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CD24 tracks divergent pluripotent states in mouse and human cells.

Shakiba N, White CA, Lipsitz YY, Yachie-Kinoshita A, Tonge PD, Hussein SM, Puri MC, Elbaz J, Morrissey-Scoot J, Li M, Munoz J, Benevento M, Rogers IM, Hanna JH, Heck AJ, Wollscheid B, Nagy A, Zandstra PW - Nat Commun (2015)

Bottom Line: Reprogramming is a dynamic process that can result in multiple pluripotent cell types emerging from divergent paths.Cell surface protein expression is a particularly desirable tool to categorize reprogramming and pluripotency as it enables robust quantification and enrichment of live cells.Thus, CD24 is a conserved marker for tracking divergent states in both reprogramming and standard pluripotent culture.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biomaterials and Biomedical Engineering (IBBME), University of Toronto, Toronto, Ontario, Canada M5S 3E1.

ABSTRACT
Reprogramming is a dynamic process that can result in multiple pluripotent cell types emerging from divergent paths. Cell surface protein expression is a particularly desirable tool to categorize reprogramming and pluripotency as it enables robust quantification and enrichment of live cells. Here we use cell surface proteomics to interrogate mouse cell reprogramming dynamics and discover CD24 as a marker that tracks the emergence of reprogramming-responsive cells, while enabling the analysis and enrichment of transgene-dependent (F-class) and -independent (traditional) induced pluripotent stem cells (iPSCs) at later stages. Furthermore, CD24 can be used to delineate epiblast stem cells (EpiSCs) from embryonic stem cells (ESCs) in mouse pluripotent culture. Importantly, regulated CD24 expression is conserved in human pluripotent stem cells (PSCs), tracking the conversion of human ESCs to more naive-like PSC states. Thus, CD24 is a conserved marker for tracking divergent states in both reprogramming and standard pluripotent culture.

No MeSH data available.


Related in: MedlinePlus

CD24 and CD40 exhibit differential expression on mouse ESCs and EpiSCs.(a) Flow cytometry analysis of CD24/SSEA1/CD40 expression in mouse ESCs and EpiSCs. Representative flow plots (n=3 technical replicates) of stained mouse ESC and control embryo-derived EpiSCs. (b) Gene expression of CD24high/CD40+ cells derived from R1 ESC culture compared with control embryo-derived EpiSCs. The gene panel is composed of pluripotency genes as well as some early differentiation markers. Values are normalized to sorted CD24low/SSEA1+ (ESC-like) cells sorted from R1 ESC culture and Gapdh. Data bars show mean±s.d. (n=3 technical replicates).
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f3: CD24 and CD40 exhibit differential expression on mouse ESCs and EpiSCs.(a) Flow cytometry analysis of CD24/SSEA1/CD40 expression in mouse ESCs and EpiSCs. Representative flow plots (n=3 technical replicates) of stained mouse ESC and control embryo-derived EpiSCs. (b) Gene expression of CD24high/CD40+ cells derived from R1 ESC culture compared with control embryo-derived EpiSCs. The gene panel is composed of pluripotency genes as well as some early differentiation markers. Values are normalized to sorted CD24low/SSEA1+ (ESC-like) cells sorted from R1 ESC culture and Gapdh. Data bars show mean±s.d. (n=3 technical replicates).

Mentions: We have shown that CD24 can be used to demarcate transgene-dependent F-class cells from ESC-like iPSCs. Given the role of CD24 in separating these iPSC states, we wondered whether CD24 would show conservation and utility in nonreprogrammed pluripotent populations. In order to investigate this possibility, we co-stained both mouse ESC and epiblast stem cell (EpiSC) populations for CD40 (a known EpiSC surface marker24) and CD24. Staining for CD24/CD40/SSEA1 levels in control mouse ESCs and EpiSCs confirmed the ability of CD24high/CD40+ staining to identify the EpiSC state, while ESCs were CD24low/CD40− (Fig. 3a). In order to better characterize the rare CD24high cells emerging in standard ESC populations, we cultured ESCs in serum and LIF conditions and sorted for the rare CD24high/CD40+ (EpiSC-like) as well as the prevalent CD24low/SSEA1+ (standard ESC) fraction of cells and conducted a survey of pluripotency and early differentiation gene expression25 (Fig. 3b). As expected, EpiSC-like CD24high/CD40+ cells exhibited lowered levels of naive pluripotency genes such as Stella, Rex1 and Nanog while also exhibiting higher levels of Foxa2, Eomes, Gata6, Sox17, Cer1 and Fgf5 (Fig. 3b). The observation of this rare EpiSC-like fraction of cells in serum-based ESC culture has been previously observed2627.


CD24 tracks divergent pluripotent states in mouse and human cells.

Shakiba N, White CA, Lipsitz YY, Yachie-Kinoshita A, Tonge PD, Hussein SM, Puri MC, Elbaz J, Morrissey-Scoot J, Li M, Munoz J, Benevento M, Rogers IM, Hanna JH, Heck AJ, Wollscheid B, Nagy A, Zandstra PW - Nat Commun (2015)

CD24 and CD40 exhibit differential expression on mouse ESCs and EpiSCs.(a) Flow cytometry analysis of CD24/SSEA1/CD40 expression in mouse ESCs and EpiSCs. Representative flow plots (n=3 technical replicates) of stained mouse ESC and control embryo-derived EpiSCs. (b) Gene expression of CD24high/CD40+ cells derived from R1 ESC culture compared with control embryo-derived EpiSCs. The gene panel is composed of pluripotency genes as well as some early differentiation markers. Values are normalized to sorted CD24low/SSEA1+ (ESC-like) cells sorted from R1 ESC culture and Gapdh. Data bars show mean±s.d. (n=3 technical replicates).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4490408&req=5

f3: CD24 and CD40 exhibit differential expression on mouse ESCs and EpiSCs.(a) Flow cytometry analysis of CD24/SSEA1/CD40 expression in mouse ESCs and EpiSCs. Representative flow plots (n=3 technical replicates) of stained mouse ESC and control embryo-derived EpiSCs. (b) Gene expression of CD24high/CD40+ cells derived from R1 ESC culture compared with control embryo-derived EpiSCs. The gene panel is composed of pluripotency genes as well as some early differentiation markers. Values are normalized to sorted CD24low/SSEA1+ (ESC-like) cells sorted from R1 ESC culture and Gapdh. Data bars show mean±s.d. (n=3 technical replicates).
Mentions: We have shown that CD24 can be used to demarcate transgene-dependent F-class cells from ESC-like iPSCs. Given the role of CD24 in separating these iPSC states, we wondered whether CD24 would show conservation and utility in nonreprogrammed pluripotent populations. In order to investigate this possibility, we co-stained both mouse ESC and epiblast stem cell (EpiSC) populations for CD40 (a known EpiSC surface marker24) and CD24. Staining for CD24/CD40/SSEA1 levels in control mouse ESCs and EpiSCs confirmed the ability of CD24high/CD40+ staining to identify the EpiSC state, while ESCs were CD24low/CD40− (Fig. 3a). In order to better characterize the rare CD24high cells emerging in standard ESC populations, we cultured ESCs in serum and LIF conditions and sorted for the rare CD24high/CD40+ (EpiSC-like) as well as the prevalent CD24low/SSEA1+ (standard ESC) fraction of cells and conducted a survey of pluripotency and early differentiation gene expression25 (Fig. 3b). As expected, EpiSC-like CD24high/CD40+ cells exhibited lowered levels of naive pluripotency genes such as Stella, Rex1 and Nanog while also exhibiting higher levels of Foxa2, Eomes, Gata6, Sox17, Cer1 and Fgf5 (Fig. 3b). The observation of this rare EpiSC-like fraction of cells in serum-based ESC culture has been previously observed2627.

Bottom Line: Reprogramming is a dynamic process that can result in multiple pluripotent cell types emerging from divergent paths.Cell surface protein expression is a particularly desirable tool to categorize reprogramming and pluripotency as it enables robust quantification and enrichment of live cells.Thus, CD24 is a conserved marker for tracking divergent states in both reprogramming and standard pluripotent culture.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biomaterials and Biomedical Engineering (IBBME), University of Toronto, Toronto, Ontario, Canada M5S 3E1.

ABSTRACT
Reprogramming is a dynamic process that can result in multiple pluripotent cell types emerging from divergent paths. Cell surface protein expression is a particularly desirable tool to categorize reprogramming and pluripotency as it enables robust quantification and enrichment of live cells. Here we use cell surface proteomics to interrogate mouse cell reprogramming dynamics and discover CD24 as a marker that tracks the emergence of reprogramming-responsive cells, while enabling the analysis and enrichment of transgene-dependent (F-class) and -independent (traditional) induced pluripotent stem cells (iPSCs) at later stages. Furthermore, CD24 can be used to delineate epiblast stem cells (EpiSCs) from embryonic stem cells (ESCs) in mouse pluripotent culture. Importantly, regulated CD24 expression is conserved in human pluripotent stem cells (PSCs), tracking the conversion of human ESCs to more naive-like PSC states. Thus, CD24 is a conserved marker for tracking divergent states in both reprogramming and standard pluripotent culture.

No MeSH data available.


Related in: MedlinePlus