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Objective To investigate the expression of P504s and claudin-4 in prostate cancer tissues and assess the value of their combined detection in prognostic evaluation of prostate cancer. Methods We analyzed a total of 154 surgical specimens or biopsy samples of prostate tissues collected between January, 2013 and July, 2015 in the pathological tissue bank of our hospital. These specimens, which consisted of 84 prostate cancer tissues from patients aged 52 to 83 years (mean 70.9±7.3 years) and 70 benign prostatic hyperplasia tissues from patients aged 51 to 85 years (mean 68.2±7.4 years), were used to construct 60-dot matrix paraffin tissue chips for detection of P504s and claudin-4 proteins using immunohistochemistry with SP method. The correlation of P504s and claudin-4 expression with the prognosis of the patients with prostate cancer was analyzed. Results The positive expression rates of P504s and claudin-4 protein were significantly lower in benign prostatic hyperplasia tissues than in prostate cancer tissues (P < 0.01). The tissue microarray data did not reveal significant difference in the positive expression rate of P504s in prostate cancer tissues with different Gleason scores or in different stages. The positive expression rates of claudin-4 protein was significantly lower in prostate cancer with a Gleason score ≤6 than in that with a score ≥7 (60.98% vs 74.42%, P < 0.05), and was also lower in prostate cancer in T2a stage or lower than that in T2b stage or higher (55.26% vs 78.26%, P < 0.01). Conclusion Abnormal expression of P504s and claudin-4 proteins may be related with the occurrence and progression of prostate cancer, and prostate cancer with a higher level of malignancy expresses higher levels of claudin-4. Combined detection of P504s and claudin-4 can improve the detection rate of prostate cancer and can be of value in prognostic evaluation of the patients.

Objective 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 (P<0.01). At 1 week, the interstitial edema was observed in SMG tissues. Nuclear pyknosis, decreased number of acinar cells, and small focal necrosis with inflammatory infiltration were also observed over time. However, these changes were most evident at 12 weeks after irradiation. In the irradiation group, the TJ ultrastructure of glands at different times appeared to be fuzzy, collapsed, and had decreased electron density. Moreover, the width of TJs was remarkably decreased (P<0.01). The expression levels of M3 and AQP5 were decreased in a time-dependent manner, and the fluorescence intensity was significantly reduced after irradiation. However, the expression levels and fluorescence intensity of claudin-4 were enhanced in different degrees. Conclusion 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. than that in the control group (P<0.01). At 1 week, the interstitial edema was observed in SMG tissues. Nuclear pyknosis, decreased number of acinar cells, and small focal necrosis with inflammatory infiltration were also observed over time. However, these changes were most evident at 12 weeks after irradiation. In the irradiation group, the TJ ultrastructure of glands at different times appeared to be fuzzy, collapsed, and had decreased electron density. Moreover, the width of TJs was remarkably decreased (P<0.01). The expression levels of M3 and AQP5 were decreased in a time-dependent manner, and the fluorescence intensity was significantly reduced after irradiation. However, the expression levels and fluorescence intensity of claudin-4 were enhanced in different degrees. Conclusion 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. [ABSTRACT FROM AUTHOR]

Objective: To investigate the expressions of claudin-4 in human clear-cell renal cell carcinoma (ccRCC), and its relationship with clinical pathology and prognosis.Methods:The expression of claudin-4 in 50 cases of ccRCC and 10 normal kidney tissues were detected by immunohistochemical staining (IHC), analyzing the relationship between the expression level of claudin-4 and clinicopathological variables. Western blot was employed to confirm the expression of claudin-4 in 4 cases of ccRCC and 4 normal kidney tissues. The relationship of claudin-4 expression level and disease-free survival (DFS) was analyzed to predict the prognosis of ccRCC. Results: The expression level of claudin-4 in ccRCC was significantly higher than that in normal kidney tissues (P<0.05) and was markedly associated with tumor diameters, T stages, clinical stages and lymph node metastasis. The result of Western blot also confirmed that the expression of claudin-4 was much higher in ccRCC than normal kidney tissues (P<0.01), which were positively correlated with the results of immunohistochemical (IHC) staining (r2=0.748, P<0.01). Furthermore, Kaplan-Meier analysis showed that elevated expression of claudin-4 was associated with worse prognosis in ccRCC. Cox proportional-hazards regression model analysis indicated that claudin-4 was not an independent prognostic factor for ccRCC patients after surgery (relative risk=1.686, 95%CI: 0.174-16.311, P=0. 652). Conclusion: Claudin-4 might play a positive role in carcinogenesis and development of ccRCC, which might become a potential therapeutic target and adverse prognostic factor. [ABSTRACT FROM AUTHOR]

Objective: To investigate the regulation of somatostatin on Hela cell growth as well as claudin-3 and claudin-4 gene expression. Methods: Hela cells were treated with 10-6, 10-8, 10-10 and 10-12M somatostatin (SST), respectively. Untreated cells were used as control. Flow cytometry was used to detect the apoptosis of Hela cells. Realtime-PCR and Western blot were performed to assay the effect of SST on m RNA and protein expression of gene claudin-3 and claudin-4. Results: After treatment with SST by differential concentrations for 12 h, it was found that 10-10 M, 10-8M and 10-6M SST could induce a significant apoptosis in the Hela cells. Under different concentrations of SST, the m RNA and protein expression of claudin-3 and claudin-4 were up-regulated. Conclusion: In Hela cells, SST could induce claudin-3 and claudin-4 gene expression and inhibit development and diffuse of cervical cancer cells. [ABSTRACT FROM AUTHOR]

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 ( P <0.01). At 1 week, the interstitial edema was observed in SMG tissues. Nuclear pyknosis, decreased number of acinar cells, and small focal necrosis with inflammatory infiltration were also observed over time. However, these changes were most evident at 12 weeks after irradiation. In the irradiation group, the TJ ultrastructure of glands at different times appeared to be fuzzy, collapsed, and had decreased electron density. Moreover, the width of TJs was remarkably decreased ( P <0.01). The expression levels of M3 and AQP5 were decreased in a time-dependent manner, and the fluorescence intensity was significantly reduced after irradiation. However, the expression levels and fluorescence intensity of claudin-4 were enhanced in different degrees., 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.

Objective: The aim of this study was to explore the possibility of creating a toxin, C-CPE-ETA', by fusing C-terminal high affinity binding domain of CPE (C-CPE) with a truncated form of Pseudomonas aeruginosa exotoxin A (ETA') and to examine whether C-CPE-ETA' could specifically target CLDN-3, 4 molecule and the targeted toxin was cytotoxic against CLDN-3,4-overexpressing ovarian cancer., Methods: CLDN-3 and CLDN-4 expressions were analyzed at the mRNA level in three ovarian cancer cell lines and epithelial ovarian cancer tissues from 20 patients. After transforming an expression plasmid of C-CPE-ETA' into E. coli BL21 (DE3) plysS strain, the recombinant protein was purified using His-Bind resin chromatography column and analyzed by Western blot and Coomassie blue staining. The specific binding, proapoptotic and cytolytic activities were evaluated by flow cytometry, fluorescence microscopy with the JC-1 probe and MTT assay in CLDN-3,4-overexpressing ovarian cancer cells., Results: Quantitive RT-PCR results showed there existed high levels of CLDN-3 and CLDN-4 in ovarian cancer cells, CAOV3, OVCAR3 and SKOV3. Moreover, high expressions of CLDN-3 and CLDN-4 were observed in 90.0% (18/20) and 60.0% (12/20) of ovarian cancer tissues, with an expression level 10-fold higher than that in the normal ovarian tissue. A 58 000 recombinant protein C-CPE-ETA' was demonstrated by Western blot and Coomassie blue staining. Purified and recombinant C-CPE-ETA' was bound with high affinity to CLDN-3,4-overexpressing ovarian cancer cells, CAOV3, OVCAR3 and SKOV3 cells. C-CPE-ETA' was strongly proapoptotic and cytotoxic towards the CLDN-3,4-overexpressing ovarian cancer cells. The concentration of IC(50) was 7.364 ng/ml for CAOV3 cells, 8.110 ng/ml for OVCAR3 cells and 22.340 ng/ml for SKOV3 cells, respectively. However, control CLDN-3,4-deficient cell line HUVEC was not susceptible to the recombinant C-CPE-ETA' at a concentration up to 10 µg/ml., Conclusions: The C-CPE-ETA' protein exhibits remarkably specific cytotoxicity for CLDN-3,4-overexpressing ovarian cancer cells. Its therapeutic potential warrants further development for ovarian cancer molecular targeted therapy.

Objective: To investigate the expressions of prostate stem cell antigen (PSCA) and Claudin-4 in human pancreatic carcinoma and to discuss its role in the ontogenesis of pancreatic cancer., Methods: Pancreatic carcinoma tissue microarray was constructed, containing 100 cores of 10 normal adult pancreas tissues, 12 chronic pancreatitis tissues, and 78 pancreatic carcinomas. The expressions of PSCA and Claudin-4 were detected using immunohistochemical method and the relationship between PSCA and Claudin-4 and the pancreatic carcinoma was analyzed., Results: The positive expression rates of PSCA and Claudin-4 protein in pancreatic carcinoma were 79. % and 88. % respectively. PSCA and Claudin-4 staining were more intense in malignant cells than in chronic pancreatic tissues and normal adult pancreas tissues. No evidence was found for an association between expressions of PSCA and Claudin-4 and other variables, including gender, age at surgery, and tumor grade., Conclusions: The expressions of PSCA and Claudin-4 are related to the pancreatic carcinomas. PSCA and Claudin-4 play a role in the development of pancreatic cancer, but PSCA and Claudin-4 are not correlated with the clinical pathology of tumor.

Objective: To investigate the expression of claudin-4 in the eutopic and ectopic endometrium of women with endometriosis and evaluate the role of claudin-4 in the pathogenesis of endometriosis., Methods: Thirty-five women with endometriosis and 35 controls were studied. Expression of claudin-4 was investigated using immunohistochemistry, western blot and RT-PCR, respectively. Morphologic change of tight junction was also observed in different kinds of endometria, Results: (1) Glandular epithelial cells of control endometrium and eutopic endometrium showed intact tight junctions in electron micrographs, whereas the morphology of tight junctions in ovarian endometriotic tissue was disrupted and collagen bundles could be easily detected. (2) The immunohistochemical staining of claudin-4 was localized to the glandular epithelial cell membrane. Deficient or weak staining was found in ovarian endometriotic tissues. In control endometrium, eutopic and ectopic endometrium of women with endometriosis, the expression of claudin-4 protein was 89 +/- 24, 84 +/- 22 and 27 +/- 14, respectively. Relative expression of claudin-4 mRNA was 14.5 +/- 6.8, 13.8 +/- 9.5 and 2.6 +/- 2.5, respectively. Expression of claudin-4 was significantly lower in the ectopic endometriotic tissue than in the eutopic endometrium and the control at both mRNA and protein levels (P<0.05). No significant difference was found between eutopic endometrium from women with endometriosis and control endometrium from women without endometriosis (P>0.05)., Conclusion: Down-regulated expression of claudin-4 might play a pathogenic role in the formation of endometriosis.

Objective: To investigate the changes of intestinal mucosal tight junction proteins claudin-1, -3, -4 in patients with irritable bowel syndrome (IBS), and elucidate its possible role in the bowel evacuation habit changes and formation in these patients., Methods: Western blotting was employed to determine tight junction protein claudin-1,-3,-4 levels in the intestinal mucosa of patients in the control group, diarrhea-predominant IBS (D-IBS) group and constipation-predominant IBS (C-IBS) group., Results: Compared with the control group, D-IBS patients showed significantly decreased claudin-1 protein levels in both the small intestinal and colonic mucosae (P<0.05), whereas C-IBS patients had significantly elevated claudin-1 protein levels (P<0.05). No significant difference was found in claudin-3 protein expression in the both small intestinal and colonic mucosae between the D-IBS group and the control group (P>0.05), but claudin-3 protein level was shown to increase significantly in C-IBS patients (P<0.05). Claudin-4 protein followed the same pattern of alteration as claudin-1., Conclusion: Down-regulated claudin-1 and -4 expressions can be associated with bowel evacuation habit changes and formation in patients with D-IBS, but up-regulated claudin-1, -3 and -4 expressions may relate to such bowel changes in patients with C-IBS.