活性维生素D_3对阿霉素肾病大鼠CD2AP的影响
|
摘要
目的探讨活性维生素D类似物骨化三醇对阿霉素(ADR)肾病大鼠足细胞CD2相关蛋白(CD2AP)表达的影响。方法随机法选取60只Sprasue-Dawley大鼠,单次尾静脉注射ADR 7.5 mg/kg复制ADR肾病模型。2周后检测24 h尿蛋白、血清总蛋白(TP)、血浆白蛋白(ALB)、总胆固醇(TC)、血清肌酐(Scr)水平。将模型大鼠随机均分为模型组(ADR)、治疗组(ADR+Vit D_3),另取30只大鼠作为对照组。于模型复制成功后2、4、6、8和10周检测24 h尿蛋白。取大鼠肾组织,用Masson染色观察肾小球的病理损伤,透射电子显微镜观察足细胞超微结构,免疫组织化学染色和Western blotting检测CD2AP定位和表达。结果 2周后3组24 h尿蛋白、TP、ALB、Scr、TC比较,差异有统计学意义(P <0.05),提示模型复制成功。大鼠24 h尿蛋白随治疗周数延长而减少(P<0.05)。组织病理学结果显示,模型组肾组织大量纤维增生,肾小管扩张和肾间质纤维化。透射电子显微镜结果显示,模型组肾足细胞结构损伤,足突广泛融合,核畸形,脂滴聚集;治疗组较模型组有所改善。免疫组织化学染色结果显示,模型组CD2AP表达较低,活性维生素D_3治疗后阳性表达面积增加(P <0.05)。各组大鼠CD2AP表达水平比较,差异有统计学意义(P <0.05);模型组降低,治疗组随时间延长表达增强(P<0.05)。结论活性维生素D_3通过上调CD2AP表达,减轻足细胞的损伤,抑制肾组织病理纤维化增生,发挥肾保护功能,减少蛋白尿的产生。
Objective To investigate the effect of active vitamin D analogue(calcitriol) on the expression of CD2 AP in podocytes of rats with adriamyicin(ADR) nephropathy. Methods Sixty Sprasue-Dawley rats were selected randomly and injected by single tail vein injection of doxorubicin(7.5 mg/kg). The levels of 24 h urine protein and total protein(TP), albumin(ALB), total cholesterol(TC) and serum creatinine(Scr) were measured to judge the model success after 2 weeks. These rats were randomly divided into model group(ADR group) and calcitriol treatment group(ADR+Vit D_3 group). The other 30 rats were used as a blank control group. 2, 4, 6, 8, 10 weeks after the injection of ADR, 24 h urine proteins were detected and kidney tissues were taken. The pathological damages of glomerulus were observed by Masson staining, and the ultrastructure of podocytes were observed by transmission electron microscope. Immunohistochemical staining and western blotting were used to detect the localization and expression of CD2-related proteins. Results There were significant differences in urine protein quantitation, TP, ALB,Scr and TC among the control group, the model group, and the treatment group at 2 weeks(P < 0.05), suggesting that the model was successful. The 24 h urine protein quantitation of rats decreased with the number of weeks of treatment(P < 0.05). Histopathology showed a large amount of fibrous hyperplasia, tubular dilatation and renal interstitial fibrosis in the renal tissue of the model group. Transmission electron microscopy showed that structural damage of the renal foot cells in the model group, extensive fusion of the foot processes, nuclear malformation, and lipid droplet aggregation. The treatment group was better than the model group. The results of immunohistochemistry showed that the expression of CD2 AP was lower in the model group than control group, and the positive expression areas were significantly increased after treatment with 1, 25-(OH)2 D_3(P < 0.05). Western blotting showed that the expression of CD2 AP in each group was significantly different(P < 0.05). and decreased in the model group. The expression of the treatment group was increased with the number of weeks(P < 0.05). Conclusions Active vitamin D_3 can upregulate the expression of CD2-related proteins, reduce the damage of podocytes, inhibit the pathological fibrosis and proliferation of renal tissues, and exert renal protective function to reduce the production of proteinuria.
Objective To investigate the effect of active vitamin D analogue(calcitriol) on the expression of CD2 AP in podocytes of rats with adriamyicin(ADR) nephropathy. Methods Sixty Sprasue-Dawley rats were selected randomly and injected by single tail vein injection of doxorubicin(7.5 mg/kg). The levels of 24 h urine protein and total protein(TP), albumin(ALB), total cholesterol(TC) and serum creatinine(Scr) were measured to judge the model success after 2 weeks. These rats were randomly divided into model group(ADR group) and calcitriol treatment group(ADR+Vit D_3 group). The other 30 rats were used as a blank control group. 2, 4, 6, 8, 10 weeks after the injection of ADR, 24 h urine proteins were detected and kidney tissues were taken. The pathological damages of glomerulus were observed by Masson staining, and the ultrastructure of podocytes were observed by transmission electron microscope. Immunohistochemical staining and western blotting were used to detect the localization and expression of CD2-related proteins. Results There were significant differences in urine protein quantitation, TP, ALB,Scr and TC among the control group, the model group, and the treatment group at 2 weeks(P < 0.05), suggesting that the model was successful. The 24 h urine protein quantitation of rats decreased with the number of weeks of treatment(P < 0.05). Histopathology showed a large amount of fibrous hyperplasia, tubular dilatation and renal interstitial fibrosis in the renal tissue of the model group. Transmission electron microscopy showed that structural damage of the renal foot cells in the model group, extensive fusion of the foot processes, nuclear malformation, and lipid droplet aggregation. The treatment group was better than the model group. The results of immunohistochemistry showed that the expression of CD2 AP was lower in the model group than control group, and the positive expression areas were significantly increased after treatment with 1, 25-(OH)2 D_3(P < 0.05). Western blotting showed that the expression of CD2 AP in each group was significantly different(P < 0.05). and decreased in the model group. The expression of the treatment group was increased with the number of weeks(P < 0.05). Conclusions Active vitamin D_3 can upregulate the expression of CD2-related proteins, reduce the damage of podocytes, inhibit the pathological fibrosis and proliferation of renal tissues, and exert renal protective function to reduce the production of proteinuria.
引文
[1]高慧,王向托,杨立志.维生素D与慢性肾脏疾病[J].中国老年学杂志,2013,33(7):1710-1712.
[2]LU M,WANG P,GE Y,et al.Activation of mineralocorticoid receptor by ecdysone,an adaptogenic and anabolic ecdysteroid,promotes glomerular injury and proteinuria involving overactive GSK3βpathway signaling[J].Sci Rep,2018,8(1):1-12.
[3]WANG Y,DEB D K,ZHANG Z,et al.Vitamin D receptor signaling in podocytes protects against diabetic nephropathy[J].American Society of Nephrology Journal,2012,23(12):179-191.
[4]HOU Y C,WU C C,LIAO M T,et al.Role of nutritional vitamin D in osteoporosis treatment[J].Clin Chim Acta,2018,5(35):179-191.
[5]PIPPIN J W,BRINKKOETTER P T,CORMACK-ABOUD F C,et al.Inducible rodent models of acquired podocyte diseases[J].Am JPhysiol Renal Physiol,2009,296(2):213-229.
[6]DEEPA P R,VARALAKSHMI P.The cytoprotective role of a lowmolecular-weight heparin fragment studied in an experimental model of glomerulotoxicity[J].Eur J Pharmacol,2003,478(2/3):199-205.
[7]ENDLICH N,SIEGERIST F,ENDLICH K.Are podocytes motile[J].Pflugers Arch,2017,469(7/8):951-957.
[8]KUTNER A,BROWN G,VITAMINS D.Relationship between structure and biological activity[J].Int J Mol Sci,2018,19(7):1-11.
[9]SONNEVELD R,FERRèS,HOENDEROP J G J,et al.Vitamin D down-regulates TRPC6 expression in podocyte injury and proteinuric glomerular disease[J].The American Journal of Pathology,2013,182(4):1196-1204.
[10]TROHATOU O,TSILIBARY E F,CHARONIS A,et al.Vitamin D3 ameliorates podocyte injury through the nephrin signalling pathway[J].J Cell Mol Med,2017,21(10):2599-2609.
[11]XIAO H,SHI W,LIU S,et al.1,25-Dihydroxyvitamin D(3)prevents puromycin aminonucleoside-induced apoptosis of glomerular podocytes by activating the phosphatidylinositol3-kinase/Akt-signaling pathway[J].Am J Nephrol,2009,30(1):34-43.
[12]RUSSO L M,SRIVATSAN S,SEAMAN M,et al.Albuminuria associated with CD2AP knockout mice is primarily due to dysfunction of the renal degradation pathway processing of filtered albumin[J].FEBS Lett,2013,587(22):3738-3741.
[13]杨芳,何泽云.禾肾丸对阿霉素肾病大鼠蛋白尿及肾组织nephrin蛋白表达影响[J].中国实验方剂学杂志,2017,23(9):158-163.
[2]LU M,WANG P,GE Y,et al.Activation of mineralocorticoid receptor by ecdysone,an adaptogenic and anabolic ecdysteroid,promotes glomerular injury and proteinuria involving overactive GSK3βpathway signaling[J].Sci Rep,2018,8(1):1-12.
[3]WANG Y,DEB D K,ZHANG Z,et al.Vitamin D receptor signaling in podocytes protects against diabetic nephropathy[J].American Society of Nephrology Journal,2012,23(12):179-191.
[4]HOU Y C,WU C C,LIAO M T,et al.Role of nutritional vitamin D in osteoporosis treatment[J].Clin Chim Acta,2018,5(35):179-191.
[5]PIPPIN J W,BRINKKOETTER P T,CORMACK-ABOUD F C,et al.Inducible rodent models of acquired podocyte diseases[J].Am JPhysiol Renal Physiol,2009,296(2):213-229.
[6]DEEPA P R,VARALAKSHMI P.The cytoprotective role of a lowmolecular-weight heparin fragment studied in an experimental model of glomerulotoxicity[J].Eur J Pharmacol,2003,478(2/3):199-205.
[7]ENDLICH N,SIEGERIST F,ENDLICH K.Are podocytes motile[J].Pflugers Arch,2017,469(7/8):951-957.
[8]KUTNER A,BROWN G,VITAMINS D.Relationship between structure and biological activity[J].Int J Mol Sci,2018,19(7):1-11.
[9]SONNEVELD R,FERRèS,HOENDEROP J G J,et al.Vitamin D down-regulates TRPC6 expression in podocyte injury and proteinuric glomerular disease[J].The American Journal of Pathology,2013,182(4):1196-1204.
[10]TROHATOU O,TSILIBARY E F,CHARONIS A,et al.Vitamin D3 ameliorates podocyte injury through the nephrin signalling pathway[J].J Cell Mol Med,2017,21(10):2599-2609.
[11]XIAO H,SHI W,LIU S,et al.1,25-Dihydroxyvitamin D(3)prevents puromycin aminonucleoside-induced apoptosis of glomerular podocytes by activating the phosphatidylinositol3-kinase/Akt-signaling pathway[J].Am J Nephrol,2009,30(1):34-43.
[12]RUSSO L M,SRIVATSAN S,SEAMAN M,et al.Albuminuria associated with CD2AP knockout mice is primarily due to dysfunction of the renal degradation pathway processing of filtered albumin[J].FEBS Lett,2013,587(22):3738-3741.
[13]杨芳,何泽云.禾肾丸对阿霉素肾病大鼠蛋白尿及肾组织nephrin蛋白表达影响[J].中国实验方剂学杂志,2017,23(9):158-163.