Yang X, Skoff R P
Department of Anatomy and Cell Biology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
J Neurosci. 1997 Mar 15;17(6):2056-70. doi: 10.1523/JNEUROSCI.17-06-02056.1997.
Proteolipid protein (PLP) has been postulated to play a critical role in the early differentiation of oligodendrocytes (OLs) in addition to its known role as a structural component of myelin. To identify this early function, we blocked the synthesis of PLP in glial cultures with antisense oligodeoxynucleotides that targeted the PLP initiation codon. Primary glial cultures were incubated with phosphorothioate-protected oligodeoxynucleotides (S-ODNs) for up to 11 d. PLP in OLs was reduced >90%. OLs treated with antisense S-ODNs appeared strikingly healthy as judged by (1) immunocytochemical staining for myelin glycolipids and myelin basic protein, (2) their prolonged survival compared with untreated cultures, and (3) their ability to re-establish membrane sheets after removal of the S-ODNs. Our studies show that PLP is required for elaboration and stability of the myelin membrane sheets made by most OLs, but it is not necessary for the network of processes established by OLs. More importantly, the number of OLs in the antisense-treated cultures was nearly sevenfold greater after a 10-11 d incubation with S-ODNs than in control cultures. The number of proliferating OL progenitors was not increased in the antisense-treated cultures, indicating that the increase in the number of OLs was attributable to prolonged OL survival. The tissue culture studies reveal that the absence of PLP/DM20 has the positive effect of promoting OL survival but the negative effect of preventing their full differentiation. This finding clarifies many of the paradoxical findings seen in the PLP mutants, the PLP overexpressers, and the PLP- animals.
除了作为髓磷脂的结构成分这一已知作用外,人们推测蛋白脂蛋白(PLP)在少突胶质细胞(OLs)的早期分化中起关键作用。为了确定这一早期功能,我们用靶向PLP起始密码子的反义寡脱氧核苷酸阻断神经胶质细胞培养物中PLP的合成。原代神经胶质细胞培养物与硫代磷酸酯保护的寡脱氧核苷酸(S-ODNs)一起孵育长达11天。OLs中的PLP减少了90%以上。通过以下几点判断,用反义S-ODNs处理的OLs看起来非常健康:(1)对髓磷脂糖脂和髓磷脂碱性蛋白进行免疫细胞化学染色;(2)与未处理的培养物相比,它们的存活时间延长;(3)去除S-ODNs后它们重新形成膜片的能力。我们的研究表明,PLP是大多数OLs形成的髓磷脂膜片的精细结构和稳定性所必需的,但对于OLs建立的突起网络则不是必需的。更重要的是,在用S-ODNs孵育10 - 11天后,反义处理培养物中的OLs数量比对照培养物中几乎多了七倍。反义处理培养物中增殖的OL祖细胞数量没有增加,这表明OLs数量的增加归因于OLs存活时间的延长。组织培养研究表明,缺乏PLP/DM20具有促进OLs存活的积极作用,但也有阻止它们完全分化的消极作用。这一发现澄清了在PLP突变体、PLP过表达体和PLP基因敲除动物中看到的许多矛盾发现。