促红细胞生成素及其受体在慢性环孢素A肾毒性大鼠肾组织中的表达

Expression of erythropoietin and its receptor in renal tissues of rats with chronic cyclosporine A nephrotoxicity

目的探讨慢性环孢素A(CsA)能否导致贫血,以及促红细胞生成素(EPO)及其受体(EPOR)在慢性CsA肾毒性大鼠肾组织中的表达变化。方法 SD大鼠给予CsA15mg/(kg·d)皮下注射4周建立慢性CsA肾毒性模型(CsA毒性组),对照组1mL/(kg·d)皮下注射橄榄油。采用全自动生化分析仪检测两组大鼠的肾功能,血红蛋白(Hb)、红细胞压积(Hct)水平,三色(Masson Trichrome)染色确定肾小管间质纤维化程度;免疫组织化学染色和蛋白质印迹法分别检测肾组织EPO及EPOR蛋白的表达;TUNEL染色和电子显微镜观察细胞凋亡。结果与对照组相比,CsA毒性组大鼠肾功能低下,肾小管间质发生纤维化,凋亡细胞增多(P<0.01);同时CsA毒性组大鼠有贫血发生,表现为Hb和Hct水平的下降(P<0.01)。免疫组织化学染色和蛋白质印迹分析结果表明,EPO在CsA毒性组肾组织中的表达减少,而EPOR的表达增加(P<0.01)。直线相关分析提示,EPO蛋白表达与肾小管间质纤维化(r=-0.729,P<0.001)和TUNEL阳性细胞数(r=-0.841,P<0.001)呈负相关。结论慢性CsA肾毒性大鼠肾组织中EPO蛋白表达减少,从而导致贫血;CsA诱导肾小管上皮细胞凋亡与EPO蛋白表达减少有关。

Objective To examine whether cyclosporine A （CsA） treatment can induce anemia and changes in expression of erythropoietin （EPO） and its receptor （EPOR） in renal tissues of rats with chronic CsA nephrotoxicity. Methods Sprague- Dawley rats were treated daily with subcutaneous injection of vehicle （olive oil, 1 mL/kg, control group） or CsA （15 mg/kg, CsA nephrotoxicity model group） for 4 weeks. The renal function and hemoglobin （Hb） and hematocrit （Hct） levels were analyzed by Automatic Biochemistry Analyzer in the two groups. Masson Trichrome staining was used to examine the extent of tubulointerstitial fibrosis. Immunohistochemistry and Western blotting analysis were used to examine the expression of EPO and EPOR protein. And cell apoptosis was observed by TUNEL assay and electron microscope. Results Compared with the control group, CsA nephrotoxicity group showed renal insufficiency, tuhulointerstitial fibrosis, and significantly more apoptotic ceils （P^0.01）. Meanwhile, CsA nephrotoxicity group also had anemia, manifested by significantly decreased Hb and Hct levels （P〈0. 01）. Immunohistochemistry and Western blotting analysis showed that CsA treatment significantly decreased EPO expression and significantly increased EPOR expression compared with the control group （P〈0. 01）. Furthermore, we also found that EPO protein expression was negatively associated with tubulointerstitial fibrosis （r= --0. 729, P〈0. 001） and the number of TUNEL-positive cells （r = --0. 841, P〈 0. 001）. Conclusion Renal EPO expression is decreased in rats with chronic CsA nephrotoxicity, which resulting in anemia; and tubular epithelial cell apoptosis induced by CsA is related to decrease of EPO protein.