Aberrant expression of the PREX2-MAGI2 axis modulates myofibroblast differentiation in pulmonary fibrosis.

Myofibroblast differentiation is a pivotal event in idiopathic pulmonary fibrosis (IPF) and is driven by transforming growth factor-β1 (TGF-β1)-dependent phosphatase and tensin homolog deleted on chromosome 10 (PTEN) destabilization; however, the intermediate regulators remain incompletely defined. We tested three lung fibroblast cell lines (1 normal and 2 IPF derived) and identified the phosphatidylinositol 3,4,5-trisphosphate-dependent Rac exchanger 2 (PREX2)-membrane-associated guanylate kinase inverted 2 (MAGI2) as a critical regulator of myofibroblast differentiation. Loss-of-function analysis of MAGI2 in normal human lung fibroblasts revealed a shift toward a myofibroblast phenotype characterized by abundant α-smooth muscle actin (α-SMA) expression. In vitro experiments revealed increased PREX2 expression and decreased MAGI2 expression in IPF-derived lung fibroblasts compared with those in normal lung fibroblasts. Consistent with this finding, in vivo experiments using bleomycin-induced murine pulmonary fibrosis revealed PREX2 upregulation in contrast to MAGI2 downregulation with the progression of lesions. Notably, PREX2 depletion in IPF-derived cells enhanced MAGI2 expression, mitigating TGF-β1-induced α-SMA expression. Conversely, PREX2 overexpression promoted myofibroblast differentiation, as evidenced by MAGI2 downregulation followed by marked α-SMA expression. The PREX2-targeting suppressive miR-338-3p upregulated MAGI2 expression in vitro and ameliorated bleomycin-induced murine pulmonary fibrosis in vivo. Immunohistochemical staining of PREX2 on lung samples obtained from patients with IPF delineated that PREX2 was exclusively expressed in the fibroblastic foci of IPF-affected lungs. Fluorescence immunostaining revealed PREX2 colocalization with α-SMA in myofibroblasts within fibroblastic foci, corroborating our hypothesis. These findings suggest that MAGI2 acts as a negative regulator of myofibroblast differentiation by stabilizing PTEN, whereas PREX2 is aberrantly expressed in the fibrotic milieu, negatively modulating MAGI2, thereby accelerating pulmonary fibrosis.NEW & NOTEWORTHY In the fibrotic milieu, upregulated phosphatidylinositol 3,4,5-trisphosphate-dependent Rac exchanger 2 (PREX2) and downregulated membrane-associated guanylate kinase inverted 2 (MAGI2) expression were observed as evidenced by in vivo and in vitro analyses. PREX2 was exclusively expressed in fibroblastic foci in lung samples derived from patients with idiopathic pulmonary fibrosis (IPF) and was colocalized with α-smooth muscle actin (α-SMA) in myofibroblasts within these foci. Thus, the aberrantly expressed PREX2-MAGI2 axis modulates myofibroblast differentiation in IPF, whereby PREX2 negatively modulates MAGI2, destabilizing phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and ultimately accelerating pulmonary fibrosis.