Bitzan M M, Wang Y, Lin J, Marsden P A
Division of Nephrology, Department of Medicine, St. Michael's Hospital and University of Toronto, Toronto, Ontario, Canada M5S 1A8.
J Clin Invest. 1998 Jan 15;101(2):372-82. doi: 10.1172/JCI522.
Interaction of bipartite Escherichia coli O157-derived verotoxins (VTs) 1 and 2 (Shiga toxin 1 and 2) with vascular endothelium is believed to play a central role in the pathogenesis of the thrombotic microangiopathy and ischemic lesions characteristic of hemolytic uremic syndrome and of E. coli O157-associated hemorrhagic colitis. We defined the effects of VTs on the expression of potent endothelial cell-derived regulators of vascular wall function, namely endothelin-1 (ET-1) and nitric oxide (NO). In quiescent bovine aortic endothelial cells, both VT1 and VT2, but not receptor-binding VT B-subunit which lacks N-glycosidase activity, induced concentration-dependent (0.1-10 nM) increases in steady state preproET-1 mRNA transcript levels, an effect that was maximal at 12-24 h. Metabolic-labeling experiments indicated that VTs increased preproET-1 mRNA transcript levels at concentrations that had trivial effects on nascent DNA, RNA, and protein synthesis. In contrast to preproET-1, endothelin converting enzyme-1 and endothelial constitutive NO synthase mRNA transcript levels remained unchanged. Consistent with these findings, VTs failed to modulate immunoreactive endothelial constitutive NO synthase expression and basal and calcium-dependent L-[14C]arginine to L-[14C]citrulline conversion or the NO chemiluminescence signal. The plant-derived toxin ricin, which shows a similar molecular mechanism of enzymatic ribosomal modification to VTs, caused comparable effects on these endothelial vasomediators and metabolite incorporation, at 3 log orders lower concentrations. Nuclear transcription and actinomycin D chase experiments indicated that VTs stabilize labile preproET-1 mRNA transcripts in endothelial cells. Therefore, VTs potently increase select mRNA transcript levels in endothelial cells at concentrations of toxins that have minimal effects on protein synthesis. Perturbed expression of endothelial-derived vasomediators may play a pathophysiologic role in the microvascular dysfunction that is the hallmark of hemolytic uremic syndrome and hemorrhagic colitis.
源自大肠杆菌O157的双分型志贺毒素1和2（VTs）（志贺毒素1和2）与血管内皮的相互作用被认为在血栓性微血管病的发病机制以及溶血性尿毒症综合征和大肠杆菌O157相关出血性结肠炎的特征性缺血性病变中起核心作用。我们确定了VTs对血管壁功能的强效内皮细胞衍生调节因子即内皮素-1（ET-1）和一氧化氮（NO）表达的影响。在静止的牛主动脉内皮细胞中，VT1和VT2，但缺乏N-糖苷酶活性的受体结合型VT B亚基均未诱导浓度依赖性（0.1 - 10 nM）的前内皮素原-1（preproET-1）mRNA转录本稳态水平增加，该效应在12 - 24小时时达到最大。代谢标记实验表明，VTs在对新生DNA、RNA和蛋白质合成影响极小的浓度下增加了preproET-1 mRNA转录本水平。与preproET-1相反，内皮素转换酶-1和内皮组成型一氧化氮合酶mRNA转录本水平保持不变。与这些发现一致，VTs未能调节免疫反应性内皮组成型一氧化氮合酶的表达以及基础和钙依赖性L-[14C]精氨酸向L-[14C]瓜氨酸的转化或NO化学发光信号。植物源性毒素蓖麻毒素，其显示出与VTs类似的酶促核糖体修饰分子机制，在低3个对数级的浓度下对这些内皮血管介质和代谢物掺入产生了类似的影响。核转录和放线菌素D追踪实验表明，VTs使内皮细胞中不稳定的preproET-1 mRNA转录本稳定。因此，VTs在对蛋白质合成影响极小的毒素浓度下能有效增加内皮细胞中特定mRNA转录本水平。内皮衍生血管介质的表达紊乱可能在溶血性尿毒症综合征和出血性结肠炎的标志性微血管功能障碍中发挥病理生理作用。
