Submitted to: Infection and Immunity
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/23/2004
Publication Date: 11/20/2004
Citation: Winter, K.R., Stoffregen, W.C., Mengeling, W.L., Nystrom, E.A. 2004. Shiga toxin binding to isolated porcine tissues and peripheral blood leukocytes. Infection and Immunity. 72(11):6680-6684.
Interpretive Summary: Shiga toxin (Stx)-producing Escherichia coli (STEC) O157:H7 and other STEC cause severe diarrhea and sometimes kidney failure and death in humans. Because they are susceptible to both natural (edema disease) and experimental infection with STEC, pigs are useful models for studying how STEC cause disease. Exposure to Stxs is toxic for cultured cells and causes damage to blood vessels, paralysis and death in experimental animals. However, the mechanisms of Stx-mediated damage are not understood. The objective of this study was to identify sites in pigs where Stxs bind. For these studies, sections of tissues were obtained from small and large intestines, liver, kidney and brains of neonatal pigs and white blood cells were obtained from the lungs and blood of 3- to 6-week-old pigs. Stx binding was determined by incubating tissue sections or white blood cell preparations with purified Stx1 or Stx2, and using immunohistochemical assays to detect bound Stxs. Both Stx1 and Stx2 bound to all of the tissues tested. Stx binding sites included some sites that contain white blood cells. Both Stx1 and Stx2 also bound to white cells from the lungs (macrophages) and from peripheral blood (granulocytes and mononuclear cells) of pigs. Globotriaosylceramide (Gb3), a receptor for Stx, was identified by immunoassay in all of the tissues and cells where Stx bound. The results of this study provide the first evidence that Stxs bind to porcine white blood cells and support the hypothesis that white blood cells have a role in Stx disease. Blood cells from pigs will be useful for Stx binding studies and for determining if Stx binding to white cells plays a role in STEC disease. Understanding how Stxs bind to cells and tissues and are transported to target tissues will help identify ways to prevent or reduce serious complications associated with STEC infections.
Technical Abstract: Shiga toxin (Stx)-producing Escherichia coli (STEC) cause severe diarrhea and hemolytic uremic syndrome in humans. STEC strains that produce Shiga toxin 2 (Stx2) are associated with more severe disease than Stx1-producing STEC strains. Identification of differences in Stx1 and Stx2 binding sites in tissues may help explain why Stx2-producing STEC are more virulent. In this study, immunohistochemical assays were used to identify and compare the distribution of Stx1 and Stx2 binding sites in neonatal porcine tissues, localize the distribution of the Stx receptor (globotriaosylceramide [Gb3]), and determine if Stxs bind to porcine leukocytes. Stx1 and Stx2 bound to all tissues tested (small and large intestines, brain, kidney, and liver), and Gb3 was localized to all sites where Stx1 and Stx2 bound. There were no apparent differences in the distributions of Stx1 and Stx2 binding that might explain differences in virulence between Stx1- and Stx2-producing STEC. Stx1 and Stx2 bound to several sites on tissues where leukocytes reside, and Stx2 bound to porcine alveolar macrophages and peripheral blood leukocytes. Gb3 was found in the same types of leukocytes to which Stx2 bound (i.e., those that look like polymorphonuclear leukocytes and monocytes), but not in other leukocytes. Porcine leukocytes will be useful for evaluating the mechanisms of Stx binding and the role leukocytes may play in STEC disease.