Glycogen synthase kinase-3 (GSK-3) regulates TGF-1-induced differentiation of pulmonary fibroblasts

Background Chronic lung diseases such as asthma, COPD and pulmonary fibrosis are characterized by abnormal extracellular matrix (ECM) turnover. TGF- is a key mediator stimulating ECM production by recruiting and activating lung fibroblasts and initiating their differentiation process into more active myofibroblasts. Glycogen synthase kinase-3 (GSK-3) regulates various intracellular signalling pathways; its role in TGF-1-induced myofibroblast differentiation is currently largely unknown. Purpose To determine the contribution of GSK-3 signalling in TGF-1-induced myofibroblast differentiation. Experimental Approach We used MRC5 human lung fibroblasts and primary pulmonary fibroblasts of individuals with and without COPD. Protein and mRNA expression were determined by immunoblotting and RT-PCR analysis respectively. Results Stimulation of MRC5 and primary human lung fibroblasts with TGF-1 resulted in time- and dose-dependent increases of -sm-actin and fibronectin expression, indicative of myofibroblast differentiation. Pharmacological inhibition of GSK-3 by SB216763 dose-dependently attenuated TGF-1-induced expression of these myofibroblasts markers. Moreover, silencing of GSK-3 by siRNA or pharmacological inhibition by CT/CHIR99021 fully inhibited the TGF-1-induced expression of -sm-actin and fibronectin. The effect of GSK-3 inhibition on -sm-actin expression was similar in fibroblasts from individuals with and without COPD. Neither smad, NF-B nor ERK1/2 were involved in the inhibitory actions of GSK-3 inhibition by SB126763 on myofibroblast differentiation. Rather, SB216763 increased the phosphorylation of CREB, which in its phosphorylated form acts as a functional antagonist of TGF-/smad signalling. Conclusion and Implication We demonstrate that GSK-3 signalling regulates TGF-1-induced myofibroblast differentiation by regulating CREB phosphorylation. GSK-3 may constitute a useful target for treatment of chronic lung diseases