CD146 Defines Commitment of Cultured Annulus Fibrosus Cells to Express a Contractile Phenotype
CD146 Defines Commitment of Cultured Annulus Fibrosus Cells to Express a Contractile Phenotype
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DOI：
10.1002/jor.23326
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发表时间：
2016-08-01
影响因子：
2.8
通讯作者：
Watanabe, Masahiko
中科院分区：
文献类型：
--
作者：
Nakai, Tomoko;Sakai, Daisuke;Watanabe, Masahiko
细胞的表征对于促进椎间盘退行性疾病的基于细胞的治疗是重要的。为此，我们分析了小鼠纤维环细胞的流式细胞仪检测其原代培养的表型变化。在检查了16种细胞表面蛋白后，我们将重点放在仅在培养扩增期间增加的CD 146上。已知CD 146是间充质干细胞的标志物，并且是它们的血管平滑肌定型的标志物，其中SM 22 α增强了收缩表型的表达。我们对CD 146+细胞进行了分类，以阐明它们的特征以及在这种变化中发挥作用的关键因素。全细胞培养物显示出向间充质谱系三能分化的能力，而分选的CD 146+细胞则没有。加入转化生长因子β 1可提高CD 146的表达，分选后的CD 146+细胞比CD 146-细胞表达更高水平的SM 22 α和弹性蛋白mRNA。在形态学上，CD 146+细胞比CD 146-细胞更广泛地沉积细胞外I型胶原，并显示丝状肌动蛋白束穿过其细胞质和细胞-细胞连接。此外，CD 146+细胞表现出显着更高的凝胶收缩性能比CD 146-细胞时，他们包埋在胶原凝胶。人纤维环CD 146+细胞也表现出更高的收缩性。免疫组织化学确定了定位于具有SM 22 α共表达的小鼠椎间盘组织的最外层环层的CD 146+细胞。这些结果表明，CD 146表达的增加表明培养的纤维环细胞逐渐改变以表达收缩表型，并且转化生长因子β 1增强这种细胞定型。(C)2016骨科研究学会。由威利期刊公司出版
Characterization of cells is important for facilitating cell-based therapies for degenerative diseases of intervertebral discs. For this purpose, we analyzed mouse annulus fibrosus cells by flowcytometory to detect phenotypic change in their primary cultures. After examination of sixteen cell surface proteins, we focused on CD146 that solely increased during culture expansion. CD146 is known to be a marker for mesenchymal stem cells and for their vascular smooth muscle commitment with expression of contractile phenotype enhanced by SM22 alpha. We sorted CD146+ cells to elucidate their characteristics and the key factors that play a role in this change. Whole cell cultures showed the ability for tripotent differentiation toward mesenchymal lineages, whereas sorted CD146+ cells did not. Expression of CD146 was elevated by addition of transforming growth factor beta 1, and sorted CD146+ cells expressed higher levels of mRNA for SM22 alpha and Elastin than did CD146- cells. Morphologically, CD146+ cells more broadly deposited extracellular type I collagen than CD146- cells and showed filamentous actin bundles traversing their cytoplasm and cell-cell junctions. Moreover, CD146+ cells demonstrated significantly higher gel contraction properties than CD146- cells when they were embedded in collagen gels. Human annulus fibrosus CD146+ cells also showed higher contractility. Immunohistochemistry determined CD146+ cells localized to the outermost annulus layers of mouse intervertebral disc tissue with co-expression of SM22 alpha. These results suggest that increment of CD146 expression indicates gradual change of cultured annulus fibrosus cells to express a contractile phenotype and that transforming growth factor beta 1 enhances this cellular commitment. (C) 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.