Luzina Irina G, Tsymbalyuk Natalya, Choi Jung, Hasday Jeffrey D, Atamas Sergei P
Department of Medicine, University of Maryland School of Medicine, Baltimore VA Medical Center, Baltimore, Maryland 21201, USA.
J Cell Physiol. 2006 Jan;206(1):221-8. doi: 10.1002/jcp.20452.
A CC chemokine CCL18 stimulates collagen production in pulmonary fibroblasts through an unknown signaling mechanism. In this study, involvement of Sp1 and Smad3 in CCL18 signaling in primary human pulmonary fibroblast cultures was investigated. Phosphorylation of Sp1, DNA-binding by Sp1, and the activity of an Sp1-dependent reporter were all increased in response to CCL18 stimulation. CCL18 did not stimulate a detectable increase in Smad3 phosphorylation or Smad3/4 DNA-binding activity, although some basal phosphorylation and DNA binding by Smad3/4 were noted. Transient overexpression of dominant negative mutants of Sp1 and Smad3 abrogated CCL18-dependent upregulation as well as basal production of collagen. These observations suggested that CCL18 activates collagen production in pulmonary fibroblasts through an Sp1-dependent pathway that also requires basal Smad3 activity. Possible involvement of autocrine TGF-beta in CCL18 signaling was considered. CCL18 stimulated increases in collagen mRNA and protein production without detectable changes in TGF-beta1, -beta2, and -beta3 mRNA or protein levels. Neutralizing anti-TGF-beta antibodies, latency-associated peptide, ALK5-specific inhibitor SD431542, and an inhibitor of the protease-dependent TGF-beta activation aprotinin, each failed to block CCL18-stimulated collagen production. These observations suggest that both CCL18 signaling in pulmonary fibroblasts and basal Smad3 activity are independent of autocrine TGF-beta.
CC趋化因子CCL18通过未知的信号传导机制刺激肺成纤维细胞中的胶原蛋白生成。在本研究中，研究了Sp1和Smad3在原代人肺成纤维细胞培养物的CCL18信号传导中的作用。响应CCL18刺激，Sp1的磷酸化、Sp1的DNA结合以及Sp1依赖性报告基因的活性均增加。尽管注意到Smad3/4有一些基础磷酸化和DNA结合，但CCL18并未刺激Smad3磷酸化或Smad3/4 DNA结合活性的可检测增加。Sp1和Smad3显性负突变体的瞬时过表达消除了CCL18依赖性上调以及胶原蛋白的基础生成。这些观察结果表明，CCL18通过Sp1依赖性途径激活肺成纤维细胞中的胶原蛋白生成，该途径也需要基础Smad3活性。考虑了自分泌TGF-β在CCL18信号传导中的可能作用。CCL18刺激胶原蛋白mRNA和蛋白质生成增加，而TGF-β1、-β2和-β3 mRNA或蛋白质水平未检测到变化。中和抗TGF-β抗体、潜伏相关肽、ALK5特异性抑制剂SD431542以及蛋白酶依赖性TGF-β激活抑制剂抑肽酶，均未能阻断CCL18刺激的胶原蛋白生成。这些观察结果表明，肺成纤维细胞中的CCL18信号传导和基础Smad3活性均独立于自分泌TGF-β。
