Generation of kisspeptin-responsive GnRH neurons from human pluripotent stem cells. (15th May 2017)

• Record Type: Journal Article
• Authors: Poliandri, Ariel; Miller, Duncan; Howard, Sasha; Nobles, Muriel; Ruiz-Babot, Gerard; Harmer, Stephen; Tinker, Andrew; McKay, Tristan; Guasti, Leonardo; Dunkel, Leo
• Is Part Of: Molecular and cellular endocrinology. Volume 447(2017)
• Page Start: 12
• Page End: 22
• Publication Date: 2017-05-15
• Subjects: GnRH neurons -- Pluripotent stem cells -- Embryonic stem cells -- Kisspeptin; Endocrinology -- Periodicals; Molecular biology -- Periodicals; Cytology -- Periodicals; Endocrinology -- Periodicals; Hormones -- Periodicals; Endocrinologie -- Périodiques; Cytology; Endocrinology; Molecular biology; Periodicals; 573.4
• DOI: 10.1016/j.mce.2017.02.030
• Languages: English
• ISSNs: 0303-7207

Abstract

Abstract: GnRH neurons are fundamental for reproduction in all vertebrates, integrating all reproductive inputs. The inaccessibility of human GnRH-neurons has been a major impediment to studying the central control of reproduction and its disorders. Here, we report the efficient generation of kisspeptin responsive GnRH-secreting neurons by directed differentiation of human Embryonic Stem Cells and induced-Pluripotent Stem Cells derived from a Kallman Syndrome patient and a healthy family member. The protocol involves the generation of intermediate Neural Progenitor Cells (NPCs) through long-term Bone morphogenetic protein 4 inhibition, followed by terminal specification of these NPCs in media containing Fibroblast Growth Factor 8 and a NOTCH inhibitor. The resulting GnRH-expressing and -secreting neurons display a neuroendocrine gene expression pattern and present spontaneous calcium transients that can be stimulated by kisspeptin. These in vitro generated GnRH expressing cells provide a new resource for studying the molecular mechanisms underlying the development and function of GnRH neurons. Graphical abstract: Highlights: A protocol that consistently and efficiently generates GnRH-expressing neurons from human Pluripotent Stem Cells. An intermediate step involving Neural Progenitor Cells production allows for expansion before terminal differentiation. Terminally differentiated GnRH-expressing neurons present spontaneous calcium transients. Spontaneous calcium transients are increased by kisspeptin, both in frequency and amplitude.