Direct isolation of myofibroblasts and fibroblasts from bleomycin-injured lungs reveals their functional similarities and differences.
Direct isolation of myofibroblasts and fibroblasts from bleomycin-injured lungs reveals their functional similarities and differences.
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DOI：
10.1186/1755-1536-6-15
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发表时间：
2013-08-08
期刊：
影响因子：
--
通讯作者：
Iwashita T
中科院分区：
文献类型：
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作者：
Akamatsu T;Arai Y;Kosugi I;Kawasaki H;Meguro S;Sakao M;Shibata K;Suda T;Chida K;Iwashita T
肌成纤维细胞在组织修复中起重要作用。肌成纤维细胞和成纤维细胞之间的功能异同尚不完全清楚，因为它们尚未从活体中分离出来。本研究的目的是建立一种利用荧光激活细胞分选直接分离损伤肺中肌成纤维细胞和成纤维细胞的方法，并比较它们的功能。我们证明了谱系特异性细胞表面标记（lin），如CD31、CD45、CD146、EpCAM （CD326）、TER119和Lyve-1在肌成纤维细胞或成纤维细胞中不表达。博来霉素损伤肺和盐水处理肺的成纤维细胞被证明富含衬层Sca-1high，博来霉素损伤肺的肌成纤维细胞被证明富含衬层Sca-1low CD49ehigh。体外增殖试验结果表明，博莱霉素损伤肺的成纤维细胞在体外增殖，而肌成纤维细胞在体外增殖，而盐处理肺的成纤维细胞在体外增殖。然而，成纤维细胞和肌成纤维细胞在体内可能具有较低的增殖能力。通过qRT-PCR对胶原蛋白和胶原合成酶基因的分析发现，博莱霉素损伤肺成纤维细胞和肌成纤维细胞中约一半基因的表达水平显著高于盐水处理肺成纤维细胞。相比之下，11种趋化因子基因中8种在肌成纤维细胞中的表达水平明显低于成纤维细胞。这是第一个直接从损伤肺中分离肌成纤维细胞和成纤维细胞的研究。我们证明了肌成纤维细胞和成纤维细胞在增殖能力和胶原蛋白、胶原合成酶和趋化因子基因表达水平方面的功能相似性和差异性。因此，这种直接分离方法有很大的潜力从肌成纤维细胞和成纤维细胞中获得有用的信息。
Myofibroblasts play a crucial role in tissue repair. The functional similarities and differences between myofibroblasts and fibroblasts are not fully understood because they have not been separately isolated from a living body. The purpose of this study was to establish a method for the direct isolation of myofibroblasts and fibroblasts from injured lungs by using fluorescence-activated cell sorting and to compare their functions. We demonstrated that lineage-specific cell surface markers (lin), such as CD31, CD45, CD146, EpCAM (CD326), TER119, and Lyve-1 were not expressed in myofibroblasts or fibroblasts. Fibroblasts of bleomycin-injured lungs and saline-treated lungs were shown to be enriched in linneg Sca-1high, and myofibroblasts of bleomycin-injured lungs were shown to be enriched in linneg Sca-1low CD49ehigh. Results from in-vitro proliferation assays indicated in-vitro proliferation of fibroblasts but not myofibroblasts of bleomycin-injured lungs and of fibroblasts of saline-treated lungs. However, fibroblasts and myofibroblasts might have a low proliferative capacity in vivo. Analysis of genes for collagen and collagen synthesis enzymes by qRT-PCR showed that the expression levels of about half of the genes were significantly higher in fibroblasts and myofibroblasts of bleomycin-injured lungs than in fibroblasts of saline-treated lungs. By contrast, the expression levels of 8 of 11 chemokine genes of myofibroblasts were significantly lower than those of fibroblasts. This is the first study showing a direct isolation method of myofibroblasts and fibroblasts from injured lungs. We demonstrated functional similarities and differences between myofibroblasts and fibroblasts in terms of both their proliferative capacity and the expression levels of genes for collagen, collagen synthesis enzymes, and chemokines. Thus, this direct isolation method has great potential for obtaining useful information from myofibroblasts and fibroblasts.