Effect and Mechanism of "Liver-Soothing and Spleen-Invigorating" Therapies Against Liver Injury in Rats： An Exploration Based on TGR5 and Intestinal Mucosal Barrier Function / 中国实验方剂学杂志

ObjectiveTo explore the effects of different treatment methods of "soothing liver， invigorating spleen， soothing liver and invigorating spleen， soothing liver first and then soothing liver and invigorating spleen， as well as invigorating spleen first and then soothing liver and invigorating spleen" on liver depression combined with liver injury in rats and their action mechanisms. MethodA six-week rat model of liver depression combined with liver injury was established by restraint stress and subcutaneous injection of carbon tetrachloride （CCl4， 5.89 g·kg-1， once every three days）. At the same time， the drugs were given by gavage. Forty-eight male SD rats of clean grade were randomly divided into eight groups， namely the normal group， model group， bicyclol （0.2 g·kg-1） group， Sinisan （4.32 g·kg-1） group， Liu Junzitang （9.26 g·kg-1） group， Chaishao Liu Junzitang A （Chai A， soothing liver and invigorating spleen，13.57 g·kg-1） group， Chaishao Liu Junzitang B （Chai B， soothing liver first and then soothing liver and invigorating spleen， 13.57 g·kg-1） group， and Chaishao Liu Junzitang C （Chai C， invigorating spleen first and then soothing liver and invigorating spleen， 13.57 g·kg-1） group， with six rats in each group. The pathological changes in liver and colon tissues of each group were observed under light microscope and electron microscope. The serum biochemical indexes of the liver were detected using an automatic biochemical analyzer. The relative mRNA expression levels of Takeda G protein-coupled receptor 5 （TGR5） and intestinal mucosal zona occluden-1 （ZO-1）， Occludin， and Claudin-1 in the liver and colon were detected by reverse-transcription polymerase chain reaction （RT-PCR）. The positive expression rate of proliferating cell nuclear antigen （PCNA） in the colon was detected by immunohistochemistry. ResultCompared with normal group， the model group exhibited significantly elevated serum alkaline phosphatase （ALP）， alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， total bilirubin （TBIL）， and direct bilirubin （DBIL） （P<0.01）， lowered TGR5 mRNA expression in liver tissue， up-regulated TGR5 mRNA expression in the colon tissue （P<0.05，P<0.01）， and down-regulated ZO-1， Occludin， and tight junction protein-1 (Claudin-1) mRNA expression and PCNA in the colon tissue （P<0.01）. Compared with the model group， bicyclol and Chai C remarkably decreased the levels of serum ALP， ALT， AST， TBIL， and DBIL （P<0.05，P<0.01）， while Liu Junzitang， Chai A， Chai B， and Chai C significantly up-regulated the TGR5 mRNA expression in the liver and down-regulated its expression in the colon （P<0.01）. Bicyclol， Chai A， Chai B， and Chai C enhanced the ZO-1 and Claudin-1 mRNA expression in the colon （P<0.05，P<0.01）. Bicyclol， Sinisan， and Chai C increased PCNA expression （P<0.01）. The comparison with the Chai C group showed that the TGR5 mRNA expression in the liver and ZO-1 mRNA expression in the colon of the bicyclol and Sinisan groups were lower， whereas the TGR5 mRNA expression in the colon was higher （P<0.01）. However， the PCNA expression in the colon of the Liu Junzitang and Chai B groups declined significantly （P<0.05）. ConclusionIn the presence of liver injury， invigorating spleen first helps to relieve the liver injury， and the efficacy of "spleen-invigorating" therapy in increasing the intestinal mucosal tight junction proteins and improving the gastrointestinal function is related to its activation of TGR5 to improve the intestinal mucosal barrier function， promote the renewal of intestinal stem cells， and drive the regeneration after injury.

Relationship between serum claudin-1 level and papillary thyroid carcinoma / 中华内分泌代谢杂志

Objective:To analyze the correlation between serum claudin-1(Cla-1) level and the risk of papillary thyroid carcinoma(PTC) in patients with thyroid nodules.Methods:The clinical data of 345 patients with thyroid nodules were retrospectively analyzed. According to the pathological results, they were divided into PTC group and benign thyroid nodule(BTN) group. The difference of serum Cla-1 level between 2 groups and its correlation with the risk of PTC were analyzed.Results:In groups of PTC( n=225) and BTN( n=120), the median value of serum Cla-1 level was 14.03(10.30, 20.40) ng/mL. The differences in the median value[17.90(14.00, 22.93)ng/mL vs 9.40(8.15, 11.20) ng/mL] of serum Cla-1 level in the PTC and BTN were statistically significant by Wilcoxon signed-rank test. The prevalence of PTC in thyroid nodules increased gradually with increasing of serum Cla-1 level. Receiver operated characteristic curve analysis showed that the best diagnostic cut-off value of the PTC was 13.02 ng/ml of which the sensitivity was 81.8%, the specificity was 89.2%, and the area under curve(AUC) was the largest(AUC=0.944, P<0.01, 95% CI 0.922-0.965). Logistic regression analysis showed that elevated serum Cla-1 level increased the risk of PTC, and it was statistically significant( OR=4.334, 95% CI 1.662-11.303, P=0.003). There was a significant correlation among the serum Cla-1 level and gender, age, location of involvement, number and diameter of cancer nodules, extracapsular invasion of thyroid, lymph node metastasis, tumor stage and combined with Hashimoto′s thyroiditis( P<0.01). Conclusion:The serum level of Cla-1 may be one of risk factors to predict PTC, and it is related to the total amount of PTC tumor cells in vivo, but it was not related to the aggressive behavior of tumor.

Claudin-1 and Claudin-3 as Molecular Regulators of Myelination in Leukoaraiosis Patients

OBJECTIVES: Leukoaraiosis is described as white matter lesions that are associated with cognitive dysfunction, neurodegenerative disorders, etc. Myelin depletion is a salient pathological feature of, and the loss of oligodendrocytes is one of the most robust alterations evident in, white matter degeneration. Recent studies have revealed that claudin proteins are aberrantly expressed in leukoaraiosis and regulate oligodendrocyte activity. However, the roles of claudin-1 and claudin-3 in oligodendrocytes and leukoaraiosis are still not well-defined. METHODS: Quantitative polymerase chain reaction was used to measure the expression of claudin-1 (CLDN1), claudin-3 (CLDN3), and myelinogenesis-related genes such as myelin basic protein (MBP), proteolipid protein (PLP), oligodendrocyte transcription factor 2 (OLIG2), and SRY-box transcription factor 10 (SOX10) in leukoaraiosis patients (n=122) and healthy controls (n=122). The expression of claudin-1 and claudin-3 was either ectopically silenced or augmented in Oli-neu oligodendrocytes, and colony formation, apoptosis, and migration assays were performed. Finally, the expression of myelin proteins was evaluated by western blotting. RESULTS: Our results revealed that in addition to SOX10, the expression levels of claudin-1, claudin-3, and myelinogenesis-related proteins were prominently downregulated in leukoaraiosis patients, compared to those in healthy controls. Furthermore, the growth and migration of Oli-neu cells were downregulated upon silencing claudin-1 or claudin-3. However, the overexpression of claudin-1 or claudin-3 resulted in the reduction of the degree of apoptosis in Oli-neu cells. In addition, claudin-1 and claudin-3 promoted the expression of MBP, OLIG2, PLP, and SOX10 at the translational level. CONCLUSION: Our data has demonstrated that the abnormal expression of claudin-1 and claudin-3 regulates the pathological progression of leukoaraiosis by governing the viability and myelination of oligodendrocytes. These findings provide novel insights into the regulatory mechanisms underlying the roles of claudin-1 and claudin-3 in leukoaraiosis.

Jiawei Baitouweng Decoction affects intestinal mucosal tight junction proteins in rats with ulcerative colitis through p38 MAPK-MLCK signaling pathway / 中国中药杂志

The aim of this paper was to explore the effect and mechanism of Jiawei Baitouweng Decoction(JWBTW) against ulcerative colitis(UC) from the perspective of intestinal mucosal tight junction proteins. From 60 SPF-grade male SD rats, 10 were randomly selected as the blank control, and the remaining 50 were treated with 3% dextran sodium sulfate(DSS) solution to induce UC and then randomized into the model group, mesalazine group, and low-, medium-, and high-dose JWBTW( L-JWBTW, M-JWBTW and H-JWBTW) groups, with 10 rats in each group. After successive medication for 14 days, the rat general conditions like body weight and stool were observed and the disease activity index(DAI) was calculated. The pathological changes in colon tissue was observed under a microscope for injury severity scoring and histopathological scoring. The serum endotoxin content was determined by limulus assay, followed by the measurement of protein expression levels of ZO-1, occludin, claudin-1, p38 MAPK, MLCK, MLC2 and p-MLC in colon tissue by Western blot. The results showed that compared with the blank group, the model group exhibited significantly reduced body weight, elevated DAI, injury severity and histopathological scores and serum endotoxin content, up-regulated protein expression levels of p38 MAPK, MLCK, MLC2 and p-MLC, and down-regulated ZO-1, occludin and claudin-1. Compared with the model group,mesalazine and JWBTW at each dose obviously increased the body weight, lowered the DAI, injury severity and histopathological scores and serum endotoxin content, down-regulated the protein expression levels of p38 MAPK, MLCK, MLC2 and p-MLC, and up-regulated the ZO-1, occludin and claudin-1, with the most obvious changes noticed in the H-JWBTW group. All these have indicated that JWBTW exerts the therapeutic effect against UC by inhibiting the activation of p38 MAPK/MLCK pathway, reversing the protein expression levels of occludin, claudin-1 and ZO-1, decreasing the serum endotoxin content, promoting the repair of intestinal mucosal mechanical barrier, maintaining the integrity of tight junctions, and reducing the permeability of intestinal mucosa.

Effect of electroacupuncture on intestinal epithelial mucosal barrier function in rats with diarrhea-predominant irritable bowel syndrome / 针刺研究

OBJECTIVE: To observe the effect of electroacupuncture (EA) on intestinal epithelial mucosal barrier function in diarrhea-predominant irritable bowel syndrome(IBS-D) rats, so as to explore its mechanisms underlying improvement of IBS-D. METHODS: Forty SD rats (half male and half female) were randomly divided into control, model, EA and medication (Pinaverium Bromide, PB) groups, with 10 rats in each group. The IBS-D model was established by chronic unpredictable mild stress combined with gavage of Senna-leaf solution. EA (2 Hz/15 Hz，0.1-1 mA) was applied to unilateral "Zusanli"(ST36),"Tianshu" (ST25), "Sanyinjiao"(SP6) and "Taichong"(LR3) alternatively for 15 min, once daily for 14 days. Rats of the medication group was treated by gavage of PB (10 mL·kg-1·d-1) for 14 days. The visceral sensitivity (pain) was assessed by using the pressure threshold which the inserted rectal balloon catheter air-inflation (connected to a blood pressure gauge) induced stronger abdominal muscular contraction to force the rat's abdomen to lift the experimental stand surface. The diarrhea index was used to evaluate loose stool grade. The expression of Claudin-1 and Occludin (intestinal epithelial tight junction proteins) of colon tissue was detected by immunohistochemistry. The activity of plasma diamine oxidase (DAO) was assayed by using spectrophotometry. RESULTS: Compared with the control group, the diarrhea index and plasma DAO activity in the model group were significantly increased (P0.05). CONCLUSION: Electroacupuncture can significantly improve abdominal pain and diarrhea in IBS-D model rats, which may be closely associated with its effects in up-regulating the expression of intestinal epithelial tight junction proteins Claudin-1 and Occludin to restore the function of intestinal epithelial mucosal barrier.

Prognostic Role of Claudin-1 Immunohistochemistry in Malignant Solid Tumors: A Meta-Analysis

BACKGROUND: Although the correlation between low claudin-1 expression and worse prognosis has been reported, details on the prognostic implications of claudin-1 expression in various malignant tumors remain unclear. The present study aimed to elucidate the prognostic roles of claudin- 1 immunohistochemistry (IHC) in various malignant tumors through a meta-analysis. METHODS: The study included 2,792 patients from 22 eligible studies for assessment of the correlation between claudin-1 expression and survival rate in various malignant tumors. A subgroup analysis based on the specific tumor and evaluation criteria of claudin-1 IHC was conducted. RESULTS: Low claudin-1 expression was significantly correlated with worse overall survival (OS) (hazard ratio [HR], 1.851; 95% confidence interval [CI], 1.506 to 2.274) and disease-free survival (DFS) (HR, 2.028; 95% CI, 1.313 to 3.134) compared to high claudin-1 expression. Breast, colorectal, esophageal, gallbladder, head and neck, and lung cancers, but not cervical, liver or stomach cancers, were significantly correlated with worse OS. Breast, colorectal, esophageal, and thyroid cancers with low claudin-1 expression were associated with poorer DFS. In the lower cut-off subgroup (< 25.0%) with respect to claudin-1 IHC, low claudin-1 expression was significantly correlated with worse OS and DFS. CONCLUSIONS: Taken together, low claudin-1 IHC expression is significantly correlated with worse survival in various malignant tumors. More detailed criteria for claudin-1 IHC expression in various malignant tumors are needed for application in daily practice.

Effect of Modified Sijunzi Tang on Protein and mRNA Expressions of Occludin, ZO-1 and Claudin-1 in Cerebral Ischemia Rats / 中国实验方剂学杂志

Objective:To investigate the effect of modified Sijunzi Tang on protein and mRNA expressions of Occludin, Claudin-1 and zonula occludens-1(ZO-1) in cerebral ischemia rats. Method:Totally 60 male Sprague-Dawley rats were randomly divided into sham operation group, model group, edaravone group, small-dose modified Sijunzi Tang group, middle-dose modified Sijunzi Tang group and high-dose modified Sijunzi Tang group. The middle cerebral artery occlusion (MCAO) model was prepared by suture method. After 7 days of treatment, the modeling group was put to death. Western blot was used to detect the contents of Occludin, Claudin-1 and ZO-1 in the ischemic cerebral cortex of rats. Detection of Occludin, ZO-1, Claudin-1 mRNA expression in the ischemic cortex of rats by Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR). Result:Compared with the sham operation group, protein expressions of Occludin, Claudin-1, and ZO-1 in the model group were significantly down-regulated (PPPPConclusion:Modified Sijunzi Tang may protect the blood-brain barrier and reduce brain edema in ischemic stroke rats by increasing the expression of tight junction proteins Occludin, ZO-1 and Claudin-1.

Study on Improvement Effects of Fuling Gancao Decoction on Functional Dyspepsia Model Rats / 中国药房

OBJECTIVE： To study improvement effects of Fuling gancao decoction on functional dyspepsia （FD） model rats. METHODS： Sixty SD rats were randomly divided into normal group， model group， Fuling gancao decoction high-dose group （20 g/kg）， Fuling gancao decoction low-dose group （10 g/kg）， drug combination group （Fuling gancao decoction 10 g/kg+domperidone 0.004 g/kg）， domperidone group （0.004 g/kg）， with 10 rats in each group. Except for normal group， other groups were givenicy dilute hydrochloric acid intragastrically to establish FD model.  After modeling， normal group and model group were given constant volume of distilled water at room temperature intragastrically， and other groups were given relevant drug solution （1 mL/100 g） intragastrically， once a day， for consecutive 7 d. The amount of gastric residue was measured to evaluate the gastric emptying function. Immunohistochemistry SP method was used to detect the protein expression of AQP3， Ghrelin and Substance P in gastric mucosa tissue. The protein expression of Claudin-1， Occludin and VIP were detected by Western blot assay. RESULTS： Compared with normal group， the amount of gastric residue was increased significantly in model group （P＜0.01）； positive expression of AQP3 and Ghrelin protein in gastric mucosa tissue was decreased significantly， while the positive expression of Substance P protein tended to increase； the protein expression levels of AQP3， Ghrelin， Claudin-1 and Occludin were decreased significantly， while those of Substance P and VIP were increased significantly （P＜0.05 or P＜0.01）. Compared with model group， the amount of gastric residue was decreased significantly in administration groups （P＜0.05 or P＜0.01）； the positive expression of AQP3 and Ghrelin protein tended to increase， while less positive expression of Substance P was found； the protein expression levels of AQP3 and Occludin in gastric mucosa tissue were increased significantly in administration groups， while those of VIP were decreased significantly  （P＜0.05 or P＜0.01）； the protein expression levels of Ghrelin and Claudin-1 in gastric mucosa tissue were increased significantly in Fuling gancao decoction high-dose group， Fuling gancao decoction low-dose group and drug combination group， while those of Substance P were decreased significantly （P＜0.05）. CONCLUSIONS： Fuling gancao decoction may improve the symptom of fluid retention in FD model rats by up-regulating the protein expression of AQP3， Ghrelin， Claudin-1 and Occludin and down-regulating the protein expression of Substance P and VIP in gastric mucosa tissue of rats.

Effect of alteplase on expressions of Claudin-1 and Claudin-5 proteins in vascular endothelial cells of rats with acute cerebral infarction / 吉林大学学报(医学版)

Objective:To investigate the effect of alteplase on the expressions of Claudin-1 and Claudin-5 proteins in the vascular endothelial cells after thrombolysis in the rats with acute cerebral infarction and its mechanism,and to provide experimental evidence for its clinical application. Methods: The models of acute thrombosis of middle cerebral artery of the rats were established.Fifty-four SD rats were randomly divided into sham operation group,model group and alteplase group (n= 18).The ultrastructure of brain endothelial cells of the rats was observed under transmission electron microscope.The expression levels of Claudin-1 and Claudin-5 proteins in the vascular endothelial cells of the rats were detected by immunoflurorescence and Western blotting methods.Results:The transmission electron microscope results showed that the brain volume in the ischemic area of the rats in model group was significantly increased and the endothelial cells were swollen,some of the cortical and surrounding brain tissues had obvious boundaries,and the thickness of the basement membrane was uniform and the tightly connected structure was very loose and disappeared;the swelling condition of the capillary endothelial cells in infarcted area of the rats in alteplase group was significantly reduced and the thickness of the basement membrane was improved,and most of the tightly connected structures between the brain capillary endothelial cells and the endothelial cells were loose and the fracture was lost and the structure disappeared.The immunofluorescence results showed that the expressions of Claudin-1 and Claudin-5 proteins in the vascular endothelial cells of the rats in alteplase group were significantly improved compared with model group. The Western blotting results showed the expression levels of Claudin-1 and Claudin-5 proteins in the vascular endothelial cells of the rats in model group were significantly decreased compared with sham operation group (P < 0.01 ); compared with model group, the expression levels of Claudin-1 and Claudin-5 proteins in the vascular endothelial cells of the rats in alteplase group at different time points were increased (P < 0.01 ). Conclusion: Alteplase can improve the structure of brain endothelial cells in the rats with acute cerebral infarction,and the mechanism may be related to decreasing the expressions of Claudin-1 and Claudin-5 proteins in the vascular endothelial cells of the rats induced by alteplase.

Effects of berberine extracted from Chinese Goldthread Rhizome on the expression of claudin-1 in ulcerative colitis mice / 预防医学

Objective To investigate the effect of berberine (BBR) extracted from Chinese Goldthread Rhizome on the expression of claudin-1 in ulcerative colitis mice. Methods UC model was established by dextran sulphate sodium (DSS) . Sixty BALB/c mice were randomly divided into a blank control group, a model control group, a low dose BBR group, a high dose BBR group and a salazosulfapyridine (SASP) positive control group. Then daily gavage administration was given for 7 days. During the course of experiment, the general conditions of mice were observed, and disease activity index (DAI) were assessed. At the end of treatment, colon tissue were dissected, its macroscopic damage index (CMDI) and tissue damage index (TDI) were evaluated, and the contents of claudin-1 protein and mRNA were detected by immunohistochemical method and real-time PCR separately. Results Compared with model control group, the clinical and pathological symptoms of colitis in BBR groups and SASP group were improved to various degrees, the DAI, CMDI and TDI in high dose BBR group and SASP group were all reduced (P<0.01) . The expression levels of claudin-1 mRNA in five groups were 1.00±0.03、 0.12±0.02、 0.21±0.02、 0.63±0.03 and 0.81±0.03, respectively. The expression levels of claudin-1 and its mRNA of colonic tissue in UC mice were all markedly lower than normal mice (P<0.01), which were all increased after BBR or SASP treatment when comparing with blank control group, whose difference was statistically significant. Conclusion Berberine can effectively relieve the colitis in UC mice by raising the expression of claudin-1, but the therapeutic effect was a little weaker than SASP.

STC2 Suppresses Breast Cancer Cell Migration and Invasion Via the PKC-Claudin-1-mediated Signaling / 医学研究杂志

Objective To study the effect of STC2 on invasion and metastasis of breast cancer cells and its mechanism.Methods By 231 HM shRNA interference cell STC2,we observed fiber cells expressing form,number plate cloning experiment statistical analysis.Immune co-precipitation (CO-IP) for testing related protein expression.Results By 231 HM STC2 cells expressing shRNA interference,231 HM presents high transfer characteristics of fiber cell morphology,cell invasion and metastasis ability of increase at the same time;On the contrary,after 231 cells expressing STC2,cell invasion and metastasis ability induced.In addition,after reducing STC2 expression of 231 HM cells,its resistance to radiation-inducled apoptosis and expression of STC2 after 231 reduced,ability to resist radiation induced apoptosis of cells induled too.Conclusion Collectively,these results indicate that STC2 may inhibit EMT at least partially through the PKC/Claudin-1-mediated signaling in human breast cancer cells.

Expression of inflammatory cytokines and its correlation with peptide YY and claudin-1 in patients with irritable bowel syndrome / 中华消化杂志

Objective To explore the pathogenesis of irritable bowel syndrome (IBS) by detecting serum levels and the colonic mucosa expression of inflammatory cytokines,peptide YY (PYY),and claudin-1,and to analyze their correlation.Methods From April 2013 to April 2015,50 outpatients with IBS and 20 healthy controls were selected.Serum levels of PYY,interleukin (IL)-10,tumor necrosis factor (TNF)-α and claudin-1 were detected by enzyme-linked immunosorbent assay (ELISA).The expression of IL-10,TNF-α,PYY and claudin-1 in colonic mucosa was determined by immunohistochemistry.Single factor analysis of variance,least significant difference (LSD) method,chi-square test,and Pearson correlation analysis were performed for statistical analysis.Results Among the 50 patients with IBS,27 cases were diarrhea-type irritable bowel syndrome (D-IBS),and 23 cases were constipated-type irritable bowel syndrome (C-IBS).The serum level and the positive expression rate of PYY in colonic mucosa of D-IBS group were significantly higher than those of healthy control group ((16.28± 2.75) ng/L vs (10.12± 1.55) ng/L;66.7 ％ (18/27) vs 30.0 ％ (6/20)),and the differences were statistically significant (LSD-t=10.19,x2 =6.182,both P＜0.05).The serum level and the positive expression rate of IL-10 in colonic mucosa of D-IBS group were both significantly lower than those of healthy control group ((2.95 ±0.24) ng/L vs (3.58±0.35) ng/L;22.2％(6/27) vs 50.0％ (10/20)),and the differences were statistically significant (LSD-t =4.52,x2=3.948,both P＜0.05).The serum level and the positive expression rate of TNF-α in colonic mucosa of D-IBS group were both significantly higher than those of healthy control group ((8.73±0.41) ng/L vs (7.73±0.51) ng/L;66.7％(18/27) vs 30.0％(6/20)),and the differences were statistically significant (LSD-t=8.43,x2 =6.182,both P＜0.05).There was no statistically significant difference between C-IBS group and healthy control group in the serum levels of PYY ((10.24±1.34) ng/L vs (10.12± 1.55) ng/L),IL-10 ((3.43 ± 0.71) ng/L vs (3.58 ± 0.35) ng/L),TNF-α ((7.81±0.26) ng/L vs (7.73 ±0.51) ng/L),and thus the positive expression rate in colonic mucosa (39.1％(9/23) vs 30.0％(6/20),56.5％(13/23) vs 50.0％(10/20),34.8％ (8/23) vs 30.0％(6/20);all P＞0.05).The serum level of claudin-1 and its positive expression rate of PYY,IL-10,TNF-α in colonic mucosa in D-IBS group were both lower than those of healthy control group ((94.44 ± 6.61) ng/Lvs (103.64 ± 5.47) ng/L;11.1％ (3/27) vs 40.0％ (8/20)),and the differences were statistically significant (LSD-t=5.76,x2 =5.349;both P＜0.05).However,the serum level of claudin-1 and its positive expression rate in colonic mucosa in C-IBS group were both higher than those of healthy control group ((115.54±3.42) ng/L vs (103.64±5.47) ng/L;73.9％ (17/23) vs 40.0％(8/20)),and the differences were statistically significant (LSD-t=5.56,x2 =5.055;both P＜0.05).The serum levels of IL-10 and PYY,TNF-α and claudin-1 were negatively correlated in the D-IBS group (r=-0.874 and -0.863,both P＜0.05).While the serum levels of TNF-α and PYY,IL-10 and claudin-1 were positively correlated (r =0.865 and 0.876,both P＜ 0.05).Conclusions There may be the imbalance of proinflammatory factors and anti-inflammatory factors in IBS patients.PYY may decrease the expression of claudin-1 by promoting IL-10 and inhibiting TNF-α,and thus ameliorate the inflammation reaction of IBS patients.

Impact of tumor necrosis factor-α and its antagonist on intestinal mucosa claudin-1, Zonula Occludens-1 and myosin light chain kinase in rat models of acute liver failure / 中华传染病杂志

Objective To study the impact of tumor necrosis factor-α (TNF-α) and its antagonist on the expressions of intestinal mucosa claudin-1,Zonula Occludens-1 (ZO-1) and myosin light chain kinase (MLCK) in rat models of acute liver failure.Methods Fifty four healthy male SpragueDawley (SD) rats were randomly divided into normal control group,model group and intervention group according to a random number table.Rats in normal control (n=6) group were intraperitoneally injected with 0.9％ saline (12 mL/kg).Rats in model group (n=24) and intervention group (n=24) were intraperitoneally injected with a full dose of D-galactosamine (D-GalN) at a dose of 1 200 mg/kg to establish model of acute liver failure,while rats in intervention group were intraperitoneally injected with TNF-α antagonists (rhTNFR∶Fc) at a dose of 12.5 mg/kg before 24 hours given D-GalN.At each time point of hour 8,24,48 and 72,six rats in both model group and the intervention group were sacrificed,respectively,while the normal control group were all anesthetized and sacrificed at 72 h.Models were repeated five times.Serum liver function was detected by biochemical method,and serum TNF-α level was detected by enzyme-linked immunosorbent assay (ELISA).Hematoxylin-eosin (HE) stained sections of liver and terminal ileum were examined under an optical microscope for pathological changes;and protein expression of the terminal ileum Claudin-1,ZO-1 protein and MLCK were determined by immunohistochemistry and Western blot.Means among groups were compared with t test.Results Acute liver failure was successfully induced in the D-GalN injected rats.In the model group,alanine aminotransferase (ALT) began to decline,total bilirubin continued to rise,and enzyme-jaundice separation developed at hour 72.But total bilirubin in intervention group at hour 72 was decreased.Light microscope showed that at hour 72,villus lodged at terminal ileum in the model group with part of villus tip failing off in the model group.Villus mucosa and submucosa interstitial were edema and infiltrated with numerous neutrophils.The terminal ileum kept integrate in the intervention group,and villus mucosa and submucosa were mild edema and only infiltrated with a small amount of neutrophil.Expressions of tumor necrosis necrosis factor (TNF)-α in rats of model group and intervention group were gradually increased and peaked at hour 24 ([239.83 ± 15.81] and [182.71± 17.08] ng/L,respectively),which were significantly higher than that of the control group ([24.19±3.57] ng/L,t=22.68and 15.73,respectively;both P＜0.01).Expression of serumTNF-α in the intervention group was significantly lower than that of model group (t=4.58,P＜0.01).Expressions of Claudin-1 and ZO-1 in model group decreased gradually at an early stage and reached the lowest level at hour 24 (0.355 ± 0.068 and 0.387 ± 0.091,respectively),which were both significantly lower than that of control group (1.640±0.188 and 1.015±0.150,respectively;t=12.87 and 7.14,respectively;both P＜0.01).In the intervention group,expressions of Claudin-1 and ZO-1 also decreased to the lowest level at hour 24 (1.051 ± 0.370 and 0.642 ± 0.082,respectivley),which were both significantly lower than that of control group (t =2.84 and 4.36,respectively;both P＜0.05),but significantly higher than model group with stastically difference (t =3.70 and 4.15,respectively;both P＜0.01).MLCK protein levels in the model and intervention group were gradually increased,which peaked at hour 24 (1.298±0.194 and 1.033 ± 0.073,respectively),significantly higher than the control group (0.460±0.069,t=8.16 and 11.44,both P＜0.01);and MLCK in the intervention group was lower than that in the model group with statistically difference (t=2.56,P＜0.05).Conclusions Expression of serum TNF-α in rat model of acute liver failure increases,which leads to decreased expression of Claudin-1 and ZO-1,and increased expression of MLCK,makes cell shrunk and cell gap increased.TNF-α antagonist could significantly reduce the inflammation and liver cell apoptosis,improve liver function by inhibiting MLCK expression and preventing decrease of Claudin-1 and ZO-1 proteins.

Relationship between skin barrier function and claudin-1 expression in patients with atopic dermatitis / 中华皮肤科杂志

Objective To evaluate skin barrier function in patients with atopic dermatitis (AD),and to assess its relationship with claudin-1 expression.Methods Totally,11 patients with AD and 11 healthy human controls were recruited in this study.A Tewameter TM210 was used to measure transepidermal water loss (TEWL) value,and high-frequency ultrasound to determine epidermal thickness and density,in lesional and non-lesional skin of the patients and normal skin of the healthy controls.A double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) was performed to determine the serum level of free claudin-1 in these subjects.One-way analysis of variance and t test were carried out to compare these parameters in different groups,and Pearson's correlation analysis to estimate the relationship between different parameters.Results The TEWL value was significantly higher in lesional skin than in nonlesional skin of patients with AD and normal skin of the healthy controls ((36.9 ± 34.2) vs.(9.1 ± 6.0) and (4.4 ± 3.1) g·m-2·h-1,both P＜ 0.05).The epidermal thickness in AD lesions was (0.23 ± 0.04) mm,significantly higher than that in nonlesional skin ((0.18 ± 0.03) mm,P ＜ 0.01) and normal control skin ((0.18 ± 0.02) mm,P ＜ 0.01).Ultrasound images revealed a characteristic subepidermal low echo-genic band in the AD lesions.The patients with AD showed a significantly lower serum level of claudin-1 compared with the healthy controls ((0.80 ± 0.88) vs.(1.73 ± 1.85) μg/L,P ＜ 0.05).Moreover,the serum level of claudin-1 was negatively correlated with epidermal thickness (r =-0.61,P ＜ 0.01),but positively correlated with the inverse of TEWL (1/TEWL,r =0.44,P ＜ 0.05).Conclusions The impaired skin barrier function,which may be evaluated by TEWL,1/TEWL and epidermal thickness,is associated with the expression of claudin-1 in patients with AD.

Hyperthermia aggravates focal cerebral ischemia-reperfusion injury in rats by down-regulating claudin-1 expression / 国际脑血管病杂志

Objective To investigate the impact of hyperthermia on the expression of claudin-1 in focal cerebral ischemia-reperfusion injury in rats and its mechanism in ischemic cerebral injury.Methods A total of 100 male Sprague-Dawley rats were randomly divided into 3 groups:sham operation,normothermia and hyperthermia groups.Both the normothermia and hyperthermia groups were redivided into 3 time points:Ischemia (1 hour) and reperfusion for 3,6,and 24 h.A model of middle cerebral artery occlusion was induced by the intraluminal suture method.At 24 h after reperfusion,brain water content was measured by the wet and dry weight method.The volume of cerebral infarction was assessed by 2,3,5 triphenyl tetrazolium chloride staining.At 3,6,and 24 h after reperfusion,the claudin-1 expression in ischemic brain tissue was measured by Western blot and immunohistochemical staining Results At 24 h after reperfusion,the mean neurological function score in the normothermia group was significantly lower than that in the hyperthermia group (2.3 ± 0.48 vs.3.2 ± 0.63; t =3.576,P =0.002).The brain water content on the operated sides in the sham operation,normothermia and hyperthermia groups was 79.31％ ± 0.60％,81.13％ ± 0.12％,and 84.4％ ± 0.55％,respectively.There were significant differences (F=147.115,P=0.000).Western blot analysis showed that at 3,6,and 24 h after reperfusion,the expression levels of claudin-1 in the normothermia and hyperthermia groups were significantly lower than the sham operation group (all P =0.000),and the expression levels of claudin-1 progressively decreased with the extension of ischemia-reperfusion time (all P ＜ 0.05).At the same time point,the expression level of claudin-1 in the hyperthermia group was significantly lower than that in the normothermia group (all P ＜ 0.01).At 3 and 6 h after reperfusion,the positive expression of claudin-1 among the cerebrovascular endothelial cells was observed in the sham operation,normothermia and hyperthermia groups,while at 24 h after reperfusion,no claudin-1 positive cells were observed.Compared to the sham operation group,at 3 h after reperfusion,the numbers of claudin-1 positive cell and claudin-1 IOD/area (integrated optical density/accumulated positive cell area) in the normothermia and hyperthermia groups begin to decrease,they decreased significantly at 6 h and disappeared at 24 h (P=0.000).At 3 and 6 h after reperfusion,claudin-1 IOD/area in the hyperthermia group was significantly lower than that in the normothermia group (all P ＜ 0.01).Conclusions During cerebral ischemia-reperfusion,hyperthermia may aggravate ischemic brain edema and brain injury by down-regulating the expression of claudin-1 in blood-brain barrier.

Acute Modulations in Stratum Corneum Permeability Barrier Function Affect Claudin Expression and Epidermal Tight Junction Function via Changes of Epidermal Calcium Gradient

Tight junction (TJ) is recognized as a second barrier of the skin. Altered expression of TJ proteins in various skin diseases characterized by the abnormal permeability barrier such as psoriasis suggests that TJ could be affected by stratum corneum (SC) barrier status. However, the physiological relationship between SC and TJ barrier remains to be investigated. Therefore, we examined the effect of SC barrier disruption on the expression of TJ proteins, claudin (Cldn)-1 and Cldn-4, and TJ barrier function in hairless mouse skin. We also investigated whether the alterations in epidermal Ca2+ affected TJ proteins expression in vivo. Repeated tape-stripping induced a sequential change of the expression and function of TJ. As early as 15-30 minutes after tape-stripping, downregulation of Cldn-1 and Cldn-4 immunoreactivity and protein level without change in mRNA level was found. This was accompanied by the abnormal leakage of lanthanum. However, by 1 hour Cldn-1 and Cldn-4 immunolocalization recovered along with normalized lanthanum permeation pattern. Moreover, the mRNA and protein levels of Cldn-1 and Cldn-4 were increased by 1 to 6 hours after tape-stripping. Inhibition of calcium loss by immersion of barrier-disrupted skin into a high Ca2+ solution prevented the dislocation of Cldn-1 and Cldn-4. Occlusion of barrier-disrupted skin delayed the restoration of Cldn-1 and Cldn-4. Our results suggest that the alteration of epidermal Ca2+ gradient caused by SC barrier perturbation affects the TJ structure and function and the faster recovery of TJ as compared to the SC barrier may imply the protective homeostatic mechanism of skin barrier.

Expression of Claudin-1 and -7 in Clear Cell Renal Cell Carcinoma and Its Clinical Significance

PURPOSE: We investigated the correlations between the expression of claudin-1 and claudin-7 in clear cell renal cell carcinoma (clear cell RCC) and clinical parameters. MATERIALS AND METHODS: The subjects of this study were 119 patients with confirmed clear cell RCC between January 2000 and December 2007. Their RCC tissues were immunohistochemically stained for claudin-1 and claudin-7. The correlations between the expression of claudin and parameters such as sex, age, body mass index (BMI), tumor size, TNM stage, Furhman nuclear grade, postoperative distant metastasis, and cancer-specific survival were analyzed. RESULTS: Among the total 119 subjects, claudin-1 was expressed in 18 (15.1%) and claudin- 7 in 31 (26.1%). Claudin-1 was expressed in patients who were older (p=0.007), who had a greater tumor size (p=0.001), who had a higher pathologic T stage (p=0.009), who had preoperative distant metastasis (p=0.035), and who had a higher Furhman nuclear grade (p=0.004). Claudin-7 was expressed only in patients who had a higher Furhman nuclear grade (p=0.031). The risk of postoperative distant metastasis was associated with the expression of claudin-1 (p0.05). CONCLUSIONS: In clear cell RCC, claudin-1 was expressed in patients who were older and who had a greater tumor size, who had higher T or M stages, and who had a higher Furhman nuclear grade. The expression of claudin-1 was associated with a higher risk of postoperative distant metastasis.

Correlation between Claudins Expression and Prognostic Factors in Prostate Cancer

PURPOSE: The purpose of this study was to evaluate the correlation between the expression of claudins and prognostic factors in patients with prostate cancer. MATERIALS AND METHODS: The subjects of this study were 48 patients who had undergone surgery for prostate cancer. The Gleason score (6 or lower, 7 or higher), prostate-specific antigen (PSA) level, T stage, biochemical recurrence, local recurrence, and distant metastasis were compared according to the expression of claudin-1 and claudin-5 in prostate cancer. RESULTS: In the group with a low expression of claudin-1, the Gleason score was 7 points or higher in 18 cases (82%) and 6 points or lower in 4 cases (18%). In the group with a high expression of claudin-1, the Gleason score was 7 points or higher in 13 cases (50%) and 6 points or lower in 13 cases (50%). Thus, the low-expression group had more cases with a Gleason score of 7 or higher (p=0.022). The group with a low expression of claudin-5 also had more cases with a Gleason score of 7 or higher (p=0.011). The mean PSA values in the groups with a low and high expression of claudin-1 were 9.6 ng/ml and 5.6 ng/ml, respectively (p=0.007). A low expression of claudin-5 was also associated with a high PSA value (p=0.002). There was no statistical difference in the expression of claudin-1 and claudin-5 by T stage, biochemical recurrence, local recurrence, or distant metastasis. CONCLUSIONS: The low expression of claudin-1, claudin-5 was associated with a Gleason score of 7 or higher and a high PSA value in prostate cancer.

Expression of claudin-1, beta-catenin and E-cadherin in adenocarcinoma of the colon / 대한내과학회지

BACKGROUND/AIMS: Disruption of the cell-to-cell junction with changes in the expression of the junctional proteins is the hallmark of cancer invasion and metastasis. To investigate the roles of claudin-1, beta-catenin and E-cadherin in adenocarcinoma of the colon, the relationship of their expression with clinical and pathological factors were examined. METHODS: The expression of claudin-1, beta-catenin and E-cadherin were examined in 47 cases of adenocarcinoma of the colon by immunohistochemical staining. RESULTS: A reduced claudin-1 expression was associated with advanced lymph node metastasis (p=0.019) and histological dedifferentiation at the invasive front (p=0.030). A reduced expression of beta-catenin and E-cadherin were correlated with histological dedifferentiation (p=0.012, p=0.010, respectively). The reduced expression of two or more proteins was correlated with the histological findings of dedifferentiation (p=0.030). CONCLUSIONS: These results suggest that loss of claudin-1, beta-catenin and E-cadherin expression may be correlated with the progression of adenocarcinoma of the colon and associated with an advanced histological grade.

Expression of Claudin-1, p53 and E-cadherin in Pseudoepitheliomatous Hyperplasia and Squamous Cell Carcinoma of the Head and Neck

BACKGROUND: Pseudoepitheliomatous hyperplasia (PEH) is a reactive proliferation of surface epithelium and can be confused with invasive squamous cell carcinoma (SCC) in head and neck biopsy specimens. To distinguish PEH from invasive SCC, immunohistochemical staining for claudin-1, E-cadherin and p53 was performed. METHODS: Eighteen cases of PEH and 29 invasive SCC from head and neck lesions were immunostained and examined. RESULTS: The invasive SCC showed increased staining of claudin-1 (p<0.001) and p53 (p<0.001) and decreased staining of E-cadherin (p=0.005) compared to the PEH specimens. The combined score calculated by adding the positive sum of claudin-1 and p53 and subtracting E-cadherin was useful for the differentiation of SCC from PEH (89.7% sensitivity and 88.9% specificity, p<0.001). CONCLUSION: The combined immunostaining for claudin-1, p53 and E-cadherin may help differentiate PEH from invasive SCC. The results of this study suggest that the increased expression of claudin-1 and p53 and the decreased expression of E-cadherin maybe markers for the aggressive growth of invasive SCC.