Morphological changes and regulation mechanisms of oligodendrocyte migration and myelination

用谱系标记抗体O4和O1检测少突胶质前体细胞的迁移和增殖。(1)后脑(脑干和小脑)中的少突胶质前体细胞从内侧脑干、外侧部分逐渐扩散到小脑原基。为了验证OPC的迁移能力，将编码EGFP基因的质粒导入E4或E5的后脑室细胞，观察O4阳性/EGFP阳性细胞的出现和扩散情况。将该质粒转入内侧脑室层，在脑干和小脑出现O4阳性/EGFP阳性细胞。这一结果表明OPC出现在脑干内侧脑室层，至少有一部分OPC可以从脑干迁移到小脑原基。(2)用溴脱氧尿苷(BrdU)和O4或O1双标记法检测OPC的增殖情况。大约20%的O4阳性细胞含有BrdU，而40%-50%的O4阳性细胞在这些区域出现后立即在实质内标记。随着胚胎的发育，标记指数逐渐降低。最后，不到5%的成熟少突胶质细胞被BrdU标记。少突胶质细胞系细胞离开脑室层后，可能在实质内最活跃地生成它们的队列。(3)用细胞表面识别分子的中和抗体检测调控OPC迁移的分子。将分泌抗体的杂交瘤细胞注射到E5鸡胚胎的第三脑室，观察E8时O4阳性细胞在视神经中的分布。当注射产生N-钙粘素抗体的杂交瘤细胞时，O4阳性细胞数比对照胚胎减少了50%。N-钙粘附素可能参与了OPC在视神经中的迁移。

Migration and proliferation of oligodendrocyte precursor cells (OPCs) were examined with lineage marker antibodies O4 and O1.(1) OPCs in the metencephalon (brainstem and cerebellum) undergo gradual dispersal from the medial brainstem, the lateral part and finally to the cerebellar anlage. To verify the migration of OPCs, plasmid encoding EGFP gene was transfected to the metencephalon ventricular cells on E4 or E5, and appearance and dispersal of O4-positive/EGFP-positive cells were examined sequentially. Transfection of the plasmid to the medial ventricular layer resulted in appearance of O4-positive/EGFP-positive cells in the brainstem and also in the cerebellum. This result indicates that OPCs arise in the medial brainstem ventricular layer and that at least some fractions of OPCs may migrate to the cerebellar anlage from the brainstem.(2) Proliferation of OPCs was examined by double-labeling of OPCs with bromodeoxyuridine (BrdU) and with O4 or O1. Approximately 20 % of O4-positive ventricular cells incorporated BrdU, while 40-50 % of O4-positive cells was labeled in the parenchyma right after their appearance in these regions. Labeling indices gradually decreased as embryo developed. Finally, less than 5 % of mature oligodendrocytes were labeled with BrdU. Oligodendrocyte lineage cells may generate their cohorts most actively in the parenchyma just after they leave from the ventricular layer.(3) Molecules regulating the OPC migration were examined with neutralizing antibodies against cell surface recognition molecules. Hybridomas secreting antibody was injected into the third ventricle of the E5 chick embryo and O4-positive cell distribution in the optic nerve was examined at E8. When the embryos were received injection of the hybridoma generating the antibody against N-cadherin, O4-positive cell number decreased to 50 % as compared to the control embryos. N-cadherin may be involved in the OPC migration in the optic nerve.