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Rapid induction and creation of functional vascular networks is essential for the success of treating ischemic tissues. The formation of mature and functional vascular networks requires the cooperation of endothelial cells (ECs) and perivascular cells. In the study, we used a thermo-responsive hydrogel system to fabricate core-shell cell bodies composed of cord-blood mesenchymal stem cells (cbMSCs) and human umbilical vascular ECs (HUVECs) for functional vasculogenesis. When seeded on Matrigel, the shelled HUVECs attempted to interact and communicate vigorously with the cored cbMSCs initially. Subsequently, HUVECs migrated out and formed tubular structures; cbMSCs were observed to coalesce around the HUVEC-derived tubes. With time progressing, the tubular networks continued to expand without regression, indicating that cbMSCs might function as perivascular cells to stabilize the nascent networks. In the in vivo study, cbMSC/HUVEC bodies were embedded in Matrigel and implanted subcutaneously in nude mice. At day 7, visible blood-filled vessels were clearly identified within the implant containing cbMSC/HUVEC bodies, indicating that the formed vessels anastomosed with the host vasculature. The cored cbMSCs were stained positive for smooth muscle actin, suggesting that they underwent smooth muscle differentiation and formed microvessels with the shelled HUVECs, as the role of perivascular cells. These data confirm that the formation of mature vessels requires heterotypic cooperation of HUVECs and MSCs. This study provides a new strategy for therapeutic vasculogenesis, by showing the feasibility of using cbMSC/HUVEC bodies to create functional vascular networks.
Confirming our previous work [6,10] we found that the co-implantation of genetically modified human umbilical vein endothelial cells (HUVEC) harboring the firefly luciferase (FLuc) gene (HUVECFLuc) and human bone marrow derived MSC (Figure S1-S4) embedded within a reconstituted basement membrane extract (BME, also known as Matrigel) supplemented with VEGF, bFGF and heparin and inoculated subcutaneously into nude mice (Figure 1a), results in the formation of mature, stable human blood vessels. These vessels are functional, meaning that they are able to anastomose with preexisting mouse vessels, and can be assessed non-invasively and quantitatively by in vivo bioluminescent imaging (BLI) (Figure 1b). The temporal silencing of luciferase activity and subsequent recovery is related to the lapse of time required for the establishment of an incipient vascular network and its integration into the mouse vascular bed .
We have herein described a new mouse model to study metastatic colonization, and we demonstrate that disseminated human breast cancer cells efficiently colonize BME-rich organoids containing a functional microvessel network composed of human endothelial cells connected to the mouse circulatory system. Despite their apparent structural simplicity, HVO were efficiently colonized by MDA-MB-231 cells, and importantly, human breast cancer cells could be unequivocally detected at different stages of the metastatic process: initial cell arrest in human capillaries, extravasation, and growth from avascular micrometastases to vascularized micrometastases growing into the surrounding BME. In control BME-rich organoids without human cells, the number of vessels was lower than in the HVO and these were more immature and leaky. Despite increased leakiness of vessels in CO, the percentage of metastatic colonization was significantly lower than that observed in HVO, although we cannot rule out that differences in microvascular density may contribute to differences in the number of metastasis. 041b061a72