阿托伐他汀与螺内酯对腹膜纤维化的作用研究

目的探讨腹膜纤维化的药物防治措施。方法50只Wister雄性大鼠随机分成5组,每组10只。A组为对照组,每日腹腔内注入0.9%生理盐水20ml。B、C、D、E组腹腔内每日注入20ml4.25%百特透析液,并于试验第7,14,21,28天分别加入乳酸盐红霉素6.25万IU;C组螺内酯100mg/（Kg·d）灌胃;D组阿托伐他汀20mg/（Kg·d）灌胃;E组两药合用。于第1、第30天测定腹膜透析液中的转化生长因子（TGF-β1）,30天后收集大鼠腹膜作常规病理（HE染色.Masson染色）,行腹膜平衡试验（PET）,免疫组化检测eNOS表达和血管生成情况。结果C、D、E组的腹膜纤维化程度比B组轻（P〈0.05）;TGF-β1）浓度明显下降（P〈0.05）;各组腹膜间皮细胞均有eNOS表达,B、C、D、E组均有新生毛细血管表达,C、D、E组较B组表达下调（P〈0.05）。结论阿托伐他汀及螺内酯能有效的防治腹膜纤维化进程。     

氟伐他汀对单侧输尿管梗阻大鼠肾间质纤维化的影响

[目的]观察氟伐他汀对单侧输尿管梗阻后大鼠肾间质单核细胞趋化蛋白-1(MCP-1)表达、单核/巨噬细胞浸润及肾间质纤维化程度的影响,探讨他汀类药物的抗炎作用机制及对肾脏的保护作用.[方法]72只SD雌性大鼠随机分成假手术(SOR)组、单侧输尿管梗阻术(UUO)组和单侧输尿管梗阻术(UUO) 氟伐他汀治疗组[T-UUO组, 氟伐他汀20 mg/(kg·d)].于术后d3、d7、d10、d14分别处死各组大鼠.用HE及Masson 染色动态观察肾脏病理变化,免疫组织化学法测定MCP-1 的表达和巨噬细胞浸润情况.[结果]UUO组肾小管-间质MCP-1与单核巨噬细胞抗原ED-1表达较SOR组增加(P＜0.05); 在术后各时间点,T-UUO组大鼠肾小管-间质MCP-1、ED-1的表达及肾间质胶原相对面积较UUO模型组显著减少,但仍高于SOR组(P＜0.05).[结论]氟伐他汀可通过降低MCP-1表达、抑制单核-巨噬细胞浸润,减轻肾间质纤维化而对肾脏有保护作用.     

红细胞生成素对单侧输尿管梗阻大鼠肾间质纤维化的作用及对MCP-1和TGF-β1表达的影响

目的探讨红细胞生成素(erythropoietin,EPO)对单侧输尿管梗阻(unilateral ureteralobstruction,UUO)模型大鼠肾间质纤维化的作用及其对单核细胞趋化蛋白1(monocyte chemoattractant pro-tein-1,MCP-1)和转化生长因子β1(transforming growth factor-β1,TGF-β1)表达的影响。方法将60只雄性Wistar大鼠分为假手术组(20例),UUO对照组(20例)及EPO治疗组(20例),分别于手术后第3,7,14,21d处死动物留取肾组织标本。应用马松染色进行肾间质纤维化程度评分;应用免疫组化方法和Western blot方法测定肾组织中MCP-1、TGF-β1的表达程度。结果马松染色显示手术后第3,7d,EPO治疗组肾间质纤维化程度较UUO对照组明显减轻(P0.05)。手术后第3,7,14d,EPO治疗组MCP-1、TGF-β1的表达明显低于UUO对照组(P0.05)。结论 EPO可以减轻UUO模型大鼠早期肾间质纤维化程度,该作用可能与EPO抑制肾组织MCP-1和TGF-β1的表达有关。     

短发夹结构RNA对人腹膜间皮细胞转化生长因子-β1表达和超微结构的影响

目的 研究靶向转化生长因子-β1（TGF-β1）的短发夹RNA（shRNA）表达载体对腹膜透析液诱导的人腹膜间皮细胞（HPMC）表达TGF-β1的影响,并观察其对HPMC超微结构的影响. 方法 利用pcDU6质粒构建两个靶向TGF-β1的shRNA真核表达载体pcDU6-A1-A2和pcDU6-B1-B2,使用脂质体介导转染腹膜透析液刺激下的HPMC,采用半定量逆转录多聚酶链式反应（RT-PCR）和酶联免疫吸附（ELISA）检测shRNA表达载体对于TGF-β1基因和蛋白质表达水平的抑制效果,使用透射电镜观察HPMC超微结构的改变. 结果 HPMC暴露于4.25%腹膜透析液中48小时后,其上清中TGF-β1的浓度与对照组相比显著增高分别为（39.71±6.60）pg/10^5 cells和（16.32±2.71）pg/10^5 cells,P＜0.01）,1.5%腹膜透析液对于TGF-β1蛋白合成无明显影响;而预先转染了shRNA载体的两组与转染pcDU6载体（空载体）的组相比,其由4.25%腹膜透析液诱导的TGF-β1蛋白增高幅度明显减少,分别下降了39.2%和47.2%（P＜0.01）;两组转染不同的shRNA载体的HPMC之间相比TGF-β1浓度无显著差异,pcDU6载体（空载体）转染组与4.25%腹膜透析液刺激组TGF-β1浓度亦无显著差异.透射电镜显示HPMC暴露于4.25%腹膜透析液可导致细胞扁平、微绒毛减少和线粒体水肿等,而转染shRNA载体组细胞的改变相对较轻. 结论 pcDU6质粒载体介导的短发夹结构RNA（shRNA）能够明显抑制腹膜透析液刺激下的人腹膜间皮细胞TGF-β1的高表达,并减轻其超微结构的异常,提示shRNA表达载体可能有益于腹膜纤维化的防治.     

转化生长因子β1在大鼠纤维化腹膜组织中的表达及作用

目的:检测转化生长因子β1在腹膜透析大鼠腹膜内表达,并探讨其在腹膜纤维化中的意义。方法:实验于2005-06/2006-03在中南大学湘雅二医院肾内科实验室完成。①实验材料:雄性SD大鼠,体质量180～240g,由中南大学湘雅二医院动物实验中心提供。②实验方法:将28只大鼠按随机数字表随机分为4组,每组7只。正常对照组不予任何干预;生理盐水组腹腔注射20mL生理盐水;低糖透析液组腹腔注射20mL1.5%葡萄糖透析液;高糖透析液组腹腔注射20mL4.25%葡萄糖透析液,均为1次/d。4周后,向大鼠腹腔注射4.25%葡萄糖腹膜透析液20mL,4h后于大鼠右下腹缓慢插入带有多个侧孔的10号静脉留置针,缓慢低位引流腹透液,量取引流液。③实验评估:取大鼠壁层腹膜组织,以苏木素-伊红染色,镜下测量腹膜厚度,采用免疫组织化学方法检测大鼠腹膜中转化生长因子β1及纤连蛋白。结果:28只大鼠均进入结果分析。①高糖透析液组、低糖透析液组超滤量均明显低于正常对照组与生理盐水组,并且高糖透析液组超滤量明显少于低糖透析液组(P均<0.05)。②高糖透析液组腹膜明显增厚,表面粗糙,间皮细胞肿胀,脱落,间皮下有大量血管生成以及胶原纤维沉积,还可见单核细胞等炎症细胞浸润,与其他组比较,腹膜厚度明显增加(P<0.05)。③高糖透析液组转化生长因子β1、纤连蛋白表达量均明显高于其他组;低糖透析液组转化生长因子β1、纤连蛋白表达量均明显高于正常对照组与生理盐水组(P<0.05)。④大鼠腹膜组织转化生长因子β1蛋白与纤连蛋白表达量、腹膜厚度之间呈明显的正相关(r=0.86,0.83,P<0.05)。结论:葡萄糖透析液可诱导腹膜组织转化生长因子β1明显上调,腹膜转化生长因子β1高表达与腹膜透析腹膜纤维化密切相关。     

晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化

目的探讨晚期糖基化终产物对大鼠腹膜间皮细胞上皮-间叶转化(EMT)的影响,建立大鼠腹膜间皮细胞EMT模型。方法体外培养的大鼠腹膜间皮细胞经含40 mmol/L晚期糖基化终产物(AGEs)、80 mmol/L AGEs的M199培养基分别培养48、72 h;以正常M199培养基和含80 mmol/LBSA的M199培养基为对照。采用相差显微镜观察细胞形态学改变,应用实时定量PCR法检测间皮细胞E-cadherin、α-SMA、Collagen I、TGF-β1、VEGF mRNA的表达;应用Western blot检测E-cadher-in、α-SMA蛋白的表达;应用ELISA法检测间皮细胞TGF-β1、VEGF蛋白表达水平。结果①AGEs(40 mmol/L、80 mmol/L)刺激后,大鼠腹膜间皮细胞由典型的上皮细胞形态逐渐变为梭形及不规则形,类似肌成纤维细胞;②AGEs(40 mmol/L、80mmol/L)刺激后,大鼠腹膜间皮细胞α-SMA、Collagen I、TGF-β1、VEGF mRNA和α-SMA、TGF-β、VEGF蛋白表达水平显著增加(P<0.05);③AGEs(40 mmol/L、80 mmol/L)刺激后,大鼠腹膜间皮细胞E-cadherin mRNA和E-cadherin蛋白表达水平显著下降(P<0.05)。结论 AGEs能上调α-SMA、Collagen I、TGF-β、VEGF表达和下调E-cadherin表达,AGEs诱导大鼠腹膜间皮细胞EMT。     

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.     

肝细胞生长因子对腹膜透析腹膜纤维化防治作用的研究

目的探讨肝细胞生长因子对腹膜透析腹膜纤维化的防治作用及其机制。方法①MTT法测定肝细胞生长因子对腹膜间皮细胞增殖的影响；②酶联免疫（ELISA）法检测细胞培养液中纤连蛋白（FN）、转化生长因子-β1（TGF-β1）及纤溶酶原激活物抑制物（PAI-1）水平；⑧逆转录-聚合酶链反应（RTPCR）法检测FN、PAI-1及TGF-β1 mRNA的表达。结果肝细胞生长因子浓度〉30ng／ml时，可以改善高糖对腹膜间皮细胞增殖的抑制作用。腹膜透析液组FN、PAI-1、TGF-β1蛋白水平显著升高，肝细胞生长因子组FN、PAI-1、TGF-β1蛋门水平均显著减少。腹膜透析液组高糖上调腹膜间皮细胞FN、PAI-1、TGF-β1 mRNA的表达，肝细胞生长因子组使FN、PAI-1、TGF-β1 mRNA的表达水平下调。结论肝细胞生长因子能改善高糖对腹膜间皮细胞增殖的抑制作用，同时可以使腹膜间皮细胞FN、PAI-1、TGF-β1的表达水平下调。     

氯沙坦联合帕立骨化醇对糖尿病大鼠肾组织CTGF及RhoA/ROCK1表达的影响

目的研究氯沙坦联合帕立骨化醇对糖尿病大鼠肾脏组织结缔组织生长因子(CTGF)及Rho亚家族蛋白A/Rho相关卷曲螺旋形成蛋白激酶1(RhoA/ROCK1)表达的影响。方法将75只大鼠按照随机数字表法分为模型组、骨化醇组、氯沙坦组、联合组和对照组,每组15只。对照组给予注射等量的缓冲液灌胃,其余各组均给予腹腔内注射链脲霉素和柠檬酸缓冲液的混合液25 mg/kg灌胃。糖尿病肾病模型建立成功后,骨化醇组给予帕立骨化醇0.4μg/(kg·d)灌胃;氯沙坦组给予氯沙坦15 mg/(kg·d)灌胃;联合组给予帕立骨化醇0.4μg/(kg·d)和氯沙坦15 mg/(kg·d)灌胃。模型组和对照组给予等量的生理盐水灌胃。观察各组24 h尿蛋白、血肌酐、血尿素氮、RhoA及ROCK1表达值、转化生长因子(TGF-β1)及结缔组织生长因子(CTGF)表达值。结果模型组、氯沙坦组、骨化醇组及联合组的24 h尿蛋白、血肌酐、血尿素氮、RhoA及ROCK1表达值、TGF-β1及CTGF表达值均高于对照组(P0.05);氯沙坦组、骨化醇组及联合组的24 h尿蛋白、血肌酐、血尿素氮、RhoA及ROCK1表达值、TGF-β1及CTGF表达值均低于模型组(P0.05);联合组的24 h尿蛋白、血肌酐、血尿素氮、RhoA及ROCK1表达值、TGF-β1及CTGF表达值低于氯沙坦组、骨化醇组(P0.05)。结论氯沙坦联合帕立骨化醇的治疗方案能够对糖尿病肾病大鼠起到保护肾脏的作用,可减轻肾脏组织的纤维化,延缓糖尿病肾病的进展。     

骨形成蛋白-7对晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化的影响

目的探讨骨形成蛋白-7(BMP-7)对晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化(EMT)的影响。方法晚期糖基化终产物诱导发生上皮-间叶转化的体外培养大鼠腹膜间皮细胞,分别经含5 ng/mL及80 mmol/L晚期糖基化终产物的M199培养基和含10 ng/mL BMP-7及80 mmol/L晚期糖基化终产物的M199培养基培养48 h,以含80 mmol/L晚期糖基化终产物的M199培养基为对照,应用实时定量PCR法检测间皮细胞E-cadherin、α平滑肌肌动蛋白(α-SMA)、Ⅰ型胶原(Collagen I)、转化生长因子β1(TGF-β1)、血管内皮生长因子(VEGF)mRNA的表达;应用Western印迹法检测E-cadherin、α-SMA蛋白的表达;应用ELISA法检测间皮细胞TGF-β1、VEGF蛋白表达水平。结果 BMP-7作用后,上皮-间叶转化的大鼠腹膜间皮细胞E-cadherin mRNA和E-cadherin蛋白表达水平显著增加(P<0.05)。BMP-7作用后,上皮-间叶转化的大鼠腹膜间皮α-SMA、Collagen I、TGF-β1、VEGFmRNA和α-SMA、TGF-β、VEGF蛋白表达水平显著下降(P<0.05)。结论 BMP-7能上调上皮-间叶转化的大鼠腹膜间皮细胞E-cadherin表达和下调α-SMA、Collagen I、TGF-β、VEGF表达,BMP-7能逆转晚期糖基化终产物诱导大鼠腹膜间皮细胞EMT。     

Structural and functional alterations of the peritoneum after prolonged exposure to dialysis solutions: role of aminoguanidine.

OBJECTIVE: The effect of long-term use of high glucose dialysate on peritoneal structure and function, and its relation with accumulation of advanced glycosylation end-product (AGE) in the peritoneum was investigated in this study. METHODS: Dialysates with 4.25% glucose were injected into the peritoneal cavity of normal rats for 12 weeks without (PD, n = 7) and with (1 g/L, PD+AG, n = 7) aminoguanidine in their drinking water. Rats not having intraperitoneal (IP) injection served as control (n = 9). After 12 weeks of IP injection, a 2-hour peritoneal equilibration test (PET) was performed using 30 mL 4.25% glucose dialysate. Intraperitoneal volume (IPV), dialysate-to-plasma urea ratio at 2 hours (D/P2), the ratio of dialysate glucose at 2 hours to initial dialysate glucose (D2/D0), and the peritoneal fluid absorption rate (Qa) were evaluated. After the PET, samples of the parietal peritoneum were taken for hematoxylin and eosin (H&E) staining and immunohistochemical staining for AGE. RESULTS: The IPV and D2/D0 glucose were significantly lower and Qa and D2/P2 urea significantly higher in the PD group than in the control group. Aminoguanidine reversed in part the changes in IPV and D2/P2 urea in the PD group; it had no effect on Qa and D2/D0 glucose. The H&E staining showed a linear mesothelial lining with negligible cells and capillaries in the narrow submesothelial space in the control group. Mesothelial denudation and submesothelial infiltration of monocytes and capillary formation were observed in the PD group. Mesothelial denudation was relatively intact in the PD+AG group compared with the PD group. Submesothelial monocyte infiltration and capillary formation in the PD+AG group were not as prominent as in the PD group. Positive AGE staining was found in the submesothelial space, vascular walls, and endomysium in the PD group, while it was markedly attenuated in PD+AG group and negligible in the control group. CONCLUSION: Long-term use of high glucose solutions induced peritoneal AGE accumulation and mesothelial denudation, and increased peritoneal permeability and peritoneal fluid absorption rate. Inhibition of peritoneal AGE accumulation prevented those functional and structural damages to the peritoneum.     

Enhanced expression of heat shock protein 47 in rat model of peritoneal fibrosis.

OBJECTIVE: Peritoneal fibrosis is one of the serious complications of continuous ambulatory peritoneal dialysis therapy and is characterized by collagen accumulation. Heat shock protein 47 (HSP-47) is a collagen-specific molecular chaperon and is closely associated with collagen synthesis; however, the involvement of HSP-47 in the progression of peritoneal fibrosis is not fully understood. DESIGN: To examine the serial pathological alterations caused by peritoneal fibrosis, we made an experimental model of peritoneal fibrosis by daily intraperitoneal injection of chlorhexidine gluconate (CG) in rats for 28 days and examined the expression of HSP-47 together with that of types I and III collagen, alpha-smooth muscle actin (aSMA), and ED-1 (a marker for macrophages) using immunohistochemistry. Rats treated with saline containing 15% ethanol were used as the control group. RESULTS: In the control group, the peritoneal tissue was slightly thickened and HSP-47 was expressed in the peritoneum at day 28. In the CG group, the peritoneal tissue serially became thickened and fibrotic. The expression of HSP-47 was evident in mesothelial cells and submesothelial connective tissue after day 7 of treatment with CG, and increased thereafter. The expression of types I and III collagen and aSMA was proportionally strengthened during our experiments. ED-1-positive cells were present in thickened areas with abundant proliferation of collagen fiber. The number of cells positive for ED-1 increased gradually and reached a maximum at day 21. CONCLUSION: Our results indicate that, in a rat experimental model of peritoneal fibrosis, the expression of HSP-47 is associated with the progression of peritoneal fibrosis.     

肾素血管紧张素系统抑制剂对长期腹膜透析患者腹膜纤维化和残余肾功能的作用

目的探讨血管紧张素转换酶抑制剂(angiotensin converting enzyme inhibitor,ACEI)和(或)血管紧张素受体拮抗剂(angiotensin receptor blocker,ARB)在长期腹膜透析患者中的作用。方法选择山东大学附属省立医院腹膜透析中心68例新腹膜透析患者,分为两组:ACEI/ARB组39例,对照组为未使用ACEI/ARB药物,共29例。观察期12个月,观察开始和结束时均行腹膜平衡实验(peritoneal equilibration tests,PET)。酶联免疫法(ELISA)检测PET前过夜腹透液中的纤维粘连蛋白(fibronectin,Fn)、转化生长因子(transforming growth factorβ1,TGF-β1)、血管内皮生长因子(vascular endothelial growth factor,VEGF)和水通道蛋白(aquaporin1,AQP1)。结果两组患者基础临床参数没有明显差别。整个研究过程中两组腹膜炎的发生率没有明显差别。ACEI/ARB组和对照组的残余肾功能[(4.7±1.9)ml/min和(4.6±2.0)ml/min]在起始时没有明显差别,12个月后两组的残余肾功能分别降低为(3.17±1.35)ml/min和(2.78±1.41)ml/min,差异有统计学意义(P=0.014)。两组的透析液葡萄糖浓度(4h)比值(D4/D0)都表现为轻微的下降,但两组间差异无统计学意义(P0.05)。与ACE/ARB组比较,12个月后对照组的透析液/血清肌酐(4h)比值(D/Pcr)升高的速度明显增加,△D/Pcr分别为(0.08±0.031)和(0.152±0.033),超滤量也显著性降低,超滤量分别为(-87.517±42.611)ml/4h和(-161.825±61.414)ml/4h,差异均有统计学意义(P0.05)。对照组过夜腹透液中除AQP1外,TGF-β1,FN和VEGF表达明显增加(P0.05),而ACE/ARB组没有显著性变化。结论在长期腹膜透析患者中肾素血管紧张素系统抑制剂可降低超滤量和残余肾功能下降的速度,抑制腹膜纤维化的发生和发展,在抗腹膜生硬化和保护腹膜功能上起到了积极的作用。     

Role of peritoneal mesothelial cells and fibroblasts in the synthesis of hyaluronan during peritoneal dialysis.

OBJECTIVE: To assess the in vitro synthesis rate of hyaluronan (HA) by human peritoneal mesothelial cells and peritoneal fibroblasts in the presence of effluent dialysate from continuous ambulatory peritoneal dialysis (CAPD) patients. METHODS: We used primary cultures of human peritoneal mesothelial cells and peritoneal fibroblasts from nonuremic patients to study the effect of interleukin-1beta (II-1beta) and pooled effluent dialysate, from noninfected and infected CAPD patients, on the synthesis of HA by the studied cells. We also tested the effect of the exogenous HA on the synthesis rate of that glycosaminoglycan. We studied the correlation between HA concentration in effluent dialysate and the stimulatory effect of that solution on in vitro synthesis of HA by mesothelium. RESULTS: Peritoneal fibroblasts produce more HA than mesothelial cells. Noninfected effluent dialysates or dialysates from CAPD patients with peritonitis stimulate synthesis of HA by mesothelial cells and fibroblasts. Interleukin-1beta has a stimulating effect, which was synergistic with effluent dialysates, on the synthesis of HA by mesothelium and peritoneal fibroblasts. A weak correlation was demonstrated between the level of HA in effluent dialysate and the stimulatory effect of that dialysate on in vitro synthesis of HA by mesothelial cells. CONCLUSIONS: Peritoneal fibroblasts are a more potent source of HA than are mesothelial cells, but probably the latter are the main source of HA in drained dialysate. Although effluent dialysates contain factors that stimulate the production of HA by mesothelium, there is weak correlation between that stimulatory effect and the actual HA concentration in the dialysate, which, in some patients, might suggest low "responsiveness" of the membrane.     

白芍总苷对腹膜纤维化大鼠HMGB-1、Smad7的影响

目的:观察白芍总苷对腹膜纤维化大鼠HMGB-1、Smad7蛋白表达的影响。方法:选用健康SD大鼠32只,随机分为空白组、模型组、白芍总苷高剂量组、白芍总苷低剂量组,每组8只。用4.25%腹膜透析液每日腹腔注射和脂多糖间断腹腔注射进行大鼠腹膜纤维化造模。白芍总苷高剂量组每日给予白芍总苷200 mg/(kg·d)灌胃,白芍总苷低剂量组每日给予白芍总苷100 mg/(kg·d)灌胃;实验第30天结束后酶联免疫分析(ELISA)检测大鼠透析液中HMGB-1、Smad7的蛋白质表达情况。结果:模型组、白芍总苷高剂量组、白芍总苷低剂量组腹透液中HMGB-1水平与空白组相比有所增高,白芍总苷两组与模型组相比较,HMGB-1水平有所下降,但各组间差异无统计学意义(P>0.05)。模型组与空白组比较,腹透液中Smad7的含量显著升高,差异有统计学意义(P<0.05)。白芍总苷两组升高更为明显,与空白组相比,差异有统计学意义(P<0.01);与模型组相比较,白芍总苷两组腹透液中Smad7水平明显上调,差异有统计学意义(P<0.05)。余各组间两两比较,腹透液中Smad7含量无统计学差异(P>0.05)。结论:白芍总苷可一定程度下调HMGB-1水平,但作用机理不确切,可能通过上调Smad7的表达来减轻大鼠的腹膜纤维化。     

High glucose solution and spent dialysate stimulate the synthesis of transforming growth factor-beta1 of human peritoneal mesothelial cells: effect of cytokine costimulation.

OBJECTIVE: To investigate the effect of high glucose and spent peritoneal dialysate on the transforming growth factor-beta1 (TGFbeta1) synthesis of cultured human peritoneal mesothelial cells (HPMCs) and to examine the effect of costimulation with high glucose or spent dialysate, and cytokines, interleukin-1beta (IL-1beta), and tumor necrosis factor-alpha (TNFalpha) on TGFbeta1 synthesis of HPMCs. DESIGN: HPMCs were exposed to different concentrations of glucose (30, 60, and 90 mmol/L) or spent peritoneal dialysate for 48 hours in the absence or presence of IL-1beta (1 ng/mL) and TNFalpha(1 ng/mL).TGFbeta1 mRNA expression was assessed by Northern blot analysis and TGFbeta1 protein release by Western blot analysis and enzyme-linked immunosorbent assay (ELISA). RESULTS: Exposure of HPMCs to high glucose conditions (30, 60, and 90 mmol/L of D-glucose) induced 2.3-, 3.6-, and 4.0-fold increases inTGFbeta1 mRNA expression of HPMC with enhancedTGFbeta1 protein synthesis and secretion into the media, whereas there were no significant changes in TGFbeta1 synthesis with equimolar concentrations of D-mannitol. Incubation with spent dialysate also significantly increased TGFbeta1 mRNA expression and protein secretion compared to control media (p < 0.05). Stimulation with IL-1beta (1 ng/mL) or TNFalpha (1 ng/mL) resulted in a significant increase in TGFbeta1 mRNA expression after 48 hours: 2.7 and 2.1 times the control level, respectively. However,TNFalpha-induced increase in TGFbeta1 mRNA expression was not translated intoTGFbeta1 protein secretion, while IL-1beta stimulation induced a significant increase in TGFbeta1 protein secretion as well as TGFbeta1 mRNA expression. Combined stimulation by high glucose or spent dialysate, together with IL-1beta or TNFalpha, showed a greater increase in TGFbeta1 mRNA expression and protein secretion compared to stimulation by high glucose or spent dialysate alone. CONCLUSION: Our results clearly show that high glucose solution and spent dialysate themselves might be sufficient to stimulate the production of TGFbeta1 by peritoneal mesothelial cells. In peritoneal dialysis patients, this state of chronic induction of TGFbeta1 is further exacerbated in the presence of peritonitis because of the stimulatory effect of proinflammatory cytokines, resulting in augmented TGFbeta1 synthesis, thus promoting peritoneal fibrosis.     

Combination therapy with an angiotensin-converting enzyme inhibitor and an angiotensin II receptor blocker synergistically suppresses chronic pancreatitis in rats

We recently demonstrated that both lisinopril and candesartan, an angiotensin-converting enzyme inhibitor and angiotensin II type 1 receptor blocker, respectively, attenuate pancreatic inflammation and fibrosis in male Wistar Bonn/Kobori (WBN/Kob) rats. The purpose of the present study was to assess whether combination therapy with low doses of both, ineffective when given alone, might synergistically exert protective effects. Lisinopril, candesartan, or a combination of both in drinking water was administered to 10-week-old male WBN/Kob rats for 10 weeks. Parameters of inflammation and fibrosis, positive immunostaining for alpha-smooth muscle actin, and gene expression of cytokine and growth factors were assessed, as well as circulating renin-angiotensin system components. Dose-dependent effects of combination therapy were also investigated. Only combination therapy attenuated gross alterations in the pancreas, as quantitatively confirmed by increases in pancreatic weights and decreases in myeloperoxidase activity, hydroxyproline content, histologic scores, relative fibrosis area, and relative area of alpha-smooth muscle actin-positive cells. Combination therapy suppressed up-regulation of tumor necrosis factor-alpha, platelet-derived growth factor-receptor beta, and transforming growth factor-beta1 mRNA in the pancreas. Dose dependence of combination therapy was recognized with reference to improvement in these parameters. The conclusions are that combination therapy synergistically alleviated pancreatic inflammation and fibrosis in male WBN/Kob rats. This effect may be related to suppression of tumor necrosis factor-alpha, platelet-derived growth factor-receptor beta, and transforming growth factor-beta1 mRNA. Compared with the either therapy alone, combination therapy with an angiotensin-converting enzyme inhibitor and an angiotensin II type 1 receptor blocker may be more beneficial for treating chronic pancreatitis.