Intrinsic factors controlling stem cell proliferation and differentiation in the central nervous system.

Populations of neural stem cells (NSCs) residing in different locations of the central nervous system (CNS) are predicted to have similar gene expression profiles, which identify them as stem cells. The best in vitro model to study NSCs is the neurosphere - a spherical colony generated from a single NSC. Currently, there is a lack of markers to characterise NSCs and distinguish them from lineage-committed progenitor cells. One promising candidate is Sox2, which is a member of the Sox family of transcription factors and is expressed within the CNS. Sox2geo/+ mice carry a p-galactosidase/ neomycin fusion gene in the Sox2 locus. Homozygous Sox2eo mutants exhibit peri-implantation lethality since Sox2 is essential for the maintenance of the pluripotent epiblast cells in the embryo. I have established that the neurosphere-generating cellular component, throughout the developing CNS, resides within the Sox2 expressing population. Through neomycin selection, neurosphere cultures derived from Sox2Pgeol+ mice have been enriched for NSCs and have been used to characterise NSCs. Differences in the gene expression profiles of cells expressing and not expressing Sox2 have been carried out through microarray experiments. To investigate the possible function of Sox2 in NSCs I have depleted SOX2 by RNA interference, followed by microarray analysis, to identify potential targets of Sox2 that may further affect the proliferation or differentiation of stem cells. The influence of the niche surrounding stem cells within the neurosphere has been addressed through a series of culture experiments comparing NSC-enriched to non-enriched neurospheres. In order to elucidate the genes responsible for the identity of stem cells, enriched NSC cultures have been compared to their tissue of origin through microarray analysis. These studies have revealed genes expressed at significantly elevated levels within the stem cell cultures compared to the tissue samples - a predominantly differentiating population. Pair-wise comparisons between neurospheres generated from the spinal cord (11.5 dpc, 14.5 dpc) and dorsal telencephalon (14.5 dpc) were used to refine candidate genes and provide an insight into the spatial and temporal properties of NSCs. Several candidate NSC markers, including transcription factors and extracellular matrix molecules common to all populations, have been identified

STAT3-Ser/Hes3 signaling: a new molecular component of the neuroendocrine system?

The endocrine system involves communication among different tissues in distinct organs, including the pancreas and components of the Hypothalamic- Pituitary-Adrenal Axis. The molecular mechanisms underlying these complex interactions are a subject of intense study as they may hold clues for the progression and treatment of a variety of metabolic and degenerative diseases. A plethora of signaling pathways, activated by hormones and other endocrine factors have been implicated in this communication. Recent advances in the stem cell field introduce a new level of complexity: adult progenitor cells appear to utilize distinct signaling pathways than the more mature cells in the tissue they co-reside. It is therefore important to elucidate the signal transduction requirements of adult progenitor cells in addition to those of mature cells. Recent evidence suggests that a common non-canonical signaling pathway regulates adult progenitors in several different tissues, rendering it as a potentially valuable starting point to explore their biology. The STAT3- Ser/Hes3 Signaling Axis was first identified as a major regulator of neural stem cells and, subsequently, cancer stem cells. In the endocrine/neuroendocrine system, this pathway operates on several levels, regulating other types of plastic cells: (a) it regulates pancreatic islet cell function and insulin release; (b) insulin in turn activates the pathway in broadly distributed neural progenitors and possibly also hypothalamic tanycytes, cells with important roles in the control of the adrenal gland; (c) adrenal progenitors themselves operate this pathway. The STAT3-Ser/Hes3 Signaling Axis therefore deserves additional research in the context of endocrinology

The role of pituitary melanocortin receptor 3 (MC3) in regulation of hormone output

Introduction/Aim: Of the five melanocortin receptors (MCs: MC1-5) identified to date, the focus of neuroendocrine studies has principally been on MC3 and MC4 due to their role in regulating body weight: a role presumed to be mediated by hypothalamic actions in response to melanocyte stimulating hormone (MSH). Recently, MC3 has been shown to have roles in regulating both statural growth and the timing of puberty: possibly mediated by MC3 expression identified in arcuate growth hormone releasing hormone (GHRH) and kisspeptin neurones, respectively (1). It has long been known that MC3 is expressed in the anterior pituitary and may mediate paracrine interactions within the gland (2). It was the aim of this series of studies to determine the potential impact of mouse pituitary MC3 on physiology: specifically the role of MC3 in pituitary gland function, the pattern of expression of the receptor and potential modifications of function by expression of interacting proteins. Methods/Results: We have determined the impact of loss of MC3 expression on anterior pituitary function using MC3 knockout mice: in adult male mice loss of MC3 leads to a significant reduction in the pituitary content of growth hormone (GH), gonadotrophins and prolactin, with no effect on adrenocorticotrophic hormone (ACTH) or thyroid stimulating hormone; in adult females the reduction is restricted to GH, however, a trend of decrease in gonadotrophins was also found. RNAScope in situ hybridisation showed that MC3 is co-expressed in a large proportion of somatotrophs and gonadotrophs and, consistent with this, imaging showed that ACTH stimulates a rise in intracellular calcium in these cell types. We have also studied the interaction of melanocortin receptor accessory proteins (MRAPs) and found that MRAP2 modifies MC3 response to ACTH but not MSH and results in bias G-protein coupled signalling. RNAScope for MRAP2 revealed that it is co-expressed in a significant proportion of MC3 positive cells. Conclusions: The effects of global knockout of MC3 on pituitary hormone contents suggest that aspects of the body weight, growth and pubertal timing roles of the receptor may also be mediated by its pituitary expression. Furthermore, the interactions of MC3 and MRAPs with the growth hormone secretagogue receptor (3) would be consistent with a pituitary role in growth hormone regulation. (1) Lam et al. (2021) Nature 599:436-441. (2) Roudbaraki et al. (1999) Endocrinol. 140:4874-4885. (3) Rediger et al. (2011) J Biol Chem. 286:39623-31

Stress-inducible-stem cells: a new view on endocrine, metabolic and mental disease?

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