CNRS, Institut de Pharmacologie Moléculaire et Cellulaire, UMR6097, Sophia Antipolis, France.
PLoS One. 2009 Aug 24;4(8):e6718. doi: 10.1371/journal.pone.0006718.
Epithelial-mesenchymal interactions are critical in regulating many aspects of vertebrate embryo development, and for the maintenance of homeostatic equilibrium in adult tissues. The interactions between epithelium and mesenchyme are believed to be mediated by paracrine signals such as cytokines and extracellular matrix components secreted from fibroblasts that affect adjacent epithelia. In this study, we sought to identify the repertoire of microRNAs (miRNAs) in normal lung human fibroblasts and their potential regulation by the cytokines TNF-alpha, IL-1beta and TGF-beta.
METHODOLOGY/PRINCIPAL FINDINGS: MiR-155 was significantly induced by inflammatory cytokines TNF-alpha and IL-1beta while it was down-regulated by TGF-beta. Ectopic expression of miR-155 in human fibroblasts induced modulation of a large set of genes related to "cell to cell signalling", "cell morphology" and "cellular movement". This was consistent with an induction of caspase-3 activity and with an increase in cell migration in fibroblasts tranfected with miR-155. Using different miRNA bioinformatic target prediction tools, we found a specific enrichment for miR-155 predicted targets among the population of down-regulated transcripts. Among fibroblast-selective targets, one interesting hit was keratinocyte growth factor (KGF, FGF-7), a member of the fibroblast growth factor (FGF) family, which owns two potential binding sites for miR-155 in its 3'-UTR. Luciferase assays experimentally validated that miR-155 can efficiently target KGF 3'-UTR. Site-directed mutagenesis revealed that only one out of the 2 potential sites was truly functional. Functional in vitro assays experimentally validated that miR-155 can efficiently target KGF 3'-UTR. Furthermore, in vivo experiments using a mouse model of lung fibrosis showed that miR-155 expression level was correlated with the degree of lung fibrosis.
CONCLUSIONS/SIGNIFICANCE: Our results strongly suggest a physiological function of miR-155 in lung fibroblasts. Altogether, this study implicates this miRNA in the regulation by mesenchymal cells of surrounding lung epithelium, making it a potential key player during tissue injury.
上皮-间充质相互作用在调节脊椎动物胚胎发育的许多方面以及维持成人组织的体内平衡方面至关重要。上皮和间充质之间的相互作用被认为是通过旁分泌信号介导的，例如成纤维细胞分泌的细胞因子和细胞外基质成分，这些信号影响相邻的上皮细胞。在这项研究中，我们试图确定正常肺成纤维细胞中的 microRNA（miRNA）谱及其受细胞因子 TNF-α、IL-1β和 TGF-β调节的可能性。
方法/主要发现：miR-155 被炎症细胞因子 TNF-α和 IL-1β显著诱导，而被 TGF-β下调。在人成纤维细胞中异位表达 miR-155 诱导了与“细胞间信号”、“细胞形态”和“细胞运动”相关的一大组基因的调节。这与 caspase-3 活性的诱导以及转染 miR-155 的成纤维细胞中细胞迁移增加一致。使用不同的 miRNA 生物信息学靶标预测工具，我们发现 miR-155 预测靶标在下调转录本群体中存在特定富集。在成纤维细胞选择性靶标中，一个有趣的命中是角质细胞生长因子（KGF，FGF-7），它是成纤维细胞生长因子（FGF）家族的成员，在其 3'UTR 中有两个潜在的 miR-155 结合位点。荧光素酶测定实验验证了 miR-155 可以有效地靶向 KGF 3'UTR。定点突变揭示了 2 个潜在位点中只有 1 个是真正功能的。体外功能实验实验验证了 miR-155 可以有效地靶向 KGF 3'UTR。此外，使用肺纤维化小鼠模型的体内实验表明，miR-155 的表达水平与肺纤维化的程度相关。
结论/意义：我们的结果强烈表明 miR-155 在肺成纤维细胞中具有生理功能。总之，这项研究表明该 miRNA 参与了间质细胞对周围肺上皮的调节，使其成为组织损伤过程中的潜在关键参与者。
