Regulation Of The Bone Marrow-derived Endothelial Progenitor Cells And Endothelial Out-growth Cells By The Notch Signaling Pathway

Endothelial progenitor cells (EPCs) have therapeutic potentials inpromoting tissue regeneration, however EPCs is a group of heterogeneous cells,the functions and regulation mechanisms in the process of mobilization fromthe bone marrow into the circulation and homing to the sites ofneovascularization are elusive. EPCs in BM or just entering the peripheral bloodexpress stem cell markers such as CD34and CD133, together with VEGFR2(KDR). Along with in vitro culturing and maturation, the cells gradually loststem cell markers, and begin to express EC specific antigens such as plateletendothelial cell adhesion molecule1(PECAM-1or CD31) and VE-cadherin,among others.Other researchers have suggested that EPCs is composed of endotheliallineage cells at different differentiation stages. Two types of EPCs have beenidentified for in vitro cultured EPCs. Early EPCs (EEPCs) are spindle-like in shape, and have limited proliferative potential and can be cultivated no morethan4weeks in vitro. Endothelial outgrowth cells (EOCs), in contrast, have acobblestone-like appearance and maintain a high proliferative potential. Butfurther defining different subpopulations of EPCs and understanding their rolesand mechanisms in vascularization is still required.The Notch pathway is highly conserved in evolution, and plays an essentialrole in cell fate determination in multiple lineages of stem and progenitor cells.Kwon et al have shown that the Jagged1-mediated Notch signaling promoteadult neovascularization by regulating the function of EPCs. Our have alsofound that Notch-RBP-J signaling regulates the mobilization, migration andfunction of EPCs through the expression of CXCR4. However, in these studiesthe role of the Notch signaling pathway in different subpopulations of EPCs,such as EEPCs and EOCs, has not been revealed. In this study, we accessed thisquestion by using in vitro cultured EPCs and RBP-J knockout mice. Our datasuggested that Notch-RBP-J signaling regulates the functions of EEPCs andEOCs in different ways.The main finds are as follows:1．We successfully cultured the bone marrow derived EEPCs and EOCs,cells with the EEPC phenotypes, CD34+/CD133+/VEGFR2+, accounted for only0.08%of total population of cells. After cultured for10days in the EPCmedium, this percentage reached50.59%.2． We found that EEPCs failed to sprout and to form lumen-likestructures. When EOCs were cultured in the system, sprouting started onaround day2, and cord-like lumen structure grew out with the culture proceeded.Blocking the Notch-RBP-J signaling pathway in vitro, EEPCs proliferation,migration and expression of CXCR4significantly decreased. Interestingly, the proliferation migration and expression of CXCR4of EOCs increased, indicatingthat Notch-RBP-J signaling pathway may has significant different role inregulating the function of EEPCs and EOCs.3． EEPCs and EOCs might play different roles in tissue repair andregeneration. Our transplantation experiments have shown that EEPCs canpromote liver regeneration with respect to liver function and hepatocyteproliferation and apoptosis, although these cells appear incompetent in directlyparticipating in vessel formation, at least in vitro. In contrast, EOCs could sproutand form vessel-like tubes under appropriate conditions, but EOCs seem not beable to promote liver regeneration in our systems. Moreover, our results suggestthat EEPCs and EOCs might take part in liver repair and regeneration throughdifferent mechanisms.In summary, we got the RBP-J knockout mice and found that bone marrowderived EEPC and EOC take part in liver repair and regeneration and this wasrequired by Notch/RBP-J signaling. Our results show that, Notch signalsinvolved in regulating the liver function and hepatocyte proliferation andNotch-RBP-J signaling pathway regulated the proliferation and migration ofEEPCs and EOCs. EEPCs and EOCs might take part in liver repair andregeneration through different mechanisms.Our results reveal understanding ofthe function and mechanisms of the Notch signaling pathway in thedevelopment and function of endothelial stem/progenitor cells.