Department of Medicine, VA Healthcare Center, San Diego, California, United States of America.
PLoS One. 2011;6(10):e25497. doi: 10.1371/journal.pone.0025497. Epub 2011 Oct 5.
Although C/EBPβ(ko) mice are refractory to Bleomycin-induced lung fibrosis the molecular mechanisms remain unknown. Here we show that blocking the ribosomal S-6 kinase (RSK) phosphorylation of the CCAAT/Enhancer Binding Protein (C/EBP)-β on Thr217 (a RSK phosphoacceptor) with either a single point mutation (Ala217), dominant negative transgene or a blocking peptide containing the mutated phosphoacceptor ameliorates the progression of lung injury and fibrosis induced by Bleomycin in mice.
METHODOLOGY/PRINCIPAL FINDINGS: Mice expressing the non-phosphorylatable C/EBPβ-Ala217 transgene had a marked reduction in lung injury on day-13 after Bleomycin exposure, compared to C/EBPβ(wt) mice, judging by the decrease of CD68(+) activated monocytes/macrophages, bone marrow-derived CD45(+) cells and lung cytokines as well as by the normal surfactant protein-C expression by lung pneumocytes. On day-21 after Bleomycin treatment, C/EBPβ(wt) mice but not mice expressing the dominant negative C/EBPβ-Ala217 transgene developed severe lung fibrosis as determined by quantitative collagen assays. All mice were of identical genetic background and back-crossed to the parental wild-type inbreed FVB mice for at least ten generations. Treatment of C/EBPβ(wt) mice with a cell permeant, C/EBPβ peptide that inhibits phosphorylation of C/EBPβ on Thr217 (40 µg instilled intracheally on day-2 and day-6 after the single Bleomycin dose) also blocked the progression of lung injury and fibrosis induced by Bleomycin. Phosphorylation of human C/EBPβ on Thr266 (human homologue phosphoacceptor) was induced in collagen-activated human lung fibroblasts in culture as well as in activated lung fibroblasts in situ in lungs of patients with severe lung fibrosis but not in control lungs, suggesting that this signaling pathway may be also relevant in human lung injury and fibrosis.
CONCLUSIONS/SIGNIFICANCE: These data suggest that the RSK-C/EBPβ phosphorylation pathway may contribute to the development of lung injury and fibrosis.
尽管 C/EBPβ（ko）小鼠对博莱霉素诱导的肺纤维化有抗性，但分子机制尚不清楚。在这里，我们表明，用单点突变（Ala217）、显性负转移基因或含有突变磷酸受体的阻断肽阻断核糖体 S-6 激酶（RSK）对 CCAAT/增强子结合蛋白（C/EBP）-β的 Thr217 磷酸化（RSK 磷酸受体）可改善博莱霉素诱导的小鼠肺损伤和纤维化的进展。
方法/主要发现：与 C/EBPβ（wt）小鼠相比，表达非磷酸化 C/EBPβ-Ala217 转基因的小鼠在博莱霉素暴露后第 13 天肺损伤明显减少，这可以通过减少 CD68（+）激活的单核细胞/巨噬细胞、骨髓源性 CD45（+）细胞和肺细胞因子以及肺肺泡细胞正常表面活性蛋白-C 表达来判断。在用博莱霉素处理后的第 21 天，C/EBPβ（wt）小鼠而非表达显性负 C/EBPβ-Ala217 转移基因的小鼠发展为严重的肺纤维化，这可以通过定量胶原测定来确定。所有小鼠都具有相同的遗传背景，并至少回交到亲本野生型近交 FVB 小鼠十代以上。用一种细胞渗透的、抑制 C/EBPβ Thr217 磷酸化的 C/EBPβ 肽（在单次博莱霉素剂量后第 2 天和第 6 天经气管内滴注 40µg）处理 C/EBPβ（wt）小鼠也阻断了博莱霉素诱导的肺损伤和纤维化的进展。在胶原激活的人肺成纤维细胞以及严重肺纤维化患者肺组织中激活的肺成纤维细胞原位中诱导人 C/EBPβ Thr266 磷酸化（人同源磷酸受体），但在对照肺中没有诱导，这表明该信号通路在人肺损伤和纤维化中也可能相关。
结论/意义：这些数据表明，RSK-C/EBPβ 磷酸化途径可能有助于肺损伤和纤维化的发展。
