Effect of ionizing radiation on the secretion of the paracellular pathway in rat submandibular glands / 华西口腔医学杂志

OBJECTIVES@#This study aims to investigate the effects of ionizing radiation on the secretion of the paracellular pathway in rat submandibular glands (SMGs) and reveal the changes in the tight junction (TJ) protein claudin-4.@*METHODS@#A total of 24 Wistar rats were randomly divided into control and irradiation groups. The irradiation groups were further divided into 1, 4, and 12 weeks groups after irradiation. One-time 20 Gy irradiation was given to the SMG area on the experimental side of the irradiation group. At 1, 4, and 12 weeks after irradiation, the secretion of SMGs was measured using the Schirmer's test. The pathological changes in the gland tissues were observed under light microscopy after hematoxylin⁃eosin (HE) staining. The changes in the TJ ultrastructure were observed under transmission electron microscopy. The immunofluorescence staining and Western blot were used to detect the expression levels of muscarinic acetylcholine M3 receptor, aquaporin 5 (AQP5), and claudin-4 protein.@*RESULTS@#At 1, 4, and 12 weeks after irradiation, the secretion of SMGs in the irradiation group was significantly decreased and lower than that in the control group (@*CONCLUSIONS@#The changes in the TJ structure, the upregulation of the claudin-4 expression, and the damage in the paracellular pathway were involved in the hyposecretion of SMGs after irradiation.

Alteration in Claudin-4 Contributes to Airway Inflammation and Responsiveness in Asthma

PURPOSE: Claudin-4 has been reported to function as a paracellular sodium barrier and is one of the 3 major claudins expressed in lung alveolar epithelial cells. However, the possible role of claudin-4 in bronchial asthma has not yet been fully studied. In this study, we aimed to elucidate the role of claudin-4 in the pathogenesis of bronchial asthma. METHODS: We determined claudin-4 levels in blood from asthmatic patients. Moreover, using mice sensitized and challenged with OVA, as well as sensitized and challenged with saline, we investigated whether claudin-4 is involved in the pathogenesis of bronchial asthma. Der p1 induced the inflammatory cytokines in NHBE cells. RESULTS: We found that claudin-4 in blood from asthmatic patients was increased compared with that from healthy control subjects. Plasma claudin-4 levels were significantly higher in exacerbated patients than in control patients with bronchial asthma. The plasma claudin-4 level was correlated with eosinophils, total IgE, FEV1% pred, and FEV1/FVC. Moreover, lung tissues from the OVA-OVA mice showed significant increases in transcripts and proteins of claudin-4 as well as in TJ breaks and the densities of claudin-4 staining. When claudin-4 was knocked down by transfecting its siRNA, inflammatory cytokine expressions, which were induced by Der p1 treatment, were significantly increased. CONCLUSIONS: These findings thus raise the possibility that regulation of lung epithelial barrier proteins may constitute a therapeutic approach for asthma.

Expression of Claudin-4 in Cholangiocarcinoma and its Clinical Significance / 胃肠病学

Background:Cholangiocarcinoma(CCA)is a fatal digestive system tumor arising from biliary epithelium. Claudin-4,a key constituent of intercellular tight junction,is aberrantly and widely expressed in various epithelial tumors,and is correlated with tumorigenesis and tumor progression. Aims:To investigate the expression of claudin-4 in CCA and its correlation with clinicopathological characteristics of the tumor and patient’s prognosis. Methods:Immunohistochemistry was used to determine the expression rate and intensity of claudin-4 in CCA tissue and matched paracancerous tissue of 77 CCA patients. Correlation of claudin-4 expression in CCA with its clinicopathological characteristics was analyzed,and survival analysis was conducted by using Kaplan-Meier method. Results:Claudin-4 was strongly expressed in CCA tissue and mildly or weakly expressed in matched paracancerous tissue;the immunohistochemical score was significantly higher in cancerous tissue than in paracancerous tissue(9. 22 ± 3. 62 vs. 7. 12 ± 4. 26,P 0. 05). Conclusions:Claudin-4 is highly expressed in CCA and negatively correlated with tumor differentiation. It might be a novel diagnostic biomarker and therapeutic target for CCA.

Expression and clinical significance of Claudin-4 in human serous ovarian cancer / 中华内分泌外科杂志

Objective To investigate the expression and clinical significance of Claudin-4 in human se-rous ovarian cancer.Methods 43 cases of serous ovarian cancer from Oct.2008 to May 2014 were studied.Re-al-time quantitative PCR( qRT-PCR) was applied to detect mRNA expression of Claudin-4 in serous ovarian cancer ( n=43 ) in comparison to the corresponding tumor-adjacent tissues.The protein expression of Claudin-4 was measured by immunohistochemistry( IHC) .Results mRNA and protein expression level of Claudin-4 was signif-icantly higher in serous ovarian cancer tissues than in adjacent normal tissues(P<0.05).The high expression of Claudin-4 protein was also associated with large tumor size, lymphatic metastasis, and advanced TNM stage( P<0.05 ) .Conclusions The expression of Claudin-4 is significantly higher in serous ovarian cancer tissues than in the adjacent normal tissues, and it is associated with clinicopathological features.Claudin-4 may become a new marker in early diagnosis and biological target therapy.

Study of claudin-4 in the diagnosis and treatment of endometrial carcinoma / 中华妇产科杂志

Objective To clarify the role of claudin-4 in endometrial tumorigenesis and explore claudin-4 be as potentially useful agent in the treatment of endometrial carcinoma.Methods The expression of claudin-4 in 62 endonetrioid endometrial carcinoma (EEC),30 atypical hyperplasia endometrial tissue and 60 human normal endometrium was determined using immunohistochemistry and realtime PCR.Ninety female BALB/c mice were transplanted with Ishikawa endometrial cancer cells,which were divided into three groups with different intraperitoneal treatments with cisplatin,paclitaxel and saline solution.After the observation period,the tumors were extracted and stained with monoclonal antibody against claudin-4.The messenger RNA expression of claudin-4 was also detected using real-time PCR.Results Among the EEC samples,34％ (21/62) showed medium staining for claudin-4 and 66％ (41/62) showed intense staining.In atypical hyperplasia group,27％ (8/30) showed weak staining,53％ (16/30) showed medium staining and 20％ (6/30) showed intense staining for claudin-4.Of the normal endometrial tissue,47％ (28/60) showed weak staining and 53％ (32/60) showed no staining for claudin-4.According to real-time PCR,the relative quantity of claudin-4 was 170 ± 12 in EEC group,89 ± 15 in atypical hyperplasia group and 18 ± 3 in normal endometrium.Compared with those in atypical hyperplasia group and normal endometrium group,the protein and mRNA expression of claudin-4 were significantly increased in the group of EEC (all P ＜ 0.05).In the study of Ishikawa xenografts,no significant changes in tumor volume and claudin-4 expression were shown in paclitaxel group compared with that in the control group.Nevertheless,a significant reduction of the tumor growth and a significant decrease in claudin-4 expression were observed in cisplatin group.After cisplatin treatment,the tumor volume was significantly decreased [(0.51 ±0.21) versus (0.73 ±0.12) cm3],and the mRNA expression of claudin-4 was also significantly decreased (153 ± 35 versus 273 ± 27).Conclusion These results demonstrate that claudin-4 is strongly expressed in EEC,which may be a useful biomarker to monitor the effects of chemotherapy in patients with endometrial carcinoma.

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.

TNF-alpha Downregulates E-cadherin and Sensitizes Response to gamma-irradiation in Caco-2 Cells / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: The purpose of the present study was to assess the biological effects of TNF-alpha in Caco-2 well-differentiated colon adenocarcinoma cells and to determine radiation sensitivity in order to develop TNF-alpha into a cancer therapeutic agent. MATERIALS AND METHODS: A cell viability test was conducted via a colorimetric and colony forming assay after 1 day and 3 days of incubation with TNF-alpha. Western blotting analysis and immunofluorescence staining were conducted to explore TNF-alpha-induced morphological and molecular changes in the adhesion molecules, E-cadherin and claudin-4. The effects of gamma-irradiation at a dose of 2 Gy on cell survival were evaluated by a clonogenic assay. The molecular changes in apoptosis-regulatory proteins were assessed by Western blotting. RESULTS: Caco-2 cells were highly resistant to TNF alpha-induced cell death and 2 Gy of gamma-irradiation. However, we observed the downregulation of the adherens junctional protein, E-cadherin and translocation of tight junctional protein, claudin-4 from the membrane to the cytosol induced by TNF-alpha treatment which would indicate cell-cell junction disruptions. These alterations of junctional proteins influenced the regulation of cell death in response to 2 Gy of gamma-irradiation. The combined treatment of TNF-alpha with 2 Gy of gamma-irradiation reduced the survival of Caco-2 cells by down-regulating bcl-xl and activating JNK pathways. CONCLUSION: These results suggest that TNF-alpha might be potentially applied as a therapeutic agent in order to enhance sensitivity to 2 Gy of gamma-irradiation administered in radiotherapy for the treatment of human colon cancer.

Correlation of Decreased Expressions of Claudin 4 and E-cadherin Proteins and the Clinicopathologic Factors of Stomach Cancer

PURPOSE: We examined the expressions of claudin-4 and E-cadherin, which are known as cell adhesion-associated proteins, in stomach cancer. The relationship of their expression with the clinicopathologic factors was examined to investigate the roles of these proteins in the invasion or metastasis of stomach adenocarcinoma. METHODS: The expressions of claudin-4 and E-cadherin were examined in 73 cases of adenocarcinoma of the stomach by performing immunohistochemical staining. RESULTS: The expressions of claudin-4 and E-cadherin in the stomach adenocarcinoma were both correlated with the histologic grade, the T-stage and nodal metastasis, respectively (P<0.05). The expression of claudin-4 was significantly associated with the expression of E-cadherin. CONCLUSION: Our data suggests that claudin-4 and E-cadherin are involved in the processes of histologic differentiation, invasion and metastasis of stomach adenocarcinoma.

Expression of claudin-1, claudin-4 and zonula occludens-1 in cervical intraepithelial neoplasia and invasive squamous cell carcinoma / 대한산부인과학회지

OBJECTIVE: Cell to cell and cell to extracellular matrix interaction are crucial in tumor development and progression. Tight junction proteins such as claudins and zonula occludens-1 (ZO-1) play an important role in these processes. This study was performed to investigate the difference of expressions of claudin-1, claudin-4 and ZO-1 in low grade squamous intraepithelial lesion (LSIL), high grade squamous intraepithelial lesion (HSIL), and invasive squamous cell carcinoma (ISCC) of the uterine cervix. METHODS: The expressions of claudin-1, claudin-4 and ZO-1 were evaluated using immunohistochemical staining in 78 cervical tissue specimens (LSIL 22 case, HSIL 36 case, and ISCC 20 case). RESULTS: Claudin-1 expression was positive in 40.9% of LSIL, in 94.0% of HSIL and in 20.0% of ISCC. The expression of claudin-1 was significantly high in HSIL (p=0.0001). Claudin-4 expression was positive in 31.8% of LSIL, in 41.7% of HSIL and in 25.0% of ISCC. The expression of claudin-4 was high in HSIL, but it was not statistically different. ZO-1 expression was positive in 13.6% of LSIL, in 41.7% of HSIL, and in 25.5% of ISCC. The expression of ZO-1 was significantly high in HSIL (p=0.011). CONCLUSION: These results indicate increased expressions of claudin-1 and ZO-1 in the HSIL that includes cervical intraepithelial neoplasia (CIN) 2 and 3, which decrease during progression to cervical cancer.

Expression of Claudin-1 and -4 in Benign Lesions and Invasive Ductal Carcinomas of the Breast

BACKGROUND: The claudins are a family of transmembrane proteins associated with tight junctions and they are critical for maintaining cell-to-cell adhesion in sheets of epithelial cells. However, their role in the progression of cancer remains largely unexplored. The aims of this study were to evaluate the expression patterns of claudin-1 and -4 in benign lesions and invasive ductal carcinomas (IDC) of the breast, and relationships between the expression of these markers and the clinicopathological characteristics in IDC patients. METHODS: We examined the claudin-1 and -4 protein expressions by performing immunohistochemical stainings in 54 benign lesions and 120 IDCs via the tissue microarray method. We evaluated the correlation between the expression of these markers and the clinicopathological characteristics of IDC. RESULTS: The expressions of claudin-1 (p=0.099) and -4 (p=0.000) were up-regulated in IDCs as compared with benign lesions. The claudin-1 expression correlated with the loss of estrogen receptor (p=0.036) and progesterone receptor (p=0.011). The claudin-4 expression correlated with lymph node metastasis (p=0.043), the nuclear grade (p=0.030), the histologic grade (p=0.007), and the loss of estrogen receptor (p=0.001) and progesterone receptor (p= 0.029). CONCLUSIONS: These results suggest that claudin-1 and -4 may play a significant role in the carcinogenesis of IDC of the breast and these may represent novel markers for this disease.