Effect of salvianolate on intestinal epithelium tight junction protein zonula occludens protein 1 in cirrhotic rats

AIM: To study the effect of salvianolate on tight junctions (TJs) and zonula occludens protein 1 (ZO-1) in small intestinal mucosa of cirrhotic rats.METHODS: Cirrhosis was induced using carbon tetrachloride. Rats were randomly divided into the untreated group, low-dose salvianolate (12 mg/kg) treatment group, medium-dose salvianolate (24 mg/kg) treatment group, and high-dose salvianolate (48 mg/kg) treatment group, and were treated for 2 wk. Another 10 healthy rats served as the normal control group. Histological changes in liver tissue samples were observed under a light microscope. We evaluated morphologic indices of ileal mucosa including intestinal villi width and thickness of mucosa and intestinal wall using a pathological image analysis system. Ultrastructural changes in small intestinal mucosa were investigated in the five groups using transmission electron microscopy. The changes in ZO-1 expression, a tight junction protein, were analyzed by immunocytochemistry. The staining index was calculated as the product of the staining intensity score and the proportion of positive cells.RESULTS: In the untreated group, hepatocytes showed a disordered arrangement, fatty degeneration was extensive, swelling was obvious, and disorganized lobules were divided by collagen fibers in hepatic tissue, which were partly improved in the salvianolate treated groups. In the untreated group, abundant lymphocytes infiltrated the fibrous tissue with proliferation of bile ducts, and collagen fibers gradually decreased and damaged hepatic lobules were partly repaired following salvianolate treatment. Compared with the untreated group, no differences in intestinal villi width between the five groups were observed. The villi height as well as mucosa and intestinal wall thickness gradually thickened with salvianolate treatment and were significantly shorter in the untreated group compared with those in the salvianolate treatment groups and normal group (P < 0.01). The number of microvilli decreased and showed irregular lengths and arrangements in the untreated group. The intercellular space between epithelial cells was wider. The TJs were discontinuous, which indicated disruption in TJ morphology in the untreated group. In the treated groups, the microvilli in the intestinal epithelium were regular and the TJs were gradually integrated and distinct. The expression of ZO-1 decreased in the small intestine of the untreated cirrhotic rats. The high expression rate of ZO-1 in ileal mucosa in the untreated group was significantly lower than that in the medium-dose salvianolate group (21.43% vs 64.29%, χ² = 5.25, P < 0.05), high-dose salvianolate group (21.43% vs 76.92%, χ² = 8.315, P < 0.01) and normal group (21.43% vs 90%, χ² = 10.98, P < 0.01).CONCLUSION: Salvianolate improves liver histopathological changes, repairs intestinal mucosa and TJ structure, and enhances ZO-1 expression in the small intestinal mucosa in cirrhotic rats.     

血小板活化因子受体拮抗剂对幼年大鼠肠黏膜上皮细胞间紧密连接蛋白的影响

目的:探讨血小板活化因子(platelet activating factor,PAF)受体拮抗剂对肠黏膜上皮细胞间紧密连接蛋白ZO-1的影响．方法:18日龄Wistar大鼠,随机分为对照组,内毒素组(LPS组)和PAF受体拮抗剂组(预防组和治疗组)．LPS组和对照组分别腹腔注射内毒素 (5 mg/kg)和生理盐水(1 mL/kg)．预防组和治疗组分别于每一时相点注射LPS前、后30 min 腹腔注射PAF受体拮抗剂BN52021(5 mg/kg)．按时间点分别处死动物,取回肠用于电镜观察,免疫组化及RT-PCR检测ZO-1．结果:电镜下对照组肠微绒毛及细胞间紧密连接未见异常．实验组上皮细胞连接增宽;微绒毛变细、稀疏,部分断裂、脱落;细胞器受损．拮抗剂组改变较实验组减轻．紧密连接蛋白ZO-1正常时均匀一致地分布于小肠上皮细胞连接处的尖端,呈蜂巢状,实验组ZO-1 分布不均,染色变淡．免疫组化平均光密度值及RT-PCR结果可见LPS组ZO-1表达明显低于对照组,6 h表达最低,光密度从对照组 0．224 7降至LPS组0．198 5,ZO-1 mRNA从1．18 降至0．16(P<0．01)．预防组及治疗组变化趋势同LPS组,6 h预防组及治疗组光密度分别为 0．199 2和0．203 8,ZO-1 mRNA分别为0．47和 0．53,与对照组相比均有显著差异(P<0．01)．各时相点ZO-1较LPS组高,预防组较治疗组 ZO-1略低,无统计学差异．结论:PAF可降低肠道紧密连接蛋白ZO-1,从而损害肠道的屏障功能,而PAF受体拮抗剂可减轻肠道屏障功能损伤的程度．     

NO体外对肠上皮细胞表达紧密连接蛋白Occludin的影响

目的:探讨一氧化氮(NO)对肠上皮细胞表达紧密连接蛋白Occludin的影响,以研究NO对肠黏膜屏障的作用机制.方法:将NO的供体Sin1与肠上皮细胞株Caco-2共培养24 h,采用MTT方法观察NO对肠上皮细胞的作用,并分别提取细胞蛋白和总RNA,采用免疫蛋白印迹(Westem blot)蛋白半定量方法和实时定量聚合酶链式反应(RO-PCR)方法检测不同NO浓度对Caco-2细胞表达紧密连接蛋白Occludin蛋白和mRNA表达的影响.结果:随着Sin1浓度升高(125,250,500和1000μmol/L)NO对细胞的杀伤作用产生并逐渐增大,Occludin蛋白表达量和mRNA的相对表达量与无Sin1刺激时蛋白及mRNA的表达量相比明显降低(蛋白:375±0.5,374±0.8,363±0.3.363±0.7 vs 398±0.7;mRNA:0.689±0.01,0.578±0.09,0.554±0.03,0.619±0.04 vs 1,均P<0.01).结论:NO可直接损伤肠上皮细胞,同时以剂量依赖形式在蛋白和分子水平影响紧密连接蛋白Occludin的表达.     

Effects of penehyclidine hydrochloride on rat intestinal barrier function during cardiopulmonary bypass

AIM:To test the ability of penehyclidine hydrochloride (PHC) to attenuate intestinal injury in a rat cardiopulmonary bypass (CPB) model.METHODS:Male Sprague-Dawley rats were randomly divided into six groups (eight each):sham-operated control;sham-operated low-dose PHC control (0.6 mg/kg);sham-operated high-dose PHC control (2.0 mg/kg);CPB vehicle control;CPB low-dose PHC (0.6 mg/kg);and CPB high-dose PHC (2.0 mg/kg).Blood samples were collected from the femoral artery 2 h after CPB for determination of plas...     

暴发性肝衰竭时肠上皮细胞间紧密连接蛋白occludin表达下降

目的:研究暴发性肝衰竭小鼠大肠上皮细胞间紧密连接蛋白occludin表达的变化.方法:BALB/c小鼠150只随机分为生理盐水对照组(n=30)、内毒素(LPS)对照组(n=30)、D-氨基半乳糖(D-GalN)对照组(n=30)和暴发性肝衰竭(LPS GalN)组(n=60).采用D-GalN和LPS联合ip制备暴发性肝衰竭小鼠动物模型.应用免疫组织化学技术、Westernblot及实时定量PCR检测暴发性肝衰竭进程中,小鼠大肠上皮细胞间紧密连接蛋白occludin的定位、表达及mRNA的变化.结果:occludin蛋白主要沿小鼠大肠黏膜上皮细胞膜的顶端呈线状分布,在暴发性肝衰竭组小鼠,6h时occludin的阳性染色开始减少,9h时更为明显.Westernblot结果与免疫组织化学结果相一致,6h时开始下降(0.48±0.07),9h达到最低值(0.36±0.05),与生理盐水对照组(0.71±0.09)相比差异显著(P<0.05).实时定量PCR结果显示,暴发性肝衰竭小鼠2h时occludinmRNA即开始下降(0.85±0.12),6h时达到最低值(0.72±0.04),与生理盐水对照组(1.00±0.05)相比有明显差异(P<0.05),9h时有所恢复(0.93±0.10).结论:在暴发性肝衰竭过程中,大肠上皮细胞间紧密连接蛋白occludin表达下降,其mRNA也呈下调趋势.     

Coagulation and fibrinolytic protein kinetics in cardiopulmonary bypass

The development of Cardiopulmonary Bypass (CPB) catopulted the field of cardiothoracic surgery into a new dimension—one that changed the lives of individuals with congenital and acquired heart disease worldwide. Despite its contributions, CPB has clear limitations and creates unique challenges for clinicians and patients alike, stemming from profound hemostatic pertubations and accompanying risk for bleeding and possibly thrombotic complications.     

体外循环心脏手术对机体免疫功能的影响

目的　探讨体外循环心脏直视手术病人围术期细胞和体液免疫功能的变化及其临床意义。方法　选择 45例心脏直视手术病人 ,检测术前、术后 1天、3天、7天和 14天的血清 T淋巴细胞亚群 ,CD3 ( T淋巴细胞总数 )、CD4(辅助 T淋巴细胞 )、CD8(抑制 T淋巴细胞 )、自然杀伤细胞 ( NK)活性和免疫球蛋白水平。结果　术后第 1、3天血清 CD3、CD4、CD8活性明显降低 ,CD4/ CD8比值下降 ,NK细胞活性增强 ;免疫球蛋白均较术前降低。术后 7至 14天逐步恢复至术前水平。结论　体外循环心脏直视手术对患者细胞和体液免疫均有不良影响 ,临床应采取相应措施 ,改善术后病人免疫机能 ,预防术后感染     

紧密连接蛋白claudin-1在肠易激综合征患者肠黏膜中的表达及意义

目的 探讨肠易激综合征(IBS)患者肠黏膜紧密连接蛋白claudln-1表达在排便性状和习惯改变中的意义.方法 用免疫组化方法检测对照组(痔疮10例、结肠息肉10例)和观察组(43例IBS患者,腹泻型23例、便秘型20例)小肠和结肠黏膜claudin.1表达.结果 免疫组化定位显示两组肠黏膜claudin.1均分布于肠上皮紧密连接的细胞膜,胞核及核膜无表达;与对照组比较,观察组腹泻型患者小肠和结肠黏膜clandin-1表达明显下降(P＜0.05),便秘型患者clandin-1表达明显上升高(P＜0.05),腹泻型与便秘型比较有显著差异(P＜0.05).结论 claudin-1表达异常在IBS患者排便性状和习惯改变形成中可能有重要作用.     

常温和中低温体外循环对甲状腺素变化的影响

目的探讨常温及低温体外循环心脏直视手术对甲状腺素的影响。方法选择先天性心脏病患者80例,随机分为常温组及低温组,各40例,分别于术晨、体外循环结束时及术后6、12、24、48h抽取患者动脉血,测定血浆T3、T4和TSH。结果两组术前各项检查指标差异无统计学意义。常温体外循环下甲状腺素T3升高水平低于中低温,恢复时间比中低温明显缩短。结论常温体外循环心脏直视手术对甲状腺素的影响显著轻于低温组,因而术后机体的恢复优于低温方法。     

肠内营养对增龄大鼠肠黏膜上皮屏障的影响

目的 研究肠内营养（百普力）对老年大鼠增龄过程中肠黏膜上皮屏障变化的影响.方法 将20只12月龄和20只3月龄SD大鼠均随机分为肠内营养组和常规饮食组,肠内营养组给予常规饲料和肠内营养液（百普力）,常规饮食组给予标准饲料,饲养1月后取各组大鼠回肠组织常规HE染色观察形态学改变,半定量RT-PCR方法检测小肠黏膜组织中紧密连接蛋白（Occludin、ZO-1）mRNA表达水平,免疫组化法检测Occludin、ZO-1水平.统计学处理采用独立样本t检验和多元回归分析.结果 常规饮食组12月龄大鼠与3月龄大鼠相比：小肠绒毛高度降低（P＜0.05）,小肠黏膜组织中Occludin、ZO-1 mRNA表达减少（P＜0.05）,小肠黏膜组织中Occludin、ZO-1减少（P＜0.05）; 12月龄肠内营养组大鼠与常规饮食组大鼠相比,小肠黏膜厚度和绒毛高度增加（P＜0.05）,小肠黏膜组织中Occludin、ZO-1 mRNA表达增多（P＜0.05）,Occludin、ZO-1增多（P＜0.05）.结论 肠内营养（百普力）能改善老年SD大鼠肠黏膜上皮屏障中紧密连接蛋白（Occludin、ZO-1）的减少程度,对衰老时肠黏膜屏障损伤具有保护作用.     

肠黏膜细胞的紧密连接与肠壁通透性的研究进展

肠黏膜细胞的紧密连接是构成肠黏膜屏障的重要结构基础,在调节肠黏膜通透性中发挥着重要的作用.其结构的破坏,可导致肠壁通透性增高,引起细菌移位、全身炎症反应及多器官功能受损.本文就肠黏膜紧密连接的结构和功能、与通透性的影响因素及改善措施进行了综述.     

水通道蛋白3与紧密连接的相关性

目的:探讨水通道蛋白3(aquaporin 3,AQP3)对肠黏膜上皮细胞间紧密连接(tight junction,TJ)的影响,并探讨其可能的作用机制.方法:应用Caco-2细胞系在体外构建肠黏膜上皮屏障,构建沉默AQP3的shRNA慢病毒载体,建立稳定转染细胞系.实验分为3组:空白对照组(BLANK)、阴性对照组(NC)、AQP3干扰组(AQP3 shRNA).Western blot验证TJ相关蛋白Occludin以及Claudin-1的表达情况;并且采用免疫细胞化学法观察TJ相关蛋白的分布和结构变化.结果:RT-PCR及Western blot结果显示在Caco-2细胞系中成功沉默AQP3的表达.干扰组与对照组相比下降约75%.Western blot结果显示AQP3干扰组TJ相关蛋白Occludin以及Claudin-1的表达明显降低.免疫细胞化学结果显示Caco-2细胞间Occludin以及Claudin-1主要表达在细胞膜和/或胞浆中,Occludin和Claudin-1细胞间棕褐色颗粒减少,结构变模糊.相邻Caco-2细胞间TJ结构遭到破坏.结论:靶向AQP3的shRNA技术可以引起TJ的结构变化和相关蛋白的表达分布的异常.     

肿瘤坏死因子受体l在暴发性肝衰竭肠黏膜屏障功能损害中的作用

肠黏膜屏障对于抵抗肠道病原微生物的侵入具有重要作用。暴发性肝衰竭（FHF）时肠黏膜屏障的通透性增加^[1-4]，细菌及其毒性产物穿透肠壁进人体内，引起自发性腹膜炎甚至败血症。FHF时肠黏膜屏障通透性的增加与肠上皮细胞紧密连接的破坏有关。我们的前期研究发现，在FHF时，肠上皮细胞间紧密连接蛋白occludin表达下降，     

保护性机械通气和PGE1联用对婴幼儿体外循环术后急性肺损伤的保护作用

目的评价联合应用保护性机械通气和前列腺素E1(PGE1)对婴幼儿体外循环后急性肺损伤(ALI)的保护作用.方法婴幼儿体外循环术后ALI患者18例,随机分为A、B两组,每组9例.A组采用保护性机械通气策略治疗,低潮气量(6～8 ml/kg),高呼气末正压(6～12 cmH2O),限制吸气峰压(＜35 cmH2O),适当允许高碳酸血症(PaCO2≤60mmHg).B组在施行保护性机械通气的基础上联合静脉应用PGE1,剂量为30～100ng/(kg·min),连用7～10天.观察两组患儿的临床转归,并监测48小时内肺动态顺应性、氧合指数、血浆肿瘤坏死因子(TNFα)和白细胞介素-8(IL-8)的含量变化.结果死亡1例(5.6%),其余无严重并发症发生.在治疗后48小时内的各个时点,B组肺的动态顺应性显著高于A组(P＜0.05),氧合指数非常显著降低(P＜0.01),血浆TNFα和IL-8含量显著低于A组(P＜0.01).结论二者联合应用具有协同作用,可明显提高婴幼儿体外循环后ALI的治疗效果.     

急性等容血液稀释对体外循环术后凝血功能的影响

目的:探讨急性等容血液稀释(ANH)对体外循环下心脏直视手术患者凝血功能的影响。方法:选择在体外循环下行心脏直视手术的患者64例,随机分为ANH组(n=30)和非ANH组(n=34),观察两组患者手术前后凝血功能和血常规指标,并记录术中出血量及输液量。结果:与术前相比,两组患者术后凝血酶原时间(PT)、活化部分凝血活酶时间(APTT)、凝血酶时间(TT)均较术前明显延长,纤维蛋白原(FIB)、血小板计数(PLT)、血红蛋白(Hb)和红细胞压积(Hct)均较术前明显降低,但两组间PT、APTT、TT、Hb、Hct变化无统计学差异;与非ANH组相比,ANH组患者术后FIB和PLT降低更加明显(P0.05)。两组患者术中失血量无统计学差异。结论:体外循环心脏直视手术患者应用ANH时对凝血功能影响轻微,且术中失血量无明显增加。     

肠炎清对受损肠黏膜修复和二胺氧化酶的影响

[目的]探讨肠炎清对受损肠黏膜修复和二胺氧化酶（DAO）的影响。[方法]将60例结肠黏膜损伤患者,随机分为2组,每组30例。并以30例门诊健康体检者作为正常对照（正常）组。治疗组用肠炎清,对照组用美沙拉嗪肠溶片。疗程均为6周。观察2患者组治疗前后肠镜下黏膜镜像评分及DAO水平的变化,并与正常组进行比较。[结果]2患者组治疗前后肠镜下黏膜镜像评分有明显改善（P〈0．01）;2患者组治疗前后DAO活性有明显下降（P〈0．01）,但治疗组治疗后与正常组比较差异无统计学意义（P〉0．05）,而对照组治疗后与正常组比较差异有统计学意义（P〈0．05）。[结论]肠炎清能够有效减轻患者临床症状,修复受损肠黏膜,降低DAO活性,从而降低肠道通透性,改善肠黏膜屏障功能。