Identification of Novel Cell Surface Markers on Mouse and Human Th17 Cells

PBMCs were isolated, CD4-purified using the MagCellect^TM Human Naïve CD4⁺ T Cell Isolation Kit (Catalog # MAGH115), and stimulated for 5 days to induce Th17 differentiation. The resulting Th17 cells were then stained with APC-conjugated Mouse Anti-Human CD4 Monoclonal Antibody (Catalog # FAB3791A) and various human surface markers. These data show the various levels of staining with the surface markers ranging from negative to high expression. These results are representative of three experiments.

PBMCs were purified as described above, and both human cells and mouse splenocytes were stimulated for 5 days to induce Th17 differentiation. The resulting Th17 cells were stained with an APC-conjugated Mouse Anti-Human CD4 Monoclonal Antibody (Catalog # FAB3791A) or an APC-conjugated Rat Anti-Mouse CD4 Monoclonal Antibody (Catalog # FAB554A) and the surface markers shown. These data validate that known surface markers associated with Th17 cells were identified in our screen. These results are representative of three experiments.

PBMCs were purified as described above, and both human cells and mouse splenocytes were stimulated for 5 days to induce Th17 differentiation. The resulting Th17 cells were stained with an APC-conjugated Mouse Anti-Human CD4 Monoclonal Antibody or a PE-conjugated Mouse Anti-Human CD4 Monoclonal Antibody (Catalog # FAB3791A or FAB3791P, respectively) or an APC-conjugated Rat Anti-Mouse CD4 Monoclonal Antibody or a PE-conjugated Rat Anti-Mouse CD4 Monoclonal Antibody (Catalog # FAB554A or # FAB554P, respectively) and the surface markers shown. In order to identify if these markers were expressed on functional human Th17 cells, a separate experiment was carried out to examine CD200, IL-18 R alpha, BTLA, and CD99 levels in conjunction with IL-22, IL-17, and ROR gamma t. These data are representative of three and two experiments, respectively.

PBMCs were purified as described above and stimulated for 5 days to induce Th17 differentiation. The resulting Th17 cells were stained with an APC-conjugated Mouse Anti-Human CD4 Monoclonal Antibody, a Fluorescein-conjugated Mouse Anti-Human CD4 Monoclonal Antibody, or a PE-conjugated Mouse Anti-Human CD4 Monoclonal Antibody (Catalog # FAB3791A, # FAB3791F, or # FAB3791P, respectively) and the surface markers shown. Cells expressing 26/27 of these novel markers also expressed IL-22, IL-17, and ROR gamma t. The four novel markers that overlap with mouse Th17 cells are shown in blue. These results are representative of three and two experiments, respectively.

Mouse splenocytes were stimulated for 5 days to induce Th17 differentiation. The resulting Th17 cells were co-stained with an APC-conjugated Rat Anti-Mouse CD4 Monoclonal Antibody, a Fluorescein-conjugated Rat Anti-Mouse CD4 Monoclonal Antibody, or a PE-conjugated Rat Anti-Mouse CD4 Monoclonal Antibody (Catalog # FAB554A, # FAB554F, or # FAB554P, respectively) and the surface markers shown. The four novel markers that overlap with human Th17 cells are shown in blue. These results are representative of three experiments.

The intricate balance of immune cells prevents the development of autoimmunity. T regulatory cells (Tregs) and Th17 cells derive from CD4⁺ precursors in the presence of TGF-beta 1. The key cytokine that skews these cells toward the Th17 lineage is IL-6, while IL-2 promotes Treg differentiation. IL-6 signaling induces STAT3, which upregulates ROR gamma t, the master regulator of Th17 cells. IL-2 signaling induces STAT5, which thereby induces FoxP3, leading to Treg differentiation. FoxP3 has been shown to be antagonistic towards Th17 cells, whereas IL-6 blocks Treg differentiation. When there is a skewing of the ratio towards the Th17 lineage, inflammatory cytokines are produced, which sets the stage for autoimmune disease progression. The immunosuppressive function of Tregs helps prevent the development of autoimmunity, but this state of tolerance can hinder the immune system from destroying cancer cells. Representative surface markers for Tregs and Th17 cells illustrate these points. Thus, understanding the correct balance of Tregs and Th17 cells in autoimmunity and cancer is critical for developing immunotherapies for these patients in the clinic.