Down-regulation of collagen and connective tissue growth factor expression with hepatocyte growth factor in lung fibroblasts from white scleroderma patients via two signaling pathways
Down-regulation of collagen and connective tissue growth factor expression with hepatocyte growth factor in lung fibroblasts from white scleroderma patients via two signaling pathways
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DOI：
10.1002/art.22874
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发表时间：
2007-10-01
影响因子：
--
通讯作者：
Silver, Richard M.
中科院分区：
文献类型：
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作者：
Bogatkevich, Galina S.;Ludwicka-Bradley, Anna;Silver, Richard M.
Objective.目的研究肝细胞生长因子（HGF）下调硬皮病肺成纤维细胞胶原和结缔组织生长因子（CTGF）表达的机制。CTGF、I型胶原和I κ B α表达，以及MAPK磷酸化，通过免疫印迹法对来自白色SSc患者的肺成纤维细胞进行研究。采用酶联免疫吸附试验检测细胞培养液中基质金属蛋白酶I（MMP-1）的表达，MMP-1活性检测采用MMP-1法，NF-κ B DNA结合活性检测采用转录因子法。在白色SSc患者的肺成纤维细胞中，HGF激活MAPK（ERK-1/2）信号通路和MMP-1，同时抑制NF-κ B，并以时间和剂量依赖性方式显着下调CTGF和胶原蛋白。小干扰RNA（siRNA）介导的Grb 2表达缺失破坏了c-Met受体下游信号传导，导致HGF诱导的ERK-1/2磷酸化减少，并恢复了HGF抑制的MMP-1、NF-κ B、胶原和CTGF表达。MAPK抑制剂U 0126阻断MMP-1活性，恢复HGF抑制的胶原和CTGF积累。MMP抑制剂GM 1489抑制MMP活性和siRNA抑制MMP-1表达不能阻止HGF诱导的ERK-1/2磷酸化和NF-κ B活性，但显著恢复了HGF抑制的胶原和CTGF积累。NF-κ B抑制剂BAY 11-7082不影响MAPK磷酸化和MMP-1的表达和活化，但显著抑制NF-κ B DNA结合活性，并与HGF协同作用，完全抑制CTGF的表达。在来自白色SSc患者的肺成纤维细胞中，HGF通过MAPK/MMP-1和NF-κ B信号通路下调CTGF的积聚，而胶原下调主要由MAPK/MMP-1依赖性通路介导。
Objective. To study the mechanisms by which hepatocyte growth factor (HGF) down-regulates collagen and connective tissue growth factor (CTGF) in scleroderma (systemic sclerosis [SSc]) lung fibroblasts.Methods. CTGF, type I collagen, and I kappa B alpha expression, together with MAPK phosphorylation, were studied by immunoblotting of lung fibroblasts derived from white SSc patients. Matrix metalloproteinase I (MMP-1) expression in cell culture medium samples was measured by enzyme-linked immunosorbent assay, MMP-1 activity was studied using an MMP-1 assay, and NF-kappa B DNA binding activity was determined using a transcription factor assay.Results. In lung fibroblasts from white SSc patients, HGF activated MAPK (ERK-1/2) signaling pathways and MMP-1, while it inhibited NF-kappa B and significantly down-regulated CTGF and collagen in a time-and dose-dependent manner. Small interfering RNA (siRNA)-mediated depletion of Grb2 expression disrupted c-Met receptor downstream signaling, which resulted in diminished HGF-induced ERK-1/2 phosphorylation and the recovery of HGF-inhibited expression of MMP-1, NF-kappa B, collagen, and CTGF. The MAPK inhibitor, U0126, blocked MMP-1 activity and restored HGF-inhibited collagen and CTGF accumulation. Inhibition of MMP activity by MMP inhibitor GM1489 and inhibition of MMP-1 expression by siRNA did not prevent HGF-induced ERK-1/2 phosphorylation and NF-kappa B activity, but significantly restored HGF-inhibited collagen and CTGF accumulation. NF-kappa B inhibitor BAY 11-7082 did not interfere with MAPK phosphorylation or MMP-1 expression and activation, but significantly inhibited NF-kappa B DNA binding activity and acted synergistically with HGF to completely diminish the expression of CTGF.Conclusion. In lung fibroblasts from white SSc patients, HGF down-regulates the accumulation of CTGF via MAPK/MMP-1 and NF-kappa B signaling pathways, whereas collagen down-regulation is mediated mainly by a MAPK/MMP-1-dependent pathway.