Klebsiella Pneumoniae Mucosal Vaccination Elicits Lung CD4+ TRM cells that are resistant to CD4 depleting antibodies

Abstract

Abstract Objective Mucosal immunization with a Klebsiella pneumoniae (Kp) vaccine elicits noble CXCR6+CD4+ T cells that mediate serotype independent immunity, but the stability of this response is unclear. Thus we conducted experiments to determine if these cells circulate and whether they could be depleted by a depleting anti-GK1.5 mAb. Methods Mice were vaccinated via intratracheal with Kp serotype 2 OMPx adjuvanted with heat-labile enterotoxin A1 domain (LTA1) and boosted at 3 weeks after the initial immunization. T-cell responses were evaluated by FACS and ELISPOT. CD4+ T cell depletion in the immunized lung, lymph nodes, and spleen by administering 300ug anti-GK1.5 mAb intraperitoneally was examined using a secondary antibody for anti-GK1.5 mAb. Finally, lung and spleen CD4+ T cells of intratracheally immunized CD45.1+ C57BL/6 mice were transferred to CD45.2+ C57BL/6 mice and Rag2−/− mice. Results Immunized lung CD4+ T cells were increased equivalently in both C57BL/6 mice and Cxcr6−/− mice, demonstrating that CXCR6 was dispensable for the formation of lung CD4+ T cells. Splenic CD4+ T cells were > 95% depleted with anti-GK1.5 mAb. In contrast, < 5% of vaccine elicited lung CD4+ T cells were depleted despite being well coated by the antibody, strongly suggesting that these cells are TRM cells. Additionally, using an adoptive transfer model, we found that lung CD4+ T cells preferentially homed to lung compared to splenic CD4+ T cells. We next performed single-cell RNAseq on these cells and elucidated several unique adhesion molecules such as Embigin, Ramp1, Lgals1, and Thy1 besides Cxcr6. Conclusions These data support that vaccine strategies can be developed by targeting a unique feature of tissue-resident memory CD4+ T cells in lungs.