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Comparative analyses of erythroid cells from ES cells and cord blood CD34+ cells reveal mechanisms for defective expansion and enucleation of ES cell-derived erythroid cells

ABSTRACT: We used both RNA-seq and ATAC-seq analyses to uncovered mechanisms for the limited expansion and impaired enucleation in ES CD34+ cell-derived orthochromatic erythroblasts

ORGANISM(S): Homo sapiens

PROVIDER: GSE179778 | GEO | 2022/03/31

REPOSITORIES: GEO

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Comparative analyses of erythroid cells from ES cells and cord blood CD34+ cells reveal mechanisms for defective expansion and enucleation of ES cell-derived erythroid cells

Project description:Comparative analyses of erythroid cells from ES cells and cord blood CD34+ cells reveal mechanisms for defective expansion and enucleation of ES cell-derived erythroid cells

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Project description:Red blood cells (RBCs) generated ex vivo have the potential to be used for transfusion. Human embryonic stem cells (ES) and induced pluripotent stem cells (iPS) possess unlimited self-renewal capacity and are the preferred cell sources to be used for ex vivo RBC generation. However, their applications are hindered by the facts that the expansion of ES/iPS-derived erythroid cells is limited and the enucleation of ES/iPS-derived erythroblasts is low compared to that derived from cord blood (CB) or peripheral blood (PB). To address this, we sought to investigate the underlying mechanisms by comparing the in vitro erythropoiesis profiles of CB CD34+ and ES CD34+ cells. We found that the limited expansion of ES CD34+ cell-derived erythroid cells was associated with defective cell cycle of erythroid progenitors. In exploring the cellular and molecular mechanisms for the impaired enucleation of ES CD34+ cell-derived orthochromatic erythroblasts (ES-ortho), we found the chromatin of ES-ortho was less condensed than that of CB CD34+ cell-derived orthochromatic erythroblasts (CB-ortho). At the molecular level, both RNA-seq and ATAC-seq analyses revealed that pathways involved in chromatin modification were down-regulated in ES-ortho. Additionally, the expression levels of molecules known to play important role in chromatin condensation or/and enucleation were significantly lower in ES-ortho compared to that in CB-ortho. Together, our findings have uncovered mechanisms for the limited expansion and impaired enucleation of ES CD34+ cell-derived erythroid cells and may help to improve ex vivo RBC production from stem cells.

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Alpha 4 integrin deficiency in human CD34+ cells engenders a precocious erythroid differentiation ,but inhibits enucleation.

Project description:The functional impact of integrin expression in erythropoiesis has been previously emphasized through its decisive influence on erythroid cell-microenvironmental (matrix and cellular) interactions especially under conditions of stress. Beyond that in several in vitro studies the relationship between the two erythroid integrins, a4 and a5, has been incongruous in terms of a proliferative support, either synergistic or antagonistic, whereas a dominant influence of a4 integrin on terminal erythropoiesis in vitro and in vivo has been consistently emphasized. However, the specific cellular and molecular details of this effect have not been defined, especially for human cells. In the present study we have cultured human CD34+ progenitor cells with induced deficiency of a4 integrin (shRNAa4) under erythroid differentiation conditions, in which expanded erythroid progenitor cells are directed to terminal erythroid maturation stages in the absence of any microenvironmental influence. Our data documented that early proliferative expansion in cells lacking a4 expression is significantly limited, but, although erythroid differentiation can proceed normally to terminal stages, their enucleation is drastically impaired. This novel aspect of a4 integrin participation in the enucleation process in vitro resonates on the lack of in vivo enucleation of primitive erythroid cells lacking any integrin expression but affecting adult cells only under stress conditions.

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Alpha 4 integrin deficiency in human CD34+ cells engenders a precocious erythroid differentiation ,but inhibits enucleation.

Project description:Alpha 4 integrin deficiency in human CD34+ cells engenders a precocious erythroid differentiation ,but inhibits enucleation.

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Molecular Signature of Erythroblast Enucleation in Human Embryonic Stem Cells

Project description:While enucleation is a critical step in the terminal differentiation of human red blood cells, the molecular mechanisms underlying this unique process remain unclear. To investigate erythroblast enucleation, we studied the erythroid differentiation of human embryonic stem cells (hESCs), which provide a unique model for deeper understanding of the development and differentiation of multiple cell types. First, using a two-step protocol, we demonstrated that terminal erythroid differentiation from hESCs is directly dependent on the age of the embryoid bodies. Second, by choosing hESCs in two extreme conditions of erythroid culture, we obtained an original differentiation model which allows one to study the mechanisms underlying the enucleation of erythroid cells by analyzing the gene and miRNA (miR) expression profiles of cells from these two culture conditions. Third, using an integrated analysis of mRNA and miR expression profiles, we identified five miRs potentially involved in erythroblast enucleation. Finally, by selective knockdown of these five miRs we found miR-30a to be a regulator of erythroblast enucleation in hESCs.

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