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Epigenetic and phenotypic profile of fibroblasts derived from induced pluripotent stem cells.

Hewitt KJ, Shamis Y, Hayman RB, Margvelashvili M, Dong S, Carlson MW, Garlick JA - PLoS ONE (2011)

Bottom Line: However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES) cells remain unclear.We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK) and their hES-derived counterparts (EDK) showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts.Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

View Article: PubMed Central - PubMed

Affiliation: Program in Cell, Molecular and Developmental Biology, Tufts University School of Medicine, Boston, Massachusetts, United States of America.

ABSTRACT
Human induced pluripotent stem (hiPS) cells offer a novel source of patient-specific cells for regenerative medicine. However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES) cells remain unclear. We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK) and their hES-derived counterparts (EDK) showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts. Array-based methylation analysis was performed for EDK, iPDK and their parental hES and iPS cell lines, and hierarchical clustering revealed that EDK and iPDK had closely-related methylation profiles. DNA methylation analysis of promoter regions associated with extracellular matrix (ECM)-production (COL1A1) by iPS- and hESC-derived fibroblasts and fibroblast lineage commitment (PDGFRβ), revealed promoter demethylation linked to their expression, and patterns of transcription and methylation of genes related to the functional properties of mature stromal cells were seen in both hiPS- and hES-derived fibroblasts. iPDK cells also showed functional properties analogous to those of hES-derived and mature fibroblasts, as seen by their capacity to direct the morphogenesis of engineered human skin equivalents. Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

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Global profiling of DNA methylation indicates that EDK and iPDK cells share similar gene methylation levels that are distinct from pluripotent cell types.Cell lines were analyzed for methylation marks using the Illumina Methylation27 chip. Hierarchical clustering analysis of degrees of methylation using a Manhattan correlation showed that hES and iPS cell lines clustered in a distinct set from differentiated iPDK and EDK cells, as well as mature fibroblasts and keratinocytes (A). The methylation status of each cell type, as assessed by the percentage of CpG sites that fell within a range of β-values, showed that CpG islands are over 60% unmethylated, and pluripotent cells appear to have greater number of highly-methylated CpG sites (B). A scatter plot comparing the mean level of CpG methylation in 3 EDK and 3 iPDK cell lines showed a high degree of correlation between the two cell types (C). R2 = 0.8715.
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pone-0017128-g002: Global profiling of DNA methylation indicates that EDK and iPDK cells share similar gene methylation levels that are distinct from pluripotent cell types.Cell lines were analyzed for methylation marks using the Illumina Methylation27 chip. Hierarchical clustering analysis of degrees of methylation using a Manhattan correlation showed that hES and iPS cell lines clustered in a distinct set from differentiated iPDK and EDK cells, as well as mature fibroblasts and keratinocytes (A). The methylation status of each cell type, as assessed by the percentage of CpG sites that fell within a range of β-values, showed that CpG islands are over 60% unmethylated, and pluripotent cells appear to have greater number of highly-methylated CpG sites (B). A scatter plot comparing the mean level of CpG methylation in 3 EDK and 3 iPDK cell lines showed a high degree of correlation between the two cell types (C). R2 = 0.8715.

Mentions: Hierarchical clustering analysis of methylation data revealed that all of the EDK and iPDK cell types had closely-related methylation profiles, while all pluripotent cell types clustered together and were distinct from EDK and iPDK (Figure 2A). Normal human fibroblasts (HFF) clustered closely with the EDK and iPDK samples, while the foreskin-derived keratinocyte (NHK) methylation profile was more similar to pluripotent cells studied, unrelated to fibroblasts. Additionally, the degree of total methylation for all the sites represented in this assay was compared between cell samples by counting the number of CpG sites falling within a range of β values varying in degree from highly methylated (0.8–1.0) to unmethylated (0–0.2) (Figure 2B). Methylation was found to be reduced in differentiated EDK and iPDK cells when compared to the pluripotent cell types from which they were derived. A scatter plot illustrating the comparative methylation state of both EDK and iPDK showed a high degree of correlation (R2 = 0.87) indicating that the methylation profile was similar between these two cell types (Figure 2C).


Epigenetic and phenotypic profile of fibroblasts derived from induced pluripotent stem cells.

Hewitt KJ, Shamis Y, Hayman RB, Margvelashvili M, Dong S, Carlson MW, Garlick JA - PLoS ONE (2011)

Global profiling of DNA methylation indicates that EDK and iPDK cells share similar gene methylation levels that are distinct from pluripotent cell types.Cell lines were analyzed for methylation marks using the Illumina Methylation27 chip. Hierarchical clustering analysis of degrees of methylation using a Manhattan correlation showed that hES and iPS cell lines clustered in a distinct set from differentiated iPDK and EDK cells, as well as mature fibroblasts and keratinocytes (A). The methylation status of each cell type, as assessed by the percentage of CpG sites that fell within a range of β-values, showed that CpG islands are over 60% unmethylated, and pluripotent cells appear to have greater number of highly-methylated CpG sites (B). A scatter plot comparing the mean level of CpG methylation in 3 EDK and 3 iPDK cell lines showed a high degree of correlation between the two cell types (C). R2 = 0.8715.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3046119&req=5

pone-0017128-g002: Global profiling of DNA methylation indicates that EDK and iPDK cells share similar gene methylation levels that are distinct from pluripotent cell types.Cell lines were analyzed for methylation marks using the Illumina Methylation27 chip. Hierarchical clustering analysis of degrees of methylation using a Manhattan correlation showed that hES and iPS cell lines clustered in a distinct set from differentiated iPDK and EDK cells, as well as mature fibroblasts and keratinocytes (A). The methylation status of each cell type, as assessed by the percentage of CpG sites that fell within a range of β-values, showed that CpG islands are over 60% unmethylated, and pluripotent cells appear to have greater number of highly-methylated CpG sites (B). A scatter plot comparing the mean level of CpG methylation in 3 EDK and 3 iPDK cell lines showed a high degree of correlation between the two cell types (C). R2 = 0.8715.
Mentions: Hierarchical clustering analysis of methylation data revealed that all of the EDK and iPDK cell types had closely-related methylation profiles, while all pluripotent cell types clustered together and were distinct from EDK and iPDK (Figure 2A). Normal human fibroblasts (HFF) clustered closely with the EDK and iPDK samples, while the foreskin-derived keratinocyte (NHK) methylation profile was more similar to pluripotent cells studied, unrelated to fibroblasts. Additionally, the degree of total methylation for all the sites represented in this assay was compared between cell samples by counting the number of CpG sites falling within a range of β values varying in degree from highly methylated (0.8–1.0) to unmethylated (0–0.2) (Figure 2B). Methylation was found to be reduced in differentiated EDK and iPDK cells when compared to the pluripotent cell types from which they were derived. A scatter plot illustrating the comparative methylation state of both EDK and iPDK showed a high degree of correlation (R2 = 0.87) indicating that the methylation profile was similar between these two cell types (Figure 2C).

Bottom Line: However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES) cells remain unclear.We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK) and their hES-derived counterparts (EDK) showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts.Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

View Article: PubMed Central - PubMed

Affiliation: Program in Cell, Molecular and Developmental Biology, Tufts University School of Medicine, Boston, Massachusetts, United States of America.

ABSTRACT
Human induced pluripotent stem (hiPS) cells offer a novel source of patient-specific cells for regenerative medicine. However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES) cells remain unclear. We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK) and their hES-derived counterparts (EDK) showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts. Array-based methylation analysis was performed for EDK, iPDK and their parental hES and iPS cell lines, and hierarchical clustering revealed that EDK and iPDK had closely-related methylation profiles. DNA methylation analysis of promoter regions associated with extracellular matrix (ECM)-production (COL1A1) by iPS- and hESC-derived fibroblasts and fibroblast lineage commitment (PDGFRβ), revealed promoter demethylation linked to their expression, and patterns of transcription and methylation of genes related to the functional properties of mature stromal cells were seen in both hiPS- and hES-derived fibroblasts. iPDK cells also showed functional properties analogous to those of hES-derived and mature fibroblasts, as seen by their capacity to direct the morphogenesis of engineered human skin equivalents. Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

Show MeSH
Related in: MedlinePlus