足细胞与糖尿病肾病关系的若干研究进展

足细胞在维持肾小球滤过屏障完整性及限制血浆蛋白的滤出方面发挥重要作用,足细胞的损伤可部分解释肾小球滤过屏障结构及功能的改变的原因。在糖尿病肾病早期,足细胞损伤已出现,并可进一步导致。肾脏损伤。本文就足细胞与糖尿病肾病中若干研究进展作简要综述。     

Analysis of intra-GBM microstructures in a SLE case with glomerulopathy associated with podocytic infolding

BACKGROUND: Systemically podocytic infolding into the GBM which causes nonargyrophilic holes in the GBM in association with intra-GBM microstructures has been considered as a new pathological entity. However, its pathomechanisms are largely unknown. METHODS: We analyzed intra-GBM microstructures in an SLE patient with glomerulopathy associated with podocytic infolding by immunoelectron microscopy for vimentin (a marker for both podocyte and endothelium) and C5b-9 and by 3D reconstruction of transmission electron microscopy (TEM) images by computer tomography method. RESULTS: Immunofluorescent study showed immunoglobulin deposition in a diffuse, capillary pattern; however, electron-dense deposits like stage 3 membranous nephropathy could be found only in some capillary loops by TEM in spite of the systemic existence of podocytic infolding and the intra-GBM microstructures. Three-dimensional reconstructed images of the TEM images revealed that some of the intra-GBM microstructures made connections with the podocyte. The clustered microstructures underneath the podocyte and their surroundings looked as a whole like the degraded part of podocyte in 3D reconstructed images. Immunoelectron microscopy showed that vimentin was positive in most intra-GBM microstructures. C5b-9 was positive along the entire epithelial side of the GBM and in some microstructures, suggesting that the podocytes may be attacked by C5b-9 and that the microstructures may contain C5b-9 bound cellular membranes. CONCLUSION: Intra-GBM microstructures may be originated mainly from the podocyte. Podotyte and GBM injuries caused by C5b-9 attack to podocytes might contribute in part to podocytic infolding and intra-GBM microstructures in this case.     

Is mesangial hypercellularity with glomerular immaturity a variant of glomerulosclerosis?

Our aim was to correlate an immunohistochemical pattern of selected podocyte cytoskeleton-associated proteins in children diagnosed with focal segmental glomerulosclerosis (FSGS) and diffuse mesangial proliferation accompanied by glomerular immaturity (Im-DMP) with the clinical courses of both diseases. The material included 33 renal biopsies obtained from children diagnosed with DMP with or without signs of glomerular immaturity (ten and 15 participants, respectively) or FSGS (eight patients). Ezrin, podocalyxin, synaptopodin and nephrin expression was evaluated by immunohistochemical assay. A positive reaction for ezrin, podocalyxin, synaptopodin and nephrin was observed in the most superficial, continuous 'layer' of podocytes in Im-DMP patients. This distribution closely mimicked the immunohistochemical pattern observed in FSGS. The severe initial course of Im-DMP was transient. Resistance to steroids (six children) and renal insufficiency (two patients) in these subjects subsided, whilst, in the FSGS patients, the resistance to steroids recognized in all the children and the renal insufficiency diagnosed in three of them were still present. Mimicry between the immunohistochemical pattern of glomerular immaturity in DMP and focal segmental glomerulosclerosis might explain the severe initial course of this nephrotic syndrome in children. The transient clinical character of the former may also indicate that it is not a variant of FSGS.     

黄萸方防治大鼠早期糖尿病肾病足细胞损伤的研究

目的 观察中药复方黄萸方对实验性糖尿病大鼠微量白蛋白尿的疗效,及对足细胞的影响.方法 采用高糖高脂喂饲加低剂量STZ诱导伴胰岛素抵抗的大鼠2型糖尿病,给予黄萸方治疗12周,观察血糖及尿微量白蛋白,采用透射电镜及图像分析技术,观察对足细胞形态的影响.结果 黄萸方可降低血糖、减少尿微量白蛋白,同时可改善足细胞损伤,疗效优于ARB.结论 黄萸方可改善实验性大鼠2型糖尿病肾脏病变,主要是改善足细胞损伤,减少微量白蛋白尿.     

替米沙坦对老年糖尿病大鼠足细胞黏附的保护作用

目的 探讨替米沙坦对足细胞黏附的保护作用。 方法 用20月龄SD大鼠制备糖尿病模型，随机分为糖尿病组和替米沙坦治疗组，另设老年和6周龄对照组。于实验第4、8、12周末收集大鼠24 h尿，监测血压。12周后行组织病理学检查，检测尿白蛋白（24 h）和血、尿肌酐。以间接免疫荧光-hoechst 33342双染检测尿中足细胞数量。RT-PCR、免疫组化和Western印迹等方法检测肾小球中整合素α3、整合素连接激酶(ILK) mRNA和蛋白表达。 结果 随实验进程，老年糖尿病大鼠尿白蛋白、血压和尿中足细胞数量明显高于对照组(P <0.05)，且出现较为明显的病理学损害。肾小球整合素α3 mRNA及蛋白表达量减少， ILK表达明显增加(P < 0.05)。第12周，整合素α3蛋白表达与ILK表达呈负相关 (P < 0.05)。尿足细胞数量、尿白蛋白程度与ILK表达呈正相关 (P < 0.05)。替米沙坦明显减轻肾脏病理损害，增加整合素α3表达，降低ILK表达，减少尿足细胞数量。 结论 老年糖尿病大鼠足细胞的黏附能力下降，尿足细胞排泄增加。替米沙坦可增强足细胞黏附，对足细胞存活起重要作用。     

高糖对小鼠足细胞向间充质细胞 转分化的影响

目的 通过观察高糖环境下小鼠足细胞表达NEPH1、desmin、TGF-β1 和Smad7 的变化,探讨高糖损伤足细胞 的机制。方法 将培养成熟的小鼠足细胞用RandA 1.0 软件完全随机分为高糖1、2、3 组（葡萄糖浓度分别为15、25 和30mmol/L）和对照组（葡萄糖浓度5mmol/L）,倒置显微镜下观察高糖干预前后足细胞的形态变化,采用RT-PCR 和Western blot技术分别检测足细胞NEPH1、desmin、TGF-β1 和Smad7 的mRNA 和蛋白表达变化。结果 与对照组相比,高糖环境下培养48h 后,足细胞形态发生不同程度变化。NEPH1 和Smad7 的mRNA 表达较对照组显著减少（P＜0.05 或0.01）,desmin 和TGF-β1的mRNA 表达较对照组增加（P＜0.05 或0.01）。其中高糖2 组的NEPH1 和Smad7 的蛋白表达较对照组显著减少（均P＜0.01）,而desmin 和TGF-β1的蛋白表达较对照组显著增加（P＜0.01）。小鼠足细胞在高糖培养下NEPH1 mRNA 和desmin mRNA 的表达呈显著负相关（r=-0.993,P＜0.01）;NEPH1 mRNA 和TGF-β1 mRNA 的表达呈显著负相关（r=-0.989,P＜0.01）;TGF-β1 mRNA 的表达和Smad7 mRNA 的表达呈显著负相关（r=-0.996,P＜0.01）。结论 高糖可能通过激活TGF-β/Smad信号通路诱导小鼠足细胞发生表型转化。     

Mindin:糖尿病肾病足细胞损伤的新型标记物

Mindin(也称spondin-2)是一个高度同源的细胞外基质蛋白,既往研究发现,mindin可作为病原微生物模式识别分子和整合素配体参与非特异性和特异性免疫反应,并有促进神经元的黏附和生长的功能.近年研究发现,mindin可由损伤的足细胞合成,其参与糖尿病肾病的发生、发展,并与糖尿病足细胞损伤密切相关.与健康人相比,2型糖尿病患者的尿液mindin表达增加,并能反映糖尿病肾病严重程度.因此,mindin有望成为糖尿病肾病足细胞损伤新标记物.     

2型糖尿病患者尿液mindin水平与糖尿病肾病的相关性

目的探讨2型糖尿病（T2DM）患者尿液mindin的表达,分析其与糖尿病肾损伤的关系。方法选取2010年10月至2011年6月于广西医科大学第一附属医院代谢糖尿病中心及内分泌科住院的T2DM患者124例为病例组,选取同期年龄、性别匹配的健康体检者30名作为对照组。根据24h尿白蛋白定量（24h-UAE）将T2DM患者分为3组,即24h—UAE〈30mg的正常蛋白尿组60例,30mg≤24h-UAE〈300mg的微量蛋白尿组32例,24h-UAE≥300mg的大量蛋白尿组32例。应用ELISA法测定尿液mindin的水平,常规方法测定血、尿肌酐、血尿素氮、胱抑素C、空腹及餐后血糖、糖化血红蛋白（HbA1c）、C-肽等临床指标,并估算肾小球滤过率（eGFR）。组间比较采用单因素方差分析及q检验,指标间的相关分析采用Spearman相关分析及偏相关分析。结果T2DM患者尿mindin含量高于对照组f正常蛋白尿组、微量蛋白尿组、大量蛋白尿组、对照组分别为161（119～255）、332（181—445）、426（230—937）和93（51—121）ng·ml^-1·μmol^-1·L^-1,F=30．196,P〈0．05],并与24h-UAE呈正相关（r=0．453,P〈0．05）,T2DM患者各组mindin水平随24h-UAE的升高而递增;相关分析显示尿mindin与尿素氮、血肌酐、胱抑素C、血糖、24h-UAE、病程呈正相关（r=0．426、0．2336、0．383,0344、0．453、0．511,均P〈0．05）,而与eGFR呈负相关（r=-0．383,P〈0．05）;结果显示尿液mindin（OR=1．481,95％CI：1.187—1．847）、空腹血糖（OR=0．842,95％CI：0．756～0．939）、舒张压（OR=1．051,95％CI：1．003～1．101）是T2DM患者肾损伤的危险因素。结论尿液mindin可反映糖尿病肾病早期肾功能损伤并与肾功能损伤的程度相关,尿液mindin的检测对糖尿病肾病患者早期诊断、病情观察、疗效判断方面具有较好的临床应用前景。