Responses of growth performance,antioxidant function,small intestinal morphology and mRNA expression of jejunal tight junction protein to dietary iron in yellow-feathered broilers

This study aimed to investigate the dose-effect of iron on growth performance,antioxidant function.intestinal morphology,and mRNA expression of jejunal tight junction protein in 1-to21-d-old yellow-feathered broilers.A total of 7201-d-old yellow-feathered maleb roilers were allocated to 9 treatments with 8 replicate cages of 10 birds per cage.The dietary treatments were consisted of a basal diet(contained 79.6 mg Fe kg^(-1))supplemented with 0,20,40,60,80,160,320,640,and 1,280 mg Fe kg^(-1)in the form of FeSO_(4)·7H_(2)O.Compared with the birds in the control group,birds supplemented with 20mg Fe kg^(-1)had higher average daily gain(ADG)(P<0.0001).Adding 640 and 1,280 mg Fe kg^(-1)significantly decreased ADG(P<0.0001)and average daily feed intake(ADFI)(P<0.0001)compared with supplementation of 20mg Fe kg^(-1).Malondialdehyde(MDA)concentration in plasma and duodenum increased linearly(P<0.0001),but MDA concentration in liver and jejunum increased linearly(P<0.05)or quadratically(P<0.05)with increased dietary Fe concentration.The villus height(VH)in duodenum and jejunum,and the ratio of villus height to crypt depth(V/C)in duodenum decreased linearly(P?0.05)as dietary Feincreased.As dietary Fe increased,the jejunal relative mRNA abundance of claudin-1 decreased linearly(P=0.001),but the jejunal relative mRNA abundance of zona occludens-1(ZO-1)and occludin decreased linearly(P?0.05)or quadratically(P?0.05).Compared with the supplementation of 20 mg Fe kg^(-1),the supplementation of640 mg Fe kg^(-1)or higher increased(P?0.05)MDA concentrations in plasma,duodenum,and jejunum,decreased VH in the duodenum and jejunum,and the addition of 1,280 mg Fe kg^(-1)reduced(P?0.05)the jejunal tight junction protein(claudin-1,ZO-1,occludin)mRNA abundance.In summary,640 mg of supplemental Fe kg^(-1)or greater was associated with decreased growth performance,increased oxidative stress,disrupted intestinal morphology,and reduced mRNA expression of jejunal tight junction protein.

Research status and hotspots of tight junctions and colorectal cancer:A bibliometric and visualization analysis

BACKGROUND Colorectal cancer(CRC)is the third most common cancer worldwide and the second leading cause of cancer-related death.Over the past two decades,numerous researchers have provided important evidence regarding the role of tight junction(TJ)proteins in the occurrence and progression of CRC.The causal relationship between the presence of specific TJ proteins and the development of CRC has also been confirmed.Despite the large number of publications in this field,a bibliometric study to review the current state of research and highlight the research trends and hotspots in this field has not yet been performed.AIM To analyze research on TJs and CRC,summarize the field’s history and current status,and predict future research directions.METHODS We searched the Science Citation Index Expanded database for all literature on CRC and TJs from 2001-2023.We used bibliometrics to analyze the data of these papers,such as the authors,countries,institutions,and references.Co-authorship,co-citation,and co-occurrence analyses were the main methods of analysis.CiteSpace and VOSviewer were used to visualize the results.RESULTS A total of 205 studies were ultimately identified.The number of publications on this topic has steadily increased since 2007.China and the United States have made the largest contributions to this field.Anticancer Research was the most prolific journal,publishing 8 articles,while the journal Oncogene had the highest average citation rate(68.33).Professor Dhawan P was the most prolific and cited author in this field.Co-occurrence analysis of keywords revealed that“tight junction protein expression”,“colorectal cancer”,“intestinal microbiota”,and“inflammatory bowel disease”had the highest frequency of occurrence,revealing the research hotspots and trends in this field.CONCLUSION This bibliometric analysis evaluated the scope and trends of TJ proteins in CRC,providing valuable research perspectives and future directions for studying the connection between the two.It is recommended to focus on emerging research hotspots,such as the correlations among intestinal microbiota,inflammatory bowel disease,TJ protein expression,and CRC.

Lactobacillus rhamnosus GR-1 attenuates foodborne Bacillus cereus-induced NLRP3 inflammasome activity in bovine mammary epithelial cells by protecting intercellular tight junctions

Background:Bacillus cereus is an important pathogen that causes human food poisoning,specifically diarrhea and vomiting.B.cereus can also induce mastitis in dairy cows and has a strong survival ability in milk,as it cannot be inactivated by high-temperature short-time pasteurization.Therefore,B.cereus can enter the market through pasteurized milk and other dairy products,imposing enormous hidden dangers on food safety and human health.Results:In this study,B.cereus 2101(BC)was isolated from milk samples of cows with mastitis.BC grew rapidly with strong hemolysis,making it difficult to prevent mastitis and ensure food security.MAC-T cells were treated with BC and/or Lactobacillus rhamnosus GR-1(LGR-1).Pretreatment with LGR-1 protected the integrity of tight junctions and the expression of zonula occludens-1(ZO-1)and occludin destroyed by BC.Furthermore,LGR-1 pretreatment reduced the expression of NOD-like receptor family member pyrin domain-containing protein 3(NLRP3),caspase recruitment and activation domain(ASC),Caspase-1 p20,gasdermin D(GSDMD)p30,inflammatory factors(interleukin(IL)-1βand IL-18),and cell death induced by BC.Moreover,LGR-1 pretreatment reduced NLRP3 inflammasome activity and increased expressions of ZO-1 and occludin induced by lipopolysaccharides(LPS)+ATP stimulation.MAC-T cells were transfected with NLRP3 si RNA or MCC950 and/or treated with BC and/or LGR-1.NLRP3-si RNA transfection and MCC950 attenuated BC-induced NLRP3 inflammasome activity.Expression of inflammatory cytokines and cell death suggested that the inflammatory pathway might play an important role in the induction of the NLRP3 inflammasome by BC and the protection of LGR-1.Conclusions:These results suggest that LGR-1 might be a probiotic alternative to antibiotics and could be administered to prevent mastitis in dairy cows,thus ensuring food security.

Tight junction disruption and the pathogenesis of the chronic complications of diabetes mellitus:A narrative review

The chronic complications of diabetes mellitus constitute a major public health problem.For example,diabetic eye diseases are the most important cause of blindness,and diabetic nephropathy is the most frequent cause of chronic kidney disease worldwide.The cellular and molecular mechanisms of these chronic complications are still poorly understood,preventing the development of effective treatment strategies.Tight junctions(TJs)are epithelial intercellular junctions located at the most apical region of cell-cell contacts,and their main function is to restrict the passage of molecules through the paracellular space.The TJs consist of over 40 proteins,and the most important are occludin,claudins and the zonula occludens.Accumulating evidence suggests that TJ disruption in different organs,such as the brain,nerves,retina and kidneys,plays a fundamental pathophysiological role in the development of chronic complications.Increased permeability of the blood-brain barrier and the blood-retinal barrier has been demonstrated in diabetic neuropathy,brain injury and diabetic retinopathy.The consequences of TJ disruption on kidney function or progression of kidney disease are currently unknown.In the present review,we highlighted the molecular events that lead to barrier dysfunction in diabetes.Further investigation of the mechanisms underlying TJ disruption is expected to provide new insights into therapeutic approaches to ameliorate the chronic complications of diabetes mellitus.

Advanced glycation end-products change placental barrier function and tight junction in rats with gestational diabetes mellitus via the receptor for advanced glycation end products/nuclear factor-κB pathway

The placenta plays an important role in nutrient transport to maintain the growth and development of the embryo.Gestational diabetes mellitus(GDM),the most common complication during pregnancy,highly affects placental function in late gestation.Advanced glycation end-products(AGEs),a complex and heterogeneous group of compounds engaged by the receptor for AGEs(RAGE),are closely associated with diabetes-related complications.In this study,AGEs induced a decrease in the expression of tight junction(TJ)proteins in BeWo cells and increased the paracellular permeability of trophoblast cells by regulating RAGE/NF-κB.Sprague-Dawley(SD)rats injected with 100 mg/kg AGEs-rat serum albumin(RSA)via the tail vein from embryo day 2 were set as the placental barrier dysfunction model group(n=10).The effect of AGEs on placental permeability was determined using the Evans-Blue dye extravasation method.The ultrastructure of the placenta samples was observed by transmission electron microscopy.The effects of AGEs on the placenta were confirmed by treating rats with RAGE antagonist FPS-ZM1 and soluble forms of RAGE(sRAGE).AGEs treatment increased placental permeability and disrupted the tight junctions in pregnant rat placenta,but has no effect on blood glucose.The expression of TJ-related proteins,including ZO-1,Occludin,and Claudin 5,were downregulated after AGEs treatment.Further,AGEs treatment increased the expression of RAGE and nuclear factor-κB in the placenta of rats and upregulated the levels of vascular endothelial growth factor.The effects of AGEs on the placenta were blocked by RAGE antagonist FPS-ZM1 and sRAGE.This study demonstrates the mechanism underlying AGEs-induced disturbance in placental function in pregnant rats and highlights the potential of AGEs in the treatment of GDM.

In vitro and in vivo evaluation of effects of Mg-6Zn alloy on tight junction of intestinal epithelial cell

The effects of biodegradable Mg?6Zn alloy on tight junction of intestinal epithelial cells (IEC-6) were investigated. In the in vitro experiments, the cells were exposed to Mg?6Zn alloy extracts with different concentrations (0, 20% and 40%) for 1, 3 and 5 d. The real-time polymerase chain reaction (PCR) results show that when the cells are treated with 40% and 20% extracts, the expression of Zona Occludens 1 (ZO-1) and Occludin increase as compared with those in the control group. In the in vivo experiments, Mg?6Zn alloy and titanium staples were implanted into rabbits’ intestinal tract for 1, 2 and 3 weeks. By immunohistochemical staining of peri-implant intestinal tissue, increased expression of Occludin and ZO-1 are observed in the Mg?6Zn alloy groups as compared with those in the titanium and control groups. The results show that Mg?6Zn alloy in intestine may promote the regeneration of tight junction, and the extract with a certain concentration can induce the expression of tight junction related genes in IEC-6 cells.

Tight junctions in inflammatory bowel diseases and inflammatory bowel disease associated colorectal cancer

Inflammatory bowel diseases are characterised by inflammation that compromises the integrity of the epithelial barrier. The intestinal epithelium is not only a static barrier but has evolved complex mechanisms to control and regulate bacterial interactions with the mucosal surface. Apical tight junction proteins are critical in the maintenance of epithelial barrier function and control of paracellular permeability. The characterisation of alterations in tight junction proteins as key players in epithelial barrier function in inflammatory bowel diseases is rapidly enhancing our understanding of critical mechanisms in disease pathogenesis as well as novel therapeutic opportunities. Here we give an overview of recent literature focusing on the role of tight junction proteins, in particular claudins, in inflammatory bowel diseases and inflammatory bowel disease associated colorectal cancer.

Moxibustion down-regulates colonic epithelial cell apoptosis and repairs tight junctions in rats with Crohn's disease

AIM: To investigate the effects of moxibustion on down-regulation of the colonic epithelial cell apoptosis and repair of the tight junctions in rats with Crohn's disease (CD). METHODS: Sixty male Sprague-Dawley rats were randomly divided into a normal control (NC) group, a model control (MC) group, an herbs-partitioned moxibustion (HPM) group, a mild-warm moxibustion (MWM) group and a salicylazosulphapyridine (SASP) group, with 12 rats in each group. The CD model rats were treated with trinitrobenzene sulphonic acid to induce intestinal inflammation. The rats in the HPM and MWM groups were treated at the Tianshu (ST25) and Qihai (CV6) acupoints once daily for 14 d, and the SASP group was fed SASP twice daily for 14 d. No additional treatment was given to the MC and NC groups. Themicrostructure of the colonic epithelium was observed under a transmission electron microscope, the transepithelial resistance was measured using a shortcircuit current, colonic epithelial cell apoptosis was determined by terminal deoxynucleotidyl transferasemediated dUTP-biotin nick end labelling assay, and the expression of occludin, claudin-1 and zonula occludens-l (ZO-1) in the colonic epithelial junction was determined by Western blotting and immunofluorescence staining. RESULTS: Compared with the MC group, the microstructure of the colonic epithelial barrier was signifi-cantly improved in rats treated with HPM, MWM or SASP, meanwhile, the current flow was reduced signifi-cantly, with values of 168.20 ± 6.14 vs 99.70 ± 3.13, 99.10 ± 4.28 and 120.30 ± 3.65 mA, respectively (P = 0.001). However, the HPM and MWM groups had higher current flow rates than the SASP group (99.70 ± 3.13, 99.10 ± 4.28 vs 120.30 ± 3.65 mA, P = 0.001). The number of the apoptotic colonic epithelial cells in HPM, MWM and SASP groups was largely reduced (61.5 ± 16.91 vs 15.5 ± 8.89, 14.8 ± 6.27 and 24.7 ± 9.68, respectively (P = 0.001); and the expression of occlu- din, claudin-1 and ZO-1 in the MWM and HPM groups was signifi cantly enhanced (0.48 ± 0.10, 0.64 ± 0.09 vs 0.18 ± 0.05 for occludin, 0.12 ± 0.02, 0.17 ± 0.03 vs 0.05 ± 0.01 for claudin-1, and 0.08 ± 0.01, 0.11 ± 0.01 vs 0.02 ± 0.01 for ZO-1). And in SASP group, the expression of occludin and ZO-1 was also signifi cantly increased (0.27 ± 0.04 vs 0.18 ± 0.05 for occludin and 0.05 ± 0.01 vs 0.02 ± 0.01 for ZO-1), but there was no significant difference for claudin-1. The HPM and MWM groups had higher expression of occludin, claudin-1 and ZO-1 than the SASP group. CONCLUSION: HPM and MWM treatment can down-regulate apoptosis of colonic epithelial cells, repair tight junctions and enhance colonic epithelial barrier function in rats with CD.

Dietary protocatechuic acid ameliorates inflammation and up-regulates intestinal tight junction proteins by modulating gut microbiota in LPS-challenged piglets

Background: Weaning is one of the major factors that cause stress and intestinal disease in piglets. Protocatechuic acid(PCA) is an active plant phenolic acid which exists in Chinese herb, Duzhong(Eucommia ulmoides Oliver), and is also considered as the main bioactive metabolite of polyphenol against oxidative stress and inflammation. This study aimed to investigate the effect of PCA on growth performance, intestinal barrier function, and gut microbiota in a weaned piglet model challenged with lipopolysaccharide(LPS).Methods: Thirty-six piglets(Pig Improvement Company line 337 × C48, 28 d of age, 8.87 kg ± 0.11 kg BW) were randomly allocated into 3 treatments and fed with a basal diet(CTL), a diet added 50 mg/kg of aureomycin(AUR), or a diet supplemented with 4000 mg/kg of PCA, respectively. The piglets were challenged with LPS(10 μg/kg BW) on d 14 and d 21 by intraperitoneal injection during the 21-d experiment. Animals(n = 6 from each group) were sacrificed after being anesthetized by sodium pentobarbital at 2 h after the last injection of LPS. The serum was collected for antioxidant indices and inflammatory cytokines analysis, the ileum was harvested for detecting mRNA and protein levels of tight junction proteins by PCR and immunohistochemical staining, and the cecum chyme was collected for intestinal flora analysis using 16 S rRNA gene sequencing.Results: Dietary supplementation of PCA or AUR significantly increased the expression of tight junction proteins including ZO-1 and claudin-1 in intestinal mucosa, and decreased the serum levels of thiobarbituric acid reactive substances(TBARS) and IL-6, as compared with CTL group. In addition, PCA also decreased the serum levels of IL-2 and TNF-α(P < 0.05). Analysis of gut microbiota indicated that PCA increased the Firmicutes/Bacteroidetes ratio(P < 0.05). Spearman's correlation analysis at the genus level revealed that PCA reduced the relative abundance of Prevotella 9, Prevotella 2, Holdemanella, and Ruminococcus torques group(P < 0.05), and increased the relative abundance of Roseburia and Desulfovibrio(P < 0.05), whereas AUR had no significant effect on these bacteria.Conclusions: These results demonstrated that both PCA and AUR had protective effect on oxidative stress, inflammation and intestinal barrier function in piglets challenged with LPS, and PCA potentially exerted the protective function by modulating intestinal flora in a way different from AUR.

Acanthopanax senticosus polysaccharides-induced intestinal tight junction injury alleviation via inhibition of NF-κB/MLCK pathway in a mouse endotoxemia model

AIM To examine the effects of Acanthopanax senticosus polysaccharides(ASPS) on intestinal tight junction(TJ) disruption and nuclear factor-kappa B(NF-κB)/myosin light chain kinase(MLCK) activation in endotoxemia.METHODS BALB/C mice(6-8-weeks-old) received continuous intragastric gavage of ASPS for 7 d before injection of lipopolysaccharide(LPS), or received ASPS once after LPS injection. Blood and intestinal mucosal samples were collected 6 h after LPS challenge. Clinical symptoms, histological injury, intestinal permeability,TJ ultrastructure, and TJ protein expression were determined.RESULTS Compared with mice in the LPS group, pretreatment with ASPS improved clinical and histological scores by 390.9%(P < 0.05) and 57.89%(P < 0.05), respectively, and gut permeability change in endotoxemic mice was shown by a 61.93% reduction in reduced leakage of fluorescein isothiocyanate-dextran 6 h after LPS injection(P < 0.05). ASPS pretreatment also prevented LPS-induced TJ ultrastructure breakdown supported by increased electron dense materials between adjoining cells, sustained redistribution and expression of occludin(0.597 ± 0.027 vs 0.103 ± 0.009, P < 0.05) and zonula occludens-1(0.507 ± 0.032 vs 0.125 ± 0.019, P < 0.05), and suppressed activation of the NF-κB/MLCK pathway indicated by reduced expression of NF-κB, phospho-inhibitor kappa B-alpha, MLCK and phospho-myosin light-chain-2 by 16.06%(P < 0.05), 54.31%(P < 0.05), 66.10%(P < 0.05) and 64.82%(P < 0.05), respectively. CONCLUSION ASPS pretreatment may be associated with inhibition of the NF-κB/MLCK pathway and concomitant amelioration of LPS-induced TJ dysfunction of intestinal epithelium in endotoxemia.

Effects of cardiopulmonary bypass on tight junction protein expressions in intestinal mucosa of rats

AIM： To investigate the tight junction protein expressions of intestinal mucosa in an experimental model of cardiopulmonary bypass （CPB） in rats. METHODS： Thirty anesthetized rats were randomly divided into two groups： Group S （n = 10） served as sham operation and group C （n = 20） served as CPB which underwent CPB for 1 h. Expression of occludin and zonula occludens-1 （ZO-1） were determined by Western blotting and immunotochemistry, respectively. Plasma levels of diamine oxidase （DAO） and d-lactate were determined using an enzymatic spectrophotometry. RESULTS： Immunohistochemical localization of occludin and ZO-1 showed disruption of the tight junctions in enterocytes lining villi at the end of CPB and 2 h after CPB. The intensities of the occludin and ZO-i at the end of CPB were lower than those of control group （76.4% ± 22.5% vs 96.5% ± 28.5% and 62.4% ± 10.1% vs 85.5% ±25.6%, P 〈 0.05） and were further lower at 2 h after CPB （50.5% ± 10.5% and 45.3% ± 9.5%, P 〈 0.05）. Plasma d-lactate and DAO levels increased significantly （8.688 ± 0.704 vs 5.745 ± 0.364 and 0.898 ± 0.062 vs 0.562 ± 0.035, P 〈 0.05） at the end of CPB compared with control group and were significantly higher at 2 h after CPB than those at the end of CPB （9.377 ± 0.769 and 1.038 ± 0.252, P 〈 0.05）. There were significant negative correlations between occludin or ZO-1 expression and DAO （r^2 = 0.5629,r^2 = 0.5424, P 〈 0.05） or d-lactate levels （r^2 = 0.6512,r^2 = 0.7073, P 〈 0.05） both at the end of CPB and 2 h after CPB. CONCLUSION： CPB markedly down-regulates the expression of occludin and ZO-1 proteins in intestinal mucosa of rats. The close correlation between expression of tight junctions （TJs） and plasma levels of DAO or d-lactate supports the hypothesis that intestinal permeability increases during and after CPB because of decreases in the expressions of TJs.

Hepatic tight junctions:From viral entry to cancer metastasis

The tight junction (TJ) is a critical cellular component for maintenance of tissue integrity, cellular interactions and cell-cell communications, and physiologically functions as the "great wall" against external agents and the surrounding hostile environment. During the host-pathogen evolution, viruses somehow found the key to unlock the gate for their entry into cells and to exploit and exhaust the host cells. In the liver, an array of TJ molecules is localized along the bile canaliculi forming the blood-biliary barrier, where they play pivotal roles in paracellular permeability, bile secretion, and cell polarity. In pathology, certain hepatic TJ molecules mediate virus entry causing hepatitis infection; deregulation and functional abnormality of the TJ have also been implicated in triggering liver cancer development and metastasis. All these findings shed new insights on the understanding of hepatic TJs in the development of liver disease and provide new clues for potential intervention.

New tight junction protein 2 variant causing progressive familial intrahepatic cholestasis type 4 in adults: A case report

BACKGROUND Progressive familial intrahepatic cholestasis(PFIC)encompasses a group of autosomal recessive disorders with high morbidity and mortality.Variants in the gene encoding tight junction protein-2(TJP2)have been linked to PFIC type 4(PFIC4),which predominantly presents in childhood.However,there are only limited data from adults with TJP2-related PFIC4.We report a family with an autosomal recessive disorder with a novel variant in the TJP2 gene in adults with very variable expression of PFIC4.CASE SUMMARY The index patient presented at 19 years old with liver cirrhosis and variceal bleeding and was treated with endoscopic banding and beta-blockers.In 2018,he developed primary liver cancer that was treated with radiofrequency ablation followed by liver transplantation in 2019.Genetic testing revealed a novel homozygous TJP2 variant causing PFIC4(TJP2([NM_004817.3]:c.[3334C>T];[3334C>T])).The consanguineous family consists of the father and mother(both heterozygous)and their 12 children,of which five carry the variant in a homozygous state;however,these five siblings have highly variable expression of PFIC4.Two homozygous brothers had cirrhosis and portal hypertension at diagnosis at the ages of 19 and 36.Two other homozygous brothers,age 23 and 19,and the homozygous sister,age 21,have elevated liver enzymes but presently no cirrhosis,which may suggest an age-dependent penetrance.In addition,five sisters had severe and mild intrahepatic cholestasis of pregnancy and carry the TJP2 variant in a homozygous and heterozygous state,respectively.CONCLUSION This novel TJP2 variant is associated with PFIC4 causing severe liver disease with cirrhosis and primary liver cancer in adolescents/adults.

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%,χ 2 = 5.25,P < 0.05),high-dose salvianolate group(21.43% vs 76.92%,χ 2 = 8.315,P < 0.01) and normal group(21.43% vs 90%,χ 2 = 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.

Targeting tight junctions during epithelial to mesenchymal transition in human pancreatic cancer

Pancreatic cancer continues to be a leading cause of cancer-related death worldwide and there is an urgent need to develop novel diagnostic and therapeutic strategies to reduce the mortality of patients with this disease. In pancreatic cancer, some tight junction proteins, including claudins, are abnormally regulated and therefore are promising molecular targets for diagnosis, prognosis and therapy. Claudin-4 and-18 are overexpressed in human pancreatic cancer and its precursor lesions. Claudin-4 is a high affinity receptor of Clostridium perfringens enterotoxin(CPE). The cytotoxic effects of CPE and monoclonal antibodies against claudin-4 are useful as novel therapeutic tools for pancreatic cancer. Claudin-18 could be a putative marker and therapeutic target with prognostic implications for patients with pancreatic cancer. Claudin-1,-7, tricellulin and marvelD3 are involved in epithelial to mesenchymal transition(EMT) of pancreatic cancer cells and thus might be useful as biomarkers during disease. Protein kinase C is closely related to EMT of pancreatic cancer and regulates tight junctions of normal human pancreatic duct epithelial cells and the cancer cells. This review focuses on the regulation of tight junctions via protein kinase C during EMT in human pancreatic cancer for the purpose of developing new diagnostic and therapeutic modalities for pancreatic cancer.

Effects of Simulated Weightlessness on Tight Junction Protein Occludin and Zonula Occluden-1 Expression Levels in the Intestinal Mucosa of Rats

This study investigated the tight junction（TJ） protein expression of the intestinal mucosa in a rat tail-suspension model under simulated weightlessness.Twenty-four Wistar rats were randomly divided into three groups：CON group（n=8）,control; SUS-14 d group（n=8）,tail-suspension for 14 days; SUS-21 d group（n=8）,tail-suspension for 21 days.Occludin and Zonula Occluden-1（ZO-1） expression levels were determined by immunohistochemical analysis and mRNA fluorescent quantitative PCR.Plasma levels of diamine oxidase（DAO） and d-lactate were determined using enzymatic spectrophotometry.Immunohistochemical results for occludin and ZO-1 showed disruption of the TJs in the intestinal mucosa in SUS-14 d and SUS-21 d groups.The expression levels of occludin and ZO-1 in SUS-21 d group were lower than those in SUS-14 d group,and significantly lower than those in CON group（Occldin：0.86±0.02 vs 1.01±0.03 vs 1.63±0.03 and ZO-1：0.82±0.01 vs 1.00±0.02 vs 1.55±0.01,P〈0.01）.Moreover,the levels of plasma DAO and d-lactate in SUS-21 d group were higher than those in SUS-14 d group,and significantly higher than those in CON group（DAO：27.58±0.49 vs 20.74±0.49 vs 12.94±0.21 and d-lactate：37.86±0.74 vs 28.26±1.01 vs 17.76±0.91,P〈0.01）.There were significant negative correlations between occludin or ZO-1 expression levels and DAO（r2=0.9014,r2=0.9355,P〈0.01） or d-lactate levels（r2=0.8989,r2=0.9331,P〈0.01）.Occludin and Zo-1 were reduced in intestinal mucosa both in mRNA and protein levels in the rat tail-suspension model.The significant negative correlations between expression levels of TJs and plasma levels of DAO or d-lactate support the hypothesis that intestinal permeability is increased due to a decrease in TJ protein expression during tail-suspension from 14 days to 21 days.

Analysis of differential expression of tight junction proteins in cultured oral epithelial cells altered by Porphyromonas gingivalis,Porphyromonas gingivalis lipopolysaccharide,and extracellular adenosine triphosphate

Tight junctions （TJs） are the most apical intercellular junctions of epithelial cells formed by occludin, claudins, junctional adhesion molecules （JAMs）, and zonula occludens （ZO）. Tight junction proteins can sense the presence of bacteria and regulate the transcription of target genes that encode effectors and regulators of the immune response. The aim of this study was to determine the impact of TJ proteins in response to Porphyromonas gingivalis （P. gingivalis）, P. gingivalis lipopolysaccharide （P. gingivalis LPS）, and extracellular adenosine triphosphate （ATP） in the oral epithelial cell culture model. Quantified real time- polymerase chain reaction （RT-PCR）, immunoblots, and immunostaining were performed to assess the gene and protein expression in TJs. It was found that P. gingivalis infection led to transient upregulation of the genes encoding occludin, claudin- 1, and claudin-4 but not JAM-A, claudin-15, or ZO-1, while P. gingivalis LPS increased claudin-1, claudin-15, and ZO-1 and decreased occludin, JAM-A, and claudin-4. Tight junction proteins showed significant upregulation in the above two groups when cells were pretreated with ATP for 3 h. The findings indicated that P. gingivalis induced the host defence responses at an early stage. P. gingivalis LPS exerted a more powerful stimulatory effect on the disruption of the epithelial barrier than P. gingivalis. ATP stimulation enhanced the reaction of TJ proteins to P. gingivalis invasion and LPS destruction of the epithelium.

Cinnamicaldehyde regulates the expression of tight junction proteins and amino acid transporters in intestinal porcine epithelial cells

Background： Cinnamicaldehyde（CA） is a key flavor compound in cinnamon essential oil possessing various bioactivities. Tight junction（TJ） proteins are vital for the maintenance of intestinal epithelial barrier function,transport, absorption and utilization of dietary amino acids and other nutrients. In this study, we tested the hypothesis that CA may regulate the expression of TJ proteins and amino acid transporters in intestinal porcine epithelial cells（IPEC-1） isolated from neonatal pigs.Results： Compared with the control, cells incubated with 25 μmol/L CA had increased transepithelial electrical resistance（TEER） and decreased paracellular intestinal permeability. The beneficial effect of CA on mucosal barrier function was associated with enhanced protein abundance for claudin-4, zonula occludens（ZO）-1, ZO-2, and ZO-3. Immunofluorescence staining showed that 25 μmol/L CA promoted the localization of claudin-1 and claudin-3 to the plasma membrane without affecting the localization of other TJ proteins, including claudin-4, occludin,ZO-1, ZO-2, and ZO-3, compared with the control cells. Moreover, protein abundances for rBAT, xCT and LAT2 in IPEC-1 cells were enhanced by 25 μmol/L CA, while that for EAAT3 was not affected.Conclusions： CA improves intestinal mucosal barrier function by regulating the distribution of claudin-1 and claudin-3 in enterocytes, as well as enhancing protein abundance for amino acid transporters rBAT, xCT and LAT2 in enterocytes. Supplementation with CA may provide an effective nutritional strategy to improve intestinal integrity and amino acid transport and absorption in piglets.

Influence of puerarin,paeoniflorin,and menthol on structure and barrier function of tight junctions in MDCK and MDCK-MDR1 Cells

Objective:In this study,the influence of puerarin,paeoniflorin,and menthol on the structure and barrier function of tight junctions(TJs)in MadineDarby canine kidney epithelial(MDCK)and MDCK-multi-drug resistance 1(MDR1)cells was evaluated to determine the mechanisms by which the drugs cross the bloodebrain barrier(BBB).Method:Cells were treated with puerarin,paeoniflorin,and menthol followed by immunohistochemical staining with occludin,claudin-1,and F-actin.The cells were then observed using laser-scanning confocal microscopy.Average optical density(AOD)of the immunofluorescence images of the proteins were analyzed using ImageJ software while Transepithelial electrical resistance(TEER)was measured using an epithelial voltohmmeter.Results:Confocal microscopy revealed that puerarin-and paeoniflorin-treated tight junction proteins were conspicuous while menthol suppressed their expression.Correspondingly,AOD values of cells treated with puerarin or paeoniflorin,or both showed no difference compared to the control group(P>.05)while the menthol group value was downregulated.In 3 h,TEER of cells not treated with menthol were similar to the control group,while treatment with menthol significantly decreased TEER value(P<.05).In addition,application of menthol decreased TEER in MDCK cells earlier than in MDCK-MDR1 cells.Conclusion:Menthol but not puerarin and paeoniflorin may enhance paracellular transport and improve drug penetration of the BBB by disrupting the structure and,thereby,weakening the barrier function of TJs.

Hyperosmolarity disrupts tight junction via TNF-α/MMP pathway in primary human corneal epithelial cells

AIM:To investigate the mechanism of the tight junction(TJ) disruption and the association between tumor necrosis factor(TNF)-α and matrix metalloproteinase(MMPs) under hyperosmotic condition in primary human corneal epithelial cells(HCECs).METHODS:The cultured HCECs were exposed to media which adding sodium chloride(Na Cl) for hyperosmolar stress or adding rh-TNF-α(10 ng/m L). NF-κB inhibitor(5 μmol/L) or GM-6001(potent and broad spectrum MMP inhibitor, 20 μmol/L)was added 1 h before that treatment. The integrity of TJ proteins was determined by immunofluorescent(IF) staining. The m RNA levels of TNF-α and MMPs were evaluated by quantitative reverse transcription polymerase chain reaction(RT-q PCR) and the protein expression by enzyme-linked immunosorbent assay(ELISA).RESULTS:TJ proteins ZO-1 and Occludin were disrupted in primary HCECs exposed to hyperosmotic medium. The m RNA expression and protein production of TNF-α increased significantly in hyperosmotic media at 500 m Os M. TNF-α mediated the expression and production of MMP-1, MMP-13, MMP-9, and MMP-3 stimulated by hyperosmotic stress. The production of MMPs in hyperosmolar media were increased through the increase of TNF-α. GM-6001 prevent the destruction of ZO-1 and Occludin in hyperosmolar stress and rh-TNF-α treated medium. TNF-α induced activation of MMPs was involved in the TJ disruption by hyperosmolarity.CONCLUSION:TJ proteins ZO-1 and Occludin are disrupted by hyperosmolar stress and TNF-α, but protected by MMP inhibitor(GM-6001). It suggests that TNF-α/MMP pathway mediates the TJ disruption in primary HCECs exposed to hyperosmotic stress.