糖尿病性勃起功能障碍大鼠阴茎海绵体Cx43 mRNA的表达

目的探讨糖尿病性勃起功能障碍（DMED）大鼠阴茎海绵体缝隙连接蛋白43（Cx43）mRNA的表达。方法 W istar大鼠150只,随机分为Ⅰ、Ⅱ、Ⅲ三组,每组分为糖尿病（DM）30只,非糖尿病（NDM）20只。DM大鼠腹腔注射链尿菌素（55 mg/kg）建立DM动物模型后,注射阿朴吗啡,Ⅰ、Ⅱ、Ⅲ三组分别于8、12及16周观察大鼠阴茎勃起情况,筛选DM性ED大鼠模型,测定其阴茎海绵体组织Cx43 mRNA表达。结果 DMED模型成功Ⅰ组23只、Ⅱ组21只、Ⅲ组20只。Ⅰ、Ⅱ、Ⅲ三组NDM大鼠Cx43 mRNA表达量分别为0.845±0.176、0.836±0.120、0.845±0.136,DM大鼠分别为0.493±0.157、0.426±0.164、0.378±0.253,DM大鼠Cx43 mRNA表达与NDM相比差异有统计学意义（P〈0.05）;随DM病程延长,Cx43 mRNA表达明显下降（P〈0.05）。结论 Cx43 mRNA在DMED大鼠海绵体中表达降低,Cx43 mRNA水平下调可能是DMED的发病机制之一。     

扩张性心肌病患者心房Cx43蛋白的表达及其意义

目的:观察扩张性心肌病（dilated cardiomyopathy,DCM）患者心房连接蛋白43（Cx43）的表达、磷酸化的Cx43（pCx43）及其分布特征。探讨Cx43与房颤（atrial fibrillation,AF）之间的关系。方法: 18例接受心脏移植术的DCM患者中,8例无AF史且手术时为窦性心律（SR,SR组）,6例有AF史但手术时为SR（AF+SR组）,4例有AF史且手术时有AF（AF+AF组）。用超声心动图(ECG)测定左心房大小与左心室收缩功能;用免疫印迹法及免疫荧光染色法检测Cx43、pCx43及其分布。结果: DCM患者的左心房直径(LAD)、左室射血分数(LVEF)在SR、AF+SR、AF+AF组间均没有显著差异。与SR组相比,AF+SR组心房组织中Cx43和pCx43无显著差异;AF+AF组总Cx43的表达量及pCx43显著升高(P<0.01及P<0.05)。病理观察提示,AF+AF组Cx43和pCx43升高,Cx43向细胞两侧分布,且pCx43升高。结论: DCM患者的房颤节律伴随Cx43的表达、磷酸化及其分布异常。     

晚期扩张型心肌病患者心肌Cx43蛋白表达的研究

目的:探讨晚期扩张型心肌病(DCM)患者心肌连接蛋白43(Cx43)表达变化及其意义。方法:运用免疫组化和图像分析技术,对5例晚期DCM患者和4例因其他心脏疾病(A对照组)行心脏移植手术切除的心脏及4例机械性损伤死者(B对照组)的左心室心肌Cx43蛋白表达进行定位和定量研究。结果:DCM患者心肌Cx43蛋白表达较清晰,主要位于心肌闰盘处,呈条状、斑点或颗粒状散在分布,有的散在分布于心肌细胞侧边;A对照组表达清楚,位于心肌闰盘处,主要呈条状,少数呈斑点状散在分布,而心肌侧边分布很少;B对照组阳性着色较淡,在闰盘处呈条状分布,少有斑点状散在分布,心肌侧边几无表达。定量检测和统计分析发现,DCM组心肌Cx43蛋白表达的量低于A对照组但高于B对照组。心肌Cx43蛋白表达的平均光密度,DCM组与A对照组、B对照组相比,A对照组与B对照组相比,差异均有统计学意义(P<0.01);心肌Cx43蛋白表达的S值,DCM组、A对照组、B对照组间比较,差异均无统计学意义(P>0.05)。结论:晚期DCM患者心肌Cx43蛋白表达的数量和分布均发生改变,这种变化很可能是导致DCM患者发生心律失常和心功能衰竭的病理基础。     

缝隙连接蛋白Cx43的磷酸化对缝隙连接通讯的调控

缝隙连接介导细胞间通讯。磷酸化通过影响缝隙连接蛋白生命周期中的各个过程而调节细胞间通讯,这些过程涉及到缝隙连接蛋白的转运、组装/解离、降解以及通道的门控等。本文将探讨缝隙连接蛋白Cx43的磷酸化对缝隙连接通讯的调控,继而讨论Cx43蛋白的磷酸化特异性抗体,有助于我们深入研究特定位点的磷酸化事件对缝隙连接通道功能的影响。     

糖尿病性ED大鼠模型建立及阴茎海绵体超微结构变化

目的观察糖尿病（DM）对大鼠勃起功能及阴茎海绵体超微结构的影响,探讨DM引起勃起功能障碍（ED）的机理。方法 24只SPF级SD雄性大鼠随机分为4组,按不同处理因素给予大鼠腹腔注射链脲佐菌素（STZ）,分别记录给药后4 d、1周、2周、3周大鼠的体质量、阿朴吗啡（APO）诱导阴茎的勃起次数及空腹血糖值。应用光镜及透射电镜观察各组大鼠阴茎海绵体的形态结构。结果注射60 mg/kg STZ组大鼠体质量较轻、血糖较高。APO诱导阴茎勃起次数组间差异显著,注射60 mg/kg STZ组应用APO后阴茎未勃起的例数较多。DM性ED（DM＆ED）大鼠阴茎海绵体的形态结构改变明显。结论大鼠注射STZ后2周血糖〉7.2 mmol/L及APO诱导未出现阴茎勃起的大鼠可认为是DM＆ED模型;DM引起大鼠阴茎海绵体超微结构明显改变可能是导致ED的重要原因之一。     

肥厚梗阻型心肌病猝死家族史患者心肌中Cx43表达变化

目的 通过观察猝死家族史肥厚型心肌病患者心肌中Cx43和TGFβ-1表达的改变,初步研究肥厚型心肌病猝死的可能机制。方法 分别选取猝死家族史肥厚型心肌病患者心肌组织（FHSD组,n=5）,非猝死家族史肥厚型心肌病患者心肌组织（NFHSD组,n=5）和非肥厚型心肌病患者心肌组织（NHCM组,n=5）作为研究样本（均获患者家属知情同意）,应用Western blot和免疫组织化学方法检测心肌组织Cx43和TGFβ-1表达。结果 Western blot结果显示,FHSD组、NFHSD组心肌组织Cx43表达与NHCM组患者相比均显著性增高（P<0.01）,NFHSD组Cx43表达明显高于FHSD组（P<0.05）;FHSD组、NFHSD组心肌组织TGF-β1表达与NHCM相比均显著性降低（P<0.01）;NFHSD组TGF-β1表达与FHSD组相比较无显著性差异。免疫组织化学结果与Western blot相同。结论 肥厚型心肌患者猝死的机制是否与心肌细胞Cx43表达在代偿期突然降低有关,尚需进一步研究。     

血管紧张素Ⅱ下调肥厚心肌细胞缝隙连接蛋白Cx43的表达

目的 研究血管紧张素Ⅱ诱导心肌细胞肥大后连接蛋白43(Cx43)表达的变化及与细胞周期分布的关系.方法 分离培养大鼠心肌细胞,用血管紧张素Ⅱ诱导心肌细胞肥大,72 h后用RT-PCR,Western-blot和免疫荧光方法 观察心肌细胞Cx43基因和蛋白表达,用流式细胞仪测定法观察心肌细胞周期分布变化以及与Cx43表达量的关系.结果 血管紧张素Ⅱ处理后的心肌细胞表现细胞肥大且细胞活力增强,S期、G2-M期细胞百分比增加,细胞内G2-M(二倍体)DNA含量降低,Cx43蛋白表达明显低于正常对照组,呈浓度依赖性下调,Cx43 mRNA表达水平显著下调,Cx43蛋白表达下调与细胞周期分布的改变相关.结论 血管紧张素Ⅱ诱导心肌细胞肥大后Cx43基因及Cx43蛋白表达出现浓度依赖性下调,这一改变与心肌肥大过程中的细胞周期变化有关,提示血管紧张素Ⅱ可能通过调控Cx43基因的表达而参与缝隙连接重构过程,而Cx43表达的变化可能与心肌肥大的机制有关.     

血管紧张素Ⅱ下调肥厚心肌细胞缝隙连接蛋白Cx43的表达

目的研究血管紧张素Ⅱ诱导心肌细胞肥大后连接蛋白43（Cx43）表达的变化及与细胞周期分布的关系。方法分离培养大鼠心肌细胞，用血管紧张素Ⅱ诱导心肌细胞肥大，72h后用RT-PCR，Western-blot和免疫荧光方法观察心肌细胞Cx43基因和蛋白表达，用流式细胞仪测定法观察心肌细胞周期分布变化以及与Cx43表达量的关系。结果血管紧张素Ⅱ处理后的心肌细胞表现细胞肥大且细胞活力增强，S期、G2-M期细胞百分比增加，细胞内G2-M（二倍体）DNA含量降低，Cx43蛋白表达明显低于正常对照组，呈浓度依赖性下调，Cx43mRNA表达水平显著下调，Cx43蛋白表达下调与细胞周期分布的改变相关。结论血管紧张素Ⅱ诱导心肌细胞肥大后Cx43基因及Cx43蛋白表达出现浓度依赖性下调，这一改变与心肌肥大过程中的细胞周期变化有关，提示血管紧张素Ⅱ可能通过调控Cx43基因的表达而参与缝隙连接重构过程，而Cx43表达的变化可能与心肌肥大的机制有关。     

应用组织芯片技术研究Cx43在肝细胞癌中的表达及意义

目的探讨间隙连接蛋白（Cx43）在癌旁组织、肝硬化组织和肝细胞癌中的表达及其在肝细胞癌中的意义。方法应用组织芯片技术和免疫组织化学技术sP法检测81例癌旁组织、82例肝硬化组织和359例肝细胞癌中Cx43的表达。结果Cx43在肝细胞癌中的表达低于癌旁组织和肝硬化组织,差异有统计学意义（P〈0．001）;癌旁组织高于肝硬化组织,差异有统计学意义（P〈0．05）。Cx43与肝细胞癌的组织学分级、术后无瘤生存时间以及肿瘤直径有关（P〈0．001、P〈0．05和P〈0．05）,而与临床分期、淋巴结转移和脉管或门静脉癌栓无关（P〉0．05）。结论Cx43的异常表达与肿瘤的发生及预后有关,可以作为判断肝细胞癌预后有用指标之一。     

连接蛋白43表达的变化与心律失常的关系

近年研究表明,多种途径都可以改变连接蛋白43（connexin43,Cx43）的表达和分布,进而影响到心律失常的发生与进展。本文主要综述在房颤和骨骼肌成肌细胞心脏移植中,Cx43表达的变化,以及体外定向重离子流放射、缺血预处理、缝隙连接阻滞剂和阿片类物质对Cx43的影响,说明Cx43表达的变化与心律失常的关系。     

Does regular consumption of green tea influence expression of vascular endothelial growth factor and its receptor in aged rat erectile tissue? Possible implications for vasculogenic erectile dysfunction progression

Erectile dysfunction (ED) is a highly prevalent disease affecting millions of men worldwide with a tendency for widespread increase. ED is now considered an early manifestation of atherosclerosis and, consequently, a precursor of systemic vascular disease. Atherosclerosis and ED share potentially modifiable risk factors, as smoking or high-fat food intake, but it is unclear how regular consumption of anti-oxidant rich drinks, which exhibit recognised anti-atherosclerotic features, affects ED progression. The objective of this study was to evaluate the modulating effects of chronic consumption of catechin-rich beverages on the vascular structure of the rat corpus cavernosum, and how this could contribute to delay or prevention of the onset of ED. Male Wistar rats aged 12 months were treated with green tea (GT) or a green tea extract solution (GTE) as the only liquid source for 6 months. Consumption of GT and GTE led to decreased plasma androgen levels without any significant change in plasma lipid levels. A reduction in corpus cavernosum intracellular storage of lipids, associated with decreased expression of vascular endothelial growth factor (VEGF) and its receptor VEGFR2 in endothelial cells, was observed. Taken together, these results suggest diminished atherosclerotic progression in cavernous tissue. However, functional studies will be necessary to elucidate if catechin-rich beverages are useful compounds in the prevention of deleterious vascular events associated with ED. It was also demonstrated that regular consumption of catechins reduces atherosclerotic progression and mortality due to cardiovascular disease. The results reported here suggest diminished atherosclerotic progression in cavernous tissue in aged rats following chronic ingestion of catechin-rich beverages.     

微小RNA-1调控自发性高血压大鼠肥厚左室心肌组织连接蛋白43的表达

目的观察微小RNA-1(miRNA-1)对自发性高血压大鼠(SHRs)肥厚左室心肌组织中连接蛋白43(Cx43)表达的调控及其意义。方法 18只17周龄雄性SHRs随机分为干预组(n=6)、阴性对照组(n=6)和空白对照组(n=6),通过脂质体瞬时转染技术,干预组由尾静脉注射miRNA-1抑制剂,两对照组分别注入miRNA-1抑制剂阴性对照和无血清培养基混合物,并通过实时荧光定量PCR、免疫组织化学法及western blot等技术,检测大鼠左室心肌组织miRNA-1及Cx43蛋白表达水平的改变。结果与阴性对照组和空白对照组比较,干预组miRNA-1表达水平明显降低,伴随着Cx43蛋白表达水平增高(0.45±0.15 vs 0.27±0.14,0.23±0.10,P均<0.05)。结论抑制miRNA-1表达能使SHR肥厚左室心肌组织Cx43蛋白水平升高。     

Diminished penile expression of vascular endothelial growth factor and its receptors at the insulin-resistant stage of a type II diabetic rat model: a possible cause for erectile dysfunction in diabetes

Erectile dysfunction (ED) is commonly experienced in men with diabetes mellitus. Vascular endothelial growth factor (VEGF) has been extensively documented for its pathogenic significance in different complications of diabetes. We hypothesized that expressions of VEGF, its receptors and its signaling pathway Akt may be drastically altered in diabetic penile tIssues and their alterations may modulate penile expression of the molecules that are believed to play a role in diabetic ED. Otsuka Long-Evans Fatty (OLETF) rats, a type II (non-insulin-dependent) diabetes mellitus, were used at the insulin-resistant stage of type II diabetes (20 weeks of age). We determined protein and mRNA expressions of VEGF, its receptors, Akt, nitric oxide synthase isoforms, and apoptosis-related molecules in the penis using immunohistochemistry, Western blotting, in situ hybridization, and real-time quantitative PCR analyses. The penile sections were also submitted to the Tdt-mediated dUTP nick end labeling assay for apoptosis. OLETF rats showed marked reductions in penile expression of VEGF, its two receptors and Akt. In OLETF rat penises, endothelial and neuronal nitric oxide synthase isoforms were expressed less abundantly. Furthermore, while anti-apoptotic markers, Bcl-2 and phosphorylated Bad, were down-regulated, pro-apoptotic markers, active caspase-3 and Bax, were up-regulated, resulting in the appearance of apoptotic cells in the penile tIssues of OLETF rats. The VEGF signaling system would work less well in diabetic penile tIssues as a result of the reduced expression, leading to diminished endothelial production of nitric oxide and apoptosis-related erectile tIssue damage. We propose that the abnormalities of the VEGF signaling system in the penis may play a role in the pathophysiology of diabetic ED.     

Phosphodiesterase-5A dysregulation in penile erectile tissue is a mechanism of priapism

The molecular mechanism for priapism is not well characterized. Although the nitric oxide (NO) pathway is known to mediate penile erection under normal conditions, we hypothesized that the mechanism of priapism rests in aberrant downstream signaling of this pathway based on our previous findings that mice lacking the gene for endothelial nitric oxide synthase (eNOS-/-) and mice lacking both neuronal NOS (nNOS) and eNOS (nNOS-/-, eNOS-/-) have a tendency for priapic activity. We investigated the role of downstream guanylate cyclase and phosphodiesterase type 5 (PDE5A) expression and function in mediating these responses in eNOS-/- and nNOS-/-, eNOS-/- mice. Erectile responses to both cavernous nerve stimulation and intracavernosal injection of the NO donor diethylamine-NONOate were augmented in eNOS-/- and nNOS-/-, eNOS-/- mice but not in WT or nNOS-/- mice. PDE5A protein expression and activity and cGMP levels were significantly lower in eNOS-/- and nNOS-/-, eNOS-/- mice, and this effect was reproduced in WT corpus cavernosum exposed to NOS inhibitors. Moreover, cavernous nerve stimulation was associated with a marked augmentation of cavernosal cGMP levels, suggesting that, although lower at baseline, the production of cGMP is unchecked in eNOS-/- and nNOS-/-, eNOS-/- mice upon neurostimulation. Transfection of eNOS-/- mice with an adenovirus encoding eNOS resulted in a normalization of PDE5A protein and activity as well as a correction of priapic activity. Coupled with the observation that sickle cell disease mice (which show a priapism phenotype) evince dysregulated PDE5A expression/activity, these data suggest that PDE5A dysregulation is a fundamental mechanism for priapism.     

肝肺综合征大鼠肺组织因子mRNA的表达及其意义

目的 检测肝肺综合征(HPS)大鼠肺组织中组织因子(TF) mRNA的表达,探讨其意义. 方法 Sprague-Dawley雄性大鼠40只,均分为肝硬化组、肝肺综合征组,检测血气分析,用qRT-PCR检测肺组织TF mRNA的表达,光镜观察肺组织切片病理改变.两组间均数比较采用非参数分析Mann-Whitney U检验,两因素之间的相关分析采用Spearman秩相关分析.结果 (1)血气分析显示肝肺综合征组PaO2 (58.20±3.19) mm Hg低于肝硬化组的(85.00±2.53) mmHg,差异有统计学意义(P＜0.05).(2)肺组织TF mRNA的表达在肝肺综合征组(相对表达量为0.77±0.22)明显高于肝硬化组(0.33±0.14),差异有统计学意义(P＜0.05);其与PaO2呈显著负相关(r=-0.565,P＜0.05).(3)肺组织病理学检查显示肝硬化组部分肺泡间隔增厚,肺泡腔大小不一,肺容量减少,炎性细胞浸润,局部肺血管内可见充血,肝肺综合征组肺泡间隔增宽明显,肺泡腔大小不一、有明显的出血,纤维素性渗出,大量炎性细胞浸润,肺容量减少,肺泡壁毛细血管扩张明显,局部增生,巨噬细胞聚集,微血栓形成.肝肺综合征组肺组织内微血栓形成,大鼠肺组织TF mRNA相对表达量为0.68±0.17,高于无血栓形成的大鼠的0.40±0.12,差异有统计学意义(P＜0.05).结论 肝肺综合征大鼠肺组织TF mRNA的表达升高,且随着缺氧程度的加重而加重,可能增加肝肺综合征肺血栓栓塞发生的风险.     

Matrix-protein-specific regulation of Cx43 expression in cardiac myocytes subjected to mechanical load

To elucidate mechanisms responsible for mechanotransduction in the heart and define the effects of remodeling of the extracellular matrix, we cultured neonatal rat ventricular myocytes on native type I collagen, fibronectin, or denatured collagen and subjected them to uniaxial, pulsatile stretch. Changes in expression of the cardiac gap junction protein, Cx43, were measured by confocal microscopy and immunoblotting. Cells grown on fibronectin or denatured collagen exhibited significantly greater Cx43 expression than cells grown on native collagen. Stretch induced a approximately 2-fold increase in Cx43 expression in cells grown on native collagen but no increase in cells grown on fibronectin or denatured collagen. Incubation of cells on native collagen with a peptide containing the arginine-glycine-aspartate (RGD) motif upregulated Cx43 expression equivalent to that induced by stretch. Nonselective activation of integrin signaling with MnCl2 also upregulated Cx43 expression in cells grown on native collagen. This effect was blocked completely by pretreatment with anti-beta1 integrin antibody but not by anti-beta3 integrin antibody. Stretch led to a marked increase in beta1 integrin immunofluorescent signal in cells grown on native collagen but not in cells grown on fibronectin or denatured collagen. Stretch-induced upregulation of Cx43 was also blocked by anti-beta1 integrin antibody. Thus, matrix protein-myocyte interactions regulate Cx43 expression via beta1 integrin signaling initiated by mechanical stimulation in cells grown on native type I collagen, or by RGD-integrin signaling independent of mechanical stress in cells grown on fibronectin or denatured collagen. Changes in the composition of the extracellular matrix may affect electrical coupling in cardiac myocytes.