Organoid Polymer Functionality and Mode of Klebsiella Pneumoniae Membrane Antigen Presentation Regulates Ex Vivo Germinal Center Epigenetics in Young and Aged B Cells

Antibiotic-resistant bacteria are a major global health threat that continues to rise due to a lack of effective vaccines. Of concern are Klebsiella pneumoniae that fail to induce in vivo germinal center B cell responses, which facilitate antibody production to fight infection. Immunotherapies using antibodies targeting antibiotic-resistant bacteria are emerging as promising alternatives, however, cannot be efficiently derived ex vivo, necessitating the need for immune technologies to develop therapeutics. Here, four-arm PEG-organoids were developed to elucidate the effects of polymer end-point chemistry, integrin ligands, and mode of K. pneumoniae antigen presentation on germinal center-like B cell epigenetics, to better define the cell-microenvironment factors regulating ex vivo germinal center dynamics. Notably, PEG vinyl sulfone or acrylate failed to sustain primary immune cells, but functionalization with maleimide (PEG-4MAL) led to B cell expansion and germinal center-like induction. RNA sequencing analysis of lymph node stromal and germinal center B cells showed niche associated heterogeneity of integrin-related genes. Incorporation of niche-mimicking bioadhesive peptides revealed that collagen 1 mimicking peptides promoted germinal center-like dynamics and epigenetics. PEG-4MAL organoids elucidated the impact of K. pneumoniae membrane embedded protein antigen versus soluble antigen presentation on germinal center-like activation and preserved the response across young and aged mice.