Abstract

Shiga toxins (Stx), produced by Stx-producing Escherichia coli (STEC), are known to target Gb3-expressing cells, contributing to organ pathology such as in the kidney and brain. However, the sensitivity of human B-lymphoblastoid cell lines to Stx2 and their Gb3 expression profiles remain poorly understood. In this preliminary study, we assessed the susceptibility of human B-lymphoblastoid cell lines to Stx2 and identified distinct resistance and sensitivity patterns. Eight representative lines were further analyzed for Gb3 expression by mass spectrometry and flow cytometry. Susceptible cell lines (e.g., GM02473 and GM07019) displayed significantly higher total and membrane-associated Gb3 levels, while resistant lines had lower or undetectable Gb3. Exosomal Gb3 quantification revealed similar expression trends, contradicting the hypothesis that Gb3-positive exosomes neutralize Stx2. Interestingly, resistant cell line GM17658 showed discordant total and exosomal Gb3 levels. Immunofluorescence microscopy and flow cytometry revealed heterogeneous Gb3 expression within cell lines, with susceptible lines having a higher proportion of Gb3-positive cells. These findings suggest that Stx2 susceptibility is associated with Gb3 expression frequency rather than intensity and raise the possibility that Gb3-positive exosomes might contribute to toxicity. Future studies need to validate the role of exosomal Stx2 transfer and the functional impact of variable levels of Gb3-positive versus Gb3-negative subpopulations in toxin response.

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