|TE LOO, MAROESKA - Radboud University|
|LOOS, SEBASTIAN - University Of Hamburg|
|SCAVIA, GAIA - Istituto Superiore Di Sanita|
|BRIGOTTI, MAURIZIO - University Of Bologna, Italy|
|LEVTCHENKO, ELENA - Radboud University|
|MONNENS, LEO - Radboud University|
Submitted to: Acta Paediatrica
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/11/2015
Publication Date: N/A
Interpretive Summary: Shiga toxin types, Stx1 and Stx2 and their variants, are considered as the cause of hemolytic uremic syndrome (HUS) by mediating damage mainly to renal glomerular endothelial cells. Epidemiologic studies have indicated that the risk of HUS development after infection by enterohemorrhagic Escherichia coli (EHEC) is significantly associated with the presence of Stx2. Several explanations have been proposed for the higher toxicity of Stx2 compared to Stx1, such as prolonged attachment to endothelial cells and higher cytokine up regulation. After access to the systemic circulation, Stxs have to be delivered to their target organs as soluble free Stx or by binding to blood components such as polymorphonuclear leukocytes, monocytes or red blood cells. A direct in vivo demonstration of the transfer of Stx bound to one of these components to target organs is lacking. It is conceivable that Stx may be transferred from the host blood cell, resulting the endocytosis into glomerular endothelial cells. In a cytotoxicity assay with human endothelial cells only negligible amounts of free Stx were detected in the sera from 34 HUS patients collected immediately after admission. In another study of 313 samples obtained from 93 confirmed infected patients with the onset of bloody diarrhea, Stx2 was only detected in less than 10 patients. Additional studies are thus needed to determine free Stx2 concentration levels in sera of patients at a specific phase of the HUS disease symptoms. In the present study, serum samples were collected immediately after admission of the patients to the clinic. Stx2 concentrations in serum collected from 16 patients with a proven EHEC infection were measured with an enzyme-linked immunosorbent assay. Additional studies were conducted to determine the functional activity of the Stx2 in sera from the family members of the HUS patients by using a novel Vero cell- based method. Our findings demonstrated that low concentrations of free serum Stx2 were present in 50% (8/16) of the patients who were suffering the acute phase of HUS. However, the free Shiga toxin concentrations were significantly higher in family members not affected by HUS but suffering mild disease symptoms. In the family members with higher Stx2 concentrations, the toxin functional activity was the most pronounced, indicating an early host cell injury during the first stages of STEC infection.
Technical Abstract: Shiga toxins (Stxs) produced by Shiga toxin-producing Escherichia coli (STEC) are considered as the main causative agent, leading to the development of the hemolytic uremic syndrome (HUS); these toxins injure endothelial cells mainly the glomeruli. After passing through the intestinal wall, Stxs have to be delivered via the systemic circulation to the target organs. The aim of this study was to measure free Stx2 in serum of children that suffered HUS after infection with STEC. Sera from patients, suffering the acute phase of the HUS, were collected immediately after admission to the clinic. The concentration of free Stx2 in serum of 16 children was measured by using a sandwich enzyme-linked immunosorbent assay. Family members of two children with HUS were also investigated, and the family members were only suffering mild disease symptoms such as watery diarrhea. In these family members, the relative toxicity of Stx2 was assessed by a Vero cell-based fluorescent assay. In 50% (8/16) of the children with HUS, Stx2 was detected in their serum. In six family members, suffering STEC infection without symptomatic HUS, Stx2 was detected in four patients. Interestingly, two patients had a significantly higher concentration of Stx2. In the two family members with the highest Stx2 concentrations, the functional toxin activity was the most pronounced. In conclusion, we have demonstrated an absent or a rather low concentration of Stx2 in serum of children admitted to the clinic with the acute phase of HUS. By contrast, the highest concentration of Stx2 in conjunction with an increased functional activity was measured in family members without HUS but mostly with watery diarrhea. Together, these findings have provided supporting evidence for an early injury caused by Stx2 during the first days of STEC infection.