Microanatomical and cellular characterization of porcine spleen and lymph node as a basis for comprehensive mapping of immune organoid models

The porcine spleen and lymph nodes exhibit distinct structural and cellular features that influence immune responses and make pigs valuable translational models. This study comprehensively characterized the microanatomy and immune cell distribution in porcine spleen and lymph nodes using histology, immunohistochemistry (IHC), and flow cytometry. Haematoxylin and eosin staining revealed a nonsinusal spleen with extensive red pulp, poorly defined marginal zones, abundant ellipsoids, and sparsely distributed germinal centres. White pulp was organized into periarteriolar lymphoid sheaths (PALS) rich in T cells, with B-cell follicles located adjesent to PALS. Lymph nodes displayed the characteristic inverted architecture of swine, with central B-cell follicles and paracortex surrounded by peripheral medullary tissue. IHC localized CD3⁺ T cells predominantly to PALS and paracortical areas, while CD79α⁺ B cells occupied follicular regions. Flow cytometry confirmed T-cell predominance (60-70% of lymphocytes) and a higher proportion of CD4⁺ helper T cells compared to CD8⁺ cytotoxic T cells, alongside a greater B-cell frequency in lymph nodes than spleen. These findings delineate the structural specialisations and compartmentalised immune architecture of porcine secondary lymphoid organs, providing a foundational reference for immune organoid development, vaccine evaluation, and comparative immunology research.