Transient Activation of PIEZO1 Mechanosensitive Channel Facilitates Ex Vivo expansion of Hematopoietic Stem Cells

Achieving long-term ex vivo expansion of functional hematopoietic stem cells (HSCs) is critical for unlocking the potential of clinical applications based on HSC transplantation. Understanding the microenvironment in which HSCs maintain their functions is a prerequisite, with mechanical perception being one of the key factors. Studies have revealed mechanosensitive ion channels (MSICs) are vital in hematopoietic system, but their roles in the expansion and functional maintenance of ex vivo cultured HSCs remain largely elusive. Here, we show that PIEZO1 is the MSIC predominantly expressed in HSCs. Unexpectedly, both absence and sustained chemical activation of PIEZO1 are detrimental to cultured HSCs, suggesting that maintaining cultured HSCs required transient PIEZO1 activation. To identify conditions that lead to such activation, we screen different microspheres and demonstrate that stiff polymer microspheres with a diameter of 500 nm transiently activate PIEZO1. PS500 (polystyrene microsphere with diameter of 500 nm) markedly promotes ex vivo expansion of murine bone marrow (BM)-derived HSCs with long-term in vivo repopulating ability, and also showed expansion effect for human umbilical cord blood (hUCB)-derived HSCs with repopulating ability in primary immunodeficient mice. Mechanistically, the transiently-activated PIEZO1 channels trigger Ca²⁺-dependent transcription of proliferative cytokines, which ultimately lead to STAT3 activation to maintain self-renewal and promote cell proliferation and survival. Collectively, our results demonstrate that PS500 transiently activates PIEZO1 mechano-sensitive channel and efficiently expands functional HSCs without any toxicity, pointing towards a promising strategy to produce functional HSCs for clinical applications.