Fig 1: Human gut-brain-vascular (hGBV) physiomimetic system for modeling gut-brain axis interactions.a Schematic representation of the Gut-Brain Axis via circulation vessel system. b Gut-Brain-Vascular-on-a-chip platform. c Stepwise assembly of the hGBV tri-organs model. d Immunofluorescence of the integrated model, showing left: large-scale view, gut epithelium-red capillary-like vascular structures-blue, and brain tissue-cyan; right: 3D confocal zoom-in of Gut-VIL1, vascular-CD31, and astrocyte-GFAP. e-i Cross-sectional view of the gut epithelium (GC). e-ii Top view of the gut epithelium (VIL1; DAPI). f Zoom-in 3D of gut villus-like structures (VIL1; DAPI) in GC. g Quantification of VIL1 enrichment on the apical side relative to the nuclear plane (DAPI), supports apical VIL1 polarity in GC. h Immunofluorescence of gut-to-brain axis showing OCLN in GC, ZO-1 in VC, astrocyte-GFAP and neuron-Tuj1 in BC. i Quantification of OCLN localization, ZO-1 localization, and GFAP expression in gut-to-brain axis. j Immunofluorescence of brain-to-gut axis showing pTau in BC, ZO-1 in VC, OCLN in GC. k Quantification of pTau expression, ZO-1 localization, OCLN localization, in brain-to-gut axis. l–n RT-qPCR validation of barrier-related genes OCLN in GC, TJP1 in VC, and astrocytic GFAP in BC, confirming changes quantified in (i). o–q RT-qPCR validation of disease- and barrier-related genes APP in BC, TJP1 in VC, OCLN in GC, confirming the changes observed in (k). Scale bars, 500 μm (d), 100 μm (d, f), 20 μm (e, h, j). Representative images are shown from one experiment; similar results are obtained in three independent experiments with comparable outcomes (d–f, h, j). Quantification is based on n = 10 independent chips per condition (g, i, k), n = 5 in independent chips per condition (l–q). Data represent Mean ± SEM; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001; ns, not significant. Statistical analysis: two-sided tests selected per dataset (Student’s t-test/Welch’s t-test or Mann-Whitney U for two-group comparisons; one-way ANOVA/Welch’s ANOVA with Tukey or Kruskal-Wallis with Dunn for multi-group comparisons), with multiple-comparisons adjustments where applicable; exact tests and full statistics (effect sizes, 95% CIs, exact P values) are reported in Supplementary Table S2. CT represents control; E.CCM+, E. Coli-conditioned media, WT wide-type, AD Alzheimer’s disease, GC gut compartment, VC vascular compartment, BC brain compartment, NPC neural progenitor cell. Created in BioRender. Vu, T. A. (2026) https://BioRender.com/702y808 (a) & https://BioRender.com/702y808 (b).