Transcriptional Levels of Intercellular Junction Proteins in an Alveolar Epithelial Cell Line Exposed to Irradiation or Bleomycin.

We performed a comparative study of the effects of X-ray irradiation and bleomycin on the mRNA levels of E-cadherin and tight junction proteins (claudin-3, claudin-4, claudin-18, ZO-2, and occludin) in an alveolar epithelial cell line L2. Irradiation decreased claudin-4 levels and increased occludin levels, while the levels of other mRNAs remained unchanged. Bleomycin increased the expression levels of all proteins examined except claudin-3. Irradiation and bleomycin have different effects on the expression level of intercellular junction proteins, indicating different reactions triggered in alveolar epithelial cells and a great prospects of further comparative studies.

Effect of sodium butyrate treatment at the basolateral membranes on the tight junction barrier function via a monocarboxylate transporter in goat mammary epithelial cells.

In lactating mammary glands, tight junctions (TJs) prevent blood from mixing with milk and maintain epithelial cell polarity, which is important for milk production. This study aimed to investigate the effect of sodium acetate and sodium butyrate (SB) stimulation direction on the TJ barrier function, which is measured with regard to transepithelial electrical resistance and fluorescein flux, in goat mammary epithelial cells. The expression and localization of the TJ proteins claudin-3 and claudin-4 were examined using Western blotting and immunofluorescence. SB treatment in the lower chamber of cell culture inserts adversely affected the TJ barrier function, whereas sodium acetate barely had any effect, regardless of stimulation direction. In addition, SB treatment in the lower chamber significantly upregulated claudin-3 and claudin-4, whereas TJ proteins showed intermittent localization. Moreover, SB induced endoplasmic reticulum (ER) stress. ARC155858, a monocarboxylate transporter-1 inhibitor, alleviated the adverse impact of SB on TJs and the associated ER stress. Interestingly, sodium ß-hydroxybutyrate, a butyrate metabolite, did not affect the TJ barrier function. Our findings indicate that sodium acetate and SB influence the TJ barrier function differently, and excessive cellular uptake of SB can disrupt TJs and induce ER stress.

Epithelial Tight Junction Anomalies in Nasal Inverted Papilloma.

OBJECTIVES: As a critical component of the epithelial barrier, tight junctions (TJs) are essential in nasal mucosa against pathogen invasion. However, the function of TJs has rarely been reported in nasal inverted papilloma (NIP). This study aims to investigate the potential factors of TJs' abnormality in NIP. METHODS: We assessed the expression of ZO-1, occludin, claudin-1, claudin-3, and claudin-7 in healthy controls and NIP by real-time quantitative polymerase chain reaction and immunofluorescent staining. The correlation between TJs expression and neutrophil count, TH 1/TH 2/TH 17 and regulatory T cell biomarkers, and the proportion of nasal epithelial cells was investigated. RESULTS: Upregulation of ZO-1, occludin, claudin-1, and claudin-7, along with downregulation of claudin-3, was found in NIP compared to control (all p < 0.05). An abnormal proportion with a lower number of ciliated cells (control vs. NIP: 37.60 vs. 8.67) and goblet cells (12.52 vs. 0.33) together with a higher number of basal cells (45.58 vs. 124.00) in NIP. Meanwhile, claudin-3 was positively correlated with ciliated and goblet cells (all p < 0.01). Additionally, neutrophils were excessively infiltrated in NIP, negatively correlated with ZO-1, but positively with claudin-3 (all p < 0.05). Furthermore, FOXP3, IL-10, TGF-ß1, IL-5, IL-13, and IL-22 levels were induced in NIP (all p < 0.01). Occludin level was negatively correlated with IL-10, IL-5, IL-13, and IL-22, whereas ZO-1 was positively with TGF-ß1 (all p < 0.05). CONCLUSION: Nasal epithelial barrier dysfunction with TJs anomalies is commonly associated with abnormal proliferation and differentiation of epithelial cells and imbalance of immune and inflammatory patterns in NIP. LEVEL OF EVIDENCE: NA Laryngoscope, 134:552-561, 2024.

Effect of Infliximab on Radiation-Induced Submandibular Gland Dysfunction in Rats.

Inflammatory response is one of the essential parts of various pathogenic mechanisms of radiation-induced salivary dysfunction. The effect of decreasing the levels of inflammatory cytokines on alleviating submandibular gland injuries after irradiation is unclear. This study aimed to explore the effect of the antibody against tumor necrosis factor-alpha, infliximab, on radiation-induced submandibular gland dysfunction in rats. Male Wistar rats received a single 20 Gy dose to the right submandibular gland region or sham irradiated. Meanwhile, the irradiated group was divided into infliximab treatment groups or untreated groups. Animals were euthanized at 1, 6, and 12 weeks postirradiation, and the irradiated submandibular gland was dissected for subsequent detection. Submandibular gland exposure caused obvious pathological changes. The increased levels of inflammatory cytokines, including tumor necrosis factor-alpha, interleukin-1ß, and interleukin-6, represent an aggravated inflammatory response. The results of the western blot, reverse transcription-quantitative polymerase chain reaction, and immunofluorescence staining showed upregulated levels of claudin-1, claudin-3, and aquaporin 5 and downregulated levels of claudin-4. Moreover, nuclear factor kappa-B phosphorylation levels were also up-regulated. In subsequent experiments, we found that infliximab alleviated inflammatory response, up-regulated tumor necrosis factor-alpha, interleukin-1ß, and interleukin-6 levels, and improved claudin-1, claudin-3, claudin-4, and aquaporin 5 expression. Our results indicate that infliximab might improve the para-cellular pathway and trans-cellular pathway destruction by reducing the inflammatory.

Application of Intestinal Barrier Molecules in the Diagnosis of Acute Cellular Rejection After Intestinal Transplantation.

Diagnosing acute rejection after intestinal transplantation currently heavily relies on histopathological analysis of graft biopsies. However, the invasive risks associated with ileoscopic examination and the inaccessibility for biopsy after ileostomy closure hinder real-time detection of rejection responses. Molecules comprising the intestinal barrier have been identified as physiological and molecular biomarkers for various bowel conditions and systemic diseases. To investigate the potential of barrier function-related molecules in diagnosing rejection after intestinal transplantation, plasma samples were collected longitudinally from transplant recipients. The samples were categorized into "indeterminate for rejection (IND)" and "acute rejection (AR)" groups based on clinical diagnoses at each time point. The longitudinal association between plasma levels of these barrier function-related molecules and acute rejection was analyzed using the generalized estimating equations (GEE) method. Logistic GEE models revealed that plasma levels of claudin-3, occludin, sIgA, and zonulin were independent variables correlated with the clinical diagnosis of acute rejection. The subsequent prediction model demonstrated moderate ability in discriminating between IND and AR samples, with a sensitivity of 76.0%, specificity of 89.2%, and accuracy of 84.6%. In conclusion, monitoring plasma levels of claudin-3, occludin, sIgA, and zonulin shows great potential in aiding the diagnosis of acute rejection after intestinal transplantation.

X-ray radiation-induced intestinal barrier dysfunction in human epithelial Caco-2 cell monolayers.

Radiation therapy and unwanted radiological or nuclear exposure, such as nuclear plant accidents, terrorist attacks, and military conflicts, pose serious health issues to humans. Dysfunction of the intestinal epithelial barrier and the leakage of luminal antigens and bacteria across the barrier have been linked to various human diseases. Intestinal permeability is regulated by intercellular structures, termed tight junctions (TJs), which are disrupted after radiation exposure. In this study, we investigated radiation-induced alterations in TJ-related proteins in an intestinal epithelial cell model. Caco-2 cells were irradiated with 2, 5, and 10 Gy and harvested 1 and 24 h after X-ray exposure. The trypan blue assay revealed that cell viability was reduced in a dose-dependent manner 24 h after X-ray exposure compared to that of non-irradiated cells. However, the WST-8 assay revealed that cell proliferation was significantly reduced only 24 h after radiation exposure to 10 Gy compared to that of non-irradiated cells. In addition, a decreased growth rate and increased doubling time were observed in cells irradiated with X-rays. Intestinal permeability was significantly increased, and transepithelial electrical resistance values were remarkably reduced in Caco-2 cell monolayers irradiated with X-rays compared to non-irradiated cells. X-ray irradiation significantly decreased the mRNA and protein levels of ZO-1, occludin, claudin-3, and claudin-4, with ZO-1 and claudin-3 protein levels decreasing in a dose-dependent manner. Overall, the present study reveals that exposure to X-ray induces dysfunction of the human epithelial intestinal barrier and integrity via the downregulation of TJ-related genes, which may be a key factor contributing to intestinal barrier damage and increased intestinal permeability.

Claudin-3 facilitates the progression and mediates the tumorigenic effects of TGF-ß in glioblastoma multiforme.

Glioblastoma multiforme (GBM) is a significantly malignant and lethal brain tumor with an average survival time of less than 12 months. Several researches had shown that Claudin-3 (CLDN3) is overexpressed in various cancers and might be important in their growth and spread. In this study, we used qRT-PCR, western blotting, immunohistochemistry, and immunofluorescence staining assays to investigate the expression levels of various proteins. To explore the proliferation abilities of GBM cells, we conducted the CCK-8 and EdU-DNA formation assays. Wound healing and transwell assays were used to investigate the capacities of invasion and migration of GBM cells. Additionally, we constructed an intracranial xenograft model of GBM to study the in vivo role of CLDN3. Our study devoted to investigate the function of CLDN3 in the pathogenesis and progression of GBM. Our study revealed that CLDN3 was upregulated in GBM and could stimulate tumor cell growth and epithelial-mesenchymal transition (EMT) in both laboratory and animal models. We also discovered that CLDN3 expression could be triggered by transforming growth factor-ß (TGF-ß) and reduced by specific inhibitors of the TGF-ß signaling pathway, such as ITD-1. Further analysis revealed that increased CLDN3 levels enhanced TGF-ß-induced growth and EMT in GBM cells, while reducing CLDN3 levels weakened these effects. Our study demonstrated the function of CLDN3 in facilitating GBM growth and metastasis and indicated its involvement in the tumorigenic effects of TGF-ß. Developing specific inhibitors of CLDN3 might, therefore, represent a promising new approach for treating this devastating disease.

Immunoexpression of Claudins -3, -4 and -7 in prostate adenocarcinomas.

Claudins are a family of essential tight junction proteins, abnormally expressed in human carcinomas. The studies that indicated the involvement of claudins in tumor biology and progression suggest the possibility of their utility as markers for diagnosis or prognosis, but also as possible targets for therapy. We investigated 50 prostate adenocarcinomas (PAs) for which we followed the expression of Claudins -3, -4 and -7 in relation to International Society of Urological Pathology (ISUP) grades. We observed the positivity for Claudin-3, Claudin-4, and Claudin-7 in 76%, 74% and 46% of cases. Analysis of the immunoexpression pattern revealed the cytoplasmic and nuclear translocation for Claudins -3 and -4, and only cytoplasmic for Claudin-7. For all claudins investigated, we noted a final staining score with significantly higher values or at the limit of statistical significance for PA belonging to ISUP groups 1-4. The internalization of Claudins -3, -4 and -7 expression, regardless of the degree of PA, indicates their involvement in prostate carcinogenesis. In addition, the similar immunoexpression patterns of the three investigated claudins and their positive linear correlation suggest a coordinated regulation and indicate the possibility of a targeted treatment strategy.

Picroside III, an Active Ingredient of Picrorhiza scrophulariiflora, Ameliorates Experimental Colitis by Protecting Intestinal Barrier Integrity.

This study aims to explore the protective effects of Picroside III, an active ingredient of Picrorhiza scrophulariiflora, on the intestinal epithelial barrier in tumor necrosis factor-α (TNF-α) induced Caco-2 cells and dextran sulfate sodium (DSS) induced colitis in mice. Results show that Picroside III significantly alleviated clinical signs of colitis including body weight loss, disease activity index increase, colon shortening, and colon tissue damage. It also increased claudin-3, ZO-1 and occludin expressions and decreased claudin-2 expression in the colon tissues of mice with colitis. In vitro, Picroside III also significantly promoted wound healing, decreased the permeability of cell monolayer, upregulated the expressions of claudin-3, ZO-1 and occludin and downregulated the expression of claudin-2 in TNF-α treated Caco-2 cells. Mechanism studies show that Picroside III significantly promoted AMP-activated protein kinase (AMPK) phosphorylation in vitro and in vivo, and blockade with AMPK could significantly attenuate the upregulation of Picroside III in ZO-1 and occludin expressions and the downregulation of claudin-2 expression in TNF-α treated Caco-2 cells. In conclusion, this study demonstrates that Picroside III attenuated DSS-induced colitis by promoting colonic mucosal wound healing and epithelial barrier function recovery via the activation of AMPK.

Claudin-4: A New Molecular Target for Epithelial Cancer Therapy.

Claudin-4 (CLDN4) is a key component of tight junctions (TJs) in epithelial cells. CLDN4 is overexpressed in many epithelial malignancies and correlates with cancer progression. Changes in CLDN4 expression have been associated with epigenetic factors (such as hypomethylation of promoter DNA), inflammation associated with infection and cytokines, and growth factor signaling. CLDN4 helps to maintain the tumor microenvironment by forming TJs and acts as a barrier to the entry of anticancer drugs into tumors. Decreased expression of CLDN4 is a potential marker of epithelial-mesenchymal transition (EMT), and decreased epithelial differentiation due to reduced CLDN4 activity is involved in EMT induction. Non-TJ CLDN4 also activates integrin beta 1 and YAP to promote proliferation, EMT, and stemness. These roles in cancer have led to investigations of molecular therapies targeting CLDN4 using anti-CLDN4 extracellular domain antibodies, gene knockdown, clostridium perfringens enterotoxin (CPE), and C-terminus domain of CPE (C-CPE), which have demonstrated the experimental efficacy of this approach. CLDN4 is strongly involved in promoting malignant phenotypes in many epithelial cancers and is regarded as a promising molecular therapeutic target.

Effect of eribulin on epithelial-mesenchymal transition plasticity in metastatic breast cancer: An exploratory, prospective study.

Epithelial-mesenchymal transition (EMT) plays a pivotal role in cancer metastasis and treatment resistance, which worsens prognosis. In phase III trials, eribulin improved overall survival in metastatic breast cancer (MBC) patients. In preclinical studies, eribulin suppressed EMT. However, clinical data on the use of eribulin for MBC patients are limited. In this exploratory, prospective study, we examined the effect of eribulin on EMT in MBC patients. Twenty-two patients aged 44-82 years with recurrent breast cancer or MBC were treated with eribulin. Breast cancer tissue samples were obtained before treatment and on Day 15 ± 5 of the first cycle of eribulin treatment. EMT markers (E-cadherin, claudin-3, vimentin, and N-cadherin) were analyzed using western blotting. EMT changes were evaluated based on the ratio of epithelial to mesenchymal markers before and after treatment in individual tumors. E-cadherin/vimentin, claudin-3/vimentin, E-cadherin/N-cadherin, and claudin-3/N-cadherin ratios were significantly higher after treatment (p = .007, p = .005, p = .006, and p = .011, respectively). Based on E-cadherin/vimentin, 65.0% of tumors shifted to an epithelial phenotype, as compared to 66.7% based on claudin-3/vimentin, 84.6% based on E-cadherin/N-cadherin, and 71.4% based on claudin-3/N-cadherin ratios. Thus, our results showed that eribulin suppressed EMT in breast cancer tissues.

PPARγ activation inhibits endocytosis of claudin-4 and protects against deoxynivalenol-induced intestinal barrier dysfunction in IPEC-J2 cells and weaned piglets.

The role of peroxisome proliferator activated receptor gamma (PPARγ) in the regulation of adipocyte differentiation has been well characterized. Besides adipose tissue, PPARγ is also highly expressed in the intestine. However, the functional role of PPARγ in the regulation of intestinal function still remains poorly understood. In the present study, we sought to understand the role of PPARγ activation on regulation of intestinal barrier function in intestinal porcine epithelial cells (IPEC-J2) and weaned piglets exposed to the mycotoxin, deoxynivalenol (DON). PPARγ activation by rosiglitazone and troglitazone, two pharmacological PPARγ ligands, increased the protein expression of tight junction proteins (TJP), claudin-3 and 4. PPARγ inhibition increased endocytosis of claudin-4 which was reversed by its activation with troglitazone. DON exposure decreased the protein expression of TJP, and also significantly suppressed PPARγ transcriptional activity. Interestingly, PPARγ activation reversed the reduction of claudin-3 and 4 caused by DON in vitro and in vivo. PPARγ activation also partially restored the transepithelial electrical resistance (TEER) and reduced the permeability of fluorescein isothiocyanate-dextran (FITC-dextran) that have been negatively impacted by DON. These effects were lost in the presence of a specific PPARγ antagonist or in PPARγ knockout cells, confirming the importance of PPARγ in the regulation of intestinal barrier function and integrity. Likewise, in weaned pigs exposed to DON, the PPARγ agonist pioglitazone mitigated the impaired villus-crypt morphology caused by DON. Therefore, pharmacological and natural bioactive compounds with PPARγ stimulatory activities could be effective in preventing DON-induced gut barrier dysfunction.

Claudin-3 Loss of Expression Is a Prognostic Marker in Castration-Resistant Prostate Cancer.

Castration-resistant prostate cancer (CRPC) development is the foremost concern after treatment of patients with high risk with locally advanced or metastatic prostate cancer. Androgen receptor (AR) is the main driver of CRPC development, through its interaction with epigenetic modifier genes, placing epigenetics modifications in the forefront of CRPC development. Comparing the DNA methylation and expression profile of androgen-sensitive and -refractory prostate cancer cells, we describe the epigenetic silencing of claudin-3 (CLDN3) in AR positive cells resistant to androgen deprivation (LNCaP-abl). CLDN3 silencing was associated with DNA methylation, loss of histone acetylation and H3K27 methylation, and was re-expressed by the combined treatment with the epigenetic modulators Aza and SAHA. From a functional point of view, CLDN3 loss was associated with increased cellular invasion. Immunohistochemical analysis showed decreased CLDN3 expression in samples from CRPC patients. Interestingly, CLDN3 expression was significantly decreased in samples from patients with high total Gleason score (≥8) and locally advanced tumors. Finally, CLDN3 loss of expression was associated with worse disease-free survival and time to clinical progression. In conclusion, our findings strongly indicate that epigenetic silencing of CLDN3 is a common event in CRPC that could be useful as a molecular marker for the prognosis of prostate cancer patients and to discriminate aggressive from indolent prostate tumors.

Acute arsenic exposure exacerbates lipopolysaccharide-induced lung injury possibly by compromising the integrity of the lung epithelial barrier in rats.

Inhalation of large amounts of arsenic can damage the respiratory tract and may exacerbate the development of bacterial pneumonia, but the exact mechanism remains unclear. In this study, male Wistar rats were randomly divided into control, arsenic trioxide (16.0 µg/kg ATO), lipopolysaccharide (0.5 mg/kg LPS), and ATO combined with LPS (16.0 µg/kg ATO + 0.5 mg/kg LPS) groups. Blood and lung tissue samples were collected from each group 12 h after exposure. The results showed that exposure to ATO or LPS alone produced different effects on leukocytes and inflammatory factors, while combined exposure significantly increased serum interleukin-6, interleukin-10, lung water content, lung lavage fluid protein, and p38 protein phosphorylation levels. Alveolar interstitial thickening, alveolar membrane edema, alveolar type I and II cell matrix vacuolization, and nuclear pyknosis were observed in rats exposed to either ATO or LPS. More severe ultrastructural changes were found in the combined exposure group, and chromatin splitting was observed in alveolar type I cells. Lanthanum nitrate particles leaked from the alveolar vascular lumen in the ATO-exposed group, whereas in the combined exposure group, Evans Blue levels were increased and lanthanum nitrate particles were present in the lung parenchyma. Claudin-3 protein expression increased and claudin-4 expression decreased after ATO or LPS exposure, while claudin-18 expression was unchanged. The changes in claudin-3 and claudin-4 protein expression were further exacerbated by combined exposure. In conclusion, these results suggest that inhalation of ATO may exacerbate the development of bacterial pneumonia and that common mechanisms may exist to synergistically disrupt epithelial barrier integrity.

Dysregulated claudin expression significantly effect breast cancer disease progression.

Background: In this study, the role of claudins in cancer progression was explored among breast cancer-affected women. Methodology: Two cohorts (discovery and validated) of breast cancer-affected women were used. In discovery cohort, 90 freshly excised breast tumor tissues along with adjacent cancer free specimens were collected at the time of surgery. These specimens were processed for RNA isolation and complementary DNA synthesis. After designing primers for claudin 3, claudin 4, and claudin 7, these sequences were synthesized from Macrogen, Korea. Claudin expression in respective tumors and controls was assessed using quantitative reverse transcription polymerase chain reaction. Any probable correlation of these molecules with various clinicopathological parameters was explored. For validation, a publicly available dataset of 2088 breast cancer patients was accessed. Claudin expression of these patients was analyzed for given clinical parameters and compared with earlier findings of discovery cohort. Results: Discovery cohort comprised 17% luminal A, 63% luminal B, 8% human epidermal growth factor receptor 2 enrich, and 12% triple-negative breast cancer tumor. High claudin 3 expression was significantly correlated with tumor size >2 cm and menopausal status. Claudin 7 expression was upregulated among poorly differentiated tumor patients. Both claudins 3/4 showed significant correlation with tumor grade, stage, size, and metastasis. Claudin-low subtype was also found in 18% of the cohort. Conclusion: Claudins impart a significant role in cell differentiation and disease progression. Hence, claudin cluster can be ascertained as the disease biomarkers for breast cancer.

Clinical Significance of Claudin Expression in Oral Squamous Cell Carcinoma.

A change in claudin expression has been demonstrated in various tumors. The present study specifically compares claudin expression in oral squamous cell carcinoma (OSCC) with healthy oral epithelium from the same individual and analyzes the association between claudin expression and the clinically relevant course parameters. Our study includes tissue samples and clinically relevant follow-up data from 60 patients with primary and untreated OSCC. The oral mucosa was analyzed via Western blot for the expression of claudin-1, -2, -3, -4, -5, and -7. Importantly, the tumor and healthy tissues were obtained pairwise from patients, allowing for intraindividual comparisons. Both the healthy and tumor epithelium from the oral cavity did not express the claudin-3 protein. The intraindividual comparison revealed that, in OSCC, claudin-2 expression was higher, and the expression of claudin-4, -5, and -7 was lower than in healthy epithelium. An association was found between increased claudin-2 expression and shorter relapse-free survival. In addition, the reduced expression of claudin-4 had a negative impact on relapse-free survival. Furthermore, associations between the reduced expression of claudin-7 and the stage of a tumor, or the presence of lymph node metastases, were found. Thus, the expression level of claudin-2, -4, and -7 appears to be predictive of the diagnosis and prognosis of OSCC.

The short-term and long-term effects of intranasal mesenchymal stem cell administration to noninflamed mice lung.

Mesenchymal stem cells (mesenchymal stromal cells; MSC)-based therapies remain a promising approach to treat degenerative and inflammatory diseases. Their beneficial effects were confirmed in numerous experimental models and clinical trials. However, safety issues concerning MSCs' stability and their long-term effects limit their implementation in clinical practice, including treatment of respiratory diseases such as asthma, chronic obstructive pulmonary disease, and COVID-19. Here, we aimed to investigate the safety of intranasal application of human adipose tissue-derived MSCs in a preclinical experimental mice model and elucidate their effects on the lungs. We assessed short-term (two days) and long-term (nine days) effects of MSCs administration on lung morphology, immune responses, epithelial barrier function, and transcriptomic profiles. We observed an increased frequency of IFNγ- producing T cells and a decrease in occludin and claudin 3 as a long-term effect of MSCs administration. We also found changes in the lung transcriptomic profiles, reflecting redox imbalance and hypoxia signaling pathway. Additionally, we found dysregulation in genes clustered in pattern recognition receptors, macrophage activation, oxidative stress, and phagocytosis. Our results suggest that i.n. MSCs administration to noninflamed healthy lungs induces, in the late stages, low-grade inflammatory responses aiming at the clearance of MSCs graft.

Claudin-3 inhibits tumor-induced lymphangiogenesis via regulating the PI3K signaling pathway in lymphatic endothelial cells.

Claudin-3 is a tight junction protein that has often been associated with the progression and metastasis of various tumors. Here, the role of claudin-3 in tumor-induced lymphangiogenesis is investigated. We found an increased lymphangiogenesis in the B16F10 tumor in claudin-3 knockout mice, accompanied by augmented melanoma cell metastasis into sentinel lymph nodes. In vitro, the overexpression of claudin-3 on lymphatic endothelial cells inhibited tube formation by suppressing cell migration, resulting in restricted lymphangiogenesis. Further experiments showed that claudin-3 inhibited lymphatic endothelial cell migration by regulating the PI3K signaling pathway. Interestingly, the expression of claudin-3 in lymphatic endothelial cells is down-regulated by vascular endothelial growth factor C that is often present in the tumor microenvironment. This study indicates that claudin-3 plays an important role as a signaling molecule in lymphatic endothelial cell activity associated with tumor lymphangiogenesis, which may further contribute to melanoma metastasis.

Zearalenone-Induced Mechanical Damage of Intestinal Barrier via the RhoA/ROCK Signaling Pathway in IPEC-J2 Cells.

Zearalenone (ZEN) is a widespread contaminant of cereals and agricultural products which causes food safety issues. Ingesting food or feed contaminated with ZEN can disrupt the intestinal epithelial barrier function. The RhoA/ROCK signaling pathway plays a key role in regulating the epithelial barrier function, but studies on such roles have rarely focused on the intestine. The aim of this experiment was to investigate the exact mechanism of ZEN-induced intestinal barrier damage and whether the RhoA/ROCK signaling pathway is involved. The results showed that ZEN significantly induced alkaline phosphatase (AP) activity and FITC-dextran (4 kDa) passage across the epithelial barrier, which significantly reduced the transepithelial resistance (TEER). Meanwhile, ZEN could induce the significantly down-regulated mRNA expression of tight junction proteins (occludin, claudin-1, ZO-1, and claudin-3) and redistribution of ZO-1 immunofluorescence. Further studies demonstrated that ZEN exposure activated the RhoA/ROCK signaling pathway, significantly up-regulated the mRNA expression of ROCK1, the main effector of the signaling pathway, the protein expression of phosphorylated myosin light chain (MLC) and myosin light chain kinase (MLCK), and relatively increased the activity of ATP in cells, simultaneously remodeling the cytoskeleton (F-actin). Overall, our study indicated that ZEN induced intestinal barrier dysfunction by activating the RhoA/ROCK signaling pathway.

The Pivotal Mediating Role of Adenosine Monophosphate-Activated Protein Kinase (AMPK) in Liver Tight Junctions and Liver Regeneration of a Partial-Hepatectomy Mouse Model.

PURPOSE: This study aims to explore the pivotal mediating role of adenosine monophosphate-activated protein kinase (AMPK) in liver tight junctions and liver regeneration of a partial hepatectomy (PH) mouse model. METHODS: A 70% PH mouse model was used. Firstly, mice were randomly divided into sham, 70% PH, AMPK-activated, and AMPK-inhibited groups. Then serum levels of alanine aminotransferase, aspartate transaminase, total bilirubin, direct bilirubin, albumin, and prealbumin were tested on postoperative days 1, 2 and 3. Furthermore, the expression of tight junction proteins like occludin, claudin-3, and ZO-1, together with bile salt export pump (BSEP), which reflects liver function, and AMPK were measured by Western blot and quantitative real-time polymerase chain reaction. Moreover, the expression of tight junction proteins, BSEP, and Ki-67 were examined by immunohistochemistry. RESULTS: After 70% PH, without intervention, the changes in expression of hepatic tight junction proteins (occludin, claudin-3, and ZO-1) were consistent with that of BSEP, which could reflect liver function. After treatment with AMPK activator, the high expression status of tight junction proteins occurred in advance and was maintained stably and for a longer time. It was beneficial to liver function and liver regeneration was promoted at early periods and enhanced continuously after PH. CONCLUSIONS: Activation of AMPK could effectively enhance the expression of hepatic tight junction proteins after PH. Therefore, it could speed up the recovery of liver function and promote liver regeneration especially early after PH.