It has furthermore been speculated whether Sca plays a role

It has furthermore been speculated whether Sca-1 plays a role in stem cell self-renewal. Thus, some experiments with Sca-1 −/− knock-out mice investigating hematopoietic and mesenchymal progenitor mg115 have pointed to a role for Sca-1 in self-renewal (Bonyadi et al., 2003; Ito et al., 2003). However, other studies on hematopoietic stem cells and muscle stem cells indicate that Sca-1 does not seem to affect self-renewal (Bradfute et al., 2005; Kafadar et al., 2009). A complex phenotype has been observed in Sca-1 −/− knock-out mice including defects in the regulation of lineage commitment in hemapoietic stem cells (Bradfute et al., 2005), osteoporosis, reduced muscle size in aging KO mice (Bonyadi et al., 2003) and delayed muscle repair after injury (Epting et al., 2008b). In mammary tumor cells, Sca-1 has been shown to regulate cell migration, cell adhesion to several extracellular matrix substrates and tumor development in early lesions (Batts et al., 2011) and a study using a mammary adenocarcinoma cell line could demonstrate that Sca-1 suppresses GDF10-dependent TGF-β signaling by disrupting the heterodimerization of the TGF-β receptors (Upadhyay et al., 2011). Additionally, TGF-β acts as a negative regulator of Sca-1 expression in both myoblasts and splenic T cells (Long et al., 2011).
Materials and methods
Results
Discussion Sca-1, encoded by the gene Ly6a, has been known for >30years and is being used extensively for isolating hematopoietic stem cells in the mouse. The role of Sca-1 is still shrouded in uncertainty although a great number of publications have tried to address this issue (for review see (Holmes and Stanford, 2007)). We have previously shown that Sca-1 is useful when studying mouse hair follicle stem cells (Gunnarsson et al., 2016; Jensen et al., 2008) as it allows you to separate cells below the infundibulum of the hair follicles from cells of the infundibulum and interfollicular epidermis which express high levels of Sca-1. Colony forming efficiency is often used to access the growth potential of epidermal sub populations and can be considered an indirect measure of stemness (Barrandon and Green, 1987; Jensen et al., 2008; Jones and Watt, 1993). One of the striking features of testing FACS sorted keratinocytes for colony forming efficiency is that Sca-1 positive cells normally perform inefficiently. However, bulge region keratinocytes from Rosa:Cre mice that express Sca-1 showed a slightly increased clonogenic capacity in colony forming assays. This was somewhat surprising and may suggest a permissive role of Sca-1 action in terms of growth. Thus, bulge stem cells expressing Sca-1 may have a higher growth potential when cultured as compared to normal bulge stem cells and Sca-1 is normally expressed at high levels in proliferating cells in the skin. This may reflect that Sca-1 plays a role in controlling proliferation and that Sca-1 does not affect stemness directly. In other tissues like the hematopoietic system, muscle, mammary tissue, and liver, stem cells express Sca-1 in contrast to the hair follicle (Bradfute et al., 2005; Epting et al., 2008a; Epting et al., 2004; Ito et al., 2003; Mitchell et al., 2005; Welm et al., 2002; Wright et al., 2008). Therefore, expression of Sca-1 per se is not incompatible with a stem cell phenotype. We also know that colony forming efficiency does not always reflect potency as Sca-1 negative and CD34 negative cells that express high levels of alpha6 integrin do well in the colony forming assay but do not form hair in a hair reconstitution assay (Jensen et al., 2008). It could be speculated that while there was no impact on hair follicle homeostasis, there may be impact of Sca-1 induction on regeneration following wounding or grafting since bulge stem cells expressing Sca-1 were more clonogenic. We also tested the Rosa:Cre mice in label retaining experiments. Since label retaining cells (LRCs) are traditionally thought to reflect the number of stem cells, we measured the number of LRCs and found an increased number of label retaining cells (LRCs) in Rosa:Cre mice as compared to control littermates.