肾康注射液对单侧输尿管梗阻大鼠肾间质纤维化的影响

目的研究肾康注射液对梗阻性肾病大鼠肾小管间质纤维化的防治作用。方法采用单侧输尿管梗阻(UUO)诱导肾间质纤维化的动物模型,以血管紧张素Ⅱ受体拮抗剂(ARB)作为阳性对照,将大鼠随机分为:模型组(UUO)、肾康组、缬沙坦组,术后第14天处死大鼠,收集血清测定肌酐、尿素氮水平,肾组织用HE染色及免疫组化半定量检测各组肾间质的α-平滑肌肌动蛋白(α-SMA)、转化生长因子-β1(TGF-β1)的表达。结果肾康组、缬沙坦组的肾小管间质TGF-β1、α-SMA的表达均明显低于UUO组;两治疗组组间比较,肾康组对TGF-β1、α-SMA表达的抑制作用同于缬沙坦组。结论中药肾康可减轻肾间质纤维化。     

维甲酸对肾间质纤维化大鼠肾间质结缔组织生长因子表达的影响

目的探讨全反式维甲酸对肾间质纤维化大鼠肾间质结缔组织生长因子(CTGF)的影响及其可能机制.方法采用单侧输尿管结扎(UUO)大鼠模型.将40只SD大鼠随机分为5组:假手术组(A组)、UUO组(B组)、UUO 苯那普利组(C组)、UUO 维甲酸小剂量组(D组)、UUO 维甲酸大剂量组(E组).于造模前1d分别给C、D、E组苯那普利10mg/(kg·d)、全反式维甲酸10mg/(kg·d)和20mg/(kg·d).A、B组给予同等剂量的生理盐水.于术后第14d取术侧肾组织行HE、Masson染色评定各组肾小管间质损害程度,免疫组织化学半定量检测各组肾间质CTGF、转化生长因子-β1(TGF-β1)、α-平滑肌肌动蛋白(α-SMA)及Ⅲ型胶原(colⅢ)的表达.结果各造模组CTGF、TGF-β1、α-SMA及colⅢ的表达及肾小管间质损伤指数明显高于A组(P＜0.05),而全反式维甲酸或苯那普利均能明显抑制此变化.结论维甲酸及苯那普利可能通过下调CTGF而减轻肾间质纤维化.     

姜黄素对单侧输尿管梗阻大鼠肾小管上皮细胞转分化影响的研究

目的:探讨姜黄素对单侧输尿管梗阻(UUO)大鼠肾小管上皮细胞转分化的作用。方法:采用UUO致肾间质纤维化大鼠模型。将24只大鼠随机均分为假手术组、模型组和姜黄素组。从制模后2 d起,姜黄素组给予100 mg.kg-1.d-1姜黄素腹腔注射。术后4周心脏穿刺抽血,测定各组大鼠血清尿素氮(BUN)和血肌酐(SCr)值。处死大鼠,用苏木素-伊红(HE)、Masson染色评定肾小管间质纤维化程度;用免疫组化方法检测肾组织内转化生长因子-β1(TGF-β1)、α-平滑肌肌动蛋白(α-SMA)、波形蛋白(Vimentin)的蛋白表达部位及表达水平。结果:与假手术组比较,模型组大鼠BUN水平显著增加(P<0.01),肾间质纤维组织相对面积显著扩大(P<0.01),肾组织内TGF-β1、α-SMA和Vimentin的蛋白表达均显著上调(P均<0.01)。姜黄素干预后,上述上调指标都被显著抑制(P<0.05或P<0.01)。结论:姜黄素可能通过抑制UUO大鼠肾小管上皮细胞转分化而减轻肾间质纤维化。     

活性维生素D_3对大鼠肾小管间质纤维化的影响及其机制研究

目的:观察活性维生素D_3对大鼠肾小管间质纤维化的影响及其可能机制。方法:将40只雄性SD大鼠随机分为4组:假手术组(Sham组)、模型组(UUO组)、UUO+低剂量活性维生素D_3组(LV组)、UUO+大剂量活性维生素D_3组(HV组),每组10只。采用单侧(左)输尿管结扎法建立梗阻性肾小管间质纤维化大鼠模型。Sham组只游离左侧输尿管而不结扎。LV组与HV组分别予以0.03μg/(kg·d)、0.06μg/(kg·d)活性维生素D_3(溶于花生油)腹腔注射,Sham组和UUO组均予以等体积花生油腹腔注射。术后14 d处死大鼠,采集血、肾脏标本,并检测血清钙、磷、肌酐(SCr)、甲状旁腺激素(PTH)。制作肾脏病理组织切片,行H-E、Masson染色,在光镜下观察肾小管间质损伤及纤维化情况。采用免疫组织化学法检测肾小管间质中α平滑肌肌动蛋白(α-SMA)、E钙黏蛋白(E-cadherin)与转化生长因子β(TGF-β)的表达。结果:与UUO组相比,LV组和HV组SCr水平[(39.0±1.83)μmol/L、(36.0±2.11)μmol/L比(43.1±5.55)μmol/L]、肾小管间质纤维化指数(2.00±0.12、1.70±0.10比2.80±0.11)、α-SMA表达(0.22±0.02、0.20±0.03比0.24±0.02)、TGF-β表达(0.26±0.03、0.25±0.03比0.32±0.04)均显著下降(P0.05),E-cadherin表达(0.30±0.08、0.34±0.11比0.22±0.07)明显升高。与LV组相比,HV组SCr水平肾间质纤维化指数、肾间质损伤指数、α-SMA表达、TGF-β表达明显下降(P0.05),E-cadherin表达上升(P0.05)。相关性分析显示,TGF-β、α-SMA表达与肾间质纤维化指数正相关,E-cadherin表达与肾间质纤维化指数负相关;TGF-β表达与α-SMA表达正相关,E-cadherin与TGF-β负相关;α-SMA表达与E-cadherin表达负相关。结论:活性维生素D_3可改善UUO大鼠肾小管间质纤维化,大剂量活性维生素D_3改善作用优于小剂量,其机制可能是:通过抑制TGF-β的过度表达从而抑制UUO大鼠肾小管上皮细胞-肌成纤维细胞转分化进而改善肾小管间质纤维化。     

环氧合酶-2抑制剂对梗阻性肾病大鼠肾组织结缔组织生长因子表达的影响

目的：探讨环氧合酶-2(COX-2)抑制剂塞来昔布(celecoxib)对梗阻性肾病大鼠模型肾组织结缔组织生长因子(CTGF)和转化生长因子β1(TGF-β1)表达的影响及机制。方法：采用单侧输尿管结扎(UUO)建立梗阻性肾病模型,并随机分为模型组和治疗组,另设假手术组作对照。治疗组每日给予塞来昔布100mg／kg,模型组、假手术组每日给予等体积的生理盐水于造模前1d至造模后14d灌胃,分别于UUO后3d、7d、14d分批处死,逆转录聚合酶链式反应(RT-PCR)检测TGF-β1、CTGF的基因表达,免疫组化技术检测TGF-β1、CTGF、α-SMA、FN的蛋白表达,放免法检测组织内cAMP的水平。结果：与假手术组相比,模型组TGF-β1、CTGF的基因表达显著上调,cAMP的水平下降(P<0．001)。塞来昔布对梗阻肾TGF-β1的表达无显著改变,但能升高组织内cAMP水平,抑制CTGF的表达,纤维化程度减轻。结论：在梗阻性肾病大鼠中TGF-β1、CTGF的表达均显著升高,选择性COX-2抑制剂通过升高组织内cAMP的水平来抑制TGF-β1的下游因子——CTGF的表达延缓肾间质纤维化。     

洛伐他汀联合厄贝沙坦对慢性肾小管间质纤维化大鼠HIF-1α的影响

目的:探讨HIF-1α与CTGF在慢性肾小管间质纤维化中的表达及药物干预的影响。方法:建立UUO模型,将SD大鼠分4组:假手术组、模型组、ARB组、他汀联合ARB组。观察各组大鼠第2周肾功能、24h尿蛋白和肾组织病理学改变,检测HIF-1α、CTGF的表达。结果:模型组肾功能及尿蛋白定量均显著增高(P<0.05),应用他汀联合ARB治疗后肾功能较模型组显著降低,肾小管间质病变面积减少,肾小管间质细胞HIF-1αmRNA和CTGFmRNA表达显著减少(P<0.05)。模型组肾组织有高水平HIF-1αmRNA、CTGFmRNA,且两者的表达呈正相关(r=0.697,P<0.01),结论:低氧上调了HIF-1α及相关因子的表达;他汀联合ARB治疗可能降低HIF-1α和CTGF在肾小管间质中的表达而发挥肾脏保护作用。     

糖尿病大鼠肾小管上皮细胞-肌成纤维细胞转分化及氯沙坦对其的影响

【目的】观察糖尿病大鼠肾小管上皮细胞-肌成纤维细胞转分化(TEMT)及氯沙坦干预对其的影响。【方法】雄性Wistar大鼠随机分为正常对照组和糖尿病模型组;后者经STZ诱导糖尿病模型成功后再随机分为糖尿病组和氯沙坦干预组[氯沙坦20mg/(kg·d)]。分别于第8周和第16周时每组各处死5只大鼠。测定24h尿蛋白排泄量、血肌酐;留取肾组织作HE和Masson染色,观察肾小管间质损伤指数、肾间质胶原面积;免疫组织化学法检测肾小管上皮细胞α-平滑肌肌动蛋白(α-SMA)、波形蛋白(Vimentin)和转化生长因子-β1(TGF-β1)表达,并作半定量分析。【结果】①与对照组相比,糖尿病模型组大鼠肾小管间质损伤指数和肾间质胶原面积明显增加(P<0.01);②糖尿病组大鼠肾小管上皮细胞α-SMA、Vimentin和TGF-β1阳性表达均显著高于对照组,α-SMA表达和TGF-β1表达呈正相关(rs=0.810,P<0.01)。③氯沙坦组尿蛋白排泄量、肾小管间质损伤和间质纤维化程度较糖尿病组减轻,肾小管上皮细胞α-SMA、Vimentin与TGF-β1表达强度较糖尿病组显著下调(P<0.01)。【结论】①糖尿病大鼠肾脏病理进程中存在TEMT;②氯沙坦可下调糖尿病大鼠肾小管上皮细胞TGF-β1、Vimentin、α-SMA表达,阻抑糖尿病肾小管上皮细胞发生TEMT而发挥肾脏保护作用。     

冬虫夏草对糖尿病大鼠肾小管上皮细胞ILK表达的影响

[目的]观察糖尿病大鼠肾小管上皮细胞整合素连接激酶(ILK)的表达及冬虫夏草对其表达的影响.[方法]雄性Wistar大鼠随机分为3组:正常对照组(N组,n=12)、糖尿病模型组(D组,n=12)、糖尿病冬虫夏草干预组[C组,n=12,冬虫夏草1000 mg/(kg·d)灌胃].成模后第8周末及16周末各组分别处死6只大鼠,测定24 h尿蛋白排泄量、血肌酐;HE及MASSON染色法观察肾脏病理改变;免疫组织化学法检测ILK、E钙粘蛋白(E-cad)、α平滑肌肌动蛋白(α-SMA)及转化生长因子β1(TGF-β1)的表达.[结果]①与N组比较,D组大鼠肾小管间质损伤指数、肾间质胶原面积明显增加(P<0.01).②D组大鼠肾小管上皮细胞E-cad阳性表达显著低于N组,TGF-β1、ILK和α-SMA阳性表达均显著高于N组;E-cad表达和TGF-β1表达呈负相关(rs=-0.60,P<0.05);ILK、α-SMA表达和TGF-β1表达呈正相关(rs=0.88,P<0.01;rs=0.91,P<0.05).③C组大鼠肾小管间质损伤指数、肾闻质胶原面积较D组明显减弱,其肾小管上皮细胞的E-cad的表达较D组明显增加,TGF-β1、ILK和α-SMA表达较D组均明显减弱(P<0.01).[结论]冬虫夏草能通过下调ILK表达发挥肾脏保护作用.     

氯沙坦对梗阻性肾病大鼠肾间质固有细胞的作用

目的研究氯沙坦对梗阻性肾病大鼠肾间质固有细胞的作用,寻找可能的临床抑制肾间质纤维化的方法。方法采用单侧输尿管结扎(UUO)模型,治疗组于造模当日至造模后14天给予科素亚[16mg(kg·d)]灌胃,另设假手术组及手术组作为对照。应用免疫组织化学方法检测肾间质TGF-β的表达,组化双染色观察α-SMA、E-钙粘素的表达。激光共聚焦显微镜观察间质小管区α-SMA表达。结果手术组两周时出现近端肾小管区TGF-β表达明显增高(P<0．01),伴α-SMA表达增高(P<0．01),E-钙粘素表达下降(P<0．05),部分肾小管细胞中可见α-SMA及E-钙粘素同时表达;治疗组与手术组相比TGF-β、α-SMA的表达明显下降(P<0．05),而E-钙粘素的表达上调(P<0．05)。共聚焦显微镜下见部分小管基底膜损伤,上皮细胞表达α-SMA。结论氯沙坦可通过抑制肾间质固有细胞转分化过程的进展,从而对肾间质纤维化具有治疗作用。     

延肾口服液对肾间质纤维化大鼠肾组织转化生长因子-β和α-平滑肌肌动蛋白表达的影响

目的 探讨延肾口服液对肾间质纤维化(RIF)大鼠的影响及可能的作用机制.方法 将75只SD雄性大鼠按随机数字表法分为假手术组和模型组,采用单侧输尿管梗阻(UUO)复制RIF模型,成模大鼠再随机分为模型组、贝那普利组及延肾口服液高、低剂量组,每组15只.术前1 d治疗组开始灌胃贝那普利3.5 ml/d及延肾口服液2.8 ml/d、1.4 ml/d;假手术组、模型组给予等量生理盐水.术后8周留取血、尿标本,行血肌酐(SCr)、血尿素氮(BUN)及尿肌酐(UCr)、尿尿素氮(UUN)和24 h尿蛋白定量测定;处死大鼠取肾组织,苏木素-伊红(HE)染色观察各组RIF程度;用免疫组化二步法和半定量逆转录-聚合酶链反应(RT-PCR)测定转化生长因子-β(TGF-β)、α-平滑肌肌动蛋白(α-SMA)的蛋白和mRNA表达.结果 ①光镜下观察,模型组肾小管上皮细胞呈现不同程度的萎缩、变性及坏死,小管周围肾间质间隙增宽;贝那普利组和延肾口服液高、低剂量组部分肾小管轻度扩张,病变程度轻于模型组.②与假手术组比较,模型组UUN、UCr、BUN及TGF-β、α-SMA的蛋白和mRNA表达均明显升高(均P＜0.01);与模型组比较,贝那普利组及延肾口服液高、低剂量组TGF-β、α-SMA蛋白和mRNA[吸光度(A)值]表达均显著减少,以延肾口服液高剂量组减少更显著(TGF-β蛋白:0.72±0.24比1.78±0.33,α-SMA蛋白:0.87±0.55比1.59±0.44;TGF-β mRNA:1.68±0.53比3.14±0.44,α-SMA mRNA:1.66±0.14比3.41±0.10,均P＜0.01);贝那普利组和延肾口服液高、低剂量组UUN(mmol/L)、UCr(μmol/L)、BUN(mmol/L)均明显降低(P＜0.05或P＜0.01),以延肾口服液高剂量组降低更显著(UUN:0.56±0.19比1.34±0.22,UCr:2788.00±178.98比3652.75±188.26,BUN:7.25±0.47比16.30±0.47,均P＜0.01),延肾口服液高剂量组与贝那普利组相比差异无统计学意义(均P＞0.05),SCr及24 h尿蛋白定量在各组间比较差异均无统计学意义 (均P＞0.05).结论 延肾口服液延缓UUO 模型大鼠RIF的作用与其抑制TGF-β和α-SMA表达有关,其疗效与贝那普利组相似,并表现出明显的量效关系.     

肾间质纤维化中结缔组织生长因子（CTGF）与肾小管上皮细胞表型转化的关系

目的研究肾间质纤维化过程中CleF与肾小管上皮细胞表型转化的关系方法采用免疫组化方法,检测各组肾活检标本CTGF和α-SMA的表达。结果CTGF主要表达于肾小管上皮细胞和一些肾间质细胞胞浆,并且随间质病变的加重,表达量增加,范围增大;α-SMA在病变肾间质区域表达增强,一些管腔破坏和萎缩的肾小管上皮细胞也可见表达,阳性分布区域与CTGF相似。肾小管间质中CTGF、α-SMA表达量之间呈正相关,CTGF和α-SMA表达量与肾间质纤维化病变程度呈正相关关系。结论CTGF可能通过促进间质中肌成纤维细胞的形成而参与肾间质纤维化的形成,CTGF的表达上调对肾脏疾病有预后价值。     

去甲斑蝥素对蛋白超负荷肾病大鼠CTGF及NF-κ Bp65表达的影响

目的观察去甲斑蝥素对蛋白超负荷肾病大鼠结缔组织生长因子（CTGF）及核因子-κB（NF-κB）p65mRNA及蛋白表达的影响，初步探讨赎延缓肾间质纤维化的机制。方法所有SD大鼠均行单侧肾切除术。实验组大鼠予以腹腔注射牛血清白蛋白（BSA），建立蛋白超负荷肾病模型后随机分为模型组（BSA组，腹腔注射BSA5g／（kg·d）和去甲斑蝥素组（NCTD组，腹腔注射BSA5g／（kg·d）和NCTD0．1mgg／（kg·d）；对照组（Saline组，腹腔注射等量生理盐水）；每组8只大鼠。第9周末处死大鼠，留取肾脏标本。行光镜和电镜观察肾组织病理改变；RTPCR法检测CTGF和NF-κBp65mRNA表达；Westernblot及免疫组化法分别检测CTGF和NF-κBp65蛋白表达。结果①肾组织病理改变：BSA组可见肾小球球性硬化，肾小管灶性萎缩，肾间质大量炎症细胞浸润，伴灶状纤维化，病理半定量积分显示小管间质病理损伤（TIL）积分高于Saline组（P〈0．01）；电镜示肾小球系膜区有电子致密物沉积，足突融合。NCTD干预后，肾组织上述病理改变有所改善，且该组TIL积分较BSA组降低（P〈0．05）；②RT-PCR结果：NCTD组CTGF及NF-κBp65mRNA的表达较BSA组均减弱（P〈0.01）；③Westernblot检测结果提示NCTD组CTGF蛋白表达较BSA组下调（P〈0．05）；免疫组化显示，与BSA组比较，NCTD组NF-κBp65表达分布的阳性面积和阳性细胞数均明显降低（P〈0．05）。结论去甲斑蝥素可通过下调蛋白超负荷肾病大鼠CTGF和NF-κBp65的表达水平，延缓肾间质纤维化。     

雷公藤甲素对T2DM大鼠肾小管-间质损伤的保护作用

目的：观察雷公藤甲素（Triptolide,TP）对2型糖尿病（T2DM）大鼠肾小管-间质骨形态发生蛋白-7（BMP-7）、结缔组织生长因子（CTGF）、转化生长因子-β1（TGF-β1）表达的影响及保护作用。方法：Wistar大鼠50只,随机分为正常组（NC组）、模型组（DM组）和观察组（TP组）。DM及TP组大鼠均采用高脂高糖联合小剂量STZ建立T2DM模型。模型成功后,TP组大鼠给予TP 200μg.kg-1.d-1灌胃。结果：治疗8周后,与NC组比较,DM组及TP组大鼠肾重/体重（KW/BW）、空腹血糖（FBG）、尿白蛋白（UAL）及CTGF、TGF-β1mRNA蛋白的表达、ED-1阳性细胞数均明显增多（P〈0.05,0.01）,TP组低于DM组（P〈0.05,0.01）;肾组织BMP-7mRNA及蛋白表达下调（P〈0.01）,TP组高于DM组（P〈0.05）。相关分析表明,尿蛋白与BMP-7 mRNA存在负相关,与CTGF、TGF-β1mRNA存在正相关;肾小管-间质内ED-1阳性细胞数与TGF-β1的表达明显相关;TGF-β1与CTGF的表达呈正相关,而与BMP-7存在负相关（均P〈0.01）;萎缩肾小管比例及间质纤维化均与TGF-β1、CTGF表达明显正相关,与BMP-7负相关（均P〈0.05）。结论：TP对T2DM大鼠肾小管-间质损害具有明显的保护作用,其机制可能与下调CTGF、TGF-β1及部分恢复BMP-7的表达有关。     

结缔组织生长因子与LPS致肝纤维化的关系

目的：研究结缔组织生长因子(CTGF)表达水平与LPS致大鼠肝纤维化的关系。方法：体外培养条件下,LPS处理大鼠肝星状细胞(HSC),半定量RT-PCR方法检测不同时点不同LPS浓度处理的HSC FN mRNA和CTGF mRNA的表达水平。结果：LPS不仅短时间内即能上调HSC FN mRNA和CTGF mRNA表达,而且LPS与CTGF mRNA的表达量呈一定的剂量效应关系。恒定LPS作用浓度(1μg／ml)下,CTGF mRNA的表达水平随LPS作用时间的增加而增加,LPS作用24h-48h后CTGF mRNA的表达水平最高。结论：LPS是一个重要的致纤维化因素,可同时上调HSC的FN mRNA和CTGF mRNA表达。CTGF在LPS介导的肝纤维化的病理生理过程中可能发挥着重要的作用。     

藻黄合剂含药血清体外培养人肾小球系膜细胞结缔组织生长因子的表达

背景:结缔组织生长因子是转化生长因子β1的下游效应因子,其异常表达在肾小球硬化及纤维化发生、发展中起重要作用。目的:观察藻黄合剂含药血清对肾小球系膜细胞结缔组织生长因子表达的影响。方法:血清药理学方法制备藻黄合剂低、中、高剂量,空白对照及海昆肾喜阳性对照大鼠含药血清,分别用体积分数10%各含药血清培养基孵育高糖作用下肾小球系膜细胞48h,另设低糖组为对照。免疫细胞化学和RT-PCR方法检测各组系膜细胞结缔组织生长因子蛋白及mRNA的表达。结果与结论:结缔组织生长因子蛋白及mRNA表达在高糖组较低糖组明显升高(P<0.01);藻黄合剂各剂量组结缔组织生长因子蛋白及mRNA表达均低于高糖组,且具有剂量依赖性;藻黄合剂高剂量组与海昆肾喜组比较结缔组织生长因子蛋白及mRNA表达差异无显著性意义(P>0.05)。提示高糖可诱导肾小球系膜细胞中结缔组织生长因子的表达,而藻黄合剂可抑制高糖条件下肾小球系膜细胞中结缔组织生长因子蛋白及mRNA的表达。     

Homocysteine induces connective tissue growth factor expression in vascular smooth muscle cells.

BACKGROUND: Increased homocysteine levels in blood might be an important risk factor for the development of cardiovascular diseases. Connective tissue growth factor (CTGF) was found to be involved in atherosclerotic plaque progression. So far, the possible connection between homocysteine and CTGF has not been studied. OBJECTIVE: This study was designed to test whether homocysteine could induce CTGF expression in vascular smooth muscle cells (VSMC). METHODS AND RESULTS: Hyperhomocysteinemia was induced in Sprague-Dawley rats after 4 weeks of a high-methionine diet. CTGF mRNA and protein expression was detected in the aortas isolated from hyperhomocysteinemic rats, but not in the controls. The underlying mechanism of homocysteine-induced CTGF expression was investigated in cultured human umbilical vein smooth muscle cells (HUVSMC). CTGF mRNA expression was induced after treatment with dl-homocysteine (50 micromol L(-1)) for 1 h, which remained at the elevated level for up to 8 h. CTGF protein level increased after homocysteine treatment for 8 h, and the elevated status was maintained for up to 48 h. Several intracellular signals elicited by homocysteine are involved in CTGF synthesis, including protein kinase C (PKC) activation and reactive oxygen species (ROS). Transfection HUVSMCs with a CTGF small interference RNA (siRNA) plasmid, which specifically inhibited the expression of CTGF, decreased extracellular matrix (ECM) accumulation caused by homocysteine. CONCLUSION: Our results demonstrate that homocysteine could increase the expression of CTGF in VSMC both in vivo and in vitro. The novel findings suggest that homocysteine might contribute to accelerated progression of atherosclerotic lesions by inducing CTGF expression.     

Urinary connective tissue growth factor excretion in patients with type 1 diabetes and nephropathy

OBJECTIVE: Excretion of growth factors in the urine has been implicated in the pathogenesis of tubulointerstitial disease that characterizes proteinuric renal disease. In this cross-sectional study, we sought to examine the urinary excretion of the profibrotic cytokine connective tissue growth factor (CTGF) in type 1 diabetic patients with incipient and overt diabetic nephropathy. RESEARCH DESIGN AND METHODS: We recruited 31 subjects with type 1 diabetes from a hospital diabetes outpatient clinic. Of these, 10 subjects were normoalbuminuric, 8 were microalbuminuric and not receiving ACE inhibitor treatment, and 13 were macroalbuminuric, 8 of whom were receiving ACE inhibitor treatment. Urinary CTGF NH(2)-terminal fragment (CTGF-N) was determined by enzyme-linked immunosorbent assay and expressed relative to urinary creatinine. RESULTS: Urinary CTGF-N was closely correlated with the degree of albuminuria (r = 0.76, P < 0.001). In comparison with normoalbuminuric subjects, urinary CTGF-N was increased 10- and 100-fold in micro- and untreated macroalbuminuric subjects, respectively (CTGF-N-to-creatinine ratio: normoalbuminuria 0.23 x// 1.3 ng/mg, microalbuminuria 2.1 x// 1.7 ng/mg, untreated macroalbuminuria 203 x// 3.8 ng/mg, and geometric mean x// tolerance factor; P < 0.05 for normoalbuminuria versus microalbuminuria, P < 0.001 for microalbuminuria versus macroalbuminuria). Urinary CTGF-N was lower (<30-fold) in macroalbuminuric subjects treated with ACE inhibitors (6.5 x// 1.7 ng/mg; P < 0.01 vs. untreated macroalbuminuria) compared with their untreated counterparts. CONCLUSIONS: In this cross-sectional study, the magnitude of urinary CTGF-N excretion was related to the severity of diabetic nephropathy. In the context of its known profibrotic actions, these findings suggest that CTGF may contribute to the chronic tubulointerstitial fibrosis that accompanies proteinuric renal disease. Prospective and interventional studies will be needed to determine whether urinary CTGF-N may provide a reliable surrogate marker of renal injury and a meaningful indicator of response to therapy.     

siRNA-mediated knockdown of connective tissue growth factor prevents N-nitrosodimethylamine-induced hepatic fibrosis in rats

Hepatic fibrosis is a dynamic process that involves the interplay of different cell types in the hepatic tissue. Connective tissue growth factor (CTGF) is a highly profibrogenic molecule and plays a crucial role in the pathogenesis of hepatic fibrosis. The aim of the present investigation was three-fold. First, we studied the expression of CTGF in the cultured hepatic stellate cells using immunohistochemical technique. Second, we induced hepatic fibrosis in rats through serial intraperitoneal injections of N-nitrosodimethylamine (NDMA; dimethylnitrosamine, DMN) and studied the upregulation of CTGF and TGF-beta1 during hepatic fibrogenesis. Third, we downregulated CTGF expression using CTGF siRNA and examined the role of CTGF siRNA to prevent the progression of NDMA-induced hepatic fibrosis. The results depicted strong staining of CTGF in the transformed hepatic stellate cells in culture. Serial administrations of NDMA resulted in activation of hepatic stellate cells, upregulation of CTGF and TGF-beta1 both at mRNA and protein levels and well-developed fibrosis in the liver. Immunostaining, Western blot analysis, semiquantitative and real-time RT-PCR studies showed downregulation of CTGF and TGF-beta1 after treatment with CTGF siRNA. The results of the present study demonstrated that CTGF gene silencing through siRNA reduces activation of hepatic stellate cells, prevents the upregulation of CTGF and TGF-beta1 gene expression and inhibits accumulation of connective tissue proteins in the liver. The data further suggest that knockdown of CTGF upregulation using siRNA has potential therapeutic application to prevent hepatic fibrogenesis.     

结缔组织生长因子在大鼠心脏移植慢性排斥反应中的表达及意义

背景：慢性排斥反应限制了心脏移植患者的长期存活．有报道结缔组织生长因子因其与细胞增殖和胶原合成有关，似乎参与了此过程。 目的：假设结缔组织生长因子在大鼠心脏移植慢性排斥反应中发挥作用，实验拟验证其表达与心脏慢性排斥反应病理过程的关系。 设计、时间及地点：随机对照动物实验，于2007—04／08在中南大学湘雅二医院动物实验中心完成。 材料：取20只Wistar大鼠作为供体，20只SD大鼠作为受体，制作腹部异位心脏移植模型20例次；另选择10只Wistar大鼠作为对照。 方法：大鼠腹部异位心脏移植术后，给予环孢素A、霉酚酸酯和甲泼尼龙三联免疫抑制治疗。移植后2．8周分别按随机数字表法麻醉后处死10只受体大鼠．取心脏；10只Wistar大鼠心脏作为对照。 主要观察指标：采用苏木精-伊红染色及Van Gieson染色观察心肌病理形态、心肌血管密度及心脏血管狭窄程度：并进行心肌纤维化半定量评分。采用免疫组织化学法检测心肌组织结缔组织生长因予蛋自的表达：并分析结缔组织生长因子表达与慢性排斥反应过程中心肌血管病变及纤维化的相关性。 结果：①术后8周移植心脏血管狭窄程度、心肌纤维化评分及结缔组织生长因子蛋白表达量明显高于术后2周及正常心脏（P〈0．05-0．01）；术后8周移植心脏血管密度低于正常心脏（P〈0．05）；结缔组织生长因子在正常心脏基本没有表达。②心肌组织结缔组织生长因子蛋白表达灰度值与心肌纤维化、血管狭窄程度均呈负相关（r=0．734，-0．713，P〈0．01），表明结缔组织生长因子表达与心肌纤维化及血管狭窄程度呈正相关。结论：结缔组织生长因子在有心肌血管病变的移植心畦心肌组织中表达增强，其表达与心肌纤维化和心肌血管病变形成有关，在心肌移植排斥反应的病理过程中起重要作用。     

Increased expression of connective tissue growth factor in patients with urethral stricture

Urethral stricture (US) is a urologic disorder with a high morbidity and recurrence rate, and may arise due to external trauma, surgical procedures or urethritis. The pathogenesis of US is closely associated with scar formation, including excessive collagen-rich connective tissues. Previous studies have shown that connective tissue growth factor (CTGF) plays a key role in the fibrosis process of many tissues and organs, such as kidney and lung. We investigated the variation of CTGF expression in urethral tissues from 12 patients with US and normal urethral tissues. The stricture length range was 1 to 2.5 cm, and 4 patients underwent repeated urethral dilatation. The normal urethral tissues were obtained from 6 patients with heart disease, who were suffering from brain death. The expression of CTGF mRNA and protein was analyzed by real-time PCR, immunohistochemistry, and western blotting analyses. Results showed that the expression level of CTGF mRNA was significantly increased in US patients compared with the control. In addition, US patients who have longer history and more repeated surgical procedures have extremely high CTGF mRNA levels and its protein expression, while the age of patients, position of injury and stricture length were not found obvious relation with CTGF expression. Moreover, the degree of collagen deposition and muscle fiber proliferation in the submucosa is consistent with the increase in CTGF expression. In conclusion, our data suggest that the up-regulation of CTGF expression may be responsible for the fibrosis process of urethral tissues in US patients.