Soroceanu L, Manning T J, Sontheimer H
Department of Neurobiology, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Glia. 2001 Feb;33(2):107-17. doi: 10.1002/1098-1136(200102)33:2<107::aid-glia1010>3.0.co;2-4.
Gap junctions are an important means for intercellular communication during development, processes of tissue differentiation, and in maintenance of adult tissue homeostasis. We investigated the expression levels and distribution of connexin-43 (Cx-43), the most abundant astrocytic gap junction protein, in acutely isolated astrocytes and glioma cells from biopsy tissue obtained from patients diagnosed with glioblastoma multiforme (GBM), low-grade astrocytomas (LGAs), or mesial temporal lobe epilepsy. Western blot and immunohistochemical analyses indicated an inverse correlation between the amount of Cx-43 protein and tumor malignancy grade, as assessed by calculating tissue mitotic indexes (MI) obtained using anti-Ki-67 nuclear antigen staining. Samples from epilepsy patients had a low MI and were intensely positive for Cx-43 staining, while LGA tissue samples exhibited moderate staining for Cx-43 and average MI, and GBM biopsies showed significantly lower levels of Cx-43 and high MI. Functional coupling was assayed using fluorescence recovery after photobleach (FRAP). We found that cells from glioma cell lines and primary cultures of human astrocytes from GBM tissues displayed significantly lower degrees of gap junction intercellular communication (GJIC) as indicated by longer and less complete recovery from photobleaching. Mean recovery values were GBM 23.8% +/- 11.4%, LGA 49.4% +/- 47%, and nontumor astrocytes 67.2% +/- 8.4%. Western blot analysis of several human glioma cell lines and tissue biopsies showed variable expression levels of Cx-43, which correlated negatively with the extent of recovery in the same samples. Taken together, our findings suggest that high-grade brain tumors show reduced intercellular communication and a decrease in connexin-43 protein levels.
缝隙连接是发育过程、组织分化过程以及维持成人组织内环境稳定期间细胞间通讯的重要方式。我们研究了连接蛋白43（Cx-43）的表达水平和分布，Cx-43是星形胶质细胞中最丰富的缝隙连接蛋白，在从多形性胶质母细胞瘤（GBM）、低级别星形细胞瘤（LGA）或内侧颞叶癫痫患者活检组织中急性分离出的星形胶质细胞和胶质瘤细胞中进行研究。蛋白质免疫印迹和免疫组织化学分析表明，通过计算使用抗Ki-67核抗原染色获得的组织有丝分裂指数（MI）评估，Cx-43蛋白量与肿瘤恶性程度呈负相关。癫痫患者的样本MI较低，Cx-43染色呈强阳性，而LGA组织样本Cx-43染色呈中度，MI平均，GBM活检显示Cx-43水平显著降低且MI高。使用光漂白后荧光恢复（FRAP）测定功能偶联。我们发现，GBM组织的胶质瘤细胞系和人星形胶质细胞原代培养物中的细胞显示出明显较低程度的缝隙连接细胞间通讯（GJIC），光漂白恢复时间更长且更不完全即表明了这一点。平均恢复值为GBM 23.8%±11.4%，LGA 49.4%±47%，非肿瘤星形胶质细胞67.2%±8.4%。对几种人胶质瘤细胞系和组织活检的蛋白质免疫印迹分析显示Cx-43表达水平各异，其与同一样本中的恢复程度呈负相关。综上所述，我们的研究结果表明，高级别脑肿瘤显示细胞间通讯减少以及连接蛋白43蛋白水平降低。
