Expression Of Connexin Research Articles

Glioma–Astrocyte connexin43 confers temozolomide resistance through activation of the E2F1/ERCC1 axis

Abstract Background Glioma is the most prevalent and lethal tumor of the central nervous system. Routine treatment with Temozolomide (TMZ) would unfortunately result in inevitable recurrence and therapy resistance, severely limiting therapeutic efficacy. Tumor associated astrocytes (TAAs) are key components of the tumor microenvironment and increasing evidence has demonstrated that aberrant expression of Connexin43 (Cx43) was closely associated with glioma progression and TMZ resistance. However, the specific role of Cx43 in mediating TMZ resistance through glioma and astrocyte interactions has not been fully explored. Methods The expression and prognostic value of Cx43 were evaluated in tumor samples and clinical databases. ShRNA-medicated knockdown and Gfap-Cre Cx43flox/flox gene mouse were used to assessed the role and functional significance of Cx43 in vitro and in vivo. Moreover, we performed mass spectrometry analysis, chromatin immunoprecipitation, and other biochemical assays to define the molecular mechanisms by which Cx43 promotes TMZ resistance. Results We confirmed that upregulation of Cx43 expression between TAAs and glioma cells contributed to TMZ resistance and tumor recurrence. Genetic knockdown or pharmacological inhibition of Cx43 enhanced TMZ-induced cytotoxicity. Mechanistically, elevated Cx43 expression induced β-catenin accumulation at the cell surface of glioma cells, suppressing TCF/LEF transcription, This led to impaired miR-205-5p expression and subsequent activation of E2F1/ERCC1 axis, which eventually led to chemoresistance. Conclusions Our study reveals a novel regulatory mechanism in which the Cx43/miR-205-5p/E2F1/ERCC1 axis contributes to TMZ resistance in glioma. These findings further highlight the potential of targeting Cx43 as a therapeutic strategy in glioma.

Nppb contributes to Sepsis-Induced myocardial injury by regulating Senescence-Related genes

BackgroundSepsis, a systemic inflammatory condition, is a leading cause of mortality due to cardiovascular injury. Sepsis and cellular senescence are closely related, yet the specific mechanisms are still unclear. This study aims to identify a novel therapeutic target for mitigating sepsis-induced myocardial injury. MethodsWe initially assessed serum inflammatory markers and myocardial injury indicators in septic mice. This involved observing inflammatory cell infiltration in ventricular muscle tissue, assessing cardiac function, and recording electrocardiograms. We examined the expression of connexin 40 (Cx40) and connexin 43 (Cx43) and analyzed mitochondrial structures in ventricular tissues. A conditional heart-specific Nppb knockout mouse model was then developed in mice to evaluate these changes. Ventricular tissues were analyzed via mRNA sequencing to identify differentially expressed genes (DEGs), which were cross-referenced with senescence-related genes to identify key hub genes. The expression and distribution of hub genes were subsequently analyzed by single-cell analysis. Finally, the expression of the senescence-related gene CCL2, along with cardiac structure and function, was validated in a conditional heart-specific Nppb knockout sepsis mouse model. ResultsMyocardial injury severity positively correlated with serum brain natriuretic peptide (BNP) in septic mice. Conditional heart-specific Nppb knockout improved myocardial injury outcomes in mice with sepsis. The DEGs identified in the conditional heart-specific Nppb knockout model overlapped with senescence-related genes, identifying seven upregulated genes associated with inflammation, cardiac contraction and apoptotic pathways. Single-cell analysis confirmed high CCL2 levels and an increased macrophage presence correlating with sepsis progression. Conditional heart-specific Nppb knockout reduced CCL2 levels, which were associated with improved cardiac structure and function. ConclusionThis study confirms that conditional heart-specific Nppb knockout reduces CCL2 expression, thereby ameliorating myocardial injury in septic mice. Targeting Nppb to regulate the senescence-related gene CCL2 may represent an effective strategy for treating myocardial injury in septic patients.

Macrophages enhance sodium channel expression in cardiomyocytes.

Cardiac macrophages facilitate electrical conduction through the atrioventricular-node (AV) in mice. A possible role for cardiomyocyte-macrophage coupling on the effect of antiarrhythmic therapy has not been investigated yet. Holter monitoring was conducted in LysMCrexCsf1rLsL-DTR mice (MMDTR) under baseline conditions and after an elctrophysiological stress test by flecainide. In vivo effects were recapitulated in vitro by patch-clamp experiments. The underlying mechanism was characterized by expression and localization analysis of connexin43 (Cx43) and voltage-gated-sodium-channel-5 (Nav1.5). ECG monitoring in MMDTR mice did not show any significant conduction abnormalities but a significantly attenuated flecainide-induced extension of RR- and PP-intervals. Patch-clamp analysis revealed that the application of flecainide to neonatal rat ventricular cardiomyocytes (CMs) changed their resting-membrane-potential (RMP) to more negative potentials and decreased action-potential-duration (APD50). Coupling of macrophages to CMs significantly enhances the effects of flecainide, with a further reduction of the RMP and APD50, mediated by an upregulation of Cx43 and Nav1.5 surface expression. Macrophage depletion in mice does not correlate with cardiac electric conduction delay. Cardiac macrophages amplify the effects of flecainide on electrophysiological properties of cardiomyocytes in vivo and in vitro. Mechanistically, formation of macrophage-cardiomyocyte cell-cell-contacts via Cx43 facilitates the recruitment of Nav1.5 to the cell membrane increasing flecainide effects.

Morphological, electrophysiological, and molecular alterations in foetal noncompacted cardiomyopathy induced by disruption of ROCK signalling.

Left ventricular noncompaction cardiomyopathy is associated with heart failure, arrhythmia, and sudden cardiac death. The developmental mechanism underpinning noncompaction in the adult heart is still not fully understood, with lack of trabeculae compaction, hypertrabeculation, and loss of proliferation cited as possible causes. To study this, we utilised a mouse model of aberrant Rho kinase (ROCK) signalling in cardiomyocytes, which led to a noncompaction phenotype during embryogenesis, and monitored how this progressed after birth and into adulthood. The cause of the early noncompaction at E15.5 was attributed to a decrease in proliferation in the developing ventricular wall. By E18.5, the phenotype became patchy, with regions of noncompaction interspersed with thick compacted areas of ventricular wall. To study how this altered myoarchitecture of the heart influenced impulse propagation in the developing and adult heart, we used histology with immunohistochemistry for gap junction protein expression, optical mapping, and electrocardiography. At the prenatal stages, a clear reduction in left ventricular wall thickness, accompanied by abnormal conduction of the ectopically paced beat in that area, was observed in mutant hearts. This correlated with increased expression of connexin-40 and connexin-43 in noncompacted trabeculae. In postnatal stages, left ventricular noncompaction was resolved, but the right ventricular wall remained structurally abnormal through to adulthood with cardiomyocyte hypertrophy and retention of myocardial crypts. Thus, this is a novel model of self-correcting embryonic hypertrabeculation cardiomyopathy, but it highlights that remodelling potential differs between the left and right ventricles. We conclude that disruption of ROCK signalling induces both morphological and electrophysiological changes that evolve over time, highlighting the link between myocyte proliferation and noncompaction phenotypes and electrophysiological differentiation.

Se-Methylselenocysteine Ameliorates DEHP-Induced Ferroptosis in Testicular Sertoli Cells via the Nrf2/GPX4 Axis.

Di (2-ethylhexyl) phthalate (DEHP) is an important plasticizer in industrial production, and its toxic effects on testes are widely recognized. Se-methylselenocysteine (SMC) is a major selenium compound found in selenium-rich plants, which possesses unique biological properties such as antioxidants. However, the effect of SMC on DEHP-induced testicular injury and the specific mechanism remains unknown. In this study, 50 mice were randomly divided into 5 groups and were given corn oil (Control), DEHP, low-dose SMC (L-SMC), moderate-dose SMC (M-SMC), or high-dose SMC (H-SMC). The sperm quality of the mice in each group was determined, and HE staining and transmission electron microscopy (TEM) were applied to observe testicular morphology, and testicular tissues were collected for the subsequent molecular biological analyses. The TM4 cell line was applied in vitro for mechanism validation. Our results showed that DEHP could lead to decreased sperm quality and blood-testis barrier damage in mice, which could be alleviated by SMC. Mitochondrial damage accompanied by accumulation of total iron content, MDA, and 4-HNE, as well as downregulation of antioxidants SOD, GSH, and GSH-Px were observed after DEHP treatment, which exhibited a typical ferroptosis feature. In vitro experiments confirmed that SMC promoted upregulation of GPX4 in TM4 cells and was able to alleviate DEHP metabolite MEHP-induced ferroptosis and promote the expression of cell junction key proteins ZO-1, Occludin, and Connexin 43, which could be inhibited by the GPX4 inhibitor RSL3 or the Nrf2 inhibitor ML385. Overall, the above results suggest that SMC ameliorates the DEHP-induced ferroptosis in testicular Sertoli cells, protects the blood-testis barrier, and prevents sperm aberrations via the Nrf2/GPX4 axis.

Calmodulin kinase II inhibition suppresses atrioventricular conduction by regulating intracellular Ca2+ homeostasis

BackgroundCa2+/calmodulin-dependent protein kinase II (CaMKII) inhibition decelerates atrioventricular node (AVN) conduction, providing a potential treatment of tachycardia. However, the effectiveness of CaMKII inhibition on tachycardia and its underlying mechanism remains unclear. ObjectiveWe aimed to assess the effectiveness of CaMKII inhibition in reducing ventricular rates during atrial fibrillation and to elucidate the underlying mechanism in affecting AVN electrophysiology. MethodsCardiac CaMKII inhibition (AC3-I) mice were used. Transesophageal atrial pacing was performed to evaluate AVN conduction function and to induce atrial fibrillation. Patch-clamp techniques were employed to record action potentials and ionic currents in AVN cells. Intracellular Ca2+ transients and sarcomere length measurements were obtained with the IonOptix system. Masson trichrome stain was used to evaluate fibrosis in the AVN region. Western blotting and immunofluorescence techniques were employed to detect connexin expression and localization. ResultsCaMKII inhibition decreased the ventricular rate during atrial fibrillation and isoproterenol-induced tachycardia. Esophageal electrocardiogram results from AC3-I mice showed longer AVN conduction than in wild-type mice. AN- and N-type AVN cells from AC3-I mice exhibited slower action potential frequencies and diastolic depolarization rates than those of wild-type mice. The study revealed that CaMKII inhibition reduced AVN cell sarcoplasmic reticulum (SR) Ca2+ content, Ca2+ release rate from the SR during diastole, Ca2+ transient amplitude, and SR Ca2+ uptake rate. In addition, CaMKII inhibition prolonged the sarcomere diastole duration and enhanced the sensitivity of sarcomeres to Ca2+. ConclusionCaMKII inhibition effectively decreases the ventricular rate during atrial fibrillation and tachycardia by slowing down AVN conduction through suppressing Ca2+ overload in AVN cells.

Reactive astrocytes generated from human iPSC are pro-inflammatory and display altered metabolism

Astrocytes are the most abundant type of glial cell in the central nervous system and they play pivotal roles in both normal health and disease. Their dysfunction is detrimental to many brain related pathologies. Under pathological conditions, such as Alzheimer's disease, astrocytes adopt an activated reactive phenotype which can contribute to disease progression. A prominent risk factor for many neurodegenerative diseases is neuroinflammation which is the purview of glial cells, such as astrocytes and microglia. Human in vitro models have the potential to reveal relevant disease specific mechanisms, through the study of individual cell types such as astrocytes or the addition of specific factors, such as those secreted by microglia. The aim of this study was to generate human cortical astrocytes, in order to assess their protein and gene expression, examine their reactivity profile in response to exposure to the microglial secreted factors IL-1α, TNFα and C1q and assess their functionality in terms of calcium signalling and metabolism. The successfully differentiated and stimulated reactive astrocytes display increased IL-6, RANTES and GM-CSF secretion, and increased expression of genes associated with reactivity including, IL-6, ICAM1, LCN2, C3 and SERPINA3. Functional assessment of these reactive astrocytes showed a delayed and sustained calcium response to ATP and a concomitant decrease in the expression of connexin–43. Furthermore, it was demonstrated these astrocytes had an increased glycolytic capacity with no effect on oxidative phosphorylation. These findings not only increase our understanding of astrocyte reactivity but also provides a functional platform for drug discovery.

CYLD Alterations Are Associated With Metastasis and Poor Prognosis in Human Papilloma Virus-Positive Head and Neck Cancer.

Human papilloma virus (HPV)-associated head and neck squamous cell carcinoma (HNSCC) is an emerging epidemic and a subset of HPV-positive patients experience aggressive disease with metastases. The CYLD gene is frequently altered in HPV-positive HNSCC, but the role of these alterations in disease progression is poorly understood. We identified 11 HPV-positive HNSCC patients with CYLD alterations and assessed their clinical course. We also characterized a unique, HPV-positive, metastatic, HNSCC patient-derived xenograft (PDX). All 11 patients developed metastasis with reduced overall survival when compared with metastatic HPV-positive patients with wild-type CYLD. The metastatic PDX harbored a CYLD mutation (S371*) and exhibited reduced expression of connexin 43, a potentially antimetastatic protein. We also investigated the functional impact of the S371* mutation, as well as 2 CYLD mutations from our 11-patient cohort. Our findings indicate that alterations in CYLD in HPV-positive HNSCC are associated with metastasis and poor prognosis.

Microgravity as a Tool to Investigate Cancer Induction in Pleura Mesothelial Cells.

The present work shows that the exposure of mesothelial cells to simulated microgravity changes their cytoskeleton and adhesion proteins, leading to a cell switch from normal towards tumoral cells. Immunohistochemical and molecular data were obtained from both MeT-5A exposed to simulated microgravity and BR95 mesothelioma cell lines. Simulated microgravity was found to affect the expression of actin, vinculin, and connexin-43, altering their quantitative and spatial distribution pattern inside the cell. The analysis of the tumoral markers p27, CD44, Fibulin-3, and NANOG and the expression of genes related to cancer transformation such as NANOG, CDH-1, and Zeb-1 showed that the simulated microgravity environment led to expression patterns in MeT-5A cells similar to those observed in BR95 cells. The alteration in both quantitative expression and structural organization of the cytoskeleton and adhesion/communication proteins can thus be considered a pivotal mechanism involved in the cellular shift towards tumoral progression.

ROS responsive conductive microspheres loaded with salvianolic acid B as adipose derived stem cell carriers for acute myocardial infarction treatment

Stem cell therapy is currently the most promising strategy for the treatment of myocardial infarction. However, the development of injectable cell carriers that can scavenge reactive oxygen species (ROS) in the infarct zone to improve transplanted cell survival remains a challenge. Here, we developed a ROS responsive conductive microsphere based on chitosan (CS) and dextran (DEX) with 4-formylphenylboronic acid (4-FPBA) as a cross-linking agent and the addition of graphite oxide (GO) and the anti-inflammatory agent salvianolic acid B (SalB), as a cell delivery carrier for myocardial infarction. These microspheres were crosslinked by dual dynamic networks of Schiff base and phenylborate bonds. The relationship between CS concentration and microsphere particle size, as well as the biocompatibility, ROS responsiveness, anti-inflammatory properties, and effects on myogenic differentiation of H9C2 cells were fully investigated. The microspheres exhibit good biocompatibility, proliferation promoting, differentiation promoting, antioxidant, and anti-inflammatory properties. When applied to mice myocardial infarction models, the ROS responsive conductive microspheres loaded with SalB and adipose derived stem cells (ADSC) exhibited excellent in vivo repair ability. In addition, they reduced myocardial fibrosis and promoted ventricular wall regeneration by promoting the expression of Connexin 43 (Cx43) and CD31, ultimately reshaping the infarcted myocardium, suggesting their great potential as cell delivery carriers for myocardial infarction treatment.

Connexin Expression Is Altered in the Eye Development of Yotari Mice: A Preliminary Study.

This study aimed to explore how Dab1 functional silencing influences the expression patterns of different connexins in the developing yotari (yot) mice eyes as potential determinants of retinogenesis. Using immunofluorescence staining, the protein expression of Dab1, Reelin, and connexin 37, 40, 43, and 45 (Cx37, Cx40, Cx43, and Cx45) in the wild-type (wt) and yot eyes at embryonic days 13.5 and 15.5 (E13.5 and E15.5) were analyzed. Different expression patterns of Cx37 were seen between the wt and yot groups. The highest fluorescence intensity of Cx37 was observed in the yot animals at E15.5. Cx40 had higher expression at the E13.5 when differentiation of retinal layers was still beginning, whereas it decreased at the E15.5 when differentiation was at the advanced stage. Higher expression of Cx43 was found in the yot group at both time points. Cx45 was predominantly expressed at E13.5 in both groups. Our results reveal the altered expression of connexins during retinogenesis in yot mice and their potential involvement in retinal pathology, where they might serve as prospective therapeutic targets.

Neurotropic Murine β-Coronavirus Infection Causes Differential Expression of Connexin 47 in Oligodendrocyte Subpopulations Associated with Demyelination.

Gap junctions (GJs) play a crucial role in the survival of oligodendrocytes and myelination of the central nervous system (CNS). In this study, we investigated the spatiotemporal changes in the expression of oligodendroglial GJ protein connexin 47 (Cx47), its primary astroglial coupling partner, Cx43, and their association with demyelination following intracerebral infection with mouse hepatitis virus (MHV). Neurotropic strains of MHV, a β-coronavirus, induce an acute encephalomyelitis followed by a chronic demyelinating disease that shares similarities with the human disease multiple sclerosis (MS). Our results reveal that Cx47 GJs are persistently lost in mature oligodendrocytes, not only in demyelinating lesions but also in surrounding normal appearing white and gray matter areas, following an initial loss of astroglial Cx43 GJs during acute infection. At later stages after viral clearance, astroglial Cx43 GJs re-emerge but mature oligodendrocytes fail to fully re-establish GJs with astrocytes due to lack of Cx47 GJ expression. In contrast, at this later demyelinating stage, the increased oligodendrocyte precursor cells appear to exhibit Cx47 GJs. Our findings further highlight varying degrees of demyelination in distinct spinal cord regions, with the thoracic cord showing the most pronounced demyelination. The regional difference in demyelination correlates well with dynamic changes in the proportion of different oligodendrocyte lineage cells exhibiting differential Cx47 GJ expression, suggesting an important mechanism of progressive demyelination even after viral clearance.

Connexin 43 controls metastatic behavior in triple negative breast cancer through TGFβ1-Smad3-intergin αV signaling axis Based on optical image diagnosis

Abnormal expression of connexin 43 (Cx43) contributes to the development and progression of cancer. However, its regulation is complex and dependent on the environment. The expression of Cx43 in triple-negative cancer lesions was analyzed by immunohistochemistry and optical coherence tomography using experimental models and clinical samples. The model of TGFβ1-SMad3-in-αv signal axis was established and verified by experiments. The results show that Cx43 plays a key role in the regulation of triple-negative cancer metastasis. In vivo, over-expressed Cx43 decreased tumor volume and inhibited ITGαV, TGF-β1, Smad3 and N-cadherin expressions, but enhanced the E-cadherin. Cx43 had the lowest expression in the TNBC samples, especially in lymph node metastatic TNBC patients and had a negative correlation with ITG alpha V, TGF-β1 and Smad3.The study demonstrated Cx43 controlled metastatic behavior through TGF-β1 -Smad3-ITG αV signaling axis in MDA-MB-231 cells, providing evidence for Cx43’s function in TNBC. The optical image diagnosis method can realize the identification and quantitative evaluation of early cancer triple negative, and provide a new strategy and means for the treatment of cancer triple negative.

Gap junction protein connexin 43 and its distribution in damaged nerve cells

BACKGROUND: Connexins, particularly connexin 43, form gap junctions that mediate neuron-glial communication. To date, the expression of connexin 43 in phenotypically distinct neurolemmocytes, in particular non-myelinating (Remak cells) and repair Schwann cells, has not been studied. AIM: To determine the presence of connexin 43 in Schwann cells of the rat sciatic nerve under normal conditions and after mechanical damage by applying a ligature. MATERIALS AND METHODS: The study was carried out on Wistar rats (n = 10). In experimental rats, the sciatic nerve was damaged by applying a ligature (40 s). 7 days after surgery, sciatic nerve segments were isolated for subsequent immunohistochemical study using antibodies to connexin 43 and glial fibrillary acidic protein (GFAP). In the control group rats, segments of intact sciatic nerves were isolated in a similar way. RESULTS: It has been shown that in the endoneurium of the intact rat sciatic nerve there are no cells expressing connexin 43. It was found that 7 days after injury, a large number of connexin-43-immunopositive cells of irregular shape with several processes were identified in the endoneurium of the damaged nerve. There was a lack of expression of connexin 43 in GFAP-containing Schwann cells. CONCLUSIONS: It can be stated that nerve damage leads to active synthesis of the studied protein by endoneurium cells; however, the origin of the cells expressing connexin 43 remains to be elucidated.

Calcium Role in Gap Junction Channel Gating: Direct Electrostatic or Calmodulin-Mediated?

The chemical gating of gap junction channels is mediated by cytosolic calcium (Ca2+i) at concentrations ([Ca2+]i) ranging from high nanomolar (nM) to low micromolar (µM) range. Since the proteins of gap junctions, connexins/innexins, lack high-affinity Ca2+-binding sites, most likely gating is mediated by a Ca2+-binding protein, calmodulin (CaM) being the best candidate. Indeed, the role of Ca2+-CaM in gating is well supported by studies that have tested CaM blockers, CaM expression inhibition, testing of CaM mutants, co-localization of CaM and connexins, existence of CaM-binding sites in connexins/innexins, and expression of connexins (Cx) mutants, among others. Based on these data, since 2000, we have published a Ca2+-CaM-cork gating model. Despite convincing evidence for the Ca2+-CaM role in gating, a recent study has proposed an alternative gating model that would involve a direct electrostatic Ca2+-connexin interaction. However, this study, which tested the effect of unphysiologically high [Ca2+]i on the structure of isolated junctions, reported that neither changes in the channel's pore diameter nor connexin conformational changes are present, in spite of exposure of isolated gap junctions to [Ca2+]i as high at the 20 mM. In conclusion, data generated in the past four decades by multiple experimental approaches have clearly demonstrated the direct role of Ca2+-CaM in gap junction channel gating.

O101 THE ANGIOTENSIN II TYPE 1 RECEPTOR-BIASED AGONIST REDUCES BLOOD PRESSURE IN SPONTANEOUSLY HYPERTENSIVE RATS AND RESTORES VENTRICULAR HYPERTROPHY BY AFFECTING THE CONNEXIN 43 PATHWAY

Purpose: Hypertension is one of the common causes of cardiovascular diseases. Long-term hypertension can lead to structural and functional impairment of the heart. Therefore, active treatment of hypertension is highly significant in preventing target organ damage. Angiotensin II (AngII) plays a crucial role in hypertension-induced cardiac hypertrophy. Reportedly, TRV027, an AngII type 1 receptor (AT1R) biased agonist, can competitively bind to AT1R and inhibit the AngII-induced damage. Whether TRV027 could lower blood pressure and treat hypertensive cardiac hypertrophy remains unknown. Methods: Spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats were selected, and some of the SHRs were administered with TRV027. H9C2 cells were utilized for in vitro study to explore the underlying mechanism of TRV027. A multidisciplinary approach combining pathology and molecular techniques was used to explore the role and mechanism of TRV027 in reducing blood pressure and restoring ventricular hypertrophy. Results: Our studies revealed that TRV027 reduced blood pressure and restored hypertension-induced ventricular hypertrophy in SHRs. Moreover, the modulated expression of Connexin43 (Cx43) and phosphorylation of the MAPK/ERK pathway were normalized by TRV027. After the inhibition of Cx43 expression, cardiomyocytes recovered from hypertrophy and damage, followed by the modulation of phosphorylation of the MAPK/ERK pathway. Conclusion: TRV027 demonstrates a potential to lower the blood pressure in SHRs and ameliorate hypertension-induced cardiac hypertrophy. While AngII stimulated cardiomyocyte injury by decreasing Cx43 expression and increasing the phosphorylation of the MAPK/ERK pathway, TRV027 repaired the hypertrophy by restoring the expression of the Cx43/MAPK/ERK pathway.

Effects of prenatal stress on reproductive function of male offspring through the KISS1 system.

Prenatal stress can lead to the programming of the neuroendocrine system in male offspring, disrupting the hypothalamic testicular axis and adversely affecting the reproductive health of male offspring. This study aimed to determine the long-term effects of prenatal stress on the KISS1 system in male offspring and the effects on reproductive function in male offspring. Sixteen pregnant females were divided into a prenatal control group (PC, n = 8) and a prenatal stress group (PS, n = 8). The PS group was modeled with chronic unpredictable mild stress (CUMS) from day 1 of gestation to full-term delivery. Differences between the two groups in various maternal parameters, including glucocorticoid secretion, litter size, and the effects of male offspring birth weight, the KISS1 system, and reproductive function, were determined. Male offspring of PS dams had lower birth weights compared to prenatal controls.KISS1 gene expression is reduced at birth and in adult PS offspring, and its receptor KISS1-R protein is similarly reduced in PS offspring at birth and adulthood. In adulthood, PS male offspring show significantly reduced sex hormone production, altered testicular morphology, reduced maturation of their supporting cells, and decreased expression of connexin 43 (CX43), leading to an altered sperm microenvironment and reduced sperm quality. In conclusion, prenatal stress leads to adverse changes in the KISS1 system in male offspring and decreased reproductive function.

Tamoxifen-induced alterations in the expression of connexin 43 in the chicken ovary

Connexin 43 (Cx43) is a gap junction protein that participates in small molecule exchange between adjacent cells. It is a predominant Cx within the mammalian ovary, where is associated with proper follicle development. The expression and regulation of Cx43 in the chicken ovary is largely unknown. The aim of the present study was to examine the expression of the Cx43 gene (GJA1) and protein as well as the immunolocalization of Cx43 in the laying hen ovary in relation to follicle development, and to examine how tamoxifen (TMX; an estrogen receptor modulator) treatment affects these factors. qRT-PCR and western blotting demonstrated differences in Cx43 mRNA transcript and protein abundances in ovarian white follicles, yellowish follicles, small yellow follicles, and the largest yellow preovulatory follicles (F3–F1). In general, Cx43 was more abundant in hierarchical than prehierarchical follicles and in granulosa cells compared with theca cells. Further, the response to TMX treatment depended on the stage of follicle development and the layer of the follicular wall. Ovarian regression following TMX treatment was accompanied by an increase in Cx43 expression in most ovarian tissues, which may impact the formation and function of Cx43 hemichannels. Overall, our results showed, for the first time, the differences in Cx43 mRNA and protein levels between ovarian follicles, suggesting the potential involvement of this gap junction protein in the regulation of ovarian follicle development and function. In addition, the results indicate a possible role for estradiol in regulation of Cx43 transcription and/or translation in the chicken ovary. Understanding the contribution of Cx43 in mechanisms underlying ovarian follicle development may be of considerable importance for poultry egg production.

Comparison of Amnio-Exchange With a Novel Synthetic Amniotic Fluid Versus Commercially Used Fluids for Fetal Therapy: An In Vivo Rodent Model.

Normal Saline (NS) and Lactated Ringer's (LR) damage human amniotic epithelium in vitro when compared with a synthetic amniotic fluid (Amnio-well, AW). We sought to evaluate the effect of amnio-exchange with NS, LR, and AW in vivo. On day E17.5, pregnant rats underwent amnio-exchange with NS, LR, or AW. Fetuses in each pregnant rat that did not undergo amnio-exchange acted as controls. Amnions were harvested at E20.5 and ultrastructure evaluated via electron microscopy. Protein levels of cleaved matrix metalloproteinase 9 (MMP9) and collagen 1 (Col1a) were evaluated via Western Blot. Connexin-43 expression was evaluated via immunofluorescence (IF). There was an increase in amnion microfractures and epithelial cellular shrinkage with NS and LR compared with control and AW. The cleaved MMP9/Col1 ratio was increased 3.9-fold in NS (p<0.001) and 4.5-fold LR (p=0.0201) relative to control, whereas AW expression was similar to control (p=0.636). Connexin-43 was also increased on IF in NS and LR relative to AW (mean gray intensity 26.5±4.5, 26.5±6.7, 19.2±3.4, p<0.001). Amnio-exchange with NS and LR led to increased amniotic microfractures and collagen degradation compared with synthetic amniotic fluid. Larger models are warranted to validate or refute these findings.

Empagliflozin mitigates ponatinib-induced cardiotoxicity by restoring the connexin 43-autophagy pathway

BackgroundEmpagliflozin (EMPA), a selective sodium-glucose cotransporter type 2 (SGLT2) inhibitor, has been shown to reduce major adverse cardiovascular events in patients with heart failure of different etiologies, although the underlying mechanism still remains unclear. Ponatinib (PON) is a multi-tyrosine kinase inhibitor successfully used against myeloid leukemia and other human malignancies, but its cardiotoxicity remains worrisome. Cardiac connexins (Cxs) are both substrates and regulators of autophagy and responsible for proper heart function. Alteration in connexin expression and localization have been described in patients with heart failure. AimsTo assess whether EMPA can mitigate PON-induced cardiac dysfunction by restoring the connexin 43-autophagy pathway. Methods and ResultsMale C57BL/6 mice, randomized into four treatment groups (CNTRL, PON, EMPA, PON+EMPA) for 28 days, showed increased autophagy, decreased Cx43 expression as well as Cx43 lateralization, and attenuated systo-diastolic cardiac dysfunction after treatment with EMPA and PON compared with PON alone. Compared with CNTRL (DMSO), cardiomyocyte-differentiated H9c2 (dH9c2) cells treated with PON showed significantly reduced cell viability to approximately 20 %, decreased autophagy, increased cell senescence and reduced DNA binding activity of serum response factor (SRF) to serum response elements (SRE), which were paralleled by reduction in cardiac actin expression. Moreover, PON induced a significant increase of Cx43 protein and its S368-phosphorylated form (pS368-Cx43), as well as their displacement from the plasma membrane to the perinuclear and nuclear cellular region. All these effects were reverted by EMPA. ConclusionEMPA attenuates PON-induced cardiotoxicity by reducing senescence, enhancing the SRE-SRF binding and restoring the connexin 43-autophagy pathway. This effect may pave the way to use of SGLT2 inhibitors in attenuating tyrosine-kinase inhibitor cardiotoxicity.