Bone marrow stromal cells cause collapse of neocapillary networks in vitro

Abstract

Incorporation of bone marrow stromal cells (BMSCs) in tissues might improve tissue function. To determine effects on capillary formation in vitro, we plated GFP-expressing rat lung microvascular endothelial cells (RLMECs) in a 3-D culture system (Matrigel) that we viewed by real-time fluorescence imaging (n=4). RLMEC formed stable neocapillary networks in <24 h. We harvested BMSCs from rat tibias and femurs, stained the cells with the mitochondrial dye, Mitotracker deep red (MT), then added the cells to 1 day-old neocapillaries in a 1:1 ratio of BMEC to EC. We counted number of capillaries per high power field (HPF). Time-lapse imaging revealed BMSC intercalation between adjoining neocapillary EC within 2 days, as indicated by juxtaposition of red and green fluorescence (P<0.05). In control gels, capillary density remained stable at 7.0±1.0 capillaries/high power field (HPF) for 5 days. BMSC addition reduced capillary density to 1.6±1.1 capillaries/HPF within 2 days (P<0.05). This BMSC-induced network collapse was blocked by the gap junction blockers, glycerrhetinic acid, or GAP126/127 (P<0.05). Network collapse was also blocked when we grew BMSCs in a cell culture insert, which prevented BMSC contact with RLMEC. Our findings indicate that at high concentration, BMSCs incorporate into neocapillary networks, forming gap junctional contacts with EC and instituting capillary destruction (Support: HL36024).