Hemolytic uremic syndrome (HUS) is a vascular disease of the kidney in which the glomerular microcapillaries become occluded with fibrin and platelets. Although HUS is associated with a number of different agents and pathogens, eg. bacteria, viruses, immunosuppressive drugs, AIDS, and familial genetic traits, little is known about the mechanisms leading to the development of HUS. Most of the new information on HUS has come from studies such as this ongoing project which have exploited the recently recognized association between infections of humans with Shiga toxin-producing dysentery bacteria and the subsequent appearance of HUS those individuals. Our long-term goal is to describe, in biochemical terms, the mechanisms by which Shiga toxins elicit the HUS disease state and to use this knowledge to develop effective preventive and therapeutic intervention modalities. The goal of the present study is to delineate the role of Shiga toxin at the vascular endothelial cell level in the development of HUS. In addition to human umbilical vein endothelial cells, these studies utilize the rarely available human renal microvascular endothelial cell type which is believed to be the primary target of Shiga toxin during the development of HUS. Our research plan explores how bacterial lipopolysaccharide (LPS) or the cytokine, tumor necrosis factor (TNF) combines with Shiga toxin to elicit cytotoxic and procoagulant responses from endothelial cells. A major goal of this research is to determine which of the signal transduction mechanisms are utilized by LPS or TNF during their interaction with endothelial cells. In addition, changes in the expression of endothelial proteins is to be examined following exposure of these cells to Shiga toxin , LPS and TNF. In a related study, we will examine how LPS or TNF induce the expression of the Shiga toxin receptor, galactotriosylceramide (Gb3) on the surface of endothelial cells. A major part of our research plan is our continued examination of the procoagulant state produced on endothelial cells by Shiga toxin, LPS and TNF. Using a combination of antigen detection (ELISA) and biological activity assays, the amount and activity of endothelial-derived procoagulant and anticoagulant factors will be determined. Similarly, a study of increased adherence of platelets and leukocytes to endothelial cells is to be conducted as a result of Shiga toxin, etc. action on these cells. Finally, the basis of renal mesangial cell (a modified smooth muscle cell) proliferation in HUS is to be examined during their coculture with kidney endothelial cells. In summary, this research combines the areas of infectious disease and vascular physiology and has as its primary goal to provide an understanding of the mechanisms underlying the development of renal vascular disease in HUS.
