Deciphering the relative roles of matrix metalloproteinase‐ and plasmin‐mediated matrix degradation during capillary morphogenesis using engineered hydrogels

Extracellular matrix (ECM) remodeling is essential for the process of capillary morphogenesis. Here we employed synthetic poly(ethylene glycol) (PEG) hydrogels engineered with proteolytic specificity to either matrix metalloproteinases (MMPs), plasmin, or both to investigate the relative contributions of MMP‐ and plasmin‐mediated ECM remodeling to vessel formation in a 3D‐model of capillary self‐assembly analogous to vasculogenesis. We first demonstrated a role for both MMP‐ and plasmin‐mediated mechanisms of ECM remodeling in an endothelial‐fibroblast co‐culture model of vasculogenesis in fibrin hydrogels using inhibitors of MMPs and plasmin. When this co‐culture model was employed in engineered PEG hydrogels with selective protease sensitivity, we observed robust capillary morphogenesis only in MMP‐sensitive matrices. Fibroblast spreading in plasmin‐selective hydrogels confirmed this difference was due to protease preference by endothelial cells, not due to limitations of the matrix itself. In hydrogels engineered with crosslinks that were dually susceptible to MMPs and plasmin, capillary morphogenesis was unchanged. These findings highlight the critical importance of MMP‐mediated degradation during vasculogenesis and provide strong evidence to justify the preferential selection of MMP‐degradable peptide crosslinkers in synthetic hydrogels used to study vascular morphogenesis and promote vascularization. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B:2507–2516, 2019.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/151850/1/jbmb34341_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/151850/2/jbmb34341.pd

A protocol for visualization of murine in situ neurovascular interfaces

Summary: Mapping cranial vasculature and adjacent neurovascular interfaces in their entirety will enhance our understanding of central nervous system function in any physiologic state. We present a workflow to visualize in situ murine vasculature and surrounding cranial structures using terminal polymer casting of vessels, iterative sample processing and image acquisition, and automated image registration and processing. While this method does not obtain dynamic imaging due to mouse sacrifice, these studies can be performed before sacrifice and processed with other acquired images.For complete details on the use and execution of this protocol, please refer to Rosenblum et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics