Heterogeneous evolution of pancreatic cancer microvasculature revealed by longitudinal intravital imaging.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy with a 5-year survival rate below 10%. Despite strong pro-angiogenic signaling often activated in PDAC, clinical trials with anti-angiogenic therapies have consistently failed, underscoring the need to understand PDAC vasculature better. In this study, we established an intravital, longitudinal imaging platform using a pancreas window in an orthotopic Panc02-GFP murine PDAC model. After surgical inoculation of tumor cells and the window implantation, PDAC vasculature was labeled by intravenous injection of fluorophore-conjugated anti-CD31 antibody, and perfusion was tracked with fluorescent red blood cells (RBCs). Longitudinal imaging of same regions was performed with real-time intravital confocal microscopy. It revealed dynamic vascular remodeling during PDAC progression, including angiogenesis, vessel pruning, dilation, and regression. Real-time tracking of fluorescent RBCs further demonstrated heterogeneous perfusion: some vessels abruptly lost flow and collapsed, while neighboring vessels showed fluctuating or partially recovering perfusion. This approach enables repeated, long-term, cellular-level visualization of both vascular remodeling and perfusion dynamics in vivo. By uncovering the spatiotemporal dynamics of PDAC vasculature, our platform provides new insights into tumor development and offers a valuable tool to evaluate vessel-targeting therapies.