VSTM1-v2 does not drive human Th17 cell differentiation: A replication study

Signal inhibitory receptor on leukocytes-1 (SIRL-1) is an immune inhibitory receptor expressed on human myeloid cells. We previously showed that dendritic cell (DC)-driven Th17 cell differentiation of human naive CD4+ T cells requires presence of neutrophils, which is inhibited by SIRL-1 ligation. VSTM1-v2 is a soluble isoform of SIRL-1, which was previously proposed to function as a Th17 polarizing cytokine. Here, we investigated the effect of VSTM1-v2 on DC-driven Th17 cell development. Neutrophils induced DC-driven Th17 cell differentiation, which was not enhanced by VSTM1-v2. Similarly, we found no effect of VSTM1-v2 on cytokine-driven Th17 cell development. Thus, our results do not support a role for VSTM1-v2 in Th17 cell differentiation

VSTM1-v2 does not bind to leukocytes.

Erythrocyte-lysed whole blood was incubated with His-tagged VSTM1-v2 or sLAIR-1ecto (10–50 μg/mL), followed by detection with anti-His-AF647. (A) Lymphocytes (lower population), monocytes (middle population) and granulocytes (upper population) were gated based on forward scatter (FSC) and sideward scatter (SSC), followed by gating on single cells and AF-647+ cells. (B) Shown are representative dot plots of lymphocytes stained with 50 μg/mL VSTM1-v2 and/or anti-His-AF647. (C-E) The graphs show the quantification of the percentage of AF647+ cells of lymphocytes (C), monocytes (D), or granulocytes (E). Data are shown as mean ± SD of two donors. (TIF)</p

VSTM1-v2 does not enhance Th17 cell differentiation and activation.

CD4+ T cells were stimulated by C. albicans-activated moDCs, with or without autologous neutrophils (A-C), or with antibodies and a polarizing cytokine mix (D), with or without addition of 10 or 100 ng/mL VSTM1-v2. Intracellular IL-17 expression was determined by flow cytometry. (A) Schematic representation of the co-culture system of moDC-driven Th17 cell differentiation and activation. (B) Representative dot plots of the percentage of IL-17+ cells after co-culture of naive CD4+ T cells. The y-axis indicates the fluorescence intensity of the IL-17A staining, while the x-axis indicates the forward scatter (FSC). (C) The percentage of IL-17+ cells after co-culture of naive CD4+ T cells (n = 10), each donor represented by a different color, or memory CD4+ T cells (n = 3; mean ± SD). (D) The percentage of IL-17+ cells after stimulation of total CD4+ T cells with anti-CD3, anti-CD28, and a Th17 polarizing mix, mean ± SD, n = 3. Statistical significance was determined using a Friedman test with Dunn’s correction (C, D). ** p biorender.com.</p