Impact of the overexpression of the tyrosine kinase receptor RET in the hematopoietic potential of induced pluripotent stem cells (iPSCs)

Abstract

Introduction: Previous studies have suggested that the tyrosine kinase receptor RET plays a significant role in the hematopoietic potential in mice and could also be used to expand cord-blood derived hematopoietic stem cells (HSCs). The role of RET in human iPSC-derived hematopoiesis has not been tested so far.

Methods: To test the implication of RET on the hematopoietic potential of iPSCs, we activated its pathway with the lentiviral overexpression of RET^WT or RET^C634Y mutation in normal iPSCs. An iPSC derived from a patient harboring the RET^C634Y mutation (iRET^C634Y) and its CRISPR-corrected isogenic control iPSC (iRET^CTRL) were also used. The hematopoietic potential was tested using 2D cultures and evaluated regarding the phenotype and the clonogenic potential of generated cells.

Results: Hematopoietic differentiation from iPSCs with RET overexpression (WT or C634Y) led to a significant reduction in the number and in the clonogenic potential of primitive hematopoietic cells (CD34⁺/CD38^-/CD49f⁺) as compared to control iPSCs. Similarly, the hematopoietic potential of iRET^C634Y was reduced as compared to iRET^CTRL. Transcriptomic analyses revealed a specific activated expression profile for iRET^C634Y compared to its control with evidence of overexpression of genes which are part of the MAPK network with negative hematopoietic regulator activities.

Conclusion: RET activation in iPSCs is associated with an inhibitory activity in iPSC-derived hematopoiesis, potentially related to MAPK activation.

Keywords: CD34; HSCs; RET; hematopoietic differentiation; iPSCs.