Department of Pediatrics and Communicable Diseases, University of Michigan Medical School, Ann Arbor, Michigan 48109-5688, USA.
Am J Physiol Lung Cell Mol Physiol. 2010 Jun;298(6):L735-43. doi: 10.1152/ajplung.00347.2009. Epub 2010 Feb 26.
We have isolated mesenchymal stem cells (MSCs) from tracheal aspirates of premature infants with respiratory distress. We examined the capacity of MSCs to differentiate into myofibroblasts, cells that participate in lung development, injury, and repair. Gene expression was measured by array, qPCR, immunoblot, and immunocytochemistry. Unstimulated MSCs expressed mRNAs encoding contractile (e.g., ACTA2, TAGLN), extracellular matrix (COL1A1 and ELN), and actin-binding (DBN1, PXN) proteins, consistent with a myofibroblast phenotype, although there was little translation into immunoreactive protein. Incubation in serum-free medium increased contractile protein (ACTA2, MYH11) gene expression. MSC-conditioned medium showed substantial levels of TGF-beta1, and treatment of serum-deprived cells with a type I activin receptor-like kinase inhibitor, SB-431542, attenuated the expression of genes encoding contractile and extracellular matrix proteins. Treatment of MSCs with TGF-beta1 further induced the expression of mRNAs encoding contractile (ACTA2, MYH11, TAGLN, DES) and extracellular matrix proteins (FN1, ELN, COL1A1, COL1A2), and increased the protein expression of alpha-smooth muscle actin, myosin heavy chain, and SM22. In contrast, human bone marrow-derived MSCs failed to undergo TGF-beta1-induced myofibroblastic differentiation. Finally, primary cells from tracheal aspirates behaved in an identical manner as later passage cells. We conclude that human neonatal lung MSCs demonstrate an mRNA expression pattern characteristic of myofibroblast progenitor cells. Autocrine production of TGF-beta1 further drives myofibroblastic differentiation, suggesting that, in the absence of other signals, fibrosis represents the "default program" for neonatal lung MSC gene expression. These data are consistent with the notion that MSCs play a key role in neonatal lung injury and repair.
我们从患有呼吸窘迫症的早产儿的气管吸出物中分离出间充质干细胞 (MSCs)。我们研究了 MSCs 分化为肌成纤维细胞的能力，肌成纤维细胞参与肺的发育、损伤和修复。通过基因芯片、qPCR、免疫印迹和免疫细胞化学检测基因表达。未刺激的 MSCs 表达编码收缩（如 ACTA2、TAGLN）、细胞外基质（COL1A1 和 ELN）和肌动蛋白结合（DBN1、PXN）蛋白的 mRNA，这与肌成纤维细胞表型一致，尽管很少有翻译为免疫反应性蛋白。在无血清培养基中孵育会增加收缩蛋白 (ACTA2、MYH11) 的基因表达。MSC 条件培养基显示出高水平的 TGF-β1，并且用 I 型激活素受体样激酶抑制剂 SB-431542 处理血清剥夺的细胞会减弱编码收缩和细胞外基质蛋白的基因的表达。用 TGF-β1 处理 MSCs 会进一步诱导编码收缩（ACTA2、MYH11、TAGLN、DES）和细胞外基质蛋白（FN1、ELN、COL1A1、COL1A2）的 mRNA 的表达，并增加 α-平滑肌肌动蛋白、肌球蛋白重链和 SM22 的蛋白表达。相比之下，人骨髓来源的 MSCs 无法进行 TGF-β1 诱导的肌成纤维细胞分化。最后，气管吸出物中的原代细胞与后期传代细胞的行为方式相同。我们得出结论，人新生儿肺 MSCs 表现出肌成纤维细胞祖细胞特征的 mRNA 表达模式。自分泌产生的 TGF-β1 进一步促进肌成纤维细胞分化，这表明在没有其他信号的情况下，纤维化代表了新生儿肺 MSC 基因表达的“默认程序”。这些数据与 MSCs 在新生儿肺损伤和修复中发挥关键作用的观点一致。
