3A. Induction of ObRb mRNA in SH-SY5Y cells and the effects of leptin on STAT3 activation. ObRb mRNA was present in retinoic acid-differentiated cells (lane 1), but not in non-differentiated cells (lane 2) or the no-template negative control (lane 3).
3B. Retinoic acid-induced differentiation of SH-SY5Y cells resulted in subcellular redistribution of ObRb. Immunocytochemistry showed that ObRb was present in the cytoplasm of the undifferentiated cells (middle panel), but at the cell surface of differentiated cells (right panel). The specificity of the staining was shown by the lack of signals in the negative control with secondary antibody only (left panel).
3C. Leptin treatment (30 nM) induced pSTAT3 throughout the study period (10 min – 6 h), with a peak at 3 h. This contrasts with the lack of changes of the β-actin signal.