TNF-alpha regulation of C3 gene expression and protein biosynthesis in rat glomerular endothelial cells.
TNF-alpha regulation of C3 gene expression and protein biosynthesis in rat glomerular endothelial cells.
复制标题
TNF-α 对大鼠肾小球内皮细胞 C3 基因表达和蛋白质生物合成的调节。
作者：
N. Sheerin;W. Zhou;S. Adler;S. Sacks
肾小球内皮细胞是与炎症细胞和可溶性成分相互作用的重要部位。为了研究这些细胞合成补体的能力，我们从分离的大鼠肾小球中克隆了它们。从细胞中提取信使RNA(Messenger RNA，mRNA)，逆转录，以此为模板，通过聚合酶链式反应(PCR)鉴定特定基因转录本。在未受刺激的内皮细胞中检测到补体级联的第三组分(C3)的mRNA编码，而没有发现第四组分(C4)的消息。用半定量的聚合酶链式反应方法发现，C3的表达受细胞因子肿瘤坏死因子-α(TNF-α)的上调，但不受细胞因子干扰素-γ(干扰素-γ)和白介素1-α(IL-1α)的上调。C3mRNA水平的升高呈时间和剂量依赖关系。这种增加依赖于新的蛋白质合成。放射性标记和免疫沉淀证实了C3蛋白的产生，这也被肿瘤坏死因子-α刺激。总之，我们证明了肾小球来源的微血管内皮细胞产生补体C3，并被肿瘤坏死因子-α刺激。我们认为这种局部合成可能在疾病的发病机制中发挥作用，然而，目前这种作用的性质尚不清楚。
Glomerular endothelial cells are an important site of interaction with the cellular and soluble components of inflammation. To investigate the capacity of these cells to synthesize complement they were cloned from isolated rat glomeruli. Messenger RNA (mRNA) was extracted from the cells, reverse transcribed and used as the template to identify specific gene transcripts with the polymerase chain reaction (PCR). mRNA coding for the third component of the complement cascade (C3) was detected in unstimulated endothelial cells, whereas no message for the fourth component (C4) could be demonstrated. Using a semiquantitative method of PCR, we found that the expression of C3 is up-regulated by the cytokine tumor necrosis factor-alpha (TNF-alpha), but not by the cytokines interferon-gamma (IFN-gamma) and interleukin 1 alpha (IL-1 alpha). The increase in levels of C3 mRNA occurred in a time and dose dependent manner. This increase was dependent on new protein synthesis. Production of the C3 protein was demonstrated by radiolabeling and immunoprecipitation, and this also was stimulated by TNF-alpha. In conclusion, we demonstrate the production of C3 by microvascular endothelium of glomerular origin and its stimulation by TNF-alpha. We believe that this local synthesis could have a role in the pathogenesis of disease, however, the nature of this role at present remains unclear.