Figure 6

Overexpression of ERBB4 in MSCs unexpectedly activates a novel NRG1-ERBB4-NRG1 autocrine loop. Overexpressing ERBB4 in MSCs unexpectedly upregulated the synthesize and secretion of its ligand NRG1, detected by conducting western blotting (a) and ELISA (b), respectively. (c) Overexpressing ERBB4 in MSCs upregulated NRG1, but did not alter expression of ERBB2. ERBB3 remained negative before and after ERBB4 overexpressing. (d) Multiple transient transfections using pER4-GFP confirmed that upregulated NRG1 was attributed to exogenous ERBB4 overexpression. (d1) With increased transfection frequency shown by encircular numbers (d1), GFP signal, which indicated transfection efficiency, was escalating accordingly, from 1.2% for the first time (d1) to 32.7% for the fifth time (d1). (d2) The expression of NRG1 was upregulated along with increased transfection efficiency. (e) In human 293FT cells, NRG1 expression was upregulated after ERBB4 transduction. The aforementioned experiments were repeated three times and representative images are shown. 293FTe, 293FT expressing pGFP; 293FT-ERBB4, 293FT expressing pER4-GFP