Functional and structural phenotyping of cardiomyocytes in the 3D organization of embryoid bodies exposed to arsenic trioxide.

Chronic exposure to environmental pollutants threatens human health. Arsenic, a world-wide diffused toxicant, is associated to cardiac pathology in the adult and to congenital heart defects in the foetus. Poorly known are its effects on perinatal cardiomyocytes. Here, bioinformatic image-analysis tools were coupled with cellular and molecular analyses to obtain functional and structural quantitative metrics of the impairment induced by 0.1, 0.5 or 1.0 µM arsenic trioxide exposure on the perinatal-like cardiomyocyte component of mouse embryoid bodies, within their 3D complex cell organization. With this approach, we quantified alterations to the (a) beating activity; (b) sarcomere organization (texture, edge, repetitiveness, height and width of the Z bands); (c) cardiomyocyte size and shape; (d) volume occupied by cardiomyocytes within the EBs. Sarcomere organization and cell morphology impairment are paralleled by differential expression of sarcomeric α-actin and Tropomyosin proteins and of acta2, myh6 and myh7 genes. Also, significant increase of Cx40, Cx43 and Cx45 connexin genes and of Cx43 protein expression profiles is paralleled by large Cx43 immunofluorescence signals. These results provide new insights into the role of arsenic in impairing cytoskeletal components of perinatal-like cardiomyocytes which, in turn, affect cell size, shape and beating capacity.

Dapagliflozin attenuates arrhythmic vulnerabilities by regulating connexin43 expression via the AMPK pathway in post-infarcted rat hearts.

We have demonstrated that dapagliflozin, a sodium-glucose cotransporter (SGLT) 2 inhibitor, attenuates reactive oxygen species (ROS) production. Connexin43 playing a role in ventricular arrhythmia is sensitive to redox status. No data are available on the effects of dapagliflozin on arrhythmogenesis. This study was to determine whether dapagliflozin attenuated arrhythmias through modulating AMP-activated protein kinase (AMPK)/free radicals-induced connexin43 after myocardial infarction. After coronary ligation, normoglycemic male Wistar rats were randomized to either vehicle or dapagliflozin (0.1 mg/kg per day) for 4 weeks. Myocardial ROS levels were significantly increased (p < 0.05) and connexin43 levels were substantially decreased after myocardial infarction (p < 0.05). Dapagliflozin administration was associated with increased SGLT1, attenuated ROS and increased connexin43 levels in myocardium (all p < 0.05). During programmed electrical stimulation, arrhythmic severity was significantly improved in the dapagliflozin-treated infarcted rats than those in the vehicle-treated infarcted rats (p < 0.05). Dapagliflozin significantly increased AMPK phosphorylation compared to vehicle after infarction (p < 0.05). Inhibition of AMPK signaling by SBI-0206965 prevented increased SGLT1 and blocked the effects of dapagliflozin on attenuated ROS levels and increased connexin43 phosphorylation (all p < 0.05). SGLT1 inhibited by KGA-2727 showed attenuated ROS levels and increased connexin43 phosphorylation (both p < 0.05) although AMPK phosphorylation was not changed, implying SGLT1 activation was mediated by AMPK in dapagliflozin-treated hearts. Dapagliflozin-treated hearts had significantly increased connexin43 phosphorylation (p < 0.05), which was significantly decreased after adding 3-morpholinosydnonimine (p < 0.05). These data indicate that clinically-relevant dapagliflozin concentrations decreased free radicals content and increased connexin43 levels through AMPK-dependent and SGLT1-independent mechanisms, which attenuated ventricular arrhythmias in the normoglycemic infarcted rats.

Effects of CMYA1 overexpression on cardiac structure and function in mice.

CMYA1 (cardiomyopathy-associated protein 1, also termed Xin) localizes to the intercalated disks (ICDs) of the myocardium and functions to maintain ICD structural integrity and support signal transduction among cardiomyocytes. Our previous study showed that CMYA1 overexpression impairs the function of gap junction intercellular communication processes. Successful model generation was verified based on PCR, western blot analysis, immunohistochemistry, and immunofluorescence analysis. Myocardial CMYA1 expression was confirmed at both the mRNA and the protein levels in the CMYA1-OE transgenic mice. Masson's trichrome staining and electron microscopy revealed myocardial fibrosis and uneven bead width or the interruption of ICDs in the hearts of the CMYA1-OE transgenic mice. Furthermore, the Cx43 protein level was reduced in the CMYA1-OE mice, and co-immunoprecipitation assays of heart tissue protein extracts revealed a physical interaction between CMYA1 and Cx43. Electrocardiogram analysis enabled the detection of an obvious ventricular bigeminy for the CMYA1-OE mice. In summary, analysis of our mouse model indicates that elevated CMYA1 levels may induce myocardial fibrosis, impair ICDs, and downregulate the expression of Cx43. The observed ventricular bigeminy in the CMYA1-OE mice may be mediated by the reduced Cx43 protein level.

MiR-23a Is Involved in Myocardial Ischemia/Reperfusion Injury by Directly Targeting CX43 and Regulating Mitophagy.

Activation of CX43 signaling protects myocardial cells from myocardial ischemia/reperfusion (I/R) injury. However, the underlying mechanism remains unclear. MicroRNAs (miRNAs) are well known to play important roles in the progression of diverse diseases. Here, we first confirmed the expression profile of CX43 in rat heart tissues with I/R injury. Then, microRNAs (miRNAs) that target CX43 were predicted using miRDB, miRWalk, and TargetScan. The candidate miR-23a was selected, and its expression level in I/R samples was investigated. To determine the role of miR-23a, rat primary myocardial cells were transfected with miR-23a mimics after they were subjected to hypoxia-reoxygenation (H/R) injury. Transfection of miR-23a mimics stimulated mitophagy through the PINK1/Parkin pathway and downregulated the protein level of CX43. Treatment of miR-23a-transfected cells with NF-kB inhibitors completely abolished miR-23a-mediated mitophagy after H/R. Moreover, miR-23a transfection significantly suppressed CX43 expression and enhanced mitophagy in the model heart in vivo. Therefore, miR-23a plays a detrimental role in myocardial I/R injury by enhancing mitophagy and inhibiting CX43 mRNA.

Expression of NEDD9 and connexin-43 in neoplastic and stromal cells of gastric adenocarcinoma.

Gastric cancer is related to high mortality rates and advanced disease stage at the time of diagnosis. Its carcinogenesis is extensively studied and is associated with genetic and epigenetic changes, changed the interaction between tumor and adjacent stromal cells, and changes in the microenvironment molecule status. Neural precursor cell-expressed developmentally down-regulated 9 (NEDD9) affects different signaling proteins and pathways, apoptosis, adhesion, cell migration, and invasiveness. Connexin-43 (Cx43) also assists in intercellular communications and has several channel-independent functions. Aberrant expression of those two gap junction proteins plays an essential role in metastatic processes. Our scope was to detect the expression of Cx43 and NEDD9 in epithelial and stromal gastric cancer compartments and its relation to tumor progression and lymph node metastases. Cancer tissue from 53 cases of node-negative and 55 cases of node-positive primary gastric carcinoma patients was analyzed for Cx43 and NEDD9 expression by immunohistochemical assay, and the results were correlated with the remaining clinical and pathological findings and survival. In our cohort of patients with lymph node metastases, we detected higher expression of epithelial Cx43 in the primary tumor and stromal Cx43 expression correlated with both epithelial NEDD9 (rho = 0.453) and stromal NEDD9 (rho = 0.484). Higher epithelial Cx43 and NEDD9 expression were associated with higher mortality (HR 1.54, 95% CI 1.01-2.37, p = 0.048). Epithelial Cx43 expression, both epithelial and stromal NEDD9 expression, T and N status were all independently associated with shorter survival. In summary, our findings suggest that increased expression of both epithelial and stromal NEDD9 and epithelial Cx43 could potentially be used as prognostic gastric cancer biomarkers.

Expression of connexin 43 by atypical fibroxanthoma.

INTRODUCTION: Connexins are transmembrane channel proteins that interconnect adjacent cells and allow the exchange of signaling molecules between cells and the extracellular milieu. They have been investigated in many tumors to obtain information about tumor nature, behavior, and prognosis. METHODS: Herein, we present a study on the immunohistochemical expression of connexin (Cx) 43 in 16 cases of atypical fibroxanthoma (AFX). For the immunohistochemical staining, a tissue array was obtained from the paraffin-embedded blocks. RESULTS: The expression was membranous and cytoplasmic in all cases. Thirteen cases (81.25%) showed strong staining. In the other three cases (18.75%), the staining was medium. None of the cases showed nuclear staining. Fifteen out of 16 cases showed a diffuse pattern, and only one case showed a focal pattern. CONCLUSIONS: Our results suggest that Cx43 may play an important role in the natural behavior of AFX.

Salidroside Attenuates Hypoxia-Induced Expression of Connexin 43 in Corpus Cavernosum Smooth Muscle Cells.

INTRODUCTION: Connexin 43 (Cx43) is the major component of gap junction in corpus cavernosum smooth muscle, which allows rapid intercellular communication. Cx43 coordinates corpus cavernosum smooth muscle cells and ensures erectile function. The role of hypoxia in Cx43 dysfunction resulting in erectile dysfunction has not been well studied, and salidroside has shown cell protective effects under hypoxia. OBJECTIVE: We aimed to investigate the protective role of salidroside and the underlying mechanisms in hypoxia-induced dysfunction of Cx43. METHODS: Corpus cavernosum smooth muscle cells prepared from young male Sprague-Dawley rats were pretreated with or without salidroside and exposed to hypoxic condition for 48 h. The cell viability, expression of hypoxia-inducible factor-1α (HIF-1α) and Cx43, and Ca2+ signals were investigated. RESULTS: Pretreatment with salidroside attenuated loss of hypoxia-induced cell viability markedly and could downregulate the HIF-1α protein expression under hypoxia. Moreover, the expression of Cx43 was significantly increased by hypoxia but was decreased with salidroside pretreatment. The salidroside pretreated group exhibited enhanced release of intracellular Ca2+ in corpus cavernosum smooth muscle cells compared with the hypoxia group after stimulation. CONCLUSION: Salidroside has a protective effect against hypoxia-induced damage to corpus cavernosum smooth muscle cells.

Connexin43 Expression and Associated Chronic Inflammation Presages the Development of Cerebral Radiation Necrosis.

Cerebral radiation necrosis (CRN) is a delayed complication of radiosurgery that can result in severe neurological deficits. The biological changes leading to necrotic damage may identify therapeutic targets for this complication. Connexin43 expression associated with chronic inflammation may presage the development of CRN. A mouse model of delayed CRN was used. The left hemispheres of adult female mice were irradiated with single-fraction, high-dose radiation using a Leksell Gamma Knife. The brains were collected 1 and 4 days, and 1-3 weeks after the radiation. The expression of connexin43, interleukin-1ß (IL-1ß), GFAP, isolectin B-4, and fibrinogen was evaluated using immunohistochemical staining and image analysis. Compared with the baseline, the area of connexin43 and IL-1ß staining was increased in ipsilateral hemispheres 4 days after radiation. Over the following 3 weeks, the density of connexin43 gradually increased in parallel with progressive increases in GFAP, isolectin B-4, and fibrinogen labeling. The overexpression of connexin43 in parallel with IL-1ß spread into the affected brain regions first. Further intensified upregulation of connexin43 was associated with escalated astrocytosis, microgliosis, and blood-brain barrier breach. Connexin43-mediated inflammation may underlie radiation necrosis and further investigation of connexin43 hemichannel blockage is merited for the treatment of CRN.

An Alternatively Translated Connexin 43 Isoform, GJA1-11k, Localizes to the Nucleus and Can Inhibit Cell Cycle Progression.

Connexin 43 (Cx43) is a gap junction protein that assembles at the cell border to form intercellular gap junction (GJ) channels which allow for cell-cell communication by facilitating the rapid transmission of ions and other small molecules between adjacent cells. Non-canonical roles of Cx43, and specifically its C-terminal domain, have been identified in the regulation of Cx43 trafficking, mitochondrial preconditioning, cell proliferation, and tumor formation, yet the mechanisms are still being explored. It was recently identified that up to six truncated isoforms of Cx43 are endogenously produced via alternative translation from internal start codons in addition to full length Cx43, all from the same mRNA produced by the gene GJA1. GJA1-11k, the 11kDa alternatively translated isoform of Cx43, does not have a known role in the formation of gap junction channels, and little is known about its function. Here, we report that over expressed GJA1-11k, unlike the other five truncated isoforms, preferentially localizes to the nucleus in HEK293FT cells and suppresses cell growth by limiting cell cycle progression from the G0/G1 phase to the S phase. Furthermore, these functions are independent of the channel-forming full-length Cx43 isoform. Understanding the apparently unique role of GJA1-11k and its generation in cell cycle regulation may uncover a new target for affecting cell growth in multiple disease models.

Expression and functional regulation of gap junction protein connexin 43 in dermal mesenchymal stem cells from psoriasis patients.

BACKGROUND: Psoriasis is a chronic recurrent inflammatory disease. Mesenchymal stem cells (MSCs) can regulate the inflammatory microenvironment, thereby controlling the proliferation, differentiation, and migration of immune cells. Connexin 43(Cx43), a key gap junction protein, has been shown to form gap junctions for communication between neighboring cells. OBJECTIVE: We investigated the expression of Cx43 in dermal mesenchymal stem cells (DMSCs) derived from psoriasis patients and explored the relationship between the Cx43-mediated gap junction intercellular communication (GJIC) and DMSCs. METHODS: Human DMSCs were isolated and propagated in adherent culture. Quantitative real-time reverse transcription PCR and western blot and immunofluorescence were used to detect the expression and localization of Cx43 in DMSCs. Fluorescence redistribution after photobleaching was performed to assess adjacent DMSCs GJIC. CCK8 was used to detect the proliferation of DMSCs before and after gap junction blocker (18α-glycyrrhetinic acid; AGA) treatment. Cell energy metabolism was analyzed with an energy metabolism analyzer. RESULTS: Cx43 was located in the cytoplasm and cytomembrane, as well as partially in the nucleus of DMSCs. The expression of Cx43 in psoriasis DMSCs was higher than that in control samples and the gap junction function was enhanced. In addition, the glycolysis and mitochondrial respiration of psoriasis DMSCs were also enhanced. However, AGA inhibited the expression of Cx43, attenuated GJIC function, and inhibited the proliferation of DMSCs. CONCLUSIONS: Our results indicated that the expression of Cx43 in DMSCs from psoriasis lesions is increased and that the inhibition of Cx43 leads to the inhibition of both GJIC and DMSCs proliferation.

Connexin 43: A novel ginsenoside Rg1-sensitive target in a rat model of depression.

Our previous studies have shown that ginsenoside Rg1 (Rg1) exerts antidepressant-like effects in animal models of depression, accompanied by an improvement of astrocytic gap junction functions. However, whether connexin 43 (Cx43), the major connexin forming gap junctions between astrocytes, is the key regulator of Rg1-induced antidepressant-like effects is still unknown. In this study, we examine in vitro and in vivo the involvement of Cx43 in the antidepressant effects of Rg1. Corticosterone was used to establish an in vitro rat model of depression. Treatment with Rg1 1 h prior to corticosterone significantly improved the cell viability of astrocytes, which was significantly inhibited by carbenoxolone, a widely used gap junction inhibitor. Moreover, Rg1 treatment significantly ameliorated antidepressant-sensitive behaviours induced by infusion of carbenoxolone or Gap26, a selective inhibitor of Cx43, into the prefrontal cortex of the animals. Rg1 treatment increased the expression of Cx43 compared with Gap26 group. According to these results, the antidepressant-like effects of Rg1 were mainly mediated by Cx43-formed gap junctions.

Expression of Connexin 43 in 32 Cases of Merkel Cell Carcinoma.

INTRODUCTION: Connexins (Cxs) are channel proteins that allow direct connection among cells and between cells and the extracellular space. There is very little information in the literature on the expression of Cxs by Merkel cell carcinoma (MCC). MATERIALS AND METHODS: Thirty-two cases of MCC were recovered from our archives and studied immunohistochemically for Cx43. RESULTS: All our cases expressed several neuroendocrine markers. Most cases showed nonimmunohistochemically perceptible staining for Cx43. There was no difference between Merkel cell polyomavirus (MCPyV)-positive and MCPyV-negative cases. One case could not be evaluated. Only 2 cases showed a focal (10% of the tumor) membranous staining of Cx43. One of these cases was MCPyV-negative and, in the other, CM2B4 could not be evaluated. CM2B4 was positive in 18 cases and negative in 13 cases, and it could not be evaluated in 1 case. CONCLUSIONS: MCC shows a low Cx43 level, with no differences between MCPyV-positive and MCPyV-negative cases. Therefore, this opens the door for Cx43 targeting in therapeutic approaches to MCC.

Gap junctions and expression of Cx36, Cx43 and Cx45 in the posterodorsal medial amygdala of adult rats.

The posterodorsal medial amygdala (MePD) has an adapted synaptic organization that dynamically modulates reproduction and other social behaviors in rats. Discrete gap junctions between glial cells were previously reported in the MePD neuropil. Connexins (Cx) are components of gap junctions and indicative of cellular electrical coupling. Here, we report the ultrastructural occurrence of gap junctions between neurons in the MePD and demonstrate the expression and immunofluorescent labeling of Cx36, Cx43 and Cx45 in this subcortical area of adult male rats. Few neuronal gap junctions were found in the MePD and, when identified, occurred between dendrites. On the other hand, there is a diffuse presence and distribution of punctate labelling for the tested Cxs. Puncta were visualized isolated or forming clusters in the same focal plane of cell bodies or along the MePD neuropil. The Cx36 puncta were found in neurons, Cx43 in astrocytes and Cx45 in both neurons and astrocytes. Our data indicate the presence of few gap junctions and different Cxs composition in the MePD. Because Cxs can assemble, form hemichannel units and/or serve as transcriptional regulator, it is likely that additional modulation of intercellular communication can occur besides the chemical transmission in the MePD of adult rats.

The Effects of Fluoride on the Gap-Junctional Intercellular Communication of Rats' Osteoblast.

The gap junction protein plays an important role in the bone formation and alteration of these proteins leading to cause bone development. Aim to determine the effects of different concentration of fluoride on gap-junctional intercellular communication (GJIC) related genes and proteins in the rats' osteoblast cells. We treated the osteoblast cells with various concentrations (0, 0.01, 0.1, 0.5, and 1.0 mM) NaF for 24 and 72 h. We used the scrape loading and dye transfer technique to research the intracellular connectivity. Moreover, the mRNA expression levels of connexin 43 (Cx43), connexin45 (Cx45), collagen I, and osteocalcin (OCN) were analyzed by qRT-PCR, the protein expression levels of connexin43 (Cx43) were analyzed by western blotting and immunofluorescence. Our results suggested that the osteoblast proliferations were decreased in the 0.5 and 1 mM NaF groups, after 24 and 72 treatments. The scrape loading and dye transfer experiment showed that the GJIC were increased in the 0.01 mM NaF group and decreased in the 0.5 and 1 mM NaF groups. In addition, the mRNA expressions of Cx43, Cx45, and OCN, and the protein expressions of Cx43 were increased in the 0.01 mM NaF group and decreased in the 0.5 and 1 mM NaF groups. In summary, these results suggest that the low concentration NaF is good for the GJIC, but the high concentration NaF damages the GJIC.

The Interplay Between E-Cadherin, Connexin 43, and Zona Occludens 1 in Retinal Pigment Epithelial Cells.

Purpose: Cell-cell contact in retinal pigment epithelium (RPE) involves adherent junctions, gap junctions, and tight junctions, which are primarily composed by E-cadherin, zona occludens 1 (ZO-1), and connexin 43, respectively. Here, we aimed to explore the relationship and interplay between these junction-associated proteins. Methods: E-cadherin, connexin 43, and ZO-1 expression in human primary RPE in the early phase after TGF-ß1 stimulation was detected. The knockdown of E-cadherin, ZO-1, and connexin 43 was performed to characterize the regulatory network involving these three proteins. Dye transfer and FITC-dextran permeability assays were conducted to observe the epithelial functional alterations. Transmission electron microscopy (TEM) was used to observe the ultrastructure of the cell-cell junctions in mouse RPE. The immunofluorescence staining and coimmunoprecipitation were performed to observe the colocalization and the physical association of E-cadherin, ZO-1, and connexin 43. Results: Among these three components, E-cadherin appeared to be the first protein that was downregulated after TGF-ß1 treatment. The ultrastructures of adherent junctions, gap junctions, and tight junctions could be observed in mouse RPE by TEM. E-cadherin, ZO-1, and connexin 43 were colocalized and physically bound to each other. The knockdown of one of these three proteins led to downregulation of the other two proteins and compromised epithelial function. Conclusions: E-cadherin, ZO-1, and connexin 43 were physically associated with each other and were mutually regulated. To enhance the understanding of cell-cell contacts, a holistic view is needed. Our results provide new insights in RPE disorders such as proliferative vitreoretinopathy.

All-trans retinoic acid exposure increases connexin 43 expression in cumulus cells and improves embryo development in bovine oocytes.

In developing follicles, cellular coupling within cumulus-oocyte complexes (COCs) creates a functional syncytium allowing for the passage of small molecules. In many species, intercellular coupling between granulosa cells results from the expression of connexin 43 (CX43 or Gja1) and the formation of gap junctional plaques. Previously, our lab has shown that oocytes with a higher developmental potential had higher CX43 expression in their cumulus cells compared with developmentally incompetent oocytes. All-trans retinoic acid (ATRA) has been shown to increase CX43 expression in several different cell types. In this study we investigated the effect of ATRA treatment, during maturation, on CX43 expression and localization in cumulus cells and the developmental competence of bovine oocytes. COCs and granulosa cells exposed to ATRA during maturation had significantly higher CX43 expression and increased gap junctional coupling, respectively. In addition, there was a significant increase in the maturation, cleavage, and blastocyst rates in ATRA treated COCs. Data from these studies suggest that not only can CX43 be used as a biomarker for oocyte health, it can also potentially be manipulated using ATRA to increase the number of oocytes achieving developmental competence.

Cell-free synthesis of connexin 43-integrated exosome-mimetic nanoparticles for siRNA delivery.

Exosomes are naturally secreted nanovesicles that have emerged as a promising therapeutic nanodelivery platform, due to their specific composition and biological properties. However, challenges like considerable complexity, low isolation yield, drug payload, and potential safety concerns substantially reduce their pharmaceutical acceptability. Given that the nano-bio-interface is a crucial factor for nanocarrier behavior and function, modification of synthetic nanoparticles with the intrinsic hallmarks of exosomes' membrane to create exosome mimetics could allow for siRNA delivery in a safer and more efficient manner. Herein, connexin 43 (Cx43)-embedded, exosome-mimicking lipid bilayers coated chitosan nanoparticles (Cx43/L/CS NPs) were constructed by using cell-free (CF) synthesis systems with plasmids encoding Cx43 in the presence of lipid-coated CS NPs (L/CS NPs). The integration of de novo synthesized Cx43 into the lipid bilayers of L/CS NPs occurred cotranslationally during one-pot reaction and, more importantly, the integrated Cx43 was functionally active in transport. In addition to considerably lower cytotoxicity (

Overexpression of miR-27b-3p Targeting Wnt3a Regulates the Signaling Pathway of Wnt/ß-Catenin and Attenuates Atrial Fibrosis in Rats with Atrial Fibrillation.

MicroRNAs (miRNAs) are regarded as a potential method for the treatment of atrial fibrillation (AF) although its molecular mechanism remains unknown. We found in our previous study that the level of peripheral blood miR-27b-3p and the expression of atrial tissue CX43 were both significantly downregulated in AF patients. In the present study, we propose and test this hypothesis that overexpression of miR-27b-3p attenuates atrial fibrosis, increases CX43 expression, and regulates the signaling pathway of Wnt/ß-Catenin by targeting Wnt3a. miR-27b-3p overexpression was induced by rat tail vein injection of adeno-associated virus. Two weeks after transfection of adeno-associated virus, the rat AF model was established by tail vein injection of acetylcholine- (ACh-) CaCl2 for 7 days, and 1 ml/kg was injected daily. The incidence and duration of AF were recorded with an electrocardiogram. Cardiac function was monitored by cardiac ultrasound. Serum cardiac enzyme was detected by ELISA. The expression of atrial miR-27b-3 and Wnt3a was assayed by quantitative RT-PCR. Atrial fibrosis was determined by Masson's trichrome staining. Expression of atrial Collagen-I and Collagen-III was tested by the immunohistochemical method. Expression of CX43 was measured by immunofluorescence. The expression of Collagen-I, a-SMA, Collagen-III, TGF-ß1, CX43, Wnt3a, ß-Catenin, and p-ß-Catenin was assayed by western blot. Our results showed that miR-27b-3p overexpression could reduce the incidence and duration of AF, alleviate atrial fibrosis, increase atrial CX43 expression, and decrease the expression of Collagen-I, a-SMA, Collagen-III, TGF-ß1, Wnt3a, and p-ß-Catenin. In addition, the results of luciferase activity assay showed that Wnt3a is a validated miR-27b-3p target in HEK 293T cells. Our results provide a new evidence that miR-27b-3p regulates the signaling pathway of Wnt/ß-Catenin by targeting Wnt3a, which may play an important role in the development of atrial fibrosis and AF.

Expression of Connexin 43 (Cx43) in Benign Cutaneous Tumors With Follicular Differentiation.

INTRODUCTION: Benign cutaneous tumors with follicular differentiation are alleged to differentiate toward parts of the hair follicle. Connexin 43 (Cx43) is a gap junction protein, the tumoral role of which has been investigated in several types of tumors. OBJECTIVE: To study the pattern of expression of Cx43 in benign cutaneous tumors with follicular differentiation and to compare it with that shown by their alleged anatomical counterparts of the hair follicle. MATERIALS AND METHODS: Five cases each of trichofolliculoma, trichilemmoma, fibrofolliculoma/trichodiscoma, trichoblastoma, trichoepithelioma, pilomatrixoma, and proliferating trichilemmal tumor, 3 cases of pilar sheath acanthoma, and 1 case of tumor of the follicular infundibulum were examined. Anti-Cx43 antibody was used. RESULTS: Cx43 was expressed by all follicular tumors studied. Comparisons between trichoblastoma and trichoepithelioma and their respective normal counterparts could not be made. In 3 tumors (trichofolliculoma, pilomatrixoma, and the spectrum fibrofolliculoma/trichodiscoma), there was a parallelism between their Cx43 expression pattern and that of their alleged anatomical counterparts. In pilar sheath acanthoma, trichilemmoma, and the tumor of the follicular infundibulum, we only found partial similarities in Cx43 expression. Only the proliferating trichilemmal tumor showed a discordant pattern of expression. CONCLUSIONS: Cx43 expression is preserved in benign cutaneous tumors with follicular differentiation and the patterns of Cx43 expression in benign cutaneous tumors with follicular differentiation parallel those of their alleged anatomical counterparts in 5 types (either totally or partially). This preservation might be related to the good behavior of the entities studied.

A Logistic Regression Model for Detecting the Presence of Malignant Progression in Atypical Meningiomas.

OBJECTIVE: To develop a method to distinguish atypical meningiomas (AMs) with malignant progression (MP) from primary AMs without a clinical history. METHODS: The clinical, radiologic, and pathologic data of 33 previously Simpson grade I resected (if any) as well as no radiotherapy treated intracranial AMs between January 2008 and December 2015 were reviewed. Immunohistochemical staining for connexin 43 (Cx43) and Ki-67 was performed. Descriptive analysis and univariate and multivariate logistic regression analyses were used to explore independent predictors of MP. A multivariable logistic model was developed to estimate the risk of MP, and its diagnostic value was determined from a receiver operating characteristic curve. RESULTS: There were 11 AMs (33.3%) with histopathologically confirmed MP from benign meningiomas. The other 22 (66.7%) were initially diagnosed AMs with no histopathologically confirmed MP during a median 60.5 months (range, 42-126 months) of follow-up. Univariate and multivariate logistic analyses showed that irregular tumor shape (P = 0.010) and low Cx43 expression (P = 0.010) were independent predictors of the presence of MP, and the predicted probability was calculated by the following formula: P = 1/[1+exp.{1.218-(3.202×Shape)+(3.814×Cx43)}]. P > 0.5 for an irregularly shaped (score 1) AM with low Cx43 expression (score 0) indicated a high probability of MP. The sensitivity, specificity, positive predictive value, negative predictive value, and overall predictive accuracy were 63.6, 95.6, 87.5, 84.0, and 84.8%, respectively. CONCLUSIONS: Low Cx43 expression and irregular tumor shape were independent predictors of the presence of MP. The relevant logistic regression model was found to be effective in distinguishing MP-AMs from primary AMs.