Figure 4.

Sam68 overexpression increases the leptin-dependent activation of PI3K and MAPK pathways in human GCs from healthy donors. GCs were transfected with Sam68 plasmid and pcDNA3 (empty vector) as control (basal) as described in Materials and Methods, and incubated in absence or presence of leptin 10 nM for 10 min. GCs lysates were separated by SDS–PAGE and Western blot analysis was performed by using anti-P-IRS-1, anti-P-PKB and anti-P-ERK1/2 antibodies to study leptin activation of the PI3K and MAPK signaling pathway. Sample protein loading was controlled by using anti-GAPDH antibodies. We show the corresponding densitometric analysis of three independent experiments as means ± s.d. Statistical analyses were performed by ANOVA. Asterisks indicate significant differences from the control and Sam68 plasmid without leptin. # indicate significant differences from leptin 10 nM and ‘ns’ no statistically significant difference according to Bonferroni’s multiple comparison post hoc test.