Pituitary Progenitor Cells Tracked Down by Side Population Dissection

Abstract

The pituitary gland represents the endocrine core, governing the body's hormonal landscape by adapting its cellular composition to changing demands. It is assumed that stem/ progenitor cells are involved in this remodeling. Recently, we uncovered a candidate stem/progenitor cell population in the anterior pituitary. Here, we scrutinized this ''side population'' (SP) and show that, unexpectedly, not the subset expressing high levels of ''stem cell antigen-1'' (Sca1 high ) but the remainder non-Sca1 high fraction clusters the pituitary progenitor cells. Transcriptomal interrogation revealed in the non-Sca1 high SP upregulated expression of the pituitary stem/progenitor cell markers Sox2 and Sox9, and of multiple factors critically involved in pituitary embryogenesis. The non-Sca1 high SP encloses the cells that generate spheres and display multipotent hormone differentiation capacity. In culture conditions selecting for the nonSca1 high subset within the SP, stem cell growth factors that induce SP expansion, affect transcription of embryonic factors, suggesting impact on a developmental program that unfolds within this SP compartment. Non-Sca1 high SP cells, revealed by Sox2 expression, are observed in the postulated periluminal stem/progenitor cell niche, but also in small groups scattered over the gland, thereby advocating the existence of multiple niches. In early postnatal mice undergoing a pituitary growth wave, Sox2 1 cells are more abundant than in adults, concordant with a larger SP and higher non-Sca1 high proportion. Together, we tracked down pituitary progenitor cells by SP phenotype, and thus provide a straightforward method to isolate and scrutinize these cells from the plastic pituitary ex vivo, as well as a culture system for in-depth exploration of their regulatory network.

Pituitary tumors contain a side population with tumor stem cell-associated characteristics

“…Cell debris was removed by centrifugation through a 3% BSA layer. The resulting cell pellet was resuspended in pituitary-optimized, serum-free chemically defined medium (SFDM, Life Technologies; supplemented with 0.5% BSA; Chen et al 2005Chen et al , 2009 for further analyses.…”

“…Mice were kept or bred in the KU Leuven Animal Facility under The pituitary was isolated from killed mice as previously described (Chen et al 2005(Chen et al , 2009). The anterior lobes (anterior pituitary (AP)) of the Drd2 K/K mice that grow prolactinomas (from 6 to 14 months of age; Cristina et al 2006) and those of the respective control mice (Drd2 C/K and Drd2 C/C ) were dispersed into single cells using collagenase type IV as explained in the previous sections for human pituitary adenomas.…”

“…High efflux capacity, as mediated by ATP-binding cassette (ABC) multidrug transporters, is considered to be one functional characteristic of CSCs; this property renders the cells more resistant to chemotherapeutic drugs. In the present study, we searched for cells that possess such efflux capacity using the 'side population' (SP) technique (Chen et al 2005(Chen et al , 2009. Cells that extrude Hoechst dye are observed as a side branch of Hoechst low cells in dual-wavelength fluorescence-activated cell sorting (FACS) analysis.…”

Pituitary Stem Cells During Normal Physiology and Disease

“…Several groups have reported the presence of putative CSCs in human pituitary adenomas and from mouse pituitary tumour models Xu et al 2009;Donangelo et al 2014;Lloyd et al 2013;Mertens et al 2015;Orciani et al 2015;van Rijn et al 2013;Yunoue et al 2011;Hosoyama et al 2010). It remains unknown if CSCs arise from PSCs, although their properties can be similar; for example, both are capable of in vitro self-renewal and differentiation and PSCs are likely to be chemoresistant like CSCs, since PSCs are found within the 'side population' generated by dye efflux, utilising the same transporter properties as chemoresistance (Chen et al 2009). …”

“…The second population was characterized by marker expression similar to stem cells of other tissues (Sca1, Nanog and Oct4) and could form floating spheres (Chen et al 2005). In later studies, the same research group showed that there was enrichment in this population for the expression of SOX2, SOX9, CD44 and CD133 (Chen et al 2009). …”