Tucker R P
Department of Neurobiology and Anatomy, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, NC 27157-1010.
Development. 1993 Jan;117(1):347-58. doi: 10.1242/dev.117.1.347.
Tenascin and thrombospondin belong to the growing family of extracellular matrix glycoproteins believed to have an anti-adhesive function during development. Immunohistochemistry has been used to identify these proteins in the developing central nervous system, in the matrix surrounding peripheral neurons, and in connective tissue. The antibodies used in most of these studies, however, could not distinguish between different splice variants (tenascin) nor different genetic forms (thrombospondin). For this reason, we used the reverse transcriptase polymerase chain reaction to generate DNA probes that are specific to the transcripts of high M(r) tenascin and thrombospondin 2. These probes were then used for an in situ hybridization study to determine the cellular origins of specific tenascin and thrombospondin forms throughout the development of the chick. The mRNA encoding high M(r) tenascin was found associated with motile cells and in tissues undergoing dynamic modeling: migrating glia, epithelial glia used as a substratum for migrating neurons, the growing tips of lung buds, and during osteogenesis. In contrast, the mRNAs of low M(r) tenascin were concentrated in areas of cartilage deposition and chondrocyte proliferation. Thrombospondin 2 mRNA was not detected in the developing central nervous system at any time during development by in situ hybridization. In contrast, it was found in embryonic mesenchyme, perichondrium, epimysium, and endothelial cells. Thrombospondin 2 mRNA was detected in poly(A) RNA isolated from embryonic spinal cord and cerebellum by polymerase chain reaction, though it was not detected in poly(A) RNA from the avascular retina. Thus, thrombospondin 2 mRNA may be present in the developing brain at low levels in endothelial cells or blood cells. These data support the notion that tenascin splice variants have distinct roles during development, and that thrombospondin 2 is more likely to be playing a role associated with the morphogenesis of connective tissue than neuronal development.
腱生蛋白和血小板反应蛋白属于细胞外基质糖蛋白家族，该家族成员在发育过程中被认为具有抗黏附功能。免疫组织化学已被用于在发育中的中枢神经系统、周围神经元周围的基质以及结缔组织中鉴定这些蛋白质。然而，在大多数这些研究中使用的抗体无法区分不同的剪接变体（腱生蛋白）或不同的基因形式（血小板反应蛋白）。因此，我们使用逆转录酶聚合酶链反应来生成对高M(r)腱生蛋白和血小板反应蛋白2的转录本特异的DNA探针。然后将这些探针用于原位杂交研究，以确定在鸡的整个发育过程中特定腱生蛋白和血小板反应蛋白形式的细胞起源。发现编码高M(r)腱生蛋白的mRNA与运动细胞以及经历动态重塑的组织相关：迁移的神经胶质细胞、用作迁移神经元基质的上皮神经胶质细胞、肺芽的生长尖端以及骨发生过程中。相比之下，低M(r)腱生蛋白的mRNA集中在软骨沉积和软骨细胞增殖区域。在发育过程中的任何时候，通过原位杂交在发育中的中枢神经系统中均未检测到血小板反应蛋白2 mRNA。相反，它存在于胚胎间充质、软骨膜、肌外膜和内皮细胞中。通过聚合酶链反应在从胚胎脊髓和小脑中分离的聚腺苷酸RNA中检测到了血小板反应蛋白2 mRNA，尽管在无血管视网膜的聚腺苷酸RNA中未检测到。因此，血小板反应蛋白2 mRNA可能以低水平存在于发育中的大脑的内皮细胞或血细胞中。这些数据支持这样的观点，即腱生蛋白剪接变体在发育过程中具有不同的作用，并且血小板反应蛋白2更有可能在结缔组织的形态发生而非神经元发育中发挥作用。
