Dissecting the roles of endothelin, TGF-β and GM-CSF on myofibroblast differentiation by keratinocytes
Dissecting the roles of endothelin, TGF-β and GM-CSF on myofibroblast differentiation by keratinocytes
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DOI：
10.1160/th03-11-0669
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发表时间：
2004-08-01
影响因子：
6.7
通讯作者：
Smola, H
中科院分区：
文献类型：
--
作者：
Shephard, P;Hinz, B;Smola, H
肌成纤维细胞是一种特殊的成纤维细胞，通过产生细胞外基质和收缩肉芽组织来促进伤口愈合。当真皮与活化的表皮角质形成细胞强烈相互作用时，它们出现在伤口愈合的阶段。与角化细胞直接共培养可上调tgf - β活性，并诱导成纤维细胞分化为α -平滑肌肌动蛋白（alphaSMA）阳性的肌成纤维细胞。tgf - β活性本身并不能完全解释这些共培养中aSMA的诱导，在这里，我们分析了机械力的产生，这是该模型中肌成纤维细胞分化的另一个有效诱导剂。使用可变形的硅胶基质，我们发现在共培养1-2天后，成纤维细胞的收缩活性已经被诱导，此时成纤维细胞通常是alphaSMA阴性。内皮素-1 （ET-1）是平滑肌细胞收缩最有效的诱导剂，在共培养中被上调，而用ET受体抑制剂PD 156252阻断ET-1可以抑制这些早期共培养中的收缩。然而，在共培养的4-5天，成纤维细胞的收缩活性与alphaSMA表达的增加相关。在低血清培养基中用ET-1刺激成纤维细胞单培养物不诱导aSMA表达；然而，ET-1确实与tgf - β协同作用。令人惊讶的是，另一种刺激肉芽组织中肌成纤维细胞分化的介质GM-CSF与tgf - β和ET-1共同刺激，抑制了成纤维细胞中aSMA的表达。GM-CSF激活NFkappaB，从而干扰tgf - β信号。阻断tgf - β和ET-1在第7天的共培养中很大程度上损害了alphasMA的诱导，并且几乎完全阻止了alphasMA的诱导。我们的研究结果分析了tgf - β和ET-1在角化细胞-fib末共培养中产生机械力的作用，并确定GM-CSF是间接作用的肌成纤维细胞的诱导剂。
Myofibroblasts are specialized fibroblasts that contribute to wound healing by producing extracellular matrix and by contracting the granulation tissue. They appear in a phase of wound healing when the dermis strongly interacts with activated epidermal keratinocytes. Direct co-culture with keratinocytes upregulates TGF-beta activity and also induces fibroblast to differentiate into alpha-smooth muscle actin (alphaSMA)-positive myofibroblasts. TGF-beta activity alone cannot completely account for aSMA induction in these co-cultures, and here we analyze mechanical force generation, another potent inducer of myofibroblast differentiation in this model. Using deformable silicone substrates, we show that contractile activity of fibroblasts is already induced after 1-2-days of co-culture, when fibroblasts are generally alphaSMA negative. Endothelin-1 (ET-1), the most potent inducer of smooth muscle cell contraction, was up-regulated in co-cultures, while blocking ET-1 with the ET receptor inhibitor PD 156252 inhibited contraction in these early co-cultures. In 4-5 days of co-culture, however, fibroblast contractile activity correlated with an increased expression of alphaSMA expression. Stimulation of fibroblast mono-cultures with ET-1 in a low serum medium did not induce aSMA expression; however, ET-1 did synergize with TGF-beta. Surprisingly, GM-CSF, another mediatorstimulating myofibroblast differentiation in granulation tissue, inhibited aSMA expression in fibroblasts, co-stimulated with TGF-beta and ET-1. GM-CSF activated NFkappaB, thus interfering with TGF-beta signaling. Blocking TGF-beta and ET-1 largely impaired alphasMA induction in co-cultures at day 7 and, in combination, almost completely prevented alphaSMA induction. Our results dissect the roles of TGF-beta and ET-1 on mechanical force generation in keratinocyte-fib rob last co-cultures, and identify GM-CSF as an inducer of myofibroblasts acting indirectly.