Bacteria-mammalian cell co-culture systems mediated by spatial engineering strategies

Bacteria and mammalian cells coexist and interact in many physiological and pathological environments, yet their direct in vitro co-culture is difficult because of mismatched growth rates, nutritional demands, and oxygen requirements. This Perspective discusses spatial engineering strategies that improve co-culture stability while preserving biologically relevant communication. We compare tumor spheroid colonization models, biomimetic hydrogel barriers, bacteria-containing microcapsules, layered hydrogel beads, and multiphasic droplets that support direct or indirect interactions. We further examine how material composition, crosslinking, pore size, shell thickness, mechanical stability, and compartment dimensions regulate bacterial confinement and molecular transport. Current systems still face trade-offs among leakage prevention, efficient exchange, physiological relevance, experimental duration, and throughput. Such models may enable mechanistic studies, therapeutic evaluation, and high-throughput investigation of dynamic bacteria-mammalian cell interactions under controlled conditions.