Particulate Shiga Toxin 2 in blood is associated to the development of hemolytic uremic syndrome in children

Authors: BRIGOTTI, MAURIZIO - University Of Bologna, Italy; He, Xiaohua; CARNICELLI, DOMENICA - University Of Bologna, Italy; ARFILLI, VALENTINA - University Of Bologna, Italy; PORCELLINI, ELISA - University Of Bologna, Italy; GALASSI, ELISABETTA - University Of Bologna, Italy; TAZZARI, PIER LUIGI - University Of Bologna, Italy; RICCI, FRANCESCA - University Of Bologna, Italy; Patfield, Stephanie; TESTA, SARA - Fondazione Irccs Ca’ Granda Ospedale Maggiore Policlinico; PAGLIALONGA, FABIO - Fondazione Irccs Ca’ Granda Ospedale Maggiore Policlinico; PICICCO, DAMIANO - Fondazione Irccs Ca’ Granda Ospedale Maggiore Policlinico; CAPRIOLI, ALFREDO - Istituto Superiore Di Sanita; SCAVIA, GAIA - Istituto Superiore Di Sanita; MORABITO, STEFANO - Istituto Superiore Di Sanita; ARDISSINO, GIANLUIGI - Fondazione Irccs Ca’ Granda Ospedale Maggiore Policlinico

Submitted to: Thrombosis and Haemostasis
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/7/2019
Publication Date: 11/27/2019
Citation: Brigotti, M., He, X., Carnicelli, D., Arfilli, V., Porcellini, E., Galassi, E., Tazzari, P., Ricci, F., Patfield, S.A., Testa, S., Paglialonga, F., Picicco, D., Caprioli, A., Scavia, G., Morabito, S., Ardissino, G. 2019. Particulate Shiga Toxin 2 in blood is associated to the development of hemolytic uremic syndrome in children. Thrombosis and Haemostasis. 120(1):107-120. https://doi.org/10.1055/s-0039-3400301.
DOI: https://doi.org/10.1055/s-0039-3400301

Interpretive Summary: The paper comes from a collaboration between a network connecting 56 pediatric units in the Lombardia Region (Italy), the European Reference laboratory for Escherichia coli (Italian NIH, Rome, Italy), the Center for hemolytic uremic syndrome control in Milan (Italy), the Agricultural Research Service of U.S. Department of Agriculture (Foodborne Toxin Detection and Prevention Research Unit) and the Section of General Pathology of my Department (DIMES, School of Medicine, University of Bologna). The study is focused on hemolytic uremic syndrome (HUS) caused by pathogenic Escherichia coli strains (STEC) producing very potent exotoxins, named Shiga toxins (Stx). The syndrome is the leading cause of renal failure in children under 3 yrs and can occur in sporadic form or as community-wide foodborne outbreaks, as dramatically exemplified in recent years in Europe, USA and Japan. We have studied 20 patients during the natural course of STEC-infection from the intestinal symptoms to the syndrome which occurred in 3 of them. The task was challenging since we have screened about 2800 children with bloody diarrhea to obtain a subgroup of patients infected by this pathogen (STEC). By comparing patients developing HUS and patients who recovered, we found that the particulated form (sedimented at g-forces corresponding to microvesicles) of Stx2 is the trigger allowing the transition from bloody diarrhea to the life-threatening HUS. HUS is a well-known and very topical health problem, often debated also on newspapers. Moreover, to the best of our knowledge, our study is the first one allowing the observation of what happens during STEC-infections in humans before the onset of HUS. Our findings may open perspective for strategies aimed at blocking the binding of Stx2 to circulating cells or to hampering the formation of pathogen microvesicles during STEC infections. It is worth noting that there are no specific treatments for HUS patients who are managed with supportive care regimens. Moreover, no animal model except baboons is able to recapitulate the complex steps involved in the pathogenesis of the syndrome. In particular, the mouse models employed for the study of renal damage are not useful for the study of toxin delivery since the receptor pattern of circulating cells in mice is different with respect to humans.

Technical Abstract: Hemolytic uremic syndrome (HUS), the leading cause of acute renal failure in children (<3 years), is mainly caused by Shiga toxins (Stx)-producing Escherichia coli (STEC) infections. STEC are confined to the gut causing hemorrhagic colitis, whereas Stx are thought to reach the kidney through the bloodstream. In different investigations, Stx were found on circulating cells, free in sera or in blood cell-derived microvesicles. Here, we have studied the natural course of STEC infections before the onset of HUS in 20 hospitalized patients out of 2846 children with bloody diarrhea. In most patients (55%) Stx were associated to neutrophils, subsequently Stx rose transiently in sera corresponding to the decrease of toxins on leukocytes. During this phase, three children showing a particulated form of Stx2 in blood (sedimented at g-forces corresponding to 1 µm-microvesicles)developed HUS, hence providing direct evidence of the involvement of blood cell-derived Stx2-containing microvesicles in triggering HUS.