Abstract
Microgravity has been shown to induces many changes in proliferation, differentiation and growth behavior of stem cells. Little is known about the effect of microgravity on hematopoietic differentiation of pluripotent stem cells (PSCs). In this study, we used the random position machine (RPM) to investigate whether simulated microgravity (SMG) allows the induction of hematopoietic stem/progenitor cell (HSPC) derived from human embryonic stem cells (hESCs) in vitro. The results showed that SMG facilitates hESCs differentiate to HSPC with more efficient induction of CD34+CD31+ hemogenic endothelium progenitors (HEPs) on day 4 and CD34+CD43+ HSPC on day 7, and these cells shows an increased generation of functional hematopoietic cells in colony-forming unit assay when compared with normal gravity (NG) conditions. Additionally, we found that SMG significantly increased the total number of cells on day 4 and day 7 which formed more 3D cell clusters. Transcriptome analysis of cells identified thousands of differentially expressed genes (DEGs) between NG and SMG. DEGs down-regulated were enriched in the axonogenesis, positive regulation of cell adhesion, cell adhesion molecule and axon guidance, while SMG resulted in the up-regulation of genes were functionally associated with DNA replication, cell cycle, PI3K-Akt signaling pathway and tumorigenesis. Interestingly, some key gene terms were enriched in SMG, like hypoxia and ECM receptor interaction. Moreover, HSPC obtained from SMG culture conditions had a robust ability of proliferation in vitro. The proliferated cells also had the ability to form erythroid, granulocyte and monocyte/macrophage colonies, and can be induced to generate macrophages and megakaryocytes. In summary, our data has shown a potent impact of microgravity on hematopoietic differentiation of hPSCs for the first time and reveals an underlying mechanism for the effect of SMG on hematopoiesis development.
Highlights
Exposure to microgravity induces a variety of changes in physiology and some specialized tissue, including musculoskeletal system, nervous system, cardiovascular system, and immune system (Garrett-Bakelman et al, 2019; Bradbury et al, 2020)
We demonstrated that simulated microgravity (SMG) facilitates human embryonic stem cells (hESCs) differentiate to hematopoietic stem/progenitor cell (HSPC) with more efficient induction of CD34+CD31+ hemogenic endothelium progenitors (HEPs) on day 4 and
CD34+CD43+ HSPC on day 7, and these cells shows an increased generation of functional hematopoietic cells in colony-forming unit (CFU) assay when compared with normal gravity (NG) conditions
Summary
Exposure to microgravity induces a variety of changes in physiology and some specialized tissue, including musculoskeletal system, nervous system, cardiovascular system, and immune system (Garrett-Bakelman et al, 2019; Bradbury et al, 2020). The effect of microgravity on hematopoietic differentiation of human embryonic stem cells (hESCs) has not been yet examined. Several signaling pathways were identified to play vital roles in hematopoietic differentiation of hESCs. The activation of canonical Wnt signaling with GSK-3 inhibitor CHIR99021 in hESCs can enhance definitive hematopoiesis and decrease the number of primitive HSPC (Sturgeon et al, 2014). Inhibition of Nodal/Activin pathway with SB-431542 in hematopoietic differentiation of hESCs can induce the development of definitive HSPC and attenuate the primitive hematopoiesis process (Kennedy et al, 2012). Inhibition of transforming growth factor-β (TGF-β) have been reported to play a vital role in priming hemogenic potential in epithelial cells signaling at the HEPs stage are increased a capacity to produce HSPC through the downregulation of mesenchymal genes (Wang Y. et al, 2020). Biochemical stimuli factors are known for enhancing cell fate transitions in hematopoiesis process, there is growing appreciation on the role of biomechanical stimuli factors (North et al, 2009) or through a combination of biochemical and biomechanical factors for hematopoietic differentiation (Yang et al, 2016)
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