02644nas a2200301   4500000000100000008004100001260001500042653002100057653001500078653002900093653002600122653001800148100002300166700001700189700002600206700001700232700001800249700002700267700001600294700002100310700002000331245011200351856007200463300001100535490000700546520177500553022001402328       2025                            d  c2025-12-0110ahuman immunology10alymph node10aLymph node stromal cells10aLymphatic vasculature10aorgan-on-chip1 aAndrew I. Morrison1 aJonas Jäger1 aCharlotte M. de Winde1 aTanja Konijn1 aHenk P. Roest1 aLuc J. W. van der Laan1 aSusan Gibbs1 aJasper J. Koning1 aReina E. Mebius00aIntegration of lymphatic vasculature to a human lymph node-on-chip enhances physiological immune properties  uhttps://www.sciencedirect.com/science/article/pii/S259000642500897X  a1023260 v353 aTo study systemic human innate and adaptive immune responses in detail, competent in vitro lymph node (LN) models with LN stromal cells (LNSCs) are required to recapitulate the physiological microenvironment. The multicellular organisation of LNs possesses a challenge for designing such microphysiological systems (MPS), particularly with the structural complexity of LNs and the lymphatic vasculature. Here, we established an organotypic LN model with integrated lymphatics in an organ-on-chip (OoC) platform containing a printed sacrificial structure, and studied the influence of a perfused lymphatic endothelial cell (LEC)-lined channel on the LN-on-chip microenvironment. Upon one-week of culture under lymphatic flow, LECs lined the tubular structure forming a lymphatic vessel through the LN model, and stable metabolic conditions within the LN-on-chip were confirmed. Interestingly, LECs in the LN-on-chip displayed the phenotype found in human LNs with upregulation of LEC-specific LN markers, such as atypical chemokine receptor 4 (ACKR4). The presence of the LEC-lined perfused vessel in the LN-on-chip resulted in the increase of native immune cells, most notably B cells, and the secretion of survival and migratory signals, namely interleukin-7 (IL-7) and CC motif chemokine ligand 21 (CCL21). Likewise, LECs promoted the abundance of immune cell clusters closer to the vessel. As such, these features represent an enhanced physiological microenvironment to allow for immune cell migration and interactions for efficient LN functioning. This approach paves the way for LN integration into multi-OoC (MOC) platforms to investigate immunological crosstalk between tissue-derived factors, immune cell trafficking and organ-specific adaptive immune responses.  a2590-0064