Induction by sphingomyelinase of shiga toxin receptor and shiga toxin 2 sensitivity in human microvascular endothelial cells

Shiga toxin-producing enterohemorrhagic Escherichia coli is the major cause of acute renal failure in young children. The interaction of Shiga toxins 1 and 2 (Stx1 and Stx2) with endothelial cells is an important step in the renal coagulation and thrombosis observed in hemolytic uremic syndrome. Previous studies have shown that bacterial lipopolysaccharide and host cytokines slowly sensitize endothelial cells to Shiga toxins. In the present study, bacterial neutral sphingomyelinase (SMase) rapidly (1 h) sensitized human dermal microvascular endothelial cells (HDMEC) to the cytotoxic action of Stx2. Exposure of endothelial cells to neutral SMase (0.067 U/ml) caused a rapid increase of intracellular ceramide that persisted for hours. Closely following the change in ceramide level was an increase in the expression of globotriaosylceramide (Gb3), the receptor for Stx2. A rapid increase was also observed in the mRNA for ceramide:glucosyltransferase (CGT), the first of three glycosyltransferase enzymes of the Gb3 biosynthetic pathway. The product of CGT (glucosylceramide) was also increased. In contrast, mRNA for the third enzyme of the pathway, Gb3 synthase, was constitutively produced and was not influenced by SMase treatment of HDMEC. These results describe a rapid response mechanism by which extracellular neutral SMase derived from either bacteria or eukaryotic cells may signal endothelial cells to become sensitive to Shiga toxins.     

Effect of implant duration on in vivo sensitivity of electromagnetic flow transducers.

Twenty-three electromagnetic flow transducers with lumen diameters of 3.5-6.0 mm were implanted in rhesus monkeys and baboonss for 12 h to 120 days. Each flow transducer was calibrated 1) in vitro on dialysis tubing with saline before implantation, 2) in vivo the last day of the implant period, and 3) again in vitro after the flow transducer was recovered. Three other flow transducers were implanted on femoral arteries of baboon just central to an arteriovenous Silastic shunt, and were calibrated in vivo daily for 23-47 days. In vitro sensitivity was not affected by implant durations of up to 120 days. In vivo sensitivity fluctuated unpredictably for the first 3-4 wk of implant, after which it followed a systematic course that depended on the lumen size. In vivo sensitivity at any time during implant (after the initial period) could be accurately predicted by knowing either the in vitro sensitivity or the terminal in vivo sensitivity.