 1 uM. Therapy of cells with 0. 1, and 1.
0 uM AG1478 effectively decreased IGF 1 induced Erk1 two phosphorylation and as expected EGF induced Erk1 two phosphorylation. These data demonstrate that inhibition of EGFR kinase activity reduces IGF 1R mediated Erk1 two activity and suggest that IGF 1R and EGFR signaling crosstalk in MCF10As to regulate Erk1 two activity. Our data also demonstrate that inhibition of EGFR signaling with AG1478 does not inhibit IGF 1R induced Akt activity but does block EGF induced Akt activity.