橙皮苷对糖尿病肾病大鼠Wnt/β-catenin信号通路的调节作用研究
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摘要
目的观察橙皮苷对糖尿病肾病大鼠的治疗作用及其对肾脏组织中Wnt/β-catenin蛋白表达的影响。方法 60只雄性SD大鼠,在适应性喂养1周后,链脲佐菌素一次性法构建大鼠糖尿病肾病大鼠模型,造模成功后将大鼠分为正常组,模型组以及橙皮苷治疗组三组,每组20只,造模成功后橙皮苷治疗组给予橙皮苷注射治疗,治疗量为0.2mg/kg大鼠体重(注射时称取1mg溶于1mL的丙二醇中),正常组和模型组给予等体积的丙二醇溶液注射。分别于治疗4周和8周后处死大鼠各半,收集其血、尿及肾脏标本,检测一般生化指标、肾脏组织病理学并运用实时荧光定量PCR和Western Bloting检测肾脏组织中Wnt-4和β-catenin的表达特点。结果正常组大鼠毛色光滑,活泼好动,与正常组相比,模型组有明显的多饮多食多尿的特点,此外毛发枯燥,精神萎靡,橙皮苷治疗后,其上述情况有明显好转,但和正常组差异仍然较大。与正常组相比,模型组大鼠肌酐(SCr)、尿素氮(BUN)、24h尿蛋白(UTP)及尿白蛋白肌酐比(UACR)水平均明显升高(P<0.05),相较于模型组,使用橙皮苷治疗4周后上述所有指标明显下降(P<0.05),治疗8周后进一步降低(P<0.05);病理学结果可见,正常组大鼠肾小管结构完整清晰,无明显病理改变;模型组可见肾小管间质局部炎症细胞浸润,肾小管上皮细胞脱落,小管扩张,基底膜断裂,橙皮苷治疗4周后纤维化面积减小,治疗8周后进一步好转;与正常组比较,模型组Wnt-4和β-catenin蛋白表达水平显著上升(P<0.05),而与模型组比较,橙皮苷治疗4周后明显下降(P<0.05),治疗8周后进一步降低(P<0.05)。结论橙皮苷可通过抑制肾脏组织中Wnt-4和β-catenin蛋白和mRNA的表达,进而改善肾功能、减轻蛋白尿,最终起到对肾脏的保护作用。
Objective To investigate the regulating effect of hesperidin on diabetic nephropathy rats and analyze its effect on the expression of Wnt/β-catenin protein in renal tissue. Methods 60 male Sprague-Dawley rats were randomly divided into three groups: normal group, diabetes group and hesperidin treatment group, 20 in each group. After the model was successfully established, hesperidin treatment group was given hesperidin injection treatment(0.2 mg/kg)(when injection, lmg hesperidin dissolve the propylene glycol), normal and model groups were given an equal volume of propylene glycol solution.The rats were sacrificed each half at the 4 th and 8 th weeks after treatment, the blood, urine and kidney specimens were collected.The expression of β-catenin and Wnt-4 in renal tissue were detected by real-time quantitative PCR and Western Blotting. Results Compared with normal group, the rats in model group showed obvious polydipsia and polyuria, the hair was dry and the spirit was dispirited. After treatment with hesperidin, the above situation had improved significantly. Compared with normal group, the levels of SCr, BUN, 24 h UTP and UACR in model group were significantly higher(P<0.05). Compared with model group, all the above rations were significantly decreased after 4 weeks of treatment with hesperidin(P<0.05), and further decreased after 8 weeks(P<0.05). The pathological results showed that normal rat renal tubular structure was complete and clear,no obvious pathological changes were observed in model group; tubulointerstitial inflammatory cell infiltration, renal tubular epithelial cell shedding, tubular dilation, basement membrane rupture, hesperidin, the fibrosis area decreased after 4 weeks of treatment, further improved after 8 weeks of treatment; compared with normal group, the expression level of Wnt-4 and β-catenin protein in model group significantly increased(P<0.05), compared with model group, hesperidin decreased significantly after 4 weeks of treatment(P<0.05), further decreased after 8 weeks of treatment(P<0.05). Conclusion Hesperidin can inhibit the expression of Wnt-4, β-catenin protein and mRNA in renal tissue, thus to improve renal function, reduce proteinuria and finally protect the kidney.
Objective To investigate the regulating effect of hesperidin on diabetic nephropathy rats and analyze its effect on the expression of Wnt/β-catenin protein in renal tissue. Methods 60 male Sprague-Dawley rats were randomly divided into three groups: normal group, diabetes group and hesperidin treatment group, 20 in each group. After the model was successfully established, hesperidin treatment group was given hesperidin injection treatment(0.2 mg/kg)(when injection, lmg hesperidin dissolve the propylene glycol), normal and model groups were given an equal volume of propylene glycol solution.The rats were sacrificed each half at the 4 th and 8 th weeks after treatment, the blood, urine and kidney specimens were collected.The expression of β-catenin and Wnt-4 in renal tissue were detected by real-time quantitative PCR and Western Blotting. Results Compared with normal group, the rats in model group showed obvious polydipsia and polyuria, the hair was dry and the spirit was dispirited. After treatment with hesperidin, the above situation had improved significantly. Compared with normal group, the levels of SCr, BUN, 24 h UTP and UACR in model group were significantly higher(P<0.05). Compared with model group, all the above rations were significantly decreased after 4 weeks of treatment with hesperidin(P<0.05), and further decreased after 8 weeks(P<0.05). The pathological results showed that normal rat renal tubular structure was complete and clear,no obvious pathological changes were observed in model group; tubulointerstitial inflammatory cell infiltration, renal tubular epithelial cell shedding, tubular dilation, basement membrane rupture, hesperidin, the fibrosis area decreased after 4 weeks of treatment, further improved after 8 weeks of treatment; compared with normal group, the expression level of Wnt-4 and β-catenin protein in model group significantly increased(P<0.05), compared with model group, hesperidin decreased significantly after 4 weeks of treatment(P<0.05), further decreased after 8 weeks of treatment(P<0.05). Conclusion Hesperidin can inhibit the expression of Wnt-4, β-catenin protein and mRNA in renal tissue, thus to improve renal function, reduce proteinuria and finally protect the kidney.
引文
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[11]Zhou L,Liu Y.Wnt/β-catenin signalling and podocyte dysfunction in proteinuric kidney disease[J].Nature Reviews Nephrology,2015,11(9):535.
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[13]Madan B,Patel MB,Zhang J,et al.Experimental inhibition of porcupine-mediated Wnt O-acylation attenuates kidney fibrosis[J].Kidney International,2016,89(5):1062.
[14]史晶晶,时博.橙皮苷对髙脂血症模型大鼠脂代谢紊乱的影响[J].中医学报,2016,31(4):554-557.
[15]李雪飞,江洪,胡笑容,等.橙皮苷减轻大鼠心肌缺血/再灌注损伤所致的炎症反应[J].武汉大学学报(医学版),2016,37(3):390-394.
[16]何伟,陈明喆.橙皮苷对IgA肾病大鼠肾脏组织中PAI-1蛋白表达的影响[J].长春中医药大学学报,2017,33(3):355-357.
[17]陈明喆,何伟.橙皮苷对IgA肾病大鼠肾脏组织中Nephrin蛋白表达的影响研究[J].现代中西医结合杂志,2016,25(26):2851-2854.
[18]陈明喆,李易,何伟.橙皮苷对IgA肾病大鼠肾脏组织中JAK/STAT信号通路相关蛋白STAT3和p-STAT3表达的影响[J].解放军医药杂志,2017,29(3):10-14.
[2]唐咸玉,何柳,何嘉莉,等.天芪降糖胶囊联合西药治疗糖尿病肾病的疗效及对血脂代谢的影响[J].湖北中医药大学学报,2016,18(2):63-65.
[3]孙小会.2型糖尿病肾病发生的流行病学特征及其影响因素的分子流行病研究[D].广州:暨南大学,2013.
[4]王焱.橙皮苷对大鼠非酒精性脂肪肝脂质代谢和炎症反应的影响及部分机制的研究[D].合肥:安徽医科大学,2010.
[5]张冬松,高慧媛,吴立军.橙皮苷的药理活性研究进展[J].中国现代中药,2006,8(7):25-27.
[6]李易,田鲁,王港,等.苦瓜总皂苷对2型糖尿病大鼠肾病组织中蛋白激酶C-α和转化生长因子β表达的影响研究[J].湖北中医药大学学报,2016,18(5):5-9.
[7]张舒媛,王东超,李博,等.糖尿病肾病研究进展[J].世界中医药,2015(10):1621-1625.
[8]王晓男.中药黄芪治疗糖尿病肾病临床疗效观察[J].辽宁中医药大学学报,2015(6):182-184.
[9]Clevers H.Wnt/β-Catenin Signaling in Development and Disease[J].Cell,2006,127(3):469-480.
[10]林辉,张祥贵.Wnt/β-catenin信号通路与肾脏疾病[J].海南医学,2017,28(2):267-271.
[11]Zhou L,Liu Y.Wnt/β-catenin signalling and podocyte dysfunction in proteinuric kidney disease[J].Nature Reviews Nephrology,2015,11(9):535.
[12]万雷,黄宏兴,罗明,等.Wnt/β-catenin信号通路拮抗剂与骨质疏松[J].辽宁中医药大学学报,2014(12):73-75.
[13]Madan B,Patel MB,Zhang J,et al.Experimental inhibition of porcupine-mediated Wnt O-acylation attenuates kidney fibrosis[J].Kidney International,2016,89(5):1062.
[14]史晶晶,时博.橙皮苷对髙脂血症模型大鼠脂代谢紊乱的影响[J].中医学报,2016,31(4):554-557.
[15]李雪飞,江洪,胡笑容,等.橙皮苷减轻大鼠心肌缺血/再灌注损伤所致的炎症反应[J].武汉大学学报(医学版),2016,37(3):390-394.
[16]何伟,陈明喆.橙皮苷对IgA肾病大鼠肾脏组织中PAI-1蛋白表达的影响[J].长春中医药大学学报,2017,33(3):355-357.
[17]陈明喆,何伟.橙皮苷对IgA肾病大鼠肾脏组织中Nephrin蛋白表达的影响研究[J].现代中西医结合杂志,2016,25(26):2851-2854.
[18]陈明喆,李易,何伟.橙皮苷对IgA肾病大鼠肾脏组织中JAK/STAT信号通路相关蛋白STAT3和p-STAT3表达的影响[J].解放军医药杂志,2017,29(3):10-14.