Mechanistic Insights into the Impact of WIN 55, 212-2, a Synthetic Cannabinoid, on Adhesion Molecules PECAM-1 and VE-cadherin in HeLa Cells: Implications on Cancer Processes.

The endocannabinoid (eCB) system comprises endogenous ligands, cannabinoid receptors (CBRs) and proteins involved in their regulation; its alteration leads to many diseases including cancer. Thus, becomes a therapeutic target for synthetic cannabinoids aimed to control cancer cell proliferation, migration, adhesion and invasion. However, little is known about adhesion molecules regulation through CBRs activation. Consequently, the aim of this study was to evaluate the effects of a CB1/CB2 agonist, WIN-55, 212-2 (WIN), on the regulation of adhesion molecules PECAM-1 and VE-cadherin in HeLa cells. CBRs expression was evaluated by immunofluorescence staining in HeLa cells. Cell viability by MTT, cell adhesion by crystal violet, adhesion molecules expression and location by Western blot and immunofluorescence staining assays were assessed on cells treated with different WIN concentrations. Results show that CB1, CB2 and GPR55 receptors are expressed in HeLa cells. Additionally, biphasic effects were observed in their metabolic activity and adhesive properties: low WIN concentrations significantly increased them, in contrast, were decreased at high ones as compared to controls (p < 0.0001), demonstrating that WIN elicits opposite effects depending on the concentration and exposure time. PECAM-1 was detected in cytoplasm, membrane and perinuclear region of HeLa cells, whereas VE-cadherin had a nuclear distribution. There were not significant differences in PECAM-1 and VE-cadherin expression and location, suggesting that WIN does not modulate these proteins. These findings support the potential use of WIN due to its anticancer properties without dysregulating adhesion molecules. WIN possible contribution to inhibit cancer progression should be further investigated.

Dangerous liaisons: Loss of keratinocyte control over melanocytes in melanomagenesis.

Melanomas arise from transformed melanocytes, positioned at the dermal-epidermal junction in the basal layer of the epidermis. Melanocytes are completely surrounded by keratinocyte neighbors, with which they communicate through direct contact and paracrine signaling to maintain normal growth control and homeostasis. UV radiation from sunlight reshapes this communication network to drive a protective tanning response. However, repeated rounds of sun exposure result in accumulation of mutations in melanocytes that have been considered as primary drivers of melanoma initiation and progression. It is now clear that mutations in melanocytes are not sufficient to drive tumor formation-the tumor environment plays a critical role. This review focuses on changes in melanocyte-keratinocyte communication that contribute to melanoma initiation and progression, with a particular focus on recent mechanistic insights that lay a foundation for developing new ways to intercept melanoma development.

Listeria monocytogenes: possible mechanism of infection of goat uterus and its effects on uterine autophagy and cell apoptosis.

Listeriosis is highly prevalent in the animal farming industry, with Listeria monocytogenes as the causative pathogen. To identify potential therapeutic targets for LM infection, we investigated the mechanisms of LM infection in goat uteri. We inoculated a group of goats with LM via jugular vein injection, isolated and raised them, and subsequently collected sterile samples of their uterine tissue after they exhibited clinical symptoms of LM infection. We used Giemsa staining, immunohistochemical staining, real-time qPCR, and Western blotting as experimental methods.First, we investigated the mechanism of Listeria monocytogenes (LM) infection in the goat uterus by examining the expression levels of listeriolysin O, E-cadherin, and tyrosine kinase c-Met in the uterus.Furthermore, we investigated the impact of LM infection on uterine autophagy and cell apoptosis. The results indicate that the injection of LM into the goats' jugular veins leads to LM infection in the goats' uteri. During LM survival inside the goat uterine cells, there is a significant increase in the expression levels of LLO, E-cadherin, and c-Met in the host uterine tissue. This suggests that LM may potentially infect goat uteri through the InlA/E-cadherin and InlB/c-Met pathways. Furthermore, LM infection increases the levels of apoptosis and autophagy in goat uteri. Apoptosis genes Bcl-2 and Bax, as well as autophagy-related genes LC3B, PINK1, and Parkin, exhibit varying degrees of changes in localization and expression in goat uteri, mediating the occurrence of apoptotic and autophagic responses.

Regulation of intercellular viscosity by E-cadherin-dependent phosphorylation of EGFR in collective cell migration.

Collective cell migration is crucial in various physiological processes, including wound healing, morphogenesis, and cancer metastasis. Adherens Junctions (AJs) play a pivotal role in regulating cell cohesion and migration dynamics during tissue remodeling. While the role and origin of the junctional mechanical tension at AJs have been extensively studied, the influence of the actin cortex structure and dynamics on junction plasticity remains incompletely understood. Moreover, the mechanisms underlying stress dissipation at junctions are not well elucidated. Here, we found that the ligand-independent phosphorylation of epithelial growth factor receptor (EGFR) downstream of de novo E-cadherin adhesion orchestrates a feedback loop, governing intercellular viscosity via the Rac pathway regulating actin dynamics. Our findings highlight how the E-cadherin-dependent EGFR activity controls the migration mode of collective cell movements independently of intercellular tension. This modulation of effective viscosity coordinates cellular movements within the expanding monolayer, inducing a transition from swirling to laminar flow patterns while maintaining a constant migration front speed. Additionally, we propose a vertex model with adjustable junctional viscosity, capable of replicating all observed cellular flow phenotypes experimentally.

Oral inoculation of fusobacterium nucleatum exacerbates ulcerative colitis via the secretion of virulence adhesin FadA.

Fusobacterium nucleatum (F. nucleatum), an anaerobic resident of the oral cavity, is increasingly recognized as a contributing factor to ulcerative colitis (UC). The adhesive properties of F. nucleatum are mediated by its key virulence protein, FadA adhesin. However, further investigations are needed to understand the pathogenic mechanisms of this oral pathogen in UC. The present study aimed to explore the role of the FadA adhesin in the colonization and invasion of oral F. nucleatum in dextran sulphate sodium (DSS)-induced colitis mice via molecular techniques. In this study, we found that oral inoculation of F. nucleatum strain carrying the FadA adhesin further exacerbated DSS-induced colitis, leading to elevated alveolar bone loss, disease severity, and mortality. Additionally, CDH1 gene knockout mice treated with DSS presented increases in body weight and alveolar bone density, as well as a reduction in disease severity. Furthermore, FadA adhesin adhered to its mucosal receptor E-cadherin, leading to the phosphorylation of ß-catenin and the degradation of IκBα, the activation of the NF-κB signalling pathway and the upregulation of downstream cytokines. In conclusion, this research revealed that oral inoculation with F. nucleatum facilitates experimental colitis via the secretion of the virulence adhesin FadA. Targeting the oral pathogen F. nucleatum and its virulence factor FadA may represent a promising therapeutic approach for a portion of UC patients.

ZEB1 Promotes Epithelial-Mesenchymal Transition of Endometrial Epithelial Cells and Plays a Critical Role in Embryo Implantation in Mice.

Zinc finger E-box binding homeobox 1 (ZEB1) promotes epithelial-mesenchymal transition (EMT) in carcinogenesis, but its role in embryo implantation has not yet been well studied. In the present study we evaluated the hypothesis that ZEB1-induced EMT is essential for embryo implantation in vivo. Endometrial epithelium from female Kunming mice (non-pregnant, and pregnant from day 2.5 to 6.5) were collected for assessment of mRNA/protein expression of ZEB1, and EMT markers E-cadherin and vimentin, by employment of real-time quantitative reverse transcription PCR, Western blot, and immunohistochemical staining. To test if knockdown of ZEB1 affects embryo implantation in vivo, mice received intrauterine injection of shZEB1 before the number of embryos implanted was counted. The results showed that, ZEB1 was highly expressed at both mRNA and protein levels in the mouse endometrium on day 4.5 of pregnancy, paralleled with down-regulated E-cadherin and up-regulated vimentin expression (P < 0.05). Intrauterine injection of shZEB1 markedly suppressed embryo implantation in mice (P < 0.01). Conclusively, the present work demonstrated that ZEB1 is essential for embryo implantation under in vivo condition, and is possibly due to its effect on modulation of endometrial receptivity through EMT.

Adiponectin suppresses stiffness-dependent, pro-fibrotic activation of lung fibroblasts.

Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible respiratory disease with limited therapeutic options. A hallmark of IPF is excessive fibroblast activation and extracellular matrix (ECM) deposition. The resulting increase in tissue stiffness amplifies fibroblast activation and drives disease progression. Dampening stiffness-dependent activation of fibroblasts could slow disease progression. We performed an unbiased, next generation sequencing (NGS) screen to identify signaling pathways involved in stiffness-dependent lung fibroblast activation. Adipocytokine signaling was downregulated in primary lung fibroblasts (PFs) cultured on stiff matrices. Re-activating adipocytokine signaling with adiponectin suppressed stiffness-dependent activation of human PFs. Adiponectin signaling depended on CDH13 expression and p38 mitogen-activated protein kinase gamma (p38MAPKγ) activation. CDH13 expression and p38MAPKγ activation were strongly reduced in lungs from IPF donors. Our data suggest that adiponectin-signaling via CDH13 and p38MAPKγ activation suppresses pro-fibrotic activation of fibroblasts in the lung. Targeting of the adiponectin signaling cascade may provide therapeutic benefits in IPF.

Ubiquitination of VE-cadherin regulates inflammation-induced vascular permeability in vivo.

VE-cadherin is a major component of the cell adhesion machinery which provides integrity and plasticity of the barrier function of endothelial junctions. Here, we analyze whether ubiquitination of VE-cadherin is involved in the regulation of the endothelial barrier in inflammation in vivo. We show that histamine and thrombin stimulate ubiquitination of VE-cadherin in HUVEC, which is completely blocked if the two lysine residues K626 and K633 are replaced by arginine. Similarly, these mutations block histamine-induced endocytosis of VE-cadherin. We describe two knock-in mouse lines with endogenous VE-cadherin being replaced by either a VE-cadherin K626/633R or a VE-cadherin KallR mutant, where all seven lysine residues are mutated. Mutant mice are viable, healthy and fertile with normal expression levels of junctional VE-cadherin. Histamine- or LPS-induced vascular permeability in the skin or lung of both of these mutant mice are clearly and similarly reduced in comparison to WT mice. Additionally, we detect a role of K626/633 for lysosomal targeting. Collectively, our findings identify ubiquitination of VE-cadherin as important for the induction of vascular permeability in the inflamed skin and lung.

KLHL14 and E-Cadherin Nuclear Co-Expression as Predicting Factor of Nonfunctioning PitNET Invasiveness: Preliminary Study.

Background/Objectives. Novel diagnostic and therapeutic approaches are needed to improve the clinical management of nonfunctioning pituitary neuroendocrine tumors (NF-PitNETs). Here, the expression of two proteins controlling the epithelial-mesenchymal transition (EMT)-an underlying NF-PitNET pathogenic mechanism-were analyzed as prognostic markers: E-cadherin (E-Cad) and KLHL14. Methods. The immunohistochemistry characterization of KLHL14 and E-Cad subcellular expression in surgical specimens of 12 NF-PitNET patients, with low and high invasiveness grades (respectively, Ki67+ < and ≥3%) was carried out. Results. The analysis of healthy vs. NF-PitNET tissues demonstrated an increased protein expression and nuclear translocation of KLHL14. Moreover, both E-Cad and KLHL14 shifted from a cytoplasmic (C) form in a low invasive NF-PitNET to a nuclear (N) localization in a high invasive NF-PitNET. A significant correlation was found between E-Cad/KLHL14 co-localization in the cytoplasm (p = 0.01) and nucleus (p = 0.01) and with NF-PitNET invasiveness grade. Conclusions. Nuclear buildup of both E-Cad and KLHL14 detected in high invasive NF-PitNET patients highlights a novel intracellular mechanism governing the tumor propensity to local invasion (Ki67+ ≥ 3%). The prolonged progression-free survival trend documented in patients with lower KLHL14 expression further supported such a hypothesis even if a larger cohort of NF-PitNET patients have to be analyzed to definitively recognize a key prognostic role for KLHL14.

Immunohistochemistry identifies E-cadherin, N-cadherin and focal adhesion kinase (FAK) as predictors of stage I non-small cell lung carcinoma spread through the air spaces (STAS), and the combinations as prognostic factors.

Background: Spread through air spaces (STAS) is one of the multiple modes of lung cancer dissemination, yet its molecular and clinicopathological characterization remains poorly studied. This study aimed to investigate the effect of adhesion molecule expression levels on the incidence of STAS and postoperative recurrence in stage I lung cancer patients undergoing radical resection. Methods: E-cadherin, P-cadherin, N-cadherin, focal adhesion kinase (FAK), epithelial cell adhesion molecule (EpCAM), neural cell adhesion molecule 1 (NCAM1), vascular cell adhesion molecule 1 (VCAM1), intercellular cell adhesion molecule-1 (ICAM-1) were analyzed retrospectively using immunohistochemistry in patients undergoing radical resection for stage I non-small cell lung cancer (NSCLC). Patients were categorized into four groups based on adhesion molecule expression levels: "low/low", "high/low", "low/high", and "high/high", and the group with the lowest recurrence-free probability (RFP) was defined as high risk. Associations between those adhesion molecules' expression levels and STAS were determined by using the Chi-squared test and logistic regression model. RFP was analyzed by using the log-rank test and Cox proportional risk model. Results: As of January 1, 2024, 12 of 60 patients undergoing radical resection for stage I lung carcinoma had a disease recurrence. All 60 patients' tissue specimens were retrospectively analyzed, and there were no significant differences between patients with STAS-positive (n=30) and STAS-negative (n=30) in baseline clinicopathologic features, except for histological growth patterns. We found that low expression of E-cadherin, high expression of N-cadherin and FAK, and males were independent predictors of higher incidence of STAS. Multivariate Cox analysis showed that tumors with low E-cadherin/high N-cadherin, low E-cadherin/high FAK, and high N-cadherin/high FAK expression were important predictors of recurrence in patients with stage I lung carcinoma. In addition, females and high N-cadherin/high FAK were associated with a high risk of recurrence in patients with STAS. Conclusions: E-cadherin, N-cadherin, and FAK are predictors of STAS occurrence in stage I NSCLC, and their combinations are prognostic factors. The discovery of these molecular markers provides clinicians with a reliable means that may help in the early identification of individuals with a higher risk of recurrence in lung cancer patients, targeting personalized treatment plans such as aggressive adjuvant therapy or closer follow-up.

Unique Cohorts of Salivary Gland Cancer Cells as an in-vitro Model of Circulating Tumor Cells.

Introduction: The characterization of circulating tumor cells (CTC) and circulating tumor microemboli (CTM) has emerged as both a challenge to the standard view of metastasis, and as a valuable means for understanding genotypic and phenotypic variability shown even within the same cancer type. However, in the case of salivary gland neoplasms, limited data are available for the role that CTCs and CTMs play in metastasis and secondary tumor formation.ru.AQ1 In response to this, we propose that similarities between in vitro clusters of cultured salivary gland cancer cells may act as a surrogate model for in vivo CTCs and CTMs isolated from patients. Materials and Methods: Using techniques in immunofluorescence, immunoblotting, and 2-dimensional migration, we isolated and characterized a group of cohort cells from a commercially available cell line (HTB-41). Results: Here, cells exhibited a hybrid phenotype with simultaneous expression of both epithelial and mesenchymal markers (E-cadherin, vimentin, and α-SMA). Cohort cells also exhibited increased migration in comparison to parental cells. Conclusion: Data suggest that these isolated cell clusters may fucntion as a potential in vitro model of CTCs and CTMs.

Mechanism of the blood-brain barrier modulation by cadherin peptides.

Aim: This study was aimed at finding the binding site on the human E-cadherin for Ala-Asp-Thr Cyclic 5 (ADTC5), ADTC7, and ADTC9 peptides as blood-brain barrier modulator (BBBM) for determining their mechanism of action in modulating the blood-brain barrier (BBB). Methods: ADTC7 and ADTC9 were derivatives of ADTC5 where the Val6 residue in ADTC5 was replaced by Glu6 and Tyr6 residues, respectively. The binding properties of ADTC5, ADTC7, and ADTC9 to the extracellular-1 (EC1) domain of E-cadherin were evaluated using chemical shift perturbation (CSP) method in the two dimensional (2D) 1H-15N-heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) spectroscopy. Molecular docking experiments were used to determine the binding sites of these peptides to the EC1 domain of E-cadherin. Results: This study indicates that ADTC5 has the highest binding affinity to the EC1 domain of E-cadherin compared to ADTC7 and ADTC9, suggesting the importance of the Val6 residue as shown in our previous in vitro study. All three peptides have a similar binding site at the hydrophobic binding pocket where the domain swapping occurs. ADTC5 has a higher overlapping binding site with ADTC7 than that of ADTC9. Binding of ADTC5 on the EC1 domain influences the conformation of the EC1 C-terminal tail. Conclusions: These peptides bind the domain swapping region of the EC1 domain to inhibit the trans-cadherin interaction that creates intercellular junction modulation to increase the BBB paracellular porosity.

[Corrigendum] Hispolon inhibits breast cancer cell migration by reversal of epithelial­to­mesenchymal transition via suppressing the ROS/ERK/Slug/E­cadherin pathway.

Subsequently to the publication of the above paper, an interested reader drew to the authors' attention that, with the cell migration assay data shown in Fig. 7 on p. 901, the "TPA" and "TPA + U0126" panels were strikingly similar, such that data which were intended to show the results from differently performed experiments had apparently been derived from the same original source. In addition, it was noted that the "TPA + hispolon" and "TPA + NAC" data panels in Fig. 4B on p. 899 contained overlapping sections. Thirdly, a data panel was shared between Figs. 1 and 4, although this was intentional on the part of the authors as the same experiment was being portrayed in these figures.  The authors were able to re­examine their original data files, and realized that errors were made in asssembling Figs. 4B and 7. The revised versions of Figs. 4 and 7, now containing the correct data for the "TPA + NAC" experiment in Fig. 4B and the Control ("Ctrl") experiment in Fig. 7, are shown on the next two pages. The authors wish to emphasize that the corrections made to these figures do not affect the overall conclusions reported in the paper, and they are grateful to the Editor of Oncology Reports for allowing them the opportunity to publish this corrigendum. All the authors agree to the publication of this corrigendum, and also apologize to the readership for any inconvenience caused. [Oncology Reports 35: 896­904, 2016; DOI: 10.3892/or.2015.4445].

Angong Niuhuang Wan ameliorates LPS-induced cerebrovascular edema by inhibiting blood‒brain barrier leakage and promoting the membrane expression of AQP4.

Introduction: Angong Niuhuang Wan (AGNHW), developed during the Qing dynasty (18th century) for the treatment of consciousness disturbances caused by severe infections, has been used to treat brain edema caused by ischemia‒reperfusion. However, it remains unclear whether AGNHW can ameliorate vascular-origin brain edema caused by lipopolysaccharides (LPS). This study explored the ameliorative effects of AGNHW on LPS-induced cerebrovascular edema in mice, as well as the potential underlying mechanisms. Methods: A cerebrovascular edema model was established in male C57BL/6N mice by two intraperitoneal injections of LPS (15 mg/kg), at 0 and 24 h. AGNHW was administered by gavage at doses of 0.2275 g/kg, 0.455 g/kg, and 0.91 g/kg, 2 h after LPS administration. In control mice, normal saline (NS) or AGNHW (0.455 g/kg) was administered by gavage 2 h after intraperitoneal injection of NS. The survival rate, cerebral water content, cerebral venous FITC-dextran leakage, Evans blue extravasation, and expression of vascular endothelial cadherin (VE-cadherin), zonula occludens-1 (ZO-1), claudin-5, phosphorylated caveolin-1 (CAV-1), and cytomembrane and cytoplasmic aquaporin 1 (AQP1) and aquaporin 4 (AQP4) were evaluated. The cerebral tissue phosphoproteome, blood levels of AGNHW metabolites, and the relationships between these blood metabolites and differentially phosphorylated proteins were analyzed. Results: AGNHW inhibited the LPS-induced decrease in survival rate, increase in cerebral water content, decrease in VE-Cadherin expression and increase in phosphorylated CAV-1 (P-CAV-1). AGNHW treatment increased the expression of AQP4 on astrocyte membrane after LPS injection. AGNHW also inhibited the LPS-induced increases in the phosphorylation of 21 proteins, including protein kinase C-α (PKC-α) and mitogen-activated protein kinase 1 (MAPK1), in the cerebral tissue. Eleven AGNHW metabolites were detected in the blood. These metabolites might exert therapeutic effects by regulating PKC-α and MAPK1. Conclusion: AGNHW can ameliorate cerebrovascular edema caused by LPS. This effect is associated with the inhibition of VE-Cadherin reduction and CAV-1 phosphorylation, as well as the upregulation of AQP4 expression on the astrocyte membrane, following LPS injection.

E-cadherin staining in the diagnosis of lobular versus ductal neoplasms of the breast: the emperor has no clothes.

Categorizing breast neoplasia as ductal or lobular is a daily exercise that relies on a combination of histologic and immunohistochemical tools. The historically robust link between loss of the E-cadherin molecule and lobular neoplasia has rendered staining for E-cadherin by immunohistochemistry a staple of this diagnostic process. Unfortunately, discordances between E-cadherin expression and histomorphology, and variations in E-cadherin staining patterns and intensities abound in clinical practice, but are often neglected in favour of a binary interpretation of the E-cadherin result. In this article, we highlight the complexities of E-cadherin expression through a review of the E-cadherin protein and its associated gene (CDH1), the mechanisms leading to aberrant/absent E-cadherin expression, and the implications of these factors on the reliability of the E-cadherin immunohistochemical stain in the classification of ductal versus lobular mammary neoplasia.

Amphiregulin Downregulates E-cadherin Expression by Activating YAP/Egr-1/Slug Signaling in SKOV3 Human Ovarian Cancer Cells.

Amphiregulin (AREG) stimulates human epithelial ovarian cancer (EOC) cell invasion by downregulating E-cadherin expression. YAP is a transcriptional cofactor that has been shown to regulate tumorigenesis. This study aimed to examine whether AREG activates YAP in EOC cells and explore the roles of YAP in AREG-induced downregulation of E-cadherin and cell invasion. Analysis of the Cancer Genome Atlas (TCGA) showed that upregulation of AREG and EGFR were associated with poor survival in human EOC. Treatment of SKOV3 human EOC cells with AREG induced the activation of YAP. In addition, AREG downregulated E-cadherin, upregulated Egr-1 and Slug, and stimulated cell invasion. Using gain- and loss-of-function approaches, we showed that YAP was required for the AREG-upregulated Egr-1 and Slug expression. Furthermore, YAP was also involved in AREG-induced downregulation of E-cadherin and cell invasion. This study provides evidence that AREG stimulates human EOC cell invasion by downregulating E-cadherin expression through the YAP/Egr-1/Slug signaling.

Expression of E-cadherin in Hyperplastic Endometrium: A Retrospective Analysis.

Aim Epithelial cadherin or E-cadherin is a cell adhesion molecule that is present in all cells to promote integrity and survival of the cells. The aim of this study was to assess the immunohistochemical staining pattern of E-cadherin in hyperplastic endometrium. Methods A total of 25 blocks of formalin-fixed paraffin-embedded tissues of endometrial biopsies, from September 2020 to May 2023, were obtained from the Department of Pathology, Saveetha Medical College. Out of these 25 histologically proven cases of endometrial hyperplasia (EH), 17 cases were of EH without atypia and 8 cases were of endometrial hyperplasia with atypia (AH, or atypical hyperplasia). Results The immunohistochemical examination revealed that E-cadherin expression was downregulated in both EH without atypia and AH. But the downregulation was more pronounced in cases of AH than in EH without atypia. This was confirmed by the comparison of E-cadherin expression between EH with and without atypia by a chi-square test, which showed a p-value of 0.05 and was proven significant. Conclusion The heterogeneous expression of E-cadherin can be attributed to the impairment of cadherin-catenin complex. This impairment is seen in AH as well as EH without atypia. This shows this impairment occurs very early in the transformation process of the endometrium from hyperplastic to neoplastic.

Inter-tissue differences in oxidative stress susceptibility reveal a less stable endothelial barrier in the brain than in the retina.

Oxidative stress can impair the endothelial barrier and thereby enable autoantibody migration in Neuromyelitis optica spectrum disorder (NMOSD). Tissue-specific vulnerability to autoantibody-mediated damage could be explained by a differential, tissue-dependent endothelial susceptibility to oxidative stress. In this study, we aim to investigate the barrier integrity and complement profiles of brain and retinal endothelial cells under oxygen-induced oxidative stress to address the question of whether the pathomechanism of NMOSD preferentially affects the brain or the retina. Primary human brain microvascular endothelial cells (HBMEC) and primary human retinal endothelial cells (HREC) were cultivated at different cell densities (2.5*104 to 2*105 cells/cm2) for real-time cell analysis. Both cell types were exposed to 100, 500 and 2500 µM H2O2. Immunostaining (CD31, VE-cadherin, ZO-1) and Western blot, as well as complement protein secretion using multiplex ELISA were performed. HBMEC and HREC cell growth phases were cell type-specific. While HBMEC cell growth could be categorized into an initial peak, proliferation phase, plateau phase, and barrier breakdown phase, HREC showed no proliferation phase, but entered the plateau phase immediately after an initial peak. The plateau phase was 7 h shorter in HREC. Both cell types displayed a short-term, dose-dependent adaptive response to H2O2. Remarkably, at 100 µM H2O2, the transcellular resistance of HBMEC exceeded that of untreated cells. 500 µM H2O2 exerted a more disruptive effect on the HBMEC transcellular resistance than on HREC. Both cell types secreted complement factors H (FH) and I (FI), with FH secretion remaining stable after 2 h, but FI secretion decreasing at higher H2O2 concentrations. The observed differences in resistance to oxidative stress between primary brain and retinal endothelial cells may have implications for further studies of NMOSD and other autoimmune diseases affecting the eye and brain. These findings may open novel perspectives for the understanding and treatment of such diseases.

Mesothelin promotes the migration of endometrioid carcinoma and is associated with the MELF pattern.

Mesothelin (MSLN) is expressed in the mesothelium in normal tissues but is overexpressed in various malignant tumors. In this study, we searched for genes that were more frequently expressed in cases of endometrioid carcinoma (EC) with the MELF (microcystic, elongated, and fragmented) pattern using laser microdissection and RNA sequencing, and found that MSLN was predominantly expressed in cases with the MELF pattern. The role of MSLN in EC was analyzed by generating MSLN-knockout and -knockdown EC cell lines. MSLN promoted migration and epithelial-mesenchymal transition (EMT). Moreover, we found that cadherin-6 (CDH6) expression was regulated by MSLN. MSLN is known to bind to cancer antigen 125 (CA125), and we found that CA125 can regulate CDH6 expression via MSLN. Immunohistochemical investigations showed that MSLN, CA125, and CDH6 expression levels were considerably elevated in EC with the MELF pattern. The expression of CA125 was similar to that of MSLN not only in terms of immunohistochemical staining intensity but also the blood level of CA125. Our results showed that MSLN contributes to the migration and EMT of EC cells through upstream CA125 and downstream CDH6. Therefore, MSLN has potential as a therapeutic target for EC with the MELF pattern.