The Role of Piezo1 in Regulating Collagen Expression in Orbital Fibroblasts under High Pressure

Abstract

Purpose.To explore the effect of Pizeo 1 on the expression of collagen Ⅰ\Ⅲ\Ⅴof orbital fibroblasts in pressure culture simulated the high orbital pressure of thyroid-associated ophthalmopathy (TAO). Methods.The primary orbital fibroblasts of guinea pig were isolated and cultured by enzymatic digestion method, and the expression levels of Piezo1, α-SMA, and collagenⅠ\Ⅲ\Ⅴin the cells under different pressure (0, 1, 2, and 3Kpa) were checked by WB and PCR. And then the orbital fibroblasts were cultured under a constant pressure of 3KPa and treated with different concentrations (1, 5, and 10 μmol/L) of Piezo1 inhibitor GsMTx4, respectively. The expression levels of Piezo1, α-SMA, and collagenⅠ\Ⅲ\Ⅴ in the orbital fibroblasts were observed. CCK8 and Flow cytometry were used to observe the cell proliferation and apoptosis. Results.With the increase of pressure, the expression levels of Piezo1 and collagenⅠ\Ⅲ\Ⅴ protein and mRNA in the orbital fibroblasts increased significantly (P< 0.05). Meanwhile, inhibition of Piezo1 by GsMTx4 significantly reduced the levels of OFs transdifferentiation and collagenⅠ\Ⅲ\Ⅴ under the high pressure (P< 0.05). Furthermore, the proliferation of orbital fibroblasts was enhanced with the increase of pressure. Under the co-culture of pressure and inhibitor GsMTx4, the low concentration group promoted the proliferation of orbital fibroblasts, and the high concentration group (10μM) promoted the proliferation of orbital fibroblasts at the early stage, but less than the 5μM group. And at the later stage, the proliferation of orbital fibroblasts was inhibited. Compression alone or compression combined with inhibitor did not affect the apoptosis of orbital fibroblasts. Conclusion. Down-regulating the expression of piezo1 can inhibit the transdifferentiation of orbital fibroblasts and the synthesis and secretion of collagenⅠ\Ⅲ\Ⅴ, which provide a new idea for exploring the fibrosis mechanism in TAO.