ABSTRACT
Objective To observe the effect of JLP deficiency on the progression of renal interstitial fibrosis in mice model of unilateral ureteral obstruction (UUO),and to investigate the role and underlying mechanism of JLP in the development of renal fibrosis in obstructive nephropathy.Methods jlp Wild type (jlp+/+) and jlp deficient (jlp-/-) mice were divided into four groups:jlp+/+-and jlp-/--sham-operated groups(jlp-/--sham group and jlp+/+-sham group),jlp+/+-and jlp-/--unilateral ureteral obstruction (UUO)-operated groups (jlp/--UUO group and jlp+/+-UUO group).Mice were sacrificed at 7 days and 14 days after the operation respectively to evaluate the fibrosis by Masson staining.The expression of JLP in jlp +/+ renal tissue was assayed by immunohistochemistry staining,immunofluorescence and Western blotting.Immunohistochemical staining was used to detect the expression of α-smooth muscle actin (α-SMA),collagen Ⅰ (COL-Ⅰ),collagen Ⅲ (COL-Ⅲ) and transforming growth factor-β1 (TGF-β1) in sham and UUO groups.Besides,the α-SMA,COL-Ⅰ,COL-Ⅲ,TGF-β1,p-Smad2 and p-Smad3 protein levels were also analyzed by Western blotting in four groups.Results The expression of JLP was mainly demonstrated in the renal tubules of mice.A large amount of collagen deposition was observed in the renal interstitial area in jlp-/--UUO group compared to jlp+/+-UUO group.Similarly,the expression of α-SMA,COL-Ⅰ,COL-Ⅲ and TGF-β1 was significantly increased in the kidney cortices in jlp-/--UUO-operated groups.Meanwhile,Western blotting showed that the expression of α-SMA,COL-Ⅰ,COL-Ⅲ,and TGF-β1 protein was obviously higher in jlp-/--UUO group.Moreover,the expression of p-Smad2 and p-Smad3 protein was markedly higher in jlp-/--UUO group.Conclusion Scaffolding protein JLP is critical in preventing renal fibrosis through the inhibition of TGF-β1 expression and myo-fibroblast production.
ABSTRACT
Objective To investigate the effect of macrophage polarization on tubulointerstitial fibrosis of mouse unilateral ureteral obstruction(UUO) model.Methods Twelve male C57BL/6J mice were employed,each of which with an age of 8 to 10 weeks.UUO model was established with these mice with the method of unilateral ureteral ligation.Mice were then sacrificed on the 7th and 14th day respectively after operation,and renal tissue specimens were obtained.The authors detected collagen deposition by Masson staining,and alpha smooth muscle actin (alpha SMA) as well as collagen type Ⅰ (Coll-1) mRNA by real-time quantitative PCR.The authors also detected the degree of renal interstitial macrophages infiltration and expression changes of polarization by immunofluorescence staining.Results Compared with the mice that were observed on the 7th day after operation,the degree of renal interstitial fibrosis in mice observed on the 14th day after operation was comparatively serious,the difference shown by semi-quantitative results was statistically significant (P ＜ 0.05).Moreover,mice observed on the 14th day after operation have more M2 macrophages,the difference between two groups of mice was statistically significant (P ＜ 0.05).On the contrary,there was no statistically significant difference in the degree of M1 macrophages infiltration between these two groups of mice.Conclusions In the renal interstitial fibrosis model induced by UUO,the degree of macrophage infiltration increased significantly,mainly resulted from M2 macrophage infiltration,suggesting that M2 macrophages were involved in the formation of renal fibrosis.