Early gut-vascular barrier breakdown precedes colitis onset in murine models

BACKGROUND & AIMS: The gut–vascular barrier (GVB) plays a pivotal role in inflammatory bowel disease (IBD) pathogenesis. We introduce a novel three-dimensional (3D) multiphoton endomicroscopy (MPEM) approach for real-time and sensitive detection of vascular permeability (VP) in the colon to identify colitis-associated vascular changes in the early stages. METHODS: Using fluorescence-based MPEM, we visualized dynamic changes in VP in vivo during longitudinal observations in different experimental colitis models (DSS- and T cell transfer-induced colitis). VP changes were systematically compared with conventional inflammatory markers, including weight loss, endoscopic scoring, colon length, histopathology, and immune cell infiltration. To assess molecular regulation of barrier functions, the expression of key molecules of vascular (PV1, VEcadherin) and epithelial (E-cadherin) barriers was investigated during development of colitis at the single-cell level. RESULTS: MPEM provided unprecedented 3D visualization of VP dynamics and showed that vascular dysfunction occurs prior to epithelial barrier breakdown and the detection of traditional inflammatory markers across all colitis models. Distinct spatial VP patterns strongly correlated with mucosal damage severity, further supporting that early GVB disruption precedes mucosal barrier breakdown. Additionally, this sequence was confirmed at the molecular level, with the vascular upregulation of the trans-endothelial permeability channel PV1 occurring before downregulation of the epithelial barrier molecule E-cadherin. CONCLUSIONS: MPEM with 3D imaging demonstrated GVB dysfunction in the very early stages of experimental colitis. Further development of MPEM-based VP analysis for use in routine clinical monitoring of patients may provide new perspectives to improve diagnosis and clinical decisionmaking in IBD.