Glycogen synthase kinase‐3 ( GSK ‐3) regulates TGF ‐β 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 ‐β 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 ‐β 1 ‐induced myofibroblast differentiation is currently largely unknown. Purpose To determine the contribution of GSK ‐3 signalling in TGF ‐β 1 ‐induced myofibroblast differentiation. Experimental Approach We used MRC 5 human lung fibroblasts and primary pulmonary fibroblasts of individuals with and without COPD . Protein and m RNA expression were determined by immunoblotting and RT‐PCR analysis respectively. Results Stimulation of MRC 5 and primary human lung fibroblasts with TGF ‐β 1 resulted in time‐ and dose‐dependent increases of α‐sm‐actin and fibronectin expression, indicative of myofibroblast differentiation. Pharmacological inhibition of GSK ‐3 by SB 216763 dose‐dependently attenuated TGF ‐β 1 ‐induced expression of these myofibroblasts markers. Moreover, silencing of GSK ‐3 by si RNA or pharmacological inhibition by CT/CHIR 99021 fully inhibited the TGF ‐β 1 ‐induced expression of α‐sm‐actin and fibronectin. The effect of GSK ‐3 inhibition on α‐sm‐actin expression was similar in fibroblasts from individuals with and without COPD . Neither smad, NF‐κB nor ERK 1/2 were involved in the inhibitory actions of GSK ‐3 inhibition by SB 126763 on myofibroblast differentiation. Rather, SB 216763 increased the phosphorylation of CREB , which in its phosphorylated form acts as a functional antagonist of TGF ‐β/smad signalling. Conclusion and Implication We demonstrate that GSK ‐3 signalling regulates TGF ‐β 1 ‐induced myofibroblast differentiation by regulating CREB phosphorylation. GSK ‐3 may constitute a useful target for treatment of chronic lung diseases.