EXPRESSION AND DISTRIBUTION OF GAP-43 IN HUMAN ASTROCYTES IN CULTURE
EXPRESSION AND DISTRIBUTION OF GAP-43 IN HUMAN ASTROCYTES IN CULTURE
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DOI：
10.1111/j.1365-2990.1995.tb01071.x
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发表时间：
1995-08-01
影响因子：
5
通讯作者：
KIM, SU
中科院分区：
文献类型：
--
作者：
MORETTO, G;XU, RY;KIM, SU
通过结合 mRNA 分析和免疫细胞化学，我们研究了从人胎儿大脑混合培养物中获得的星形胶质细胞富集群中生长相关蛋白 43 (GAP-43) 的表达。从细胞沉淀中提取总细胞RNA并反转录成cDNA；使用 GAP-43 特异性引物对 cDNA 进行 PCR 扩增，并通过聚丙烯酰胺凝胶分离 PCR 产物。使用神经胶质原纤维酸性蛋白 (GFAP) 和 GAP-43 的抗体对分离的细胞培养物进行双重免疫荧光染色。结果表明，GAP-43 的转录和翻译均发生在培养的星形胶质细胞中。在体外维持的早期阶段，在 GFAP 阳性星形胶质细胞的细胞突起和细胞质中检测到 GAP-43 免疫反应物质。在较旧的培养物中，GAP-43 免疫反应性持续存在于大部分细胞中，并有在核周区域积累的趋势。这些观察结果证明 GAP-43 并不局限于神经元细胞。在星形胶质细胞中观察到的与细胞骨架成分的密切空间关联表明该蛋白质在控制细胞形状、运动和粘附过程中发挥作用。
By combining mRNA analysis and immunocytochemistry, we investigated the expression of the growth associated protein 43 (GAP-43) in enriched populations of astrocytes, obtained from mixed cultures of human fetal brains. Total cellular RNA was extracted from cell pellets and reverse transcribed into cDNA; cDNA was subjected to PCR amplification using primers specific for GAP-43 and PCR products were separated through polyacrylamide gels. Double immunofluorescence staining was performed on dissociated cell cultures using antibodies to glial fibrillary acidic protein (GFAP) and to GAP-43. Results showed that both transcription and translation for GAP-43 occur in cultured astrocytes. GAP-43 immunoreacting material was detected in the cell processes and diffusely in the cytoplasm of GFAP-positive astrocytes, during early stages of maintenance in vitro. In older cultures, GAP-43 immunoreactivity persisted in a large percentage of cells, with a tendency to accumulate in perinuclear areas. These observations provide evidence that GAP-43 is not restricted to neuronal cells. The close spatial association with cytoskeletal constituents, as observed in astrocytes, suggests a role for this protein in the control of cell shape, motility and adhesion processes.