A culture platform to study quiescent hematopoietic stem cells following genome editing

Other than genetically engineered mice, few reliable platforms are available for the study of hematopoietic stem cell (HSC) quiescence. Here we present a platform to analyze HSC cell cycle quiescence by combining culture conditions that maintain quiescence with a CRISPR-Cas9 genome editing system optimized for HSCs. We demonstrate that preculture of HSCs enhances editing efficiency by facilitating nuclear transport of ribonucleoprotein complexes. For post-editing culture, mouse and human HSCs edited based on non-homologous end joining and cultured under low-cytokine, low-oxygen, and high-albumin conditions retain their phenotypes and quiescence better than those cultured under the proliferative conditions. Using this approach, HSCs regain quiescence even after editing by homology-directed repair. Our results show that low-cytokine culture conditions for gene-edited HSCs are a useful approach for investigating HSC quiescence ex vivo.

Automated summary

This study presents a platform for analyzing the quiescence of hematopoietic stem cells (HSCs) by combining culture conditions and a CRISPR-Cas9 genome editing system. The study demonstrates that preculturing HSCs enhances editing efficiency and that low-cytokine culture conditions better preserve the phenotypes and quiescence of edited HSCs.