Connective tissue growth factor mediates growth differentiation factor 8-induced increase of lysyl oxidase activity in human granulosa-lutein cells

• GDF8 up-regulates CTGF expression and secretion.
• GDF8 up-regulates LOX expression and increases LOX activity.
• CTGF mediates the GDF8-induced increases in LOX expression and activity.
• ALK5 and SMAD2/SMAD3-SMAD4 are involved in the GDF8-induced up-regulation of CTGF.
• SMAD3 binds to the CTGF promoter.

Lysyl oxidase (LOX) is an essential enzyme for the stabilization of the extracellular matrix (ECM) and the subsequent follicle and oocyte maturation. Currently, there is limited information pertaining to the regulation of LOX activity in human ovarian tissue. Growth differentiation factor 8 (GDF8) is a unique member of the transforming growth factor-β superfamily that is expressed in human granulosa cells and has important roles in regulating a variety of ovarian functions. The aim of the present study was to investigate the effects of GDF8 on the regulation of LOX expression and activity in human granulosa cells and to examine the underlying molecular determinants. An established immortalized human granulosa cell line (SVOG) and primary granulosa-lutein cells were used as study models. Using dual inhibition approaches (TGF-β type I inhibitor SB505124 and small interfering RNAs) and ChIP analyses, we have demonstrated that GDF8 up-regulated the expression of connective tissue growth factor (CTGF) through the activin receptor-like kinase 5-mediated SMAD2/3-SMAD4 signaling pathways. In addition, the increase in CTGF expression contributed to the GDF8-induced increase in LOX expression and activity. Our findings suggest that GDF8 and CTGF may play critical roles in the regulation of ECM formation in human granulosa cells.