Abstract
Cord blood hematopoietic stem cells (CB-HSCs) are an outstanding source for transplantation approaches. However, the amount of cells per donor is limited and culture expansion of CB-HSCs is accompanied by a loss of engraftment potential. In order to analyze the molecular mechanisms leading to this impaired potential we profiled global and local epigenotypes during the expansion of human CB hematopoietic stem and progenitor cells (HPSCs). Human CB-derived CD34+ cells were cultured in serum-free medium together with SCF, TPO, FGF, with or without Igfbp2 and Angptl5 (STF/STFIA cocktails). As compared to the STF cocktail, the STFIA cocktail maintains in vivo repopulation capacity of cultured CD34+ cells. Upon expansion, CD34+ cells genome-wide remodel their epigenotype and depending on the cytokine cocktail, cells show different H3K4me3 and H3K27me3 levels. Expanding cells without Igfbp2 and Angptl5 leads to higher global H3K27me3 levels. ChIPseq analyses reveal a cytokine cocktail-dependent redistribution of H3K27me3 profiles. Inhibition of the PRC2 component EZH2 counteracts the culture-associated loss of NOD scid gamma (NSG) engraftment potential. Collectively, our data reveal chromatin dynamics that underlie the culture-associated loss of engraftment potential. We identify PRC2 component EZH2 as being involved in the loss of engraftment potential during the in vitro expansion of HPSCs.
Highlights
Leading to reduced HLA-requirement compared to bone marrow (BM) cells
This approach introduced Insulin-like growth factor-binding protein 2 (Igfbp2) and a group of angiopoietin-like (Angptl) proteins, secreted glycoproteins consisting of seven members, as alternative growth factors for Hematopoietic stem cells (HSCs) expansion[9]
Functional characterization of CD34+ cells cultured under different cytokine cocktail conditions confirmed the increased multilineage engraftment potential in primary and secondary NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) recipients in STFIA- versus STF-cultured Cord blood (CB) CD34+ cells (Fig. 1A, Fig. S1)[9]
Summary
Leading to reduced HLA-requirement compared to BM cells. limited cell numbers per isolate restrict CB transplantation. The evolutionary conserved Polycomb-group (PcG) and Trithorax-group (trxG) proteins play pivotal roles in the regulation of HSC function[13,14] Both act as multifactorial complexes that influence gene expression by adding specific modifications to histone tails. Maps of the epigenetic landscapes of HSCs and differentiated progeny revealed that combinatorial modification patterns ensure cooperative regulation of transcription supporting the notion that epigenetics accompanies HSC function and differentiation[17] This notion is increasingly translated into practice as epigenetic strategies are considered for HSC expansion and as treatment option of hematopoietic malignancies[19,20,21]. Inhibition by EZH2 inhibitor treatment increased engraftment potential of cultured HPSCs
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