Environmental Optimization Enables Maintenance of Quiescent Hematopoietic Stem Cells Ex Vivo

Abstract

Hematopoietic stem cells (HSCs) maintain lifelong hematopoiesis by remaining quiescent in the bone marrow niche. Recapitulation of a quiescent state in culture has not been achieved, as cells rapidly proliferate and differentiate in vitro. After exhaustive analysis of different environmental factor combinations and concentrations as a way to mimic physiological conditions, we were able to maintain engraftable quiescent HSCs for 1 month in culture under very low cytokine concentrations, hypoxia, and very high fatty acid levels. Exogenous fatty acids were required likely due to suppression of intrinsic fatty acid synthesis by hypoxia and low cytokine conditions. By contrast, high cytokine concentrations or normoxia induced HSC proliferation and differentiation. Our culture system provides a means to evaluate properties of steady-state HSCs and test effects of defined factors in vitro under near-physiological conditions.

Towards Mimicking the Fetal Liver Niche: The Influence of Elasticity and Oxygen Tension on Hematopoietic Stem/Progenitor Cells Cultured in 3D Fibrin Hydrogels

“…On the other hand, the LSK-SLAM cell number remained similar in hypoxic conditions compared to in normoxic conditions over the 10 days of culture. Similar effects were also reported for HSPCs cultured in suspension in hypoxia or normoxia, suggesting that normoxia enhanced the expansion of TNCs and progenitors while hypoxia preserved FLT3 − CD34 − LSK cells (a subgroup of HSPCs between LSK and LSK-SLAM cells [ 57 ]) by inducing a quiescent state [ 42 , 58 , 59 ].…”

Alpha lipoic acid promotes development of hematopoietic progenitors derived from human embryonic stem cells by antagonizing ROS signals

“…Hypoxia and a low ROS level are required for HSCs self‐renewal and maintenance 1–4 . Optimization of the levels of environmental factors such as oxygen, cytokines, and fatty acids enables maintenance of HSCs quiescence ex vivo 5 . Small molecules that can maintain low ROS levels 6 and inhibit cell apoptosis or autophagy 7 are also useful for maintenance or expansion of HSCs.…”

Awakening the HSC: Dynamic Modeling of HSC Maintenance Unravels Regulation of the TP53 Pathway and Quiescence

“…This evaluation might be quite challenging under in vitro conditions where both isolation and maintenance of HSC require much effort. Identifying methods of isolation and maintenance of HSCs is still challenging (Passegué et al, 2005 ; Weissman and Shizuru, 2008 ; Challen et al, 2009 ; Rossi et al, 2011 ; Frisch and Calvi, 2014 ; Jiang et al, 2018 ; Kobayashi et al, 2019 ). Hence, the possibility of having an in silico mathematical model for simulating the behavior of HSC is of great interest.…”

“…Similarly, if we consider our whole set of attractors, we also observe an expression of anti-apoptotic proteins. Sustaining our hypothesis of maintenance of LT-HSC, Kobayashi et al recently published a study describing the ideal setting for in vitro maintenance of quiescent HSCs (Kobayashi et al, 2019 ). In their work, the authors actually show that hypoxic condition in the presence of residual cytokine stimuli sets the best environment for the maintenance of quiescence (Kobayashi et al, 2019 ).…”