齋藤 加奈子

BACKGROUND: Genome-wide association studies have revealed the involvement of 5'-nucleotidase domain-containing protein 2 (NT5DC2) in neuropsychiatric disorders such as schizophrenia and bipolar disorder; however, its function remains unclear. We recently found that NT5DC2 downregulation in PC12D cells increases catecholamine synthesis by promoting tyrosine hydroxylase (TH) activity. In addition, affinity purification-mass spectrometry suggested a potential interaction between NT5DC2 and monoamine oxidase A (MAO A). In this study, we examined the impact of NT5DC2 on MAO A activity in PC12D cells and the related effects on catecholamine metabolism. METHODS AND RESULTS: We analyzed changes in catecholamine metabolism in siRNA-mediated NT5DC2-downregulated PC12D cells by measuring the catecholamines and major acid metabolites produced by the addition of exogenous dopamine (DA) to PC12D cells, with DOPA synthesis suppressed by the addition of a TH inhibitor. Western blot analysis revealed that NT5DC2 primarily binds to the non-phosphorylated form of MAO A. NT5DC2 downregulation reduced MAO A activity, leading to decreased dopamine metabolism and increased noradrenaline synthesis. CONCLUSION: Our findings suggest that NT5DC2 regulates MAO A activity to control catecholamine synthesis. This study provides valuable insights into disorders associated with catecholamine dysregulation, such as Parkinson's disease and neuropsychiatric disorders.

The hypothalamus is comprised of heterogenous cell populations and includes highly complex neural circuits that regulate the autonomic nerve system. Its dysfunction therefore results in severe endocrine disorders. Although recent experiments have been conducted for in vitro organogenesis of hypothalamic neurons from embryonic stem (ES) or induced pluripotent stem (iPS) cells, whether these stem cell-derived hypothalamic neurons can be useful for regenerative medicine remains unclear. We therefore performed orthotopic transplantation of mouse ES cell (mESC)-derived hypothalamic neurons into adult mouse brains. We generated electrophysiologically functional hypothalamic neurons from mESCs and transplanted them into the supraoptic nucleus of mice. Grafts extended their axons along hypothalamic nerve bundles in host brain, and some of them even projected into the posterior pituitary (PPit), which consists of distal axons of the magnocellular neurons located in hypothalamic supraoptic and paraventricular nuclei. The axonal projections to the PPit were not observed when the mESC-derived hypothalamic neurons were ectopically transplanted into the substantia nigra reticular part. These findings suggest that our stem cell-based orthotopic transplantation approach might contribute to the establishment of regenerative medicine for hypothalamic and pituitary disorders.

Human stem cell-derived organoid culture enables the in vitro analysis of the cellular function in three-dimensional aggregates mimicking native organs, and also provides a valuable source of specific cell types in the human body. We previously established organoid models of the hypothalamic-pituitary (HP) complex using human pluripotent stem cells. Although the models are suitable for investigating developmental and functional HP interactions, we consider that isolated pituitary cells are also useful for basic and translational research on the pituitary gland, such as stem cell biology and regenerative medicine. To develop a method for the purification of pituitary cells in HP organoids, we performed surface marker profiling of organoid cells derived from human induced pluripotent stem cells (iPSCs). Screening of 332 human cell surface markers and a subsequent immunohistochemical analysis identified epithelial cell adhesion molecule (EpCAM) as a surface marker of anterior pituitary cells, as well as their ectodermal precursors. EpCAM was not expressed on hypothalamic lineages; thus, anterior pituitary cells were successfully enriched by magnetic separation of EpCAM+ cells from iPSC-derived HP organoids. The enriched pituitary population contained functional corticotrophs and their progenitors; the former responded normally to a corticotropin-releasing hormone stimulus. Our findings would extend the applicability of organoid culture as a novel source of human anterior pituitary cells, including stem/progenitor cells and their endocrine descendants.