Effects of pirfenidone on renal fibrosis in mice with diabetic nephropathy and its mechanisms

Objective To investigate effects of pirfenidone (PFD) on diabetic nephropathy model in db/db mice and to explore its possible mechanisms. Methods (1) Wild-type mice were as the normal control group, and db/db mice were divided into model group and PFD group, with 6 mice in each group. In the PFD group mice were administered continuously by 250 mg·kg-1·d-1 PFD for 18 weeks, and mice in the other two groups were administered by 0.5% sodium carboxymethyl cellulose. Blood glucose and 24 h urinary albumin were measured. The pathological changes of renal tissue were evaluated by PAS staining, PASM staining, Masson staining and Sirius red staining. The expression of collagen type Ⅳ in kidney tissues was detected by immunohistochemistry. (2) Mouse mesangial cells (SV40 MES-13 cells) were cultured as research objects. They were divided into control group, hyperosmolar group, high glucose (HG) group, and 50, 100, 200, 400, 800, 1600 mg/L PFD+HG group. BrdU cell proliferation test was used to evaluate cell proliferation rate. Cells were divided into control group, hyperosmolar group, HG group and PFD+HG group. The mRNA expressions of α-smooth muscle actin (α-SMA), collagen type Ⅰ, collagen type Ⅳ, transforming growth factor-β1 (TGF-β1), interleukin (IL)-1β, IL-6 and monocyte chemotactic protein-1 (MCP-1) were detected by real-time PCR. Results (1) Compared with normal control group, the model mice had higher weight, blood glucose and 24 h urinary albumin, accompanied with glomerular hypertrophy, mesangial area expansion, tubulointerstitial fibrosis and deposition of collagen type Ⅳ (all P<0.05). Compared with those in model group, in PFD group 24 h urinary albumin decreased, glomerular hypertrophy, mesangial area expansion and tubulointerstitial fibrosis alleviated, and the protein expression of collagen type Ⅳ inhibited (all P<0.05). (2) Compared with those in HG group, MES-13 cell proliferation rates of 100, 200, 400, 800, 1600 mg/L PFD+HG groups decreased (all P<0.05), and the mRNA expressions of α-SMA, collagen type Ⅰ, collagen type Ⅳ, TGF-β1, IL-1β, IL-6 and MCP-1 reduced in 400 mg/L PFD+HG group (all P<0.05). Conclusions PFD can inhibit high glucose-induced proliferation and activation of glomerular mesangial cells, decrease the expression of TGF-β1 and proinflammatory factors, as well as reduce the synthesis of collagen, which improve renal fibrosis of db/db mice. Key words: Diabetic nephropathy; Glomerular mesangial cells; Fibrosis; Inflammatory; Pirfenidone