人胚胎干细胞诱导分化生成血管类器官的研究OA

Objective To establish and characterize vascular organoids based on the capacity of definitive differentiation and multiple cell self-organization of human embryonic stem cells(hESCs).Methods The hESCs were subjected to a stepwise induction protocol to sequentially differentiate into mesoderm progenitors,lateral plate mesoderm progenitors,and endothelial cells over a 5-day period.The resulting endothelial cell population was harvested and embedded in Matrigel to undergo three-dimensional culture of 6-8 days,allowing for self-organization into vascular organoids with branched luminal networks.The organoids were characterized via morphological analysis,flow cytometry,immunofluorescence staining,and transmission electron microscopy.Results The generated organoids formed complex networks of branched lumens,containing(41.10±4.51)%CD31+ endothelial cells and(6.88%±0.40)%CD140b+ pericytes.Among the endothelial cells,(43.00±0.21)%were CD31+CD184+ arterial endothelial cells and(3.66±0.25)%were CD31+CD184-CD73+ venous endothelial cells.Immunofluorescence analysis found that the vascular organoids were composed of VE-cadherin + Sox17 +arterial endothelial cells,VE-cadherin + NR2F2 + venous endothelial cells,PDGFR-β + pericytes,α-SMA + smooth muscle cells,and collagen I.Tight junctions were observed between endothelial cells,and characteristic Weibel-Palade body ultrastructures were identified within the cytoplasm that resembled the structures of human microvascular units.Conclusion We have established a robust method for generating hESCs-based vascular organoids that mimic the architecture of functional microvasculature.This three-dimensional vascular organoid can serve as a promising platform for research on vascular development and disease modeling.