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Updated: Jul 3, 2026
Epithelial Cell Infection Analyses with Shigella
Published on: February 9, 2024
Interactions between Shiga toxins and human polymorphonuclear leukocytes
Maurizio Brigotti1, Domenica Carnicelli, Elisa Ravanelli
1Università di Bologna, Via San Giacomo, 14, Bologna, Italy 40126. maurizio.brigotti@unibo.it
Insights
Shiga toxin (Stx) binds to mature human polymorphonuclear leukocytes (PMN), delaying their apoptosis and enabling toxin transfer to other PMN. This interaction may explain Stx persistence in children with hemolytic uremic syndrome (HUS).
Area of Science:
- Microbiology
- Immunology
- Cell Biology
Background:
- Shiga toxin-producing Escherichia coli cause severe human infections, including hemolytic uremic syndrome (HUS), a leading cause of childhood renal failure.
- The mechanism of Shiga toxin (Stx) dissemination in HUS, particularly its transport in the bloodstream, remains incompletely understood.
- Polymorphonuclear leukocytes (PMN) have been implicated as potential carriers of Stx, but the precise nature of their interaction requires further elucidation.
Purpose of the Study:
- To investigate the interaction between Shiga toxins (Stx) and human polymorphonuclear leukocytes (PMN).
- To determine if PMN play a role in the transport and persistence of Stx in the context of HUS.
Main Methods:
- Flow cytometry (direct and indirect) was used to analyze Stx binding to human PMN.
- Binding experiments with radiolabeled toxins were performed.
- The human myeloid leukemia cell line (HL-60) was utilized to model inducible granulocytic differentiation.
- Apoptosis assays (caspase 3 activation, annexin V binding) were conducted to assess the effect of Stx on PMN viability.
- Mixed PMN populations were analyzed to study toxin transfer.
Main Results:
- Shiga toxins (Stx) bind to the surface of mature human PMN, but not immature PMN.
- Toxin binding was observed only after granulocytic differentiation, confirmed using the HL-60 cell model.
- Stx binding delayed spontaneous apoptosis in PMN, evidenced by reduced caspase 3 activation and annexin V staining.
- Flow cytometry demonstrated the transfer of Stx from Stx-positive PMN to Stx-negative PMN.
Conclusions:
- Mature human PMN bind Shiga toxins (Stx) on their surface.
- Stx binding inhibits PMN apoptosis and facilitates toxin transfer between PMN.
- These findings suggest a mechanism for Stx persistence in the bloodstream of HUS patients, involving PMN as carriers and transfer agents.
Abstract:
Human intestinal infections by Shiga toxin (Stx)-producing Escherichia coli cause hemorrhagic colitis and hemolytic uremic syndrome (HUS), which represents the main cause of acute renal failure in early childhood. In HUS, Stx released in the gut enter the bloodstream and are targeted to renal endothelium. The mechanism of toxin delivery is still a matter of debate, although the role of polymorphonuclear leukocytes (PMN) as a Stx carrier has been indicated. The aim of this paper was to better define the interactions between Stx and human PMN. Direct and indirect flow cytometric analysis and binding experiments with radiolabeled toxins demonstrated that Stx bind to the surface of human mature PMN but not to immature PMN from G-CSF-treated donors. The use of the human myeloid leukemia cell (HL-60) model for inducible cell differentiation confirmed that the toxin binding occurs only after granulocytic differentiation. Stx binding caused a delay of the spontaneous apoptosis of PMN, as shown by the delayed appearance of apoptotic nuclei and activation of caspase 3 and by the higher number of cells negative to the annexin V-binding assay after 48 h. Moreover, flow cytometric analysis of mixed Stx-positive and Stx-negative PMN populations showed that the toxins were transferred from positive to negative PMN. The delayed, spontaneous apoptosis and the passage of the toxic ligand from older PMN to new, mature cells entering the circulation from the bone marrow may explain the previously reported persistence of Stx in the blood of children with HUS.
