鱼油脂肪乳对肠外营养大鼠小肠黏膜紧密连接形态结构的影响

目的:探讨鱼油脂肪乳剂(富含ω-3多不饱和脂肪酸)对肠外营养(PN)大鼠肠上皮细胞紧密连接(TJ)形态和紧密连接蛋白occludin表达的影响。方法:将大鼠随机分为四组,即对照组(正常喂食),PN(禁食5d行PN支持)组,小剂量鱼油治疗(1 ml/kg鱼油静脉注射+PN)组和大剂量鱼油治疗(2 ml/kg鱼油静脉注射+PN)组。观察大鼠术后小肠黏膜上皮细胞TJ形态,以及紧密连接蛋白occludin的表达分布。结果:PN组大鼠治疗5 d后肠上皮细胞TJ结构部分缺失、间隙增宽;大剂量鱼油治疗组TJ形态基本完整,小剂量鱼油治疗组TJ部分缺失。各组occludin蛋白总量无明显差异。提取脂筏组分,发现与对照组比,三个实验组脂筏区域中occludin蛋白表达均明显减少,但大剂量鱼油治疗组较PN组明显升高(P<0.01)。结论:长期PN可导致TJ结构的缺失,脂筏区域紧密连接蛋白occludin表达减少。大剂量鱼油脂肪乳剂可维持occludin蛋白在脂筏区域的表达,保护小肠黏膜细胞TJ的结构和功能。     

植物乳酸杆菌对阻塞性黄疸大鼠肝紧密连接蛋白表达的影响

目的: 探讨植物乳酸杆菌(LP)对阻塞性黄疸大鼠肝细胞凋亡和紧密连接(TJ)蛋白表达变化的影响. 方法: 将大鼠随机分为五组,即对照组、胆总管结扎组、胆总管结扎+LP组、内引流组和内引流+LP组,实验持续10 d.给予添加LP组大鼠每天灌胃1010 CFU.用TUNEL法测肝细胞凋亡阳性率;用透射电镜、Western blot、RT-PCR观察TJ超微结构及相关蛋白(Occludin,Claudin-1,Claudin-4,ZO-1)的表达和分布变化. 结果: 大鼠胆总管结扎10 d后肝细胞凋亡增多,口服LP后状况有所缓解.内引流+LP组较阻黄组凋亡降低(P<0.01).大鼠肝细胞TJ破坏非常明显,口服LP后有所改善,内引流+LP组改善显著.内引流+LP组TJ蛋白mRNA及蛋白的表达改善最明显. 结论: 阻塞性黄疸大鼠内引流及口服LP对受损肝有明显地改善作用,肝细胞凋亡阳性率下降,TJ蛋白及mRNA表达明显好转,有效地抑制了阻塞性黄疸大鼠肝屏障的损伤.     

痰瘀同治方药对Caco-2单层细胞胆汁酸吸收的抑制作用及机制研究

目的探究复方中药对人结肠腺癌Caco-2细胞胆汁酸吸收的抑制作用及机制。方法Transwell细胞小室接种Caco-2细胞,培养21 d以建立单层细胞模型,随机分为空白对照组、胆汁酸刺激组及复方中药+胆汁酸组,其中选取了4种胆汁酸,分别为胆酸(CA)、脱氧胆酸(DCA)、甘氨胆酸(GCA)以及牛磺胆酸(TCA)。观察各组胆汁酸跨膜转运量、相关基因mRNA及蛋白表达水平的变化。结果与相应的胆汁酸刺激组相比,复方中药干预可显著降低胆酸(CA)、甘氨胆酸(GCA)以及牛磺胆酸(TCA)在Transwell细胞小室下室中的浓度,差异有统计学意义(P 0.05),但对DCA无明显作用(P 0.05)。同时,复方中药可对抗CA、GCA以及TCA导致的Caco-2细胞顶端钠依赖性胆汁酸转运体(ASBT)、有机溶质转运体β(OSTβ) m RNA及蛋白表达水平的升高,差异有统计学意义(P 0.05),但对回肠胆汁酸结合蛋白(IBABP)的m RNA表达无显著影响(P 0.05);对脱氧胆酸(DCA)刺激所引起的ASBT和OSTβm RNA及蛋白表达水平的改变无显著影响(P 0.05),但可对抗IBABP mRNA表达水平的下降,差异有统计学意义(P 0.05)。复方中药干预可对抗GCA以及TCA所导致的OSTαm RNA表达水平的升高,差异有统计学意义(P 0.01),但对CA、DCA无显著影响(P 0.05)。结论复方中药可有效抑制肠道上皮细胞对胆汁酸的吸收,其作用可能主要是通过下调ASBT和OSTβ基因m RNA及蛋白的表达来实现,可能是复方中药抑制胆汁酸吸收的潜在作用靶点。     

Caco-2细胞模型的建立及其在真菌毒素吸收率研究中的应用

目的构建Caco-2单层细胞转运5种毒素的吸收率模型并验证。方法以Caco-2细胞系为工具,以脱氧雪腐镰刀菌烯醇(DON)、黄曲霉毒素B1(AFB1)、AFB2、黄曲霉毒素G1(AFG1)和AFG2为研究对象,通过对转运介质溶液中毒素的初始浓度、温度、p H值、抑制剂存在与否等实验条件的优化,构建Caco-2细胞转运DON和4种黄曲霉毒素的生理模型并进行验证。结果 DON、AFB1、AFB2、AFG1及AFG2透过Caco-2单层细胞的量和表观渗透系数(Papp)随着体系溶液中p H的升高有下降趋势。AFB1在p H 7.4~7.5溶液中的Papp值最高,AFB2、AFG1、AFG2及DON在p H 7.0~7.1溶液中的Papp值最高。随着体系温度的升高,DON、AFB1、AFB2、AFG1及AFG2的透膜能力均有所增加。无论体系中AFB1、AFG1及AFG2浓度低或高,均在60 min时透过率达到平台期并维持稳定;而AFB2及DON从高浓度到低浓度均在120 min时透过率达到平台期并维持稳定。结论所建5种毒素Caco-2单层细胞模型灵敏、稳定、可靠、有效。     

Bcl-2蛋白在子宫内膜异位症的表达及意义

目的 :检测正常子宫内膜及内膜异位症的在位、异位内膜 Bcl-2蛋白表达及细胞凋亡率 ,探讨其在子宫内膜异位症发生机制中的作用。方法 :采用免疫组化方法对 50例同期因宫颈癌 I期 ,输卵管节育术患者及盆腔子宫内膜异位症患者 47例的在位、异位内膜 Bcl-2蛋白表达及应用流式细胞技术检测细胞凋亡率。结果 :1对照组 :子宫内膜的 Bcl-2蛋白主要表达于子宫内膜腺上皮细胞浆和 /或细胞膜上。增生期 Bcl-2蛋白的表达显著高于分泌期 (P<0 .0 1)。 2内膜异位症组 :在位内膜的 Bcl-2蛋白亦主要表达于子宫内膜腺上皮细胞胞浆和 /或胞膜上 ,增生期 Bcl-2蛋白表达与分泌期有显著性差异 (P<0 .0 1)。异位症组异位内膜增生期、分泌期无显著性差异 (P>0 .0 5)。 3异位症组 :异位内膜与在位内膜分泌期比较 Bcl-2蛋白表达有显著性差异 (P<0 .0 5)。 4内膜异位症组与对照组比较 :其分泌期 Bcl-2蛋白表达显著高于对照组的分泌期 (P<0 .0 1)。内膜异位症在位内膜细胞凋亡率低于正常子宫内膜。异位内膜细胞 AP峰明显低于异位症在位内膜细胞和正常子宫内膜细胞。结论 :1子宫内膜周期性生理变化与 Bcl-2蛋白表达水平有关。子宫内膜的周期性变化可能与凋亡有关。 Bcl-2蛋白的表达水平越高 ,组织的抗凋亡性越强。 2异位内膜细胞凋?     

内毒素对大鼠肝细胞清蛋白基因表达的影响

目的:观察内毒素刺激后体外培养大鼠肝细胞清蛋白基因的表达,进一步了解感染时低清蛋白血症的分子机制.方法:将1μg/ml内毒素作用于大鼠肝细胞后0、2、8、12、24 h,分别用逆转录聚合酶链反应(RT-PCR)、流式细胞仪和酶链免疫检测法(EIA),检测肝细胞清蛋白mRNA、细胞质内清蛋白前体和分泌到细胞外的清蛋白变化.结果:肝细胞清蛋白mRNA、细胞质内清蛋白前体和分泌到细胞外的清蛋白的变化是一致的,即内毒素作用后2和8 h下降不明显,12 h开始下降,24 h后明显下降.清蛋白mRNA降低约30%,清蛋白前体和清蛋白均下降50%.结论:内毒素可以在体外抑制大鼠肝细胞清蛋白基因的表达,通过抑制清蛋白mRNA的表达来抑制肝细胞清蛋白的合成.     

肿瘤坏死因子和干扰素对肠上皮紧密连接的破坏作用

目的:了解肿瘤坏死因子(TNF-α)和干扰素(IFN-γ)对上皮细胞功能的作用,研究TNF-α和IFN-γ与上皮细胞屏障功能的关系. 方法:T84细胞株用加有5%胎牛血清的DMEM/F-12 混合培养液培养.实验分为四组:即对照组、TNF-α处理组、IFN-γ处理组和TNF-α与IFN-γ联合处理组.TNF-α处理组和IFN-γ处理组细胞分别用加有10 ng/mL TNF-α和100 U/mL IFN-γ的培养液处理72 h.各细胞因子联合处理组用100 U/mL IFN-γ处理19 h后,再加入10 ng/mL TNF-α.用EVOM电压电阻仪测定细胞跨膜电阻抗(TER)变化,用透射电镜观察细胞紧密连接的形态改变. 结果:IFN-γ单独处理48 h后,TER降低至78.06%,培养至72 h降低为59.31%.在单一TNF-α作用下,24 h后TER值逐渐降低,72 h 降低至56.82%.与对照细胞相比,TNF-α和IFN-γ联合处理的T84细胞株TER值显著降低,72 h TER值为20.88%,表明细胞紧密连接的结构破坏. 结论:TNF-α和IFN-γ损伤T84细胞株的紧密连接屏障功能,两者具有协同作用.     

甲状旁腺素、维生素D_3对Caco-2细胞钙结合蛋白D9k mRNA表达的影响

目的探讨甲状旁腺素(PTH)对小肠细胞钙结合蛋白(CaBP)D9k mRNA表达的影响以及探讨PTH是否影响1,25-(OH)2-D3对Caco-2细胞钙结合蛋白D9k mRNA表达的促进作用。方法以人结肠癌上皮细胞株Caco-2细胞作为小肠细胞体外模型,PTH分三个剂量10-8、10-9和10-12mol/L分别干预Caco-2细胞5、10、20、40和80min;10-8mol/L1,25-(OH)2-D3干预Caco-2细胞2、4、8、16和24h,溶剂对照为0.1%乙醇;10-8mol/L1,25-(OH)2-D3联合以上三剂量PTH干预Caco-2细胞2、4、8、16和24h,溶剂对照亦为0.1%乙醇。运用RT-PCR方法进行CaBP-D9k mRNA测定,以GAPDH作为内对照。结果10-8mol/LPTH作用20min,10-12mol/L作用10min,CaBP-D9k mRNA表达量高于空白组,其余时间点低于空白组;10-8mol/L1,25-(OH)2-D3干预Caco-2细胞2、4、8、16和24h,CaBP-D9k mRNA的表达均高于溶剂对照(0.1%乙醇);与10-8mol/L1,25-(OH)2-D3单独作用比较,10-8mol/L1,25-(OH)2-D3联合以上三剂量PTH作用,CaBP-D9k mRNA相对表达量均低于单独作用的表达量。结论10-8mol/L、10-12mol/LPTH可能促进Caco-2细胞CaBP-D9k mRNA瞬时(1~20min)合成增加;10-8mol/L1,25-(OH)2-D3可明显诱导Caco-2细胞CaBP-D9k mRNA的表达;PTH可能抵消或者抑制1,25-(OH)2-D3促进Caco-2细胞CaBP-D9k mRNA表达的作用。     

青蒿琥酯对高温复合内毒素致细胞凋亡保护作用

目的 探讨高温复合内毒素(LPS)作用下,青蒿琥酯(AR)对RAW264.7细胞热休克蛋白70(HSP70)合成及细胞凋亡的影响.方法采用体外培养的巨噬细胞系RAW264.7细胞.分为空白对照组、高温复合内毒素组、AR组(又分为4个亚组),于40℃、95%O2、5%CO2条件下共同孵育24h.用RT-PCR技术检测细胞内HSP70mRNA及其蛋白的表达量,用流式细胞术检测细胞凋亡率.结果 空白组和高温复合内毒素组HSP70m RNA的表达率分别为10%,23%,而AR组可显著提高HSP70mRNA的表达率,为54%,并与高温内毒素组差异有统计学意义(P＜0.05).HSP70蛋白的表达情况与HSPT0mRNA一致,而细胞凋亡率的变化则呈相反趋势.结果 AR进一步诱导高温复合内毒素作用下RAW264.7细胞HSP70m RNA和HSP70蛋白的表达,且可明显降低细胞的凋亡率,可能是其抗损伤保护作用的机制之一.     

银杏提取物对大鼠肝缺血再灌注损伤CO和ET含量及凋亡相关蛋白Bcl-2的影响

目的 观察银杏提取物(EGB)对大鼠肝缺血再灌注损伤(I/R)时CO、ET含量变化及对细胞凋亡相关蛋白Bcl-2的影响.方法 建立大鼠肝缺血再灌流模型,24只大鼠随机分成三组:假手术组(Sham组)、I/R组和I/R+EGB组.观察大鼠肝组织形态学改变,测定血浆中一氧化碳(CO)、内皮素(ET)含量变化、并测定凋亡相关蛋白Bcl-2的变化.结果 HE染色显示EGB组肝损伤明显减轻;EGB组肝血浆中CO含量升高、ET含量降低,与Sham组及I/R组相比有显著差异.EGB组凋亡相关蛋白Bcl-2明显高于I/R组.结论 EGB对大鼠肝I/R损伤具有保护作用,其机制可能与增强内源性CO表达、降低ET含量,及增加细胞相关蛋白Bcl-2表达有关.     

Cyperus esculentus L. Tubers (Tiger Nuts) Protect Epithelial Barrier Function in Caco-2 Cells Infected by Salmonella Enteritidis and Promote Lactobacillus plantarum Growth

Cyperus esculentus L. tubers (tiger nuts) contain different compounds with several intestinal health-promoting properties. Here, we studied the capacity of tiger nuts from Valencia, Spain, to prevent epithelial barrier function disruption induced by Salmonella enteritidis in Caco-2 cell cultures. Paracellular permeability was assessed by transepithelial electrical resistance (TER) and tight junction protein immunolocalization. Moreover, the effect of tiger nuts on S. enteritidis agglutination, oxidative stress, and Lactobacillus plantarum growth was tested. Compared to controls, tiger nuts partially restored TER in S. enteritidis-infected cultures, an effect confirmed by immunolocalization of tight junction proteins ZO-1 and occludin. The results also revealed that this protective effect may be associated with the capacity to agglutinate the pathogen, restore TER in TNFα-stimulated cultures, and reduce reactive oxygen species in H₂O₂-stimulated cultures. Moreover, they favor L. plantarum growth. In conclusion, this study demonstrates that the tiger nut protects epithelial barrier function by reducing bacterial invasion, along with counteracting TNFα and H₂O₂ effects, thus giving an additional value to this tuber as a potential functional food.     

Effect of Colonic Bacterial Metabolites on Caco-2 Cell Paracellular Permeability In Vitro

One common effect of tumor promoters is increased tight junction (TJ) permeability. TJs are responsible for paracellular permeability and integrity of the barrier function. Occludin is one of the main proteins responsible for TJ structure. This study tested the effects of physiological levels of phenol, ammonia, primary bile acids (cholic acid, CA, and chenodeoxycholic acid, CDCA), and secondary bile acids (lithocholic acid, LCA, and deoxycholic acid, DCA) on paracellular permeability using a Caco-2 cell model. Paracellular permeability of Caco-2 monolayers was assessed by transepithelial electrical resistance (TER) and the apical to basolateral flux of [ ¹⁴ C]-mannitol. Secondary, but not primary, bile acids increased permeability as reflected by significantly decreased TER and increased mannitol flux. Both phenol and ammonia also increased permeability. The primary bile acid CA significantly increased occludin expression (P < 0.05), whereas CDCA had no significant effect on occludin expression as compared to the negative control. The secondary bile acids DCA and LCA significantly increased occludin expression (P < 0.05), whereas phenol had no significant effect on the protein expression as compared to the negative control. This suggests that the increased permeability observed with LCA, DCA, phenol, and ammonia was not related to an effect on occludin expression. In conclusion, phenol, ammonia, and secondary bile acids were shown to increase paracellular permeability and reduce epithelial barrier function at doses typical of levels found in fecal samples. The results contribute to the evidence these gut microflora-generated products have tumor-promoting activity.     

Effect of colonic bacterial metabolites on Caco-2 cell paracellular permeability in vitro

One common effect of tumor promoters is increased tight junction (TJ) permeability. TJs are responsible for paracellular permeability and integrity of the barrier function. Occludin is one of the main proteins responsible for TJ structure. This study tested the effects of physiological levels of phenol, ammonia, primary bile acids (cholic acid, CA, and chenodeoxycholic acid, CDCA), and secondary bile acids (lithocholic acid, LCA, and deoxycholic acid, DCA) on paracellular permeability using a Caco-2 cell model. Paracellular permeability of Caco-2 monolayers was assessed by transepithelial electrical resistance (TER) and the apical to basolateral flux of [14C]-mannitol. Secondary, but not primary, bile acids increased permeability as reflected by significantly decreased TER and increased mannitol flux. Both phenol and ammonia also increased permeability. The primary bile acid CA significantly increased occludin expression (P < 0.05), whereas CDCA had no significant effect on occludin expression as compared to the negative control. The secondary bile acids DCA and LCA significantly increased occludin expression (P < 0.05), whereas phenol had no significant effect on the protein expression as compared to the negative control. This suggests that the increased permeability observed with LCA, DCA, phenol, and ammonia was not related to an effect on occludin expression. In conclusion, phenol, ammonia, and secondary bile acids were shown to increase paracellular permeability and reduce epithelial barrier function at doses typical of levels found in fecal samples. The results contribute to the evidence these gut microflora-generated products have tumor-promoting activity.     

Effect of eicosapentaenoic acid (EPA) on tight junction permeability in intestinal monolayer cells

The purpose of this study is to evaluate the effect of C18 and C20 long chain fatty acids on tight junction permeability in a model of intestinal epithelium. METHODS: Confluent Caco-2 cells on porous filters with double chamber system were used to measure fluorescein sulfonic acid (FS) permeability and transepithelial electrical resistance (TEER). Lactate dehydrogenase release and ultrastructure were evaluated. Effect of 200 microM eicosapentaenoic acid (EPA, C20:5 n-3), arachidonic acid (AA, C20: 4 n-6), alpha-linoleic acid (ALA, C18: 3 n-3), linoleic acid (LA, C18: 2 n-6), or oleic acid (OA, C18: 1 n-9) enrichment in the culture medium during 24 hours were compared. The effect of the cyclooxygenase inhibitor, indomethacin, lipoxygenase inhibitors, NDGA or AA861, and antioxidant, BHT, was evaluated as a mechanism to change tight junction permeability. RESULTS: Caco-2 cells formed polarized columnar epithelial cells with densely packed microvilli and well developed junctional complexes. Addition of EPA enhanced FS permeability to 3.0+/-1.6-fold and lowered TEER to 0.59+/-1.2-fold vs. control with concentration dependency without cell injury (P<0.01-0.05). OA, AA or LA did not change, but ALA enhanced tight junction permeability. Indomethacin and AA861 normalized the changes mediated by EPA. CONCLUSIONS: EPA affects tight junction permeability in intestinal monolayer cells specifically and concentration dependently via cyclooxygenase and lipoxygenase products.     

Effects of Supernatants from Escherichia coli Nissle 1917 and Faecalibacterium prausnitzii on Intestinal Epithelial Cells and a Rat Model of 5-Fluorouracil-Induced Mucositis

Faecalibacterium prausnitzii (Fp) and Escherichia coli Nissle 1917 (EcN) are probiotics, which have been reported to ameliorate certain gastrointestinal disorders. We evaluated the effects of supernatants (SN) derived from Fp and EcN on 5-fluorouracil (5-FU)-treated intestinal cells and in a rat model of mucositis. In vitro: IEC-6, Caco-2, and T-84 cells were analyzed for viability and monolayer permeability. In vivo: Female dark agouti rats were gavaged with Fp or EcN SN and injected intraperitoneally with saline (control) or 5-FU to induce mucositis. Rats were euthanized and intestinal tissues collected for myeloperoxidase assay and histological analyses. In vitro: Caco-2 cell viability was further reduced when treated with Fp SN + 5-FU compared to 5-FU controls. In both Caco-2 and T-84 cells, Fp SN partially prevented the decrease in transepithelial electrical resistance (TER) caused by 5-FU administration. In vivo: 5-FU-injected rats administered Fp SN or EcN SN partly prevented body weight loss and normalized water intake compared to 5-FU controls. These results suggest a growth inhibitory mechanism of Fp SN action on transformed epithelial cells that could be mediated by effects on tight junctions. Factors derived from Fp SN and EcN SN could have a role in reducing the severity of intestinal mucositis.     

Butyrate protects Caco-2 cells from Campylobacter jejuni invasion and translocation

Invasion in and translocation across enterocytes are major events during Campylobacter jejuni-induced enteritis in humans. C. jejuni in vitro infection of cell monolayers typically results in loss of tight junction integrity, which could contribute to translocation. In the present study, we wanted to investigate whether butyrate is able to confer protection to Caco-2 cells against C. jejuni invasion, thus reducing paracellular permeability and limiting C. jejuni translocation. Protection of Caco-2 cells against C. jejuni invasion was assessed using a gentamicin protection assay. Transwell systems were used to investigate the impact of butyrate on translocation of C. jejuni across a Caco-2 monolayer and its effect on transepithelial resistance during infection. Butyrate protected Caco-2 cells against C. jejuni invasion in a concentration-dependent manner. Differentiated Caco-2 cells were less susceptible to C. jejuni invasion than 3-d-old undifferentiated cells and higher concentrations of butyrate and longer incubation times were needed to become refractive for invasion. C. jejuni translocation over Caco-2 monolayers was reduced when monolayers were treated with butyrate and this was accompanied by an enhanced drop in transepithelial resistance. The present study showed that butyrate is able to protect Caco-2 cells from two major virulence mechanisms of C. jejuni, namely invasion and translocation, but not from a decline in transepithelial resistance.     

Kaempferol enhances intestinal barrier function through the cytoskeletal association and expression of tight junction proteins in Caco-2 cells

Kaempferol, a natural flavonoid present in fruits, vegetables, and teas, provides beneficial effects for human health. We investigated the promotive effect of kaempferol on tight junction (TJ) barrier integrity in human intestinal Caco-2 cell monolayers. Transepithelial electrical resistance (TER; a TJ integrity marker) across the monolayers rapidly and markedly increased during the first 6 h after kaempferol administration and remained elevated until 48 h without any changes in the lucifer yellow or dextran fluxes. Immunoblot analysis demonstrated that kaempferol promoted the actin cytoskeletal association of the TJ proteins, zonula occludens (ZO)-1, ZO-2, occludin, claudin-1, claudin-3, and claudin-4, which was associated with the increase in TER. Kaempferol-mediated ZO-2 and claudin-4 expression was relatively smaller or occurred later than the kaempferol-promoted cytoskeletal association. Confocal microscopy showed that kaempferol-induced assembly of occludin and claudin-3 occurred at the TJ at 6 h postadministration. Extraction of cholesterol with methyl-β-cyclodextrin suppressed the kaempferol-mediated increase in TER. Sucrose density gradient centrifugation showed that the kaempferol treatment increased the TJ protein distributions in the cholesterol-rich lipid microdomain fraction. Taken together, these results indicate that the membrane lipid microdomain is involved in the kaempferol-mediated promotion of TJ protein assembly and intestinal TJ integrity.     

Effect of gamma-linolenic acid or docosahexaenoic acid on tight junction permeability in intestinal monolayer cells and their mechanism by protein kinase C activation and/or eicosanoid formation

OBJECTIVE: Polyunsaturated fatty acids have been characterized as immunonutrients, but the effect of gamma-linolenic acid (GLA) or docosahexaenoic acid (DHA) on intestinal permeability has rarely been reported. METHODS: Confluent Caco-2 cells on porous filter were used to measure tight junction function by fluorescein sulfonic acid permeability and transepithelial electrical resistance. Treatments with 0, 10, 50, and 100 microM of GLA or DHA during 24 h were compared. Then the effects of butylated hydroxytoluene (antioxidant), 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (protein kinase C antagonist), and inhibitors of enzymatic degradation to the eicosanoids, indomethacin (cyclooxygenase inhibitor) and 2-(12-hydroxydodeca-5,10-diynyl)-3,5,6-trimethyl-p-benzoquinone (lipoxygenase inhibitor), on GLA or DHA were examined. RESULTS: GLA and DHA enhanced fluorescein sulfonic acid permeability to 8.7- and 1.4-fold, respectively, and lowered transepithelial electrical resistance to 0.52- and 0.73-fold, respectively, versus the control in a concentration-dependent manner without cell injury (P < 0.001 to 0.05). Indomethacin and 2-(12-hydroxydodeca-5,10-diynyl)-3,5,6-trimethyl-p-benzoquinone enhanced the changes mediated by GLA but did not alter the DHA effect. Butylated hydroxytoluene was ineffective. 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine facilitated the changes mediated by GLA, DHA, and eicosapentaenoic acid. The results indicated that the mechanism to change tight junction permeability via protein kinase C regulation is common but that via eicosanoid formation differs among GLA, DHA, and eicosapentaenoic acid. CONCLUSIONS: GLA and DHA affect tight junction permeability in intestinal monolayer cells specifically and in a concentration-dependent manner.     

紧密连接的结构与功能

紧密连接（tight junction,TJ）是上皮细胞之间的一种重要的连接复合体,由跨膜蛋白家族（occludin和claudin）、膜周蛋白家族（ZO蛋白）等构成,起着细胞旁通透屏障和维持细胞极性的作用。多种生理、病理因素参与改变紧密连接结构和功能,如血氧浓度（缺氧）、激素、炎症因子以及雌激素等。紧密连接结构和紧密连接相关因子在正常子宫内膜和胎盘组织中均有表达,与妊娠病理生理的发生发展密切相关。