Pirfenidone inhibits transforming growth factor β₁/transforming growth factor-activated kinase 1 pathway to improve peritoneal fibrosis in rats

Objective To observe the effect of pirfenidone（PFD） on high glucose-induced peritoneal fibrosis（PF） and explore its molecular mechanism in rats. Methods Male Sprague-Dawley rats were infused with 100 mL/（kg·d） of 4.25% glucose-based standard peritoneal dialysis fluid for 4 weeks. The rats were divided into four groups: control group（Ctr group）, peritoneal fibrosis group（PF group）, PF rats with PFD 250 mg/（kg·d）（PF-PFD250 group） and PF rats with PFD 750 mg/（kg·d）（PF-PFD750 group）. Ultrafiltration volume（UFV） and mass transfer of glucose（MTG） were used to assess peritoneal transport function. Masson staining was performed to evaluate the deposition of collagen in peritoneal. Immunohistochemistry was performed to measure the expression levels of transforming growth factor β₁（TGF-β₁）, α smooth muscle actin（α-SMA） and E-cadherin. Western-blot was performed to measure the expression levels of TGF-β₁, α-SMA, E-cadherin, collagen-Ⅰ, transforming growth factor-activated kinase 1（TAK1）/posphorylated TAK1（P-TAK1） and P38/posphorylated P38（P-P38）. In vitro, peritoneal mesothelial cells（PMCs） were cultured, which were divided into blank group, TGF-β₁ group, TGF-β₁+PFD(10^-3 mol/L) group and TGF-β₁+P38 inhibitor（SB203580 20 nmol/L） group. Cell counting kit-8（CCK-8） assay was used to measure cell viability. Western-blot was performed to measure the expression levels of α-SMA, E-cadherin, collagen-Ⅰ, TAK1/P-TAK1 and P38/P-P38. Results Compared with control group, PF group showed decreased UFV （1.588±0.955） vs（9.675±2.626） mL, P＜0.01 and increased MTG （18.625±3.042） vs（8.880±1.940） mmol/kg, P＜0.01 as well as markedly increased peritoneal thickness and higher expression of TGF-β₁, α-SMA, collagen-Ⅰ, P-TAK1 and P-P38, lower expression of E-cadherin（all P＜0.01）. Compared with PF group, PF-PFD250 group and PF-PFD750 group showed increased UFV and decreased MTG as well as markedly decreased peritoneal thickness（all P＜0.01）. The elevated TGF-β₁, α-SMA, collagen-Ⅰ, P-TAK1 and P-P38 in PF rats were significantly decreased and the declined E-cadherin in PF rats were significantly increased by PFD（all P＜0.01）. In vitro, compared with blank group, the cell viability decreased, the expression of P-TAK1, P-P38, α-SMA, collagen-Ⅰ increased and the expression of E-cadherin decreased（all P＜0.01） in TGF-β₁ group. Compared with TGF-β₁ group, the cell viability increased, the expression of α-SMA and collagen-Ⅰ were significantly decreased and the expression of E-cadherin was significantly increased in TGF-β₁+PFD group and TGF-β₁+P-P38 inhibitor group（all P＜0.01）. Both P-TAK1 and P-P38 expression decreased in TGF-β₁+PFD group（P＜0.01）, while only P-P38 expression decreased in TGF-β₁+PFD group（P＜0.01）. Conclusions Pirfenidone shows a protective effect on PF by inhibition of peritoneal extracellular matrix accumulation and peritoneal epithelial-mesenchymal transition, the underlying mechanism may be partly blockade of TGF-β₁/TAK1 pathway.