The novel monoclonal antibody 9F5 reveals expression of a fragment of GPNMB/osteoactivin processed by furin-like protease(s) in a subpopulation of microglia in neonatal rat brain.
The novel monoclonal antibody 9F5 reveals expression of a fragment of GPNMB/osteoactivin processed by furin-like protease(s) in a subpopulation of microglia in neonatal rat brain.
复制标题
DOI：
10.1002/glia.23034
复制
发表时间：
2016-11
期刊：
影响因子：
6.2
通讯作者：
Nakayama, Hitoshi
中科院分区：
文献类型：
--
作者：
Kawahara, Kohichi;Hirata, Hiroshi;Ohbuchi, Kengo;Nishi, Kentaro;Maeda, Akira;Kuniyasu, Akihiko;Yamada, Daisuke;Maeda, Takehiko;Tsuji, Akihiko;Sawada, Makoto;Nakayama, Hitoshi
为了区分小胶质细胞（MG）的亚型，我们开发了一种针对大鼠原发性MG的一种亚型（1型）的新型单克隆抗体9 F5。在Western blot和免疫细胞化学分析中，9 F5对该细胞类型显示出高选择性，并且与大鼠腹腔巨噬细胞（Mφ）无交叉反应。我们鉴定了9 F5的抗原分子：大鼠糖蛋白非转移性黑素瘤蛋白B（GPNMB）/骨激活素的50 - 70-kDa片段，起始于Lys 170。此外，9 F5与GPNMB的免疫反应性取决于弗林蛋白酶样蛋白酶的活性。更重要的是，大鼠1型MG表达GPNMB片段，而2型MG和Mφ不表达，尽管所有这些细胞都表达GPNMB的mRNA和全长蛋白。这些结果表明，9 F5与MG的反应性在很大程度上取决于GPNMB的切割，并且与2型MG和Mφ相反，1型MG可能具有用于GPNMB切割的弗林蛋白酶样蛋白酶。在新生大鼠脑中，在包括前脑脑室下区、胼胝体和视网膜的特定脑区中观察到变形样9 F5 + MG。用9 F5抗体和抗Iba 1抗体（其在整个CNS中与MG反应）进行的双免疫染色显示，9 F5 + MG是Iba 1 + MG的一部分，表明MG亚型存在于体内。我们认为，9 F5是一个有用的工具，区分大鼠1型MG和MG/Mφ的其他亚型，并揭示GPNMB片段在脑发育过程中的作用。GLIA 2016;64：1938-1961
To differentiate subtypes of microglia (MG), we developed a novel monoclonal antibody, 9F5, against one subtype (type 1) of rat primary MG. The 9F5 showed high selectivity for this cell type in Western blot and immunocytochemical analyses and no cross‐reaction with rat peritoneal macrophages (Mφ). We identified the antigen molecule for 9F5: the 50‐ to 70‐kDa fragments of rat glycoprotein nonmetastatic melanoma protein B (GPNMB)/osteoactivin, which started at Lys170. In addition, 9F5 immunoreactivity with GPNMB depended on the activity of furin‐like protease(s). More important, rat type 1 MG expressed the GPNMB fragments, but type 2 MG and Mφ did not, although all these cells expressed mRNA and the full‐length protein for GPNMB. These results suggest that 9F5 reactivity with MG depends greatly on cleavage of GPNMB and that type 1 MG, in contrast to type 2 MG and Mφ, may have furin‐like protease(s) for GPNMB cleavage. In neonatal rat brain, amoeboid 9F5+ MG were observed in specific brain areas including forebrain subventricular zone, corpus callosum, and retina. Double‐immunοstaining with 9F5 antibody and anti‐Iba1 antibody, which reacts with MG throughout the CNS, revealed that 9F5+ MG were a portion of Iba1+ MG, suggesting that MG subtype(s) exist in vivo. We propose that 9F5 is a useful tool to discriminate between rat type 1 MG and other subtypes of MG/Mφ and to reveal the role of the GPNMB fragments during developing brain. GLIA 2016;64:1938–1961
登录
查看更多内容
影响因子：
3.7
作者：
Bouchut A;Chawla AR;Jeffers V;Hudmon A;Sullivan WJ Jr
通讯作者：
Sullivan WJ Jr
影响因子：
2.5
作者：
IMAMOTO, K;LEBLOND, CP
通讯作者：
LEBLOND, CP
影响因子：
6
作者：
Abdelmagid, Samir M.;Belcher, Joyce Y.;Safadi, Fayez F.
通讯作者：
Safadi, Fayez F.
影响因子：
2.5
作者：
Bulloch, Karen;Miller, Melinda M.;McEwen, Bruce S.
通讯作者：
McEwen, Bruce S.
影响因子：
15.9
作者：
Kreutzberg, GW
通讯作者：
Kreutzberg, GW