Prenatal exposure to inflammatory conditions increases Cx43 and Panx1 unopposed channel opening and activation of astrocytes in the offspring effect on neuronal survival

Several epidemiological studies indicate that children born from mothers exposed to infections during gestation, have an increased risk to develop neurological disorders, including schizophrenia, autism and cerebral palsy. Given that it is unknown if astrocytes and their crosstalk with neurons participate in the above mentioned brain pathologies, the aim of this work was to address if astroglial paracrine signaling mediated by Cx43 and Panx1 unopposed channels could be affected in the offspring of LPS‐exposed dams during pregnancy. Ethidium uptake experiments showed that prenatal LPS‐exposure increases the activity of astroglial Cx43 and Panx1 unopposed channels in the offspring. Induction of unopposed channel opening by prenatal LPS exposure depended on intracellular Ca²⁺ levels, cytokine production and activation of p38 MAP kinase/iNOS pathway. Biochemical assays and Fura‐2AM/DAF‐FM time‐lapse fluorescence images revealed that astrocytes from the offspring of LPS‐exposed dams displayed increased spontaneous Ca²⁺ dynamics and NO production, whereas iNOS levels and release of IL‐1β/TNF‐α were also increased. Interestingly, we found that prenatal LPS exposure enhanced the release of ATP through astroglial Cx43 and Panx1 unopposed channels in the offspring, resulting in an increased neuronal death mediated by the activation of neuronal P2X₇ receptors and Panx1 channels. Altogether, this evidence suggests that astroglial Cx43 and Panx1 unopposed channel opening induced by prenatal LPS exposure depended on the inflammatory activation profile and the activation pattern of astrocytes. The understanding of the mechanism underlying astrocyte‐neuron crosstalk could contribute to the development of new strategies to ameliorate the brain abnormalities induced in the offspring by prenatal inflammation. GLIA 2015;63:2058–2072     

缝隙连接在癫痫中的作用(英文)

癫痫是最常见的神经系统疾病之一,临床主要表现为周期性和无法预见性的癫痫样发作。目前认为,癫痫的形成、同步化以及癫痫样放电的维持与细胞间的缝隙连接有关。大量的体内外实验研究表明,缝隙连接抑制剂具有较强的抗癫痫作用,提示缝隙连接可能成为开发新型抗癫痫药物的潜在靶点。此外,选择性药理学研究和基因敲除技术研究表明,神经元间与胶质细胞间的缝隙连接在癫痫中的作用需区别看待。随着条件性基因敲除技术的成熟、高选择性的新型缝隙连接调控药物的出现以及更详细的人类癫痫脑组织样本的研究,人们将更全面地了解缝隙连接在癫痫中的作用。本综述总结了目前关于缝隙连接在癫痫发病机制中作用的研究结果,并对临床和基础研究中癫痫发作导致的缝隙连接蛋白表达的变化进行讨论。     

Differential connexin expression,gap junction intercellular coupling,and hemichannel formation in NT2/D1 human neural progenitors and terminally differentiated hNT neurons

Connexin-mediated gap junctions and open hemichannels in nonjunctional membranes represent two biologically relevant mechanisms by which neural progenitors can coordinate their response to changes in the extracellular environment. NT2/D1 cells are a teratocarcinoma progenitor line that can be induced to differentiate terminally into functional hNT neurons and NT-G nonneuronal cells. Clinical transplants of hNT neurons and experimental grafts of NT2/D1 progenitors or hNT neurons have been used in cell-replacement therapy in vivo. Previous studies have shown that NT2/D1 cells express connexin 43 (Cx43) and that NT2/D1 progenitors are capable of dye transfer. To determine whether NT2/D1 progenitors and differentiated hNT cultures express other connexins, Cx26, Cx30, Cx32, Cx36, Cx37, Cx43, and Cx46.6 mRNA and protein were analyzed. NT2/D1 progenitors express Cx30, Cx36, Cx37, and Cx43. hNT/NT-G cultures express Cx36, Cx37, and de novo Cx46.6. Cx26 and Cx32 were not expressed in NT2/D1 or hNT/NT-G cells. NT2/D1 progenitors formed functional gap junctions as assessed by dye coupling as well as open hemichannels in nonjunctional membranes as assessed by dye-uptake studies. Dye coupling was inhibited by the gap junction blocker 18alpha-glycyrrhetinic acid. Hemichannel activity was inhibited by the dual-specificity chloride channel/connexin hemichannel inhibitor flufenamic acid but not by the chloride channel inhibitor 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid. Both dye coupling and dye uptake were substantially reduced following differentiation of NT2/D1 progenitors. We conclude that the pattern of connexin expression in NT2/D1 cells changes over the course of differentiation corresponding with a reduction in biochemical coupling and hemichannel activity in differentiated cells.     

胃癌和胃癌前病变Cx43、PCNA的表达及意义

目的　通过观察间隙连接蛋白 Cx4 3和增殖细胞核抗原 (PCNA )在正常胃粘膜、胃癌前病变和胃癌中的表达和分布情况 ,探讨 Cx基因表达与胃癌发生的关系。　方法　运用 SP免疫组织化学方法检测 Cx4 3、PCNA在 70例原发性胃癌 ,6 2例中、重度不典型增生和 16例正常胃粘膜中表达规律。　结果　 Cx4 3在正常胃粘膜 ,中、重度不典型增生和胃癌中表达的阳性率分别为 10 0 % ,83.9%和 17.1% ,三者比较差异有显著性 (P<0 .0 5 )。 Cx4 3表达与胃癌的分化程度相关 (P<0 .0 1)。胃癌前病变、胃癌组中的 PCNA较正常胃粘膜组增高 (P<0 .0 1)。 Cx4 3表达与 PCNA呈负相关 (P<0 .0 1)。　结论　 Cx4 3表达降低在胃癌发生发展中起重要作用 ,Cx4 3可做为胃癌早期诊断的指标     

Expression of connexin 36 in central nervous system and its role in epileptic seizure

Objective  This review discusses the experimental and clinical studies those show the expression of connexin 36 in the central nervous system and the possible role of connexin 36 in epileptic seizure.

Data sources  All articles used in this review were mainly searched from PubMed published in English from 1996 to 2012. 

Study selection   Original articles and reviews were selected if they were related to the expression of connexin 36 in the central nervous system and its role in epilepsy.

Results  The distribution of connexin 36 is developmentally regulated, cell-specific and region-specific. Connexin 36 is involved in some neuronal functions and epileptic synchronization. Changes in the connexin 36 gene and protein were accompanied by seizures. Selective gap junction blockers have exerted anticonvulsant actions in a variety of experiments examined in both humans and experimental animals.

Conclusions  Connexin 36 plays an important role in both physiological and pathological conditions in the central nervous system. A better understanding of the role of connexin 36 in seizure activity may contribute to the development of new therapeutic approaches to treating epilepsy.     

防治血管再狭窄的新靶点——缝隙连接蛋白43

缝隙连接是介导相邻细胞间离子和小分子信号物质直接交换的跨膜通道,血管平滑肌细胞中主要表达连接蛋白43(connexin43,CX43)。研究表明,CX43表达上调与平滑肌细胞的增殖、迁移及细胞外基质生成密切相关,促使血管损伤后新内膜生成,引起血管再狭窄,提示CX43可能成为血管性疾病治疗的新靶点。本文综述CX43与血管再狭窄之间关系及以CX43为靶点的药物和基因治疗的研究进展。     

激动M_3受体对间隙连接蛋白43表达的影响

目的探索激动心肌M3受体后间隙连接蛋白43的变化情况,进一步发现M3受体作为抗心律失常药物作用新靶点的作用机制。方法通过免疫组化、划痕标记荧光传输技术结合激光共聚焦显微镜和RT-PCR的方法,研究激动M3受体对间隙连接蛋白43表达的影响。结果证实激动M3受体可以改善细胞和细胞之间的通讯,改善病理条件下间隙连接蛋白43的重构,且其磷酸化水平在间隙连接蛋白的重构过程中发挥了重要作用,并且还发现激动M3受体可以恢复病理条件下下降的间隙连接蛋白43的mRNA水平。结论激动M3受体可以改善病理条件下间隙连接蛋白43的重构,并且其磷酸化形式在闰盘重构中发挥重要的作用。     

GJB2基因突变致聋儿童的人工耳蜗植入效果

背景：基因诊断技术的普及和推广,使得先天性听力障碍的儿童可以检测出致聋基因,其中在人工耳蜗植入患者中GJB2(gap junction protein beta 2,GJB2 )基因突变的发生率为26.5%。本文对31例GJB2基因突变致聋儿童人工耳蜗植入后听觉康复效果进行了随访评估,从而为评价此类患者人工耳蜗植入效果提供参考依据。     

Cardiac conduction is required to preserve cardiac chamber morphology

Electrical cardiac forces have been previously hypothesized to play a significant role in cardiac morphogenesis and remodeling. In response to electrical forces, cultured cardiomyocytes rearrange their cytoskeletal structure and modify their gene expression profile. To translate such in vitro data to the intact heart, we used a collection of zebrafish cardiac mutants and transgenics to investigate whether cardiac conduction could influence in vivo cardiac morphogenesis independent of contractile forces. We show that the cardiac mutant dco^s226 develops heart failure and interrupted cardiac morphogenesis following uncoordinated ventricular contraction. Using in vivo optical mapping/calcium imaging, we determined that the dco cardiac phenotype was primarily due to aberrant ventricular conduction. Because cardiac contraction and intracardiac hemodynamic forces can also influence cardiac development, we further analyzed the dco phenotype in noncontractile hearts and observed that disorganized ventricular conduction could affect cardiomyocyte morphology and subsequent heart morphogenesis in the absence of contraction or flow. By positional cloning, we found that dco encodes Gja3/Cx46, a gap junction protein not previously implicated in heart formation or function. Detailed analysis of the mouse Cx46 mutant revealed the presence of cardiac conduction defects frequently associated with human heart failure. Overall, these in vivo studies indicate that cardiac electrical forces are required to preserve cardiac chamber morphology and may act as a key epigenetic factor in cardiac remodeling.     

(-)-Epicatechin effects in rat liver epithelial cells: stimulation of gap junctional communication and counteraction of its loss due to the tumor promoter 12-O-tetradecanoylphorbol-13-acetate

Gap junctional intercellular communication (GJIC) is a direct signaling pathway for neighboring cells. Disturbances in GJIC are suggested to play a role in carcinogenesis and may be involved in cardiac arrhythmia. Tumor promoters like 12-O-tetradecanoylphorbol-13-acetate (TPA) are capable of inhibiting GJIC, whereas GJIC is stimulated by several micronutrients like genistein, retinoids or carotenoids. (-)-Epicatechin (4-40 microM), a major flavonoid in cocoa and green tea, exhibited stimulatory effects on GJIC in WB-F344 rat liver epithelial cells after 24-72hr of incubation; no change was observed after 90 min. However, treatment of cells for 90 min with TPA (5 or 10nM) led to complete loss of GJIC, whereas 40% loss was found with 1nM. These inhibitory effects of TPA were largely suppressed when (-)-epicatechin or genistein (40 microM) were present during the incubation. In communicating WB-F344 cells, most of the major gap junction protein connexin43 (Cx43) was located in the plasma membrane. When the cells were exposed to TPA, considerably less protein was found in the membrane. Such a delocalization of Cx43 proteins was not observed when TPA was coincubated with the flavonoids, (-)-epicatechin or genistein. It is concluded that TPA affects Cx43 trafficking between cellular compartments, and that this effect is counteracted by (-)-epicatechin or genistein.