Gavrilyuk Vitaliy, Kalinin Sergey, Hilbush Brian S, Middlecamp Andrew, McGuire Susan, Pelligrino Dale, Weinberg Guy, Feinstein Douglas L
Department of Anesthesiology, University of Illinois, Chicago, Illinois 60612, USA.
J Neurochem. 2005 Mar;92(5):1140-9. doi: 10.1111/j.1471-4159.2004.02942.x.
Brain inflammation is regulated by endogenous substances, including neurotransmitters such as noradrenaline (NA), which can increase anti-inflammatory genes. To identify NA-regulated, anti-inflammatory genes, we used TOGA (total gene expression analysis) to screen rat astrocyte-derived RNA. NA-inducible cDNA clone DST11 encodes an isoform of the complement C5a receptor (C5aR), with 39% identity at the amino acid level to the rat C5aR, and 56% identity to a recently described human C5aR variant termed C5L2 (complement 5a-like receptor). Quantitative PCR confirmed that in astrocytes, DST11 mRNA expression is increased by NA, whereas in vivo depletion of cortical NA reduced DST11 levels. Western blot analysis demonstrated basal and NA-induced expression of DST11 as a 45 kDa protein in primary astrocytes cultures. Immunocytochemical staining of adult rat brain revealed DST11-immunoreactivity throughout brain, co-localized to neurons and astrocytes. In astrocytes, induction of nitric oxide synthase type 2 was increased by treatment with antisense oligonucleotides to DST11. Reducing DST11 expression also increased nuclear factor kappaB reporter gene, and decreased cAMP response element reporter gene activation. These results demonstrate that DST11 is a C5aR isoform expressed by glia and neurons, which is regulated by NA, and exerts anti-inflammatory functions. Changes in DST11 levels in diseased brain could therefore contribute to the progression of inflammatory damage.
脑炎症由内源性物质调节，这些物质包括神经递质，如去甲肾上腺素（NA），其可增加抗炎基因。为了鉴定NA调节的抗炎基因，我们使用全基因表达分析（TOGA）筛选大鼠星形胶质细胞来源的RNA。NA诱导的cDNA克隆DST11编码补体C5a受体（C5aR）的一种异构体，在氨基酸水平上与大鼠C5aR有39%的同一性，与最近描述的人类C5aR变体C5L2（补体5a样受体）有56%的同一性。定量PCR证实，在星形胶质细胞中，NA可增加DST11 mRNA的表达，而体内皮质NA的耗竭会降低DST11水平。蛋白质印迹分析表明，在原代星形胶质细胞培养物中，DST11作为一种45 kDa的蛋白质有基础表达和NA诱导的表达。成年大鼠脑的免疫细胞化学染色显示，整个脑中都有DST11免疫反应性，其与神经元和星形胶质细胞共定位。在星形胶质细胞中，用针对DST11的反义寡核苷酸处理可增加2型一氧化氮合酶的诱导。降低DST11的表达也会增加核因子κB报告基因，并降低cAMP反应元件报告基因的激活。这些结果表明，DST11是一种由神经胶质细胞和神经元表达的C5aR异构体，受NA调节，并发挥抗炎功能。因此，患病脑中DST11水平的变化可能有助于炎症损伤的进展。
