SCA-1 Expression Level Identifies Quiescent Hematopoietic Stem and Progenitor Cells

Abstract

SummaryBlood cell generation depends on continuous cellular output by the sequential hierarchy of hematopoietic stem cell (HSC) and progenitor populations that all contain quiescent and actively cycling cells. Hematopoietic stem and progenitor cells (HSPCs) express the surface molecule Stem cell antigen 1 (SCA-1/LY6A). Using histone 2B-red fluorescent fusion protein label retention and cell-cycle reporter mice, we demonstrate that high SCA-1 expression (SCA-1hi) identifies not only quiescent HSCs but quiescent cells on all hierarchical levels within the lineage−SCA-1+KIT+ (LSK) population. Each transplanted SCA-1hi HSPC population also displayed self-renewal potential superior to that of the respective SCA-1lo population. SCA-1 expression is inducible by type I interferon (IFN). We show, however, that quiescence and high self-renewal capacity of cells with brighter SCA-1 expression at steady state were independent of type I IFN signaling. We conclude that SCA-1 expression levels can be used to prospectively isolate functionally heterogeneous HSPC subpopulations.

Jak1 Integrates Cytokine Sensing to Regulate Hematopoietic Stem Cell Function and Stress Hematopoiesis

“…Previous studies have shown that Sca1 expression increases upon IFN-α exposure of LSKs (Essers et al, 2009; Morcos et al, 2017). Consistent with a defective IFN response upon loss of Jak1, Jak1-null hematopoietic cells failed to upregulate Sca1 surface expression in response to IFN-α secretion caused by a single injection of poly(I:C) in vivo (Figure 5B).…”

Continuous mitotic activity of primitive hematopoietic stem cells in adult mice

“…PB neutrophils (PMN, CD11b + Gr-1 hi ), B- (CD19 + ) and T-lymphocytes (CD3 + ) of secondary recipients ( n = 4/group) were analyzed for donor origin at indicated time points (right panel, mean and SD are shown, significance was calculated by repeated measures two-way ANOVA with Bonferroni post-test). (I) Maximum H2B-FP fluorescence intensities (as judged by the 99th percentile) of MPPs (LSK CD48 − /lo CD150 − ) isolated from chased R26 rtTA / Col1A1 H2B-RFP (shown in red; Säwén et al, 2016, dashed line, open circles; Morcos et al, 2017, continuous line, filled circles) or R26 rtTA/rtTA / Col1A1 H2B-GFP/H2B-GFP (green; same animals as in Fig. 2 A “GFP Morcos”) individual mice are depicted.…”

“…Mean frequencies and SD of LSK CD48 −/lo CD150 + cells are shown, except for data from Oguro et al (2013), in which a weighted mean of the HSC-1 and HSC-2 subpopulations was calculated ([1× HSC-1 + 3× HSC-2]/4). (B) Maximum H2B-FP fluorescence intensities (as judged by the 99th percentile) of HSCs (LSK CD48 −/lo CD150 + ) isolated from chased R26 rtTA / Col1A1 H2B-RFP (shown in red; Säwén et al, 2016, dashed line, open circles; Morcos et al, 2017, continuous line, filled circles) or R26 rtTA/rtTA / Col1A1 H2B-GFP/H2B-GFP (green; same animals as in A “GFP Morcos”) individual mice are depicted. The data were fitted by our mathematical model (see G), and loss of maximum fluorescence was attributed to H2B-FP degradation.…”

“… Mitotic activity of HSC populations inferred from H2B-FP label dilution data. (A) Representative examples for mixtures of Gaussian distributions fitted to H2B-RFP (left panel, R26 rtTA / Col1A1 H2B-RFP animals; n = 1–9/time point, pooled from three independent experiments, data from Morcos et al, 2017) or H2B-GFP (right panel, R26 rtTA/rtTA / Col1A1 H2B-GFP/H2B-GFP mice shown in Fig. 2, A and B) label dilution data of HSCs (continuous curves: recorded data; dotted gray curves: fitted mixture distributions).…”

“…The multipotent and self-renewing hematopoietic stem cells (HSCs) reside at the top of the hematopoietic hierarchy and can give rise to all blood lineages throughout the life of an individual (Eaves, 2015). Numerous studies have unambiguously demonstrated heterogeneous cell cycle activity within the HSC population (Foudi et al, 2009; Glauche et al, 2009; Morcos et al, 2017; Passegué et al, 2005; Qiu et al, 2014; Säwén et al, 2016; Takizawa et al, 2011; Wilson et al, 2008). A concept of deeply quiescent (also termed “dormant”) and actively cycling HSCs has been firmly established from experiments investigating mitotic history by means of labeled nucleotide analogues (e.g., BrdU; Kiel et al, 2007; Wilson et al, 2008) or histone 2B (H2B) fused to either enhanced green fluorescent protein (H2B-GFP) or mCherry (hereafter referred to as H2B-RFP; Foudi et al, 2009; Qiu et al, 2014; Säwén et al, 2016; Wilson et al, 2008).…”