EPO对鼠肾缺血再灌注损伤后肾小管间质纤维化和Bcl-2/Bax表达的影响
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摘要
目的探讨促红细胞生成素(EPO)对肾缺血再灌注损伤(ischemia reperfusion injury, IRI)后肾小管间质纤维化和凋亡蛋白(Bcl-2/Bax)表达的影响。方法通过单侧肾IRI构建患侧肾小管间质纤维化模型。实验小鼠随机分为四组：假手术组(n=20)；缺血再灌注组(n=20)；EPO低剂量组(造模,EPO100u/kg,1次／周,n=20)；EPO高剂量组(造模,EPO10000u/kg,1次／周,n=20)。成模后4周处死各组小鼠,取其患侧肾脏,HE、Masson染色观察肾脏病理改变,并以免疫组化的方法检测肾组织中Bcl-2和Bax蛋白表达水平。结果①缺血再灌注组肾脏出现不同程度肾小管萎缩或扩张、间质纤维化、炎性细胞浸润,其小管间质纤维化积分明显高于假手术组(P<0.01)；两EPO干预组肾小管和间质病变较缺血再灌注组减轻,小管间质纤维化积分减少(P<0.05),其EPO高剂量组积分下降尤明显(P<0.01)。②免疫组化显示：Bcl-2和Bax主要着染于肾小管和肾间质；缺血再灌注组和两EPO干预组肾脏Bcl-2和Bax表达均较假手术组明显上调(P<0.01),但两EPO干预组肾脏Bcl-2表达上调均高于缺血再灌注组(P<0.01),而Bax表达上调却低于缺血再灌注组(P<0.01,P<0.05)；缺血再灌注组Bcl-2／Bax比值较假手术组低(P<0.01),而两EPO干预组Bcl-2／Bax比值却较缺血再灌注组高(P<0.01)；两EPO干预组比较,仅EPO高剂量组肾脏Bcl-2表达上调强于EPO低剂量组(P<0.01)。③相关分析显示小管间质纤维化积分与Bax水平呈明显正相关(r=0.571,P<0.05),与Bcl-2、Bcl-2/Bax比值呈明显负相关(F=-0.790,P<0.01；r=-0.802,P<0.01)。结论1Bcl-2/Bax蛋白异常表达参与了肾缺血再灌注损伤后期肾小管间质纤维化进程；2.EPO能减轻小鼠肾缺血再灌注损伤后期肾小管间质纤维化程度,且可能有一定的剂量关系；3.EPO对肾缺血再灌注损伤后期肾小管间质纤维化的影响与调控Bcl-2/Bax的表达有一定相关性。
Abstract
Objective:To investigate effects of erythropoietin on renal tubulointerstitial fibrosis and Bcl-2 (anti-apoptotic protein)/Bax(pro-apoptotic protein) expression after renal ischemia/reperfusion injury(IRI) in mice. Methods:80 male C57BL/6 mice with 8-12 weeks age were randomly divided into 4 groups:sham-operated control group(n=20), IRI group(n=20), treatment group with low-dose EPO (model rats treated with EPO 100u/kg, Once a week, n=20), treatment group with high-dose EPO(model rats treated with EPO 1000u/kg, once a week, n=20). Left renal artery of mice was clamped for 35min with micro-artery clamp to make model of renal IRI. The morphological changes were observed by HE and Masson stain. Meanwhile, the expression of Bcl-2 and Bax in renal tissue were assessed by immunohistochemical method. Results:①Pathological changes, which demonstrated tubular atrophy, expansion, interstitial fibrosis, inflammatory cell infiltration, and increased scores of tubulointerstitial fibrosis with varying degrees, were found in IRI group, but these pathological changes in treatment groups with low-and high-dose EPO were significantly improved as compared with the IRI group(P<0.05 or 0.01). Bcl-2 and Bax were mainly stained in renal tubular and interstitium with the standard immunohistochemistry. Compared with the sham-operated group, the levels of Bcl-2 and Bax expression were remarkably increased (P<0.01), the ratio of Bcl-2/Bax was remarkably decreased in IRI group (P<0.01). Compared with the IRI group, Bax expression was significantly reduced, Bcl-2 expression and the ratio of Bcl-2/Bax were incresead in treatment groups with low-and high-dose EPO(P<0.01). Increased espression of Bcl-2 in treatment group with high-dose EPO was higher than that in treatment group with low-dose EPO(P<0.01).③The degree of renal tubulointerstitial fibrosis was correlated with Bcl-2, Bax and Bcl-2/Bax ratio (r=-0.790, P<0.01; r=0.571, P<0.01; r=-0.802, P<0.01). Conclusions:1. Abnormal expression of Bcl-2/Bax is involved in development of renal tubulointerstitial fibrosis after ischemia/reperfusion-induced kidney injury.2. EPO can attenuate the renal tubulointerstitial fibrosis of renal IRI, and might has some relationship with EPO dose; 3. Effects of EPO on renal tubulointerstitial fibrosis after renal ischemia/reperfusion-induced kidney injury may be associated with the regulation of Bcl-2/Bax axis.
Abstract
Objective:To investigate effects of erythropoietin on renal tubulointerstitial fibrosis and Bcl-2 (anti-apoptotic protein)/Bax(pro-apoptotic protein) expression after renal ischemia/reperfusion injury(IRI) in mice. Methods:80 male C57BL/6 mice with 8-12 weeks age were randomly divided into 4 groups:sham-operated control group(n=20), IRI group(n=20), treatment group with low-dose EPO (model rats treated with EPO 100u/kg, Once a week, n=20), treatment group with high-dose EPO(model rats treated with EPO 1000u/kg, once a week, n=20). Left renal artery of mice was clamped for 35min with micro-artery clamp to make model of renal IRI. The morphological changes were observed by HE and Masson stain. Meanwhile, the expression of Bcl-2 and Bax in renal tissue were assessed by immunohistochemical method. Results:①Pathological changes, which demonstrated tubular atrophy, expansion, interstitial fibrosis, inflammatory cell infiltration, and increased scores of tubulointerstitial fibrosis with varying degrees, were found in IRI group, but these pathological changes in treatment groups with low-and high-dose EPO were significantly improved as compared with the IRI group(P<0.05 or 0.01). Bcl-2 and Bax were mainly stained in renal tubular and interstitium with the standard immunohistochemistry. Compared with the sham-operated group, the levels of Bcl-2 and Bax expression were remarkably increased (P<0.01), the ratio of Bcl-2/Bax was remarkably decreased in IRI group (P<0.01). Compared with the IRI group, Bax expression was significantly reduced, Bcl-2 expression and the ratio of Bcl-2/Bax were incresead in treatment groups with low-and high-dose EPO(P<0.01). Increased espression of Bcl-2 in treatment group with high-dose EPO was higher than that in treatment group with low-dose EPO(P<0.01).③The degree of renal tubulointerstitial fibrosis was correlated with Bcl-2, Bax and Bcl-2/Bax ratio (r=-0.790, P<0.01; r=0.571, P<0.01; r=-0.802, P<0.01). Conclusions:1. Abnormal expression of Bcl-2/Bax is involved in development of renal tubulointerstitial fibrosis after ischemia/reperfusion-induced kidney injury.2. EPO can attenuate the renal tubulointerstitial fibrosis of renal IRI, and might has some relationship with EPO dose; 3. Effects of EPO on renal tubulointerstitial fibrosis after renal ischemia/reperfusion-induced kidney injury may be associated with the regulation of Bcl-2/Bax axis.
引文
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