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Transcriptional analysis of primary adult human skin cells that express stage-specific embryonic antigen 3 (SSEA-3)

ABSTRACT: Primary human skin cells were gated based on expression of stage-specific embryonic antigen 3 (SSEA3) with the high SSEA3 expressing cells (top 10%, categorized as SSEA3-positive or SSEA3-high) and negative SSEA3 expressing cells (bottom 10%, categorized as SSEA3-negative) purified into two different subpopulations. Sorted cells were allowed to adhere overnight under DMEM/F12 + 10% FBS culture conditions before total RNA was collected and global transcriptional analysis was performed.

ORGANISM(S): Homo sapiens

PROVIDER: GSE33066 | GEO | 2012/03/01

SECONDARY ACCESSION(S): PRJNA149475

REPOSITORIES: GEO

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Project description:PurposeThe presence of multipotent human limbal stromal cells resembling mesenchymal stromal cells (MSC) provides new insights to the characteristic of these cells and its therapeutic potential. However, little is known about the expression of stage-specific embryonic antigen 4 (SSEA-4) and the embryonic stem cell (ESC)-like properties of these cells. We studied the expression of SSEA-4 surface protein and the various ESC and MSC markers in the ex vivo cultured limbal stromal cells. The phenotypes and multipotent differentiation potential of these cells were also evaluated.MethodsLimbal stromal cells were derived from corneoscleral rims. The SSEA-4(+) and SSEA-4(-) limbal stromal cells were sorted by fluorescence-activated cells sorting (FACS). Isolated cells were expanded and reanalyzed for their expression of SSEA-4. Expression of MSC and ESC markers on these cells were also analyzed by FACS. In addition, expression of limbal epithelial and corneal stromal proteins such as ATP-binding cassette sub-family G member 2 (ABCG2), tumour protein p63 (p63), paired box 6 (Pax6), cytokeratin 3 (AE5), cytokeratin 10, and keratocan sulfate were evaluated either by immunofluorecence staining or reverse transcription polymerase chain reaction. Appropriate induction medium was used to differentiate these cells into adipocytes, osteocytes, and chondrocytes.ResultsExpanded limbal stromal cells expressed the majority of mesenchymal markers. These cells were negative for ABCG2, p63, Pax6, AE-5, and keratocan sulfate. After passaged, a subpopulation of these cells showed low expression of SSEA-4 but were negative for other important ESC surface markers such as Tra-1-60, Tra-1-81, and transcription factors like octamer-binding transcription factor 4 (Oct4), SRY(sex determining region Y)-box 2 (Sox2), and Nanog. Early passaged cells when induced were able to differentiate into adipocytes, osteocytes and chondrocytes.ConclusionsThe expanded limbal stromal cells showed features of multipotent MSC. Our study confirmed the expression of SSEA-4 by a subpopulation of cultured limbal stromal cells. However, despite the expression of SSEA-4, these cells did not express any other markers of ESC. Therefore, we conclude that the cells did not show properties of ESC.

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Transcriptional analysis of primary adult human skin cells that express stage-specific embryonic antigen 3 (SSEA-3)

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