Microglia activation influences dye coupling and Cx43 expression of the astrocytic network

Under inflammatory conditions, activated microglia are capable of producing proinflammatory cytokines that are reported to influence cell-to-cell communication. The present study was performed to evaluate the influence of microglial activation on the coupling efficiency of the astroglial network. Primary astrocyte cultures of newborn rats were cocultured with either 5% (M5) or 30% (M30) microglia. Microglial activation (rounded phagocytotic phenotype) was investigated using the monoclonal anti-ED1 antibody, and immunofluorescence with a polyclonal anti-Cx43 antibody was used to study astroglial Cx43 expression and distribution. Functional coupling of astrocytes was evaluated by monitoring the transfer of microinjected Lucifer yellow into neighboring cells. The data obtained can be summarized as follows: astroglia/M30 cocultures contained significantly fewer resting microglia and significantly more activated microglia than the M5 cocultures; significantly reduced astroglial Cx43 staining was found in M30 cocultures concurrently with a reduced number of dye coupled astrocytes; and the positive correlation of percent activated microglia with reduced astroglial Cx43 expression was highly significant, indicating that the degree of intercellular communication in the astroglial network may be modulated by the activation of microglia under in vitro conditions.     

Immunohistochemical examination of c-Met protein expression in astrocytic tumors

Hepatocyte growth factor/scatter factor (HGF/ SF), which has various physiological functions, and its receptor c-Met, the human c-met proto-oncogene product, are thought to be determinant in the pathological processes of various malignancies. To investigate the possible role of HGF/SF in the progression of development of astrocytic tumors, we examined the expression of c-Met in these tumors. Immunohistochemistry using the streptavidin-biotin peroxidase complex method and immunofluorescence double staining with anti-c-Met polyclonal and anti-glial fibrillary acidic protein monoclonal antibodies were performed. Positive c-Met expression was detected in 31 of the 42 astrocytic tumors and some of the control cases analyzed. c-Met-positive cells showed morphological characteristics of astrocytes. Especially in the cases of high-grade tumors, c-Met positivity was abundant in cells in both vascular-rich and peripheral regions of the tumors but not in the cells with distinctly malignant features. Immunofluorescence double staining revealed that the c-Met-positive cells were in part of astrocytic origin. We suggest that c-Met-positive cells are affected by some factors in the lesions where the pathological processes are in a state of development. Our studies indicated that c-Met expression might take part in glioma invasion but not in the development of malignancy. Received: 18 April 1997 / Revised: 12 June 1997 / Revised, accepted: 15 October 1997     

WISP-2在人脑星形细胞瘤中的表达研究

目的探讨WISP-2mRNA及蛋白在人脑星形细胞瘤中的表达及意义。方法采用半定量PCR检测WISP-2mRNA在人脑星形细胞瘤组织与正常脑组织中的表达差异;随后利用免疫组化方法验证WISP-2蛋白在星形细胞瘤组织与正常脑组织之间的表达差异。结果半定量PCR检测发现,WISP-2mRNA在正常脑组织中无表达,在星形细胞瘤中相对表达量为0.677±0.445,两者比较有统计学意义(t=4.783,P0.05),在高级别组(Ⅲ-Ⅳ)与低级别组(Ⅰ-Ⅱ)星形细胞瘤中的相对表达量分别为0.924±0.438、0.478±0.344,两者比较有统计学意义(t=3.909,P0.05)。免疫组化检测发现,正常脑组织细胞中无WISP-2蛋白阳性表达(0/10),WISP-2蛋白在星形细胞瘤组织内呈高表达,阳性率为66.0%(31/47),两者比较有统计学意义(χ2=11.92,P0.01),WISP-2蛋白在高级别组(Ⅲ-Ⅳ)与低级别组(Ⅰ-Ⅱ)星形细胞瘤中的表达阳性率分别为85.7%(18/21)、50.0%(13/26),两者比较有统计学意义(χ2=6.60,P0.05).与WISP-2mRNA在星形细胞瘤组织内表达特征相一致,两者具有正相关性(r=0.63,P0.001)。结论WISP-2mRNA和蛋白在人脑星形细胞瘤组织中明显高表达,并与其恶性程度有关,可作为判断脑星形细胞瘤恶性程度的一个辅助指标。     

Ganglioside (GD2) expression and intermediary filaments in astrocytic tumors

The search of proliferation markers in astrocytic tumors that may serve as targets for therapeutic interventions, is in full progress. Membrane-bound signal transducers for growth factors are amongst the substances of interest. Gangliosides are lipid-sugar compounds localized on the cell membrane that are thought to modify pertinent signals and, therefore, may influence a variety of functions in normal and pathologic conditions including those that act upon tumor growth. Intracranial supratentorial astrocytic gliomas of the adult represent a tumor group, that may be divided into three grades of malignancy, the most anaplastic member being the glioblastoma. A stepwise anaplasia is assumed and accompanied by genetic events that are partly specific for these grades. In earlier investigations, it had been shown that there is a tendency towards formation of more simple members of the ganglioside family with ongoing malignancy of those tumors. Yet, the results were only partly congruent and the correlation to tumor grades rather loose. We, therefore, investigated the occurrence of triaose gangliosides within these tumors in situ by immunohistochemistry. In this paper, we corroborate our earlier observation that triaose gangliosides preferentially occur within the cytoplasm of large protoplasmic and gemistocytic astrocytes. The potency of the expression of GD2 is calculated and plotted against the expression of two markers of intermediate glial filaments, namely GFAP (glial fibrillary acid protein) and vimentine. A high interdependence of the three compounds could be demonstrated by correlation analysis. Thus, the conclusion must be drawn that the correlation of ganglioside patterns to the proliferation of astrocytic tumors is as poor as that of GFAP or vimentin expression, respectively.     

星形细胞肿瘤的预后因素分析

目的探讨影响星形细胞肿瘤预后因素,为临床治疗提供理论依据。方法复习167例星形细胞肿瘤患者的资料,使用Kaplan-Meier单因素生存分析法及多因素Cox比例风险模型进行分析。结果单因素分析结果显示性别、年龄、术前癫痫、组织学分级、术前高级神经功能状态、术后患者机能状况评分等因素与患者预后有显著相关性（P〈0.01）;肿瘤切除程度、术后高级神经功能状态、综合治疗情况与预后有相关性（P〈0.05）。高级别星形细胞肿瘤病例COX多因素分析则显示性别、术后KPS评分、术后个体化综合治疗情况与患者预后显著相关（P〈0.01）。结论患者性别、年龄、病理组织学分级、术前癫痫、高级神经功能状态对预后影响较大,多因素分析显示综合治疗与高级别星形细胞肿瘤患者预后显著相关。     

伽玛刀照射后胶质细胞CX43和GFAP表达变化及意义

目的研究伽玛刀照射后胶质细胞缝隙连接蛋白43(Connexin43,Cx43)和胶质纤维酸蛋白(glial fibrillary acid protein,GFAP)表达的变化。方法体外培养原代星形胶质细胞(astrocytes,Ast)分为正常对照组和γ刀照射组,后者经γ刀照射(边缘剂量4～36Gy),培养72小时后检测GFAP,Cx43。结果4～12Gy组与正常对照组无显著差异,至16Gy组胶质细胞开始增生GFAP表达阳性细胞数大于正常对照组且呈剂量依赖性增高至32Gy组GFAP表达达最大值。Cx43表达在12Gy组即开始明显下降且Cx43表达呈计量依赖性减低,在24～32Gy降低尤为显著至32Gy组达最低值。结论原代培养Ast经伽玛刀照射后GFAP表达增高,同时Cx43表达减低。Cx43的异常低表达可能是胶质增生及放射损伤的重要原因。     

Acute downregulation of Cx43 alters P2Y receptor expression levels in mouse spinal cord astrocytes

Propagation of intercellular calcium waves (ICW) between astrocytes depends on the diffusion of signaling molecules through gap junction channels and diffusion through the extracellular space of neuroactive substances acting on plasmalemmal receptors. The relative contributions of these two pathways vary in different brain regions and under certain pathological conditions. We have previously shown that in wild-type spinal cord astrocytes, ICW are primarily gap junction-dependent, but that deletion of the main gap junction protein (Cx43) by homologous recombination results in a switch in mode of ICW propagation to a purinoceptor-dependent mechanism. Such a compensatory mechanism for ICW propagation was related to changes in the pharmacological profile of P2Y receptors, from an adenine-sensitive P2Y(1), in wild-type, to a uridine-sensitive P2U receptor subtype, in Cx43 knockout (KO) astrocytes. Using oligonucleotide antisense to Cx43 mRNA for acute downregulation of connexin43 expression levels, we provide evidence for the molecular nature of such compensatory mechanism. Pharmacological studies and Western blot analysis indicate that there is a reciprocal regulation of P2Y(1) and P2Y(4) expression levels, such that downregulation of Cx43 leads to decreased expression of the adenine-sensitive P2Y(1) receptor and increased expression of the uridine-sensitive P2Y(4) receptor. This change in functional expression of the P2Y receptor subtype population in acutely downregulated Cx43 was paralleled by changes in the mode of ICW propagation, similar to that previously observed for Cx43 KO spinal cord astrocytes. On the basis of these results, we propose that Cx43 regulates both modes of ICW by altering P2Y receptor subtype expression in addition to providing intercellular coupling.     

Coupling of astrocyte connexins Cx26, Cx30, Cx43 to oligodendrocyte Cx29, Cx32, Cx47: Implications from normal and connexin32 knockout mice

Oligodendrocytes in vivo form heterologous gap junctions with astrocytes. These oligodendrocyte/astrocyte (A/O) gap junctions contain multiple connexins (Cx), including Cx26, Cx30, and Cx43 on the astrocyte side, and Cx32, Cx29, and Cx47 on the oligodendrocyte side. We investigated connexin associations at A/O gap junctions on oligodendrocytes in normal and Cx32 knockout (KO) mice. Immunoblotting and immunolabeling by several different antibodies indicated the presence of Cx32 in liver and brain of normal mice, but the absence of Cx32 in liver and brain of Cx32 KO mice, confirming the specificity and efficacy of the antibodies, as well as allowing the demonstration of Cx32 expression by oligodendrocytes. Oligodendrocytes throughout brain were decorated with numerous Cx30-positive puncta, which also were immunolabeled for both Cx32 and Cx43. In Cx32 KO mice, astrocytic Cx30 association with oligodendrocyte somata was nearly absent, Cx26 was partially reduced, and Cx43 was present in abundance. In normal and Cx32 KO mice, oligodendrocyte Cx29 was sparsely distributed, whereas Cx47-positive puncta were densely localized on oligodendrocyte somata. These results demonstrate that astrocyte Cx30 and oligodendrocyte Cx47 are widely present at A/O gap junctions. Immunolabeling patterns for these six connexins in Cx32 KO brain have implications for deciphering the organization of heterotypic connexin coupling partners at A/O junctions. The persistence and abundance of Cx43 and Cx47 at these junctions after Cx32 deletion, together with the paucity of Cx29 normally present at these junctions, suggests Cx43/Cx47 coupling at A/O junctions. Reductions in Cx30 and Cx26 after Cx32 deletion suggest that these astrocytic connexins likely form junctions with Cx32 and that their incorporation into A/O gap junctions is dependent on the presence of oligodendrocytic Cx32.     

Cx36, Cx43 and Cx45 in mouse and rat cerebellar cortex: species‐specific expression,compensation in Cx36 null mice and co‐localization in neurons vs. glia

Electrical synapses formed by connexin36 (Cx36)‐containing gap junctions between interneurons in the cerebellar cortex have been well characterized, including those formed between basket cells and between Golgi cells, and there is gene reporter‐based evidence for the expression of connexin45 (Cx45) in the cerebellar molecular layer. Here, we used immunofluorescence approaches to further investigate expression patterns of Cx36 and Cx45 in this layer and to examine localization relationships of these connexins with each other and with glial connexin43 (Cx43). In mice, strain differences were found, such that punctate labelling for Cx36 was differentially distributed in the molecular layer of C57BL/6 vs. CD1 mice. In mice with EGFP reporter representing Cx36 expression, Cx36‐puncta were localized to processes of stellate cells and other cerebellar interneurons. Punctate labelling of Cx45 was faint in the molecular layer of wild‐type mice and was increased in intensity in mice with Cx36 gene ablation. The vast majority of Cx36‐puncta co‐localized with Cx45‐puncta, which in turn was associated with the scaffolding protein zonula occludens‐1. In rats, Cx45‐puncta were also co‐localized with Cx36‐puncta and additionally occurred along Bergmann glial processes adjacent to Cx43‐puncta. The results indicate strain and species differences in Cx36 as well as Cx45 expression, possible compensatory processes after loss of Cx36 expression and localization of Cx45 to both neuronal and Bergmann glial gap junctions. Further, expression of both Cx43 and Cx45 in Bergmann glia of rat may contribute to the complex properties of junctional coupling between these cells and perhaps to their reported coupling with Purkinje cells.     

人脑胶质瘤Cx43基因表达及其与肿瘤细胞增殖关系的研究

目的 探讨缝隙连接蛋白43(Cx43)基因表达与胶质瘤细胞增殖之间的关系,及其在胶质瘤发生发展中的作用,拟为手术后治疗及疗效观察提供客观依据.方法 采用原位杂交和免疫组织化学染色方法检测胶质瘤细胞Cx43 mRNA、Cx43 蛋白和增殖细胞核抗原的表达水平.结果 对照脑组织和胶质瘤细胞Cx43 mRNA 阳性表达率分别为100%(10/10)和61.70%(29/47),差异具有统计学意义(Z =- 5.407,P = 0.000);低度恶性(WHOⅠ～Ⅱ级)胶质瘤细胞Cx43 mRNA 阳性表达率为100%(7/7)和93.75%(15/16),高度恶性(WHOⅢ～Ⅳ级)为33.33%(6/18)和16.67%(1/6);Cx43 mRNA 阳性表达率与胶质瘤组织病理学分级呈负相关(rs = - 0.794,P = 0.000).对照脑组织和胶质瘤细胞Cx43 蛋白表达水平与Cx43 mRNA 基本一致.对照脑组织增殖细胞核抗原表达阴性;不同级别胶质瘤细胞增殖细胞核抗原阳性表达率依次为WHOⅣ级100%(6/6)、WHOⅢ级94.44%(17/18)、WHOⅡ级62.50%(10/16)和WHOⅠ级42.86%(3/7);增殖细胞核抗原阳性表达率与胶质瘤组织病理学分级呈正相关(rs = 0.589,P = 0.000);WHOⅢ～Ⅳ级与WHOⅠ～Ⅱ级之间差异具有统计学意义(H = 13.239,P = 0.000).Cx43 mRNA 与Cx43蛋白表达水平呈正相关(rs = 0.963,P = 0.000),Cx43 mRNA 及其蛋白质与增殖细胞核抗原表达水平呈负相关(rs = - 0.621,P = 0.000;rs = - 0.913,P = 0.000).结论 胶质瘤细胞Cx43 mRNA 及其蛋白质与增殖细胞核抗原表达水平和肿瘤组织病理学分级呈负相关.提示,Cx43 基因表达与肿瘤细胞增殖活性和胶质瘤恶性进展密切相关.     

实验性自身免疫性脑脊髓炎脑内缝隙连接蛋白Cx43和Cx32的表达变化

目的 观察豚鼠实验性自身免疫性脑脊髓炎(EAE)模型脑内星形胶质细胞(AS)缝隙连接(GJ)蛋白Cx43与神经元缝隙连接蛋白Cx32表达变化及相互关系,以探讨胶质细胞和神经元表面缝隙连接与EAE模型损伤的关系.方法 采用免疫组化方法,显示EAE模型豚鼠脑内胶质细胞和神经元表面缝隙连接蛋白Cx43和Cx32的表达变化及分布规律.结果 EAE豚鼠模型建立成功后,脑内星形胶质细胞Cx43-Li细胞表达明显增加,呈斑点样或分枝样;神经元Cx32-Li细胞表达也增加,并存在一定时间相关性.结论 EAE豚鼠脑内Cx43和Cx32表达增加,提示EAE发生后星形胶质细胞与神经元间信息交流可能加强.     

MRI评分在星形细胞起源肿瘤诊断中的应用

目的观察星形细胞起源肿瘤患者的MRI表现,探讨MRI评分与病理学分级之间的相关性。方法选择经手术病理证实的幕上星形细胞起源肿瘤患者44例,其中星形细胞瘤9例、间变性星形细胞瘤14例及多形性胶质母细胞瘤21例,手术后进行MRI评分。评分标准包括肿瘤是否越过中线、有无水肿、不均质性、出血、边界清晰、囊变或坏死、血管流空效应、强化程度和强化的不均质性等9项指标。结果MRI评分随肿瘤病理分级的提高而增加,星形细胞瘤患者的不均质性、水肿、边界清晰、血管流空效应、强化程度和强化的不均质性等6项指标均低于间变性星形细胞瘤和多形性胶质母细胞瘤,其差异具有显著性意义(均P<0.05);而间变性星形细胞瘤与多形性胶质母细胞瘤之间9项指标相比,仅不均质性和强化程度2项差异有显著性意义(P<0.05)。结论MRI评分对选择星形细胞起源肿瘤的治疗方法及判断患者预后具有重要临床参考价值。     

Increased expression of podoplanin in malignant astrocytic tumors as a novel molecular marker of malignant progression

Podoplanin (aggrus) is a mucin-like transmembrane sialoglycoprotein that is expressed on lymphatic endothelial cells. Podoplanin is putatively involved in cancer cell migration, invasion, metastasis, and malignant progression and may be involved in platelet aggregation. Previously, we showed upregulated expression of podoplanin in central nervous system (CNS) germinomas, but not in non-germinomatous germ cell tumors, except for parts of immature teratomas in limited numbers. However, little information exists about its role in CNS astrocytic tumors. In this study, 188 astrocytic tumors (30 diffuse astrocytomas, 43 anaplastic astrocytomas, and 115 glioblastomas) were investigated using immunohistochemistry with an anti-podoplanin antibody, YM-1. In 11 of 43 anaplastic astrocytomas (25.6%) and in 54 of 115 glioblastomas (47.0%), podoplanin was expressed on the surface of anaplastic astrocytoma cells and glioblastoma cells, especially around necrotic areas and proliferating endothelial cells. However, the surrounding brain parenchyma was not stained by YM-1. On the other hand, podoplanin expression was not observed in diffuse astrocytoma (0/30: 0%). Furthermore, we investigated the expression of podoplanin using quantitative real-time PCR and Western blot analysis in 54 frozen astrocytic tumors (6 diffuse astrocytomas, 14 anaplastic astrocytomas, and 34 glioblastomas). Podoplanin mRNA and protein expression were markedly higher in glioblastomas than in anaplastic astrocytomas. These data suggest that podoplanin expression might be associated with malignancy of astrocytic tumors.     

星形细胞起源肿瘤的恶性进展与Topo-Ⅱα表达关系的研究

目的研究拓扑异构酶-Ⅱα(topoisomerase-Ⅱα,Topo-Ⅱα)在星形细胞起源肿瘤中的表达与病理分级的关系,并探讨Topo-Ⅱα对判断此类肿瘤患者预后的价值。方法应用免疫组织化学二步法对49例不同级别的星形细胞起源肿瘤标本Topo-Ⅱα蛋白表达进行分析,以评价阳性细胞百分数与星形细胞起源肿瘤恶性程度的关系。结果随着肿瘤病理级别的提高,细胞异型性明显增加、核染色增强、对比明显;而且Topo-Ⅱα标记指数亦随之升高,低级别(Ⅰ、Ⅱ级)与高级别(Ⅲ、Ⅳ级)肿瘤之间的差异具有显著性意义(P<0.01),但Ⅲ、Ⅳ级肿瘤之间的差异则无显著性意义(P>0.05)。结论Topo-Ⅱα可较为准确地反映星形细胞起源肿瘤的增殖潜能和恶性程度,对判断脑胶质瘤患者的预后具有重要参考价值。     

Conditional entropy as an indicator of pleomorphism in astrocytic tumors

The entropies of nuclear arrangements as an indicator of pleomorphism are assessed using a morphometric method. Sixty astrocytic tumors (grades I, II, III and IV; 15 cases each) were reviewed and analyzed. All slides were stained with HE and MIB‐1 antibody. The MIB‐1 labeling index (LI) was assessed by counting nuclei under a microscope. Images of HE‐stained slides were digitized and segmented using the watershed algorithm. Then, six nuclear parameters were measured automatically: (i) the number of total nuclei in the image, (ii) percentage of total nuclear area in the image, (iii) the mean area of the nucleus, (iv) the standard deviation of the area of the nucleus, (v) the entropy of nuclear arrangement (Entropy_simple), and (vi) conditional entropy of nuclear arrangement (Entropy_conditional). Entropy_simple was defined according to the area of the nucleus and Entropy_conditional was defined according to both the area of the nucleus and the area of its neighboring nuclei. Image processing and image analysis were performed with public domain software developed in the laboratory. Segmentation of the images resulted in inappropriate segmentation in a few percent of the images. The measurements obtained for each parameter were classified using discriminant analysis. The percentage of correct classification with Entropy_conditional was 62%, which was the highest value among all the measurements. Classification based on the combination of all measurements resulted in a rate of correct classification of 88%. Thus, conditional entropy of nuclear arrangement is useful for grading of astrocytic tumors and it is proposed as an indicator of pleomorphism.     

缝隙连接蛋白-43阻断剂辛醇对小鼠脑缺血再灌注损伤的神经保护作用

目的观察小鼠脑缺血再灌注后缝隙连接蛋白-43(Cx43)表达变化及缝隙蛋白阻断剂辛醇对小鼠梗死体积的影响。方法采用线栓法制备小鼠大脑中动脉阻塞模型(MCAO模型),应用免疫印迹(Western blot)方法观察脑缺血再灌注损伤前后Cx43表达变化和辛醇对Cx43表达的影响,并计算各组死亡率及脑梗死体积。结果小鼠脑缺血2h再灌注损伤24h条件下,缺血区Cx43表达在损伤前后无明显变化(P>0.05)。辛醇能显著抑制脑缺血再灌注损伤后Cx43表达,降低死亡率及减少脑梗死体积,具有统计学差异(P<0.05)。结论由Cx43组成的缝隙连接蛋白在脑缺血再灌注损伤过程中具有重要作用,缝隙蛋白-43阻断剂辛醇通过抑制海马及皮质Cx43表达,进而降低死亡率及梗死体积,从而发挥神经保护作用。     

多层螺旋CT灌注成像对星形细胞肿瘤分级的准确性评价

目的 评价多层螺旋CT灌注成像(CTP)参数对星形细胞肿瘤分级的敏感性和特异性.方法 对第三军医大学第三附属医院野战外科研究所收治的53例脑肿瘤患者进行CTP检查,经手术和病理学证实为星形细胞肿瘤的30例患者纳入研究对象.CTP采用GE LightSpeed 64层螺旋CT机行灌注扫描,在AW4.2P后处理工作站对原始数据进行后处理,测定肿瘤最大灌注区和对侧正常组织的脑血流量(CBF)、脑血容量(CBV)、平均通过时间(MTT)及毛细血管表面通透性(PS).手术获取脑肿瘤标本进行组织病理学检查.结果 星形细胞肿瘤高级别组CBF、CBV和PS值均明显高于低级别组,差异有统计学意义(P<0.05),MTT值比较差异无统计学意义(P>0.05).受试者工作特征曲线(ROC曲线)分析表明,CBF、CBV和PS值对鉴别高、低级别星形细胞肿瘤的ROC曲线下面积分别为0.914、0.876和0.914,而MTT无鉴别作用,其ROC曲线下面积为0.455.采用CBF=62.635 mL/(100 g·min),CBV=4.310 mL/100 g和PS=5.925 mL/(100 g·min)作为分界点鉴别高、低级别星形细胞肿瘤的敏感性均为84.2%,特异性分别是81.8%、81.8%和91.9%.结论 多层螺旋CTP参数CBF、CBV及PS值对鉴别高、低级别星形细胞肿瘤具有较高敏感性和特异性.     

实验性蛛网膜下腔出血后脑血管痉挛兔基底动脉Cx43蛋白表达的时相变化

目的 通过建立兔二次蛛网膜下腔出血实验模型,观察兔蛛网膜下腔出血后基底动脉缝隙连接蛋白Cx43表达的时相变化特点,初步探讨蛛网膜下腔出血后脑血管痉挛的形成机制.方法 选择健康新西兰大白兔30只,随机分为5组:正常对照组(n=6)和蛛网膜下腔出血模型组(1d、3d、7d和14d,n=6):建立兔二次蛛网膜下腔出血后脑血管痉挛实验模型,脑血管造影分析基底动脉的直径变化并应用Western Blot检测基底动脉Cx43蛋白的表达变化.对血管直径与Cx43表达变化情况进行相关分析.结果 成功建立兔二次蛛网膜下腔出血模型;脑血管造影显示注血后1d基底动脉即出现痉挛(85.7%±8.6%,P<0.05);7d时达高峰(66.5%±7.6%,P<0.01);14d时仍有痉挛(78.4%±8.2%,P<0.05)但程度较前缓解.Cx43蛋白表达在建立SAH模型后1d(38.6%±5.6%,P<0.05)、3d(50.2%±5.7%,P<0.05)、7d(57.8%±5.3%,P<0.01)、14d(32.4%±3.6%.P<00.05)均升高,其中7d为高峰,14d开始下降.Cx43蛋白表达的时相性变化与SAH后基底动脉直径的时相性变化相关系数为0.914.结论 实验结果 显示蛛网膜下腔出血后兔基底动脉缝隙连接蛋白Cx43的表达发生了时相性变化,并且Cx43蛋白表达强弱与蛛网膜下腔出血后脑血管痉挛程度在时程上存在正相关关系,表明缝隙连接蛋白Cx43可能参与蛛网膜下腔出血后脑血管痉挛的形成.

Abstract：

Objective The study was designed to explore the change of expression of connexin43(Cx43)protien in the model of subarachnoid hemorrhage(SAH)of rabbits,hoping to provide the basis to study the mechanism of cerebral vasospasm(CVS).Methods 30 New Zealand rabbits were divided into 5 groups:SAH group(1d,3d,7d,14d,n=6)and control group(n=6).The model of CVS following SAH was established.Digital subtraction angiography was performed to detect the change of the basilar arteries diameter.The expression of Cx43 protien in basilar arteries tissue at different time points following experimental SAH was examined by using western blotting analysis.The data were statistically analyzed using the bivariate correlations test.Results The model of SAH in rabbits was successfully established.All 30 rabbits were analyzed.Cerebral angiograms on 1d,3d,7d and 14d showed severe narrowing of the BAs,and on 7d showed the most narrowing and on 14d began to Relieve.Western blotting showed that the expression of Cx43 protein were detected in normal rabbit basilar arteries tissue.However,the expression of Cx43 protein increased gradually and significantly in models compared with that of control(P<0.05),which reached peak on 7d(P<0.01)and then decreased on 14d(P<0.05).There was positive correlation between expression of Cx43 and cerebral vasospasm.Conclusions The above results demonstrates at the first time that the Cx43 protein expression is altered after the SAH,and exhibits a time-dependent change.which might be connected with the development of CVS.In summary,our data demonstrates gap junctions may play an important role in the pathogenesis of cerebral vasospasm after SAH.