| Mesenchymal stem cells (MSCs) contribute to the maintenance and regeneration of a variety of connective tissues, including bone, cartilage, adipose, tendom, muscle and stomal cell. Characterization of MSCs in adult significantly accelerates their applications in regenerative medicine. However, little is known regarding their early embryonic ontogeny and the mechanism of migration from emerging site to new hematopoissis sites during embryonic development.During embryonic development, the first HSCs were generated in aorta-gonad-mesonephros (AGM ) region, then they migrated through the circulation into multiple hemogenic sites. MSC which Served as the HSC supportive microenviroment, whether it can also be generated in the AGM region or migrated into other hematopoietic sites, remain unknown .The Dzierzak team first explored the mesenchymal differeation ability of primary cells from AGM and found that primary adherent cells derived from AGM had the potential to differentiate into osteogenic and chondrogenic lineage but not adipogenic lineage . The same team also examined the potential of immortal AGM stromal clones to differentiate into mesenchymal lineages, the clones are only uni-potential, but one of the clones was able to form adipose droplets. From the above two articles, It can be infered that mouse AoM may exist tri-potential MSC. We successfully identify canonical tri-potential MSCs in the mouse E11.5 aorta with surrounding mesenchyme (AoM), and from E12.5-E14.5 circulation blood through optimizing the condition of in vitro culture and differentiation . The P2-P3 AoM-MSCs, Blood-MSC and Bone-MSC did not express hematopoietic and endothelial markers (CD45-CD31- Flk-1-CD34-CD11b-), expressed pericyte markers (CD140b+CD140a+α-SMA+NG2+) and mesenchymal cell markers (CD44+CD29+ CD105+), and could in vitro induced into adipogenic, osteogenic , chondrogenic differentiation. Then ,we analyze the frequency of Colony forming unit of fibroblast (CFU-F) of E11.5-E12.5 AoM derived cells (E11.5, 3.125±1.4 /2000 Cells;E12.5, 2.55±1.468 /2000 Cells),and the total CFU-F of E11.5-E12.5 embryo (E11.5,128.53±55.37;E12.5, 240.975±138.726 ).Later, we analyze the differentiation ability of single CFU-F derived from E11.5 embryo, in the total 24 CFU-F Clones , 4.17%(1/24)Clone had the tri-potential,20.8%(5/24)Clones had bi-potential ,70.83%(17/24)clones had uni-potetial .Hematopoietic stem cell (HSC) could autonomously emergence in AGM region, transmigrate through the endothelial cell layer and release in the blood flow, colonize the fetal liver , finally engraft in bone marrow. Do AoM-MSC have the same property of migration ? We found that embryonic blood and endothelial cell conditional medium (ECM) significantly induced AoM-MSC migration, but not conditional medium from primitive red cells. Previous research showed that growth factors were more potent chemotactic agents than chemokines for bone marrow-derived MSCs. Among 11 candidates pivotal in adult MSC trafficking, the highest chemotactic activities of AoM-MSCs were observed with bFGF, PDGF-BB, TGF-β1 and TGF-β3.The embryonic blood and ECM induced migration of the AoM-MSC, which was inhibited by adding small molecular inhibitors of PDGF receptor, secondly inhibited by TGF-βreceptor, but not bFGF receptor .MAPK pathways are closely related to cell migration , The embryonic blood ,ECM and PDGF-BB mediated migration of AoM-MSCs was inhibited by JNK inhibitor, SP600125 or PI3K inhibitor, LY294002 , however, ERK and P38 pathway inhibitor did not work .when the AoM-MSC migrated into the vascular cavity ,they were encircled by large numbers of primitive red blood cells , although the conditional medium from primitive red blood cells could not remarkably induced migration of AoM-MSC, but the interaction between primitive red cell and MSC significantly reduced migration ability of AoM-MSC, however, the mechanism of how to modulate the migration of MSC needs further explore.Recently, The Nishikawa team showed that the earliest wave of MSC in E9.5 embryonic trunk is generated from Sox1+ neuroepithelium but not from mesoderm. However this wave of Sox1+ derived MSC is transient and is replaced by MSCs from unknown sources. The dominant origin of late embryo and adult bone marrow remains unkown, Our experiment confirmed that tri-potential MSC existed in E11.5 AoM vascular region and the AoM-MSC could migrate via embryonic circulation in vitro. This research added new and important envidence to the hypothesis that MSC generate and locate in the perivascular zone in vivo, but the relationship between AoM-MSC and Sox1+ neuroepithelium derived MSC is unknown. We will further utilize PDGFRa-Cre and PDGFRb-Cre mouse to investigate the dominant origin of adult MSC and in vivo property of migration. |
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