*Purpose: Aquaporins are a family of transmembrane water channels implicated in a broad range of physiological functions. We previously reported that Aquaporin 4 (AQP4) is expressed by T cells and treatment with a small molecule AQP4 inhibitor (AER-270) significantly delays T cell mediated mouse heart allograft rejection. Our findings did not exclude a role for antigen presenting cells (APCs), and the purpose of this study is to investigate the requirement for AQP4 in dendritic cell functions.

*Methods: Mar TCR Tg T cells (reactive to HY^Dby epitope + I-A^b) were stimulated with WT or AQP4^-/- spleen APCs with the addition of HY^Dbyp. Bone marrow derived dendritic cells (BMDCs) were cultured from WT and AQP4^-/- mice in combination with AER-270 treatment. Changes in the expression of DC maturation markers after stimulation with either LPS or protein antigen were measured by flow cytometry. Antigen uptake was assessed following pulse of DCs with Lucifer Yellow, FITC-dextran and OVA-AF647. Antigen presentation was measured after pulse with OVA or BALB.C cell lysates with anti-SIINFEKL-H2-K^b and anti-Ea-I-A^b respectively. BALB.C (H-2D^d) to B6 (H-2D^d) skin transplants were performed to analyze donor antigen presentation in vivo. Antigen processing capacity was determined by pulsing BMDCs with DQ-OVA (an acid reactive fluorescent reporter).

*Results: Stimulation with AQP4^-/- spleen APCs resulted in reduced frequencies of IFNγ producing MAR T cells compared to WT APCs. Splenic CD11c⁺ DCs express high levels of AQP4. AQP4^-/-BMDCs have no defect in up-regulation of surface markers CD40, CD80, CD86, MHC-I, MHC-II following stimulation. The absence or inhibition of AQP4 did not affect antigen uptake, regardless of the antigen uptake pathway. However, AQP4^-/- BMDCs pulsed with OVA protein had reduced expression of SIINFEKL peptide-MHC-I complexes. In contrast, when pulsed with OVA-derived SIINFEKL peptide, AER-270 treated WT BMDCs or AQP4^-/- BMDCs showed no change in peptide-MHC complex expression. Taken together these data suggest a role for AQP4 in antigen processing. Consistent with this, the absence or inhibition of AQP4 resulted in decreased DQ-OVA signal, indicating that AQP4 is required for the antigen loading into the lysosome.

*Conclusions: Our data indicate that AQP4 plays a critical role in the processing of antigens by dendritic cells that impacts their ability to prime T cells, and can be therapeutically targeted in a transplant setting.

« Back to 2022 American Transplant Congress