Generation of hypothalamic neural stem cell-like cells in vitro from human pluripotent stem cells

When damaged, restoring the function of the hypothalamus is currently impossible. It is unclear whether neural stem cells exist in the hypothalamus. Studies have reported that adult rodent tanycytes around the third ventricle function as hypothalamic neural stem cell-like cells. However, it is currently impossible to collect periventricular cells from humans. We attempted to generate hypothalamic neural stem cell-like cells from human embryonic stem cells (ESCs). We focused on retina and anterior neural fold homeobox (RAX) because its expression is gradually restricted to tanycytes during the late embryonic stage. We differentiated RAX::VENUS knockin human ESCs (hESCs) into hypothalamic organoids and sorted RAX⁺ cells from mature organoids. The isolated RAX⁺ cells formed neurospheres and exhibited self-renewal and multipotency. Neurogenesis was observed when neurospheres were transplanted into the mouse hypothalamus. We isolated RAX⁺ hypothalamic neural stem cell-like cells from wild-type human ES organoids. This is the first study to differentiate human hypothalamic neural stem cell-like cells from pluripotent stem cells.