Lian R H, Kotwal G J, Wellhausen S R, Hunt L A, Justus D E
Department of Microbiology and Immunology, University of Louisville, KY 40292, USA.
J Immunol. 1996 Jul 15;157(2):864-73.
IFN-gamma-activated endothelial cells actively participate in initiating immune responses by interacting with immunocompetent cells via class II MHC proteins. In this study, dextran sulfate, a synthetic heparin analogue, was shown to selectively inhibit IFN-gamma-induced surface expression of HLA-DR molecules by human umbilical cord vascular endothelial cells, but not other cytokine-induced molecules such as ELAM-1 or ICAM-1. Inhibition occurred regardless of whether dextran sulfate was added 24 h before, at the same time as, or 24 h after IFN-gamma stimulation of cells. In addition, both high (500 kDa) and low (5 kDa) molecular mass dextran sulfate molecules were able to block class II expression, whereas treating cells with naturally occurring polysulfated glycosaminoglycans such as heparin, heparan, and chondroitin sulfate did not produce any suppressive effects. Radiolabeling of cells with [35S]methionine followed by radioimmunoprecipitation using anti-HLA-DR alpha mAb demonstrated that biosynthesis of class II proteins was specifically blocked. RT-PCR and Southern blotting were utilized to examine transcription of the HLA-DR alpha gene and demonstrated an absence of HLA-DR alpha mRNA from dextran sulfate-treated and IFN-gamma-induced cells. Dextran sulfate also prevented transcription of the gene encoding CIITA, a transactivator protein required for IFN-gamma-inducible expression of class II genes. Thus, dextran sulfate apparently inhibited this step or an earlier one in the intracellular signaling pathway for IFN-gamma in human endothelial cells, subsequent to IFN-gamma binding to its cell surface receptor.
γ-干扰素激活的内皮细胞通过经由II类主要组织相容性复合体（MHC）蛋白与免疫活性细胞相互作用，积极参与启动免疫反应。在本研究中，硫酸葡聚糖，一种合成的肝素类似物，被证明可选择性抑制人脐静脉血管内皮细胞中γ-干扰素诱导的HLA-DR分子的表面表达，但不影响其他细胞因子诱导的分子，如ELAM-1或ICAM-1。无论硫酸葡聚糖是在γ-干扰素刺激细胞前24小时、同时还是刺激后24小时添加，均会出现抑制作用。此外，高分子量（500 kDa）和低分子量（5 kDa）的硫酸葡聚糖分子均能够阻断II类表达，而用天然存在的多硫酸化糖胺聚糖如肝素、硫酸乙酰肝素和硫酸软骨素处理细胞则未产生任何抑制作用。用[35S]甲硫氨酸对细胞进行放射性标记，随后使用抗HLA-DRα单克隆抗体进行放射免疫沉淀，结果表明II类蛋白的生物合成被特异性阻断。利用逆转录-聚合酶链反应（RT-PCR）和Southern印迹法检测HLA-DRα基因转录，结果显示硫酸葡聚糖处理且γ-干扰素诱导的细胞中不存在HLA-DRα mRNA。硫酸葡聚糖还可阻止编码CIITA的基因转录，CIITA是一种II类基因γ-干扰素诱导表达所需的反式激活蛋白。因此，硫酸葡聚糖显然在人内皮细胞中，于γ-干扰素与其细胞表面受体结合后，抑制了γ-干扰素细胞内信号通路中的这一步骤或更早的步骤。
