Haematopoietic stem cell survival and transplantation efficacy is limited by the BH3-only proteins Bim and Bmf.
Bottom Line: Thereby, we revealed a critical role for Bim and Bmf as regulators of HSPC dynamics both during early engraftment and long-term reconstitution.HSPCs derived from wild-type donors were readily displaced by Bim- or Bmf-deficient or Bcl-2-overexpressing HSPCs as early as 10 days after engraftment.Finally, we provide proof of principle that RNAi-based reduction of BIM or BMF, or overexpression of BCL-2 in human CD34(+) cord blood cells may be an attractive therapeutic option to increase stem cell survival and transplantation efficacy.
Affiliation: Division of Developmental Immunology, Biocenter, Innsbruck Medical University, Innsbruck, Austria. firstname.lastname@example.orgShow MeSH
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Mentions: Lentivirally transduced shRNA specific for BIM or BMF was used to test the relevance of these two BH3-only proteins for the induction of human HSPC apoptosis. In addition, lentiviruses were generated for expression of their antagonists BCL-2 and BCL-XL. The RNAi-knockdown efficiency was determined by qRT-PCR for BIM and BMF mRNA (Fig 5C) and by immunoblotting for target proteins isolated from sorted GFP+ CD34+ cells (Supporting Information Fig 5C). RNA levels of BIM and BMF were reduced by ∼69 and ∼78%, respectively, at the time of analysis (Fig 5C). Culturing GFP+ CD34+ cells for 2 days in the absence of cytokines revealed that knockdown of BIM but not BMF delayed cytokine deprivation-induced apoptosis ex vivo, although this effect was not as pronounced as that seen in Bim-deficient mouse HSPCs (comp. Figs 6A, 1B). After 4 days BIM knockdown proved inefficient, whereas overexpression of BCL-2 or BCL-XL still effectively increased survival (Fig 6A, right panel).
Affiliation: Division of Developmental Immunology, Biocenter, Innsbruck Medical University, Innsbruck, Austria. email@example.com