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Global gene expression profiling reveals similarities and differences among mouse pluripotent stem cells of different origins and strains.

ABSTRACT: Pluripotent stem cell lines with similar phenotypes can be derived from both blastocysts (embryonic stem cells, ESC) and primordial germ cells (embryonic germ cells, EGC). Here, we present a compendium DNA microarray analysis of multiple mouse ESCs and EGCs from different genetic backgrounds (strains 129 and C57BL/6) cultured under standard conditions and in differentiation-promoting conditions by the withdrawal of Leukemia Inhibitory Factor (LIF) or treatment with retinoic acid (RA). All pluripotent cell lines showed similar gene expression patterns, which separated them clearly from other tissue stem cells with lower developmental potency. Differences between pluripotent lines derived from different sources (ESC vs. EGC) were smaller than differences between lines derived from different mouse strains (129 vs. C57BL/6). Even in the differentiation-promoting conditions, these pluripotent cells showed the same general trends of gene expression changes regardless of their origin and genetic background. These data indicate that ESCs and EGCs are indistinguishable based on global gene expression patterns alone. On the other hand, a detailed comparison between a group of ESC lines and a group of EGC lines identified 20 signature genes whose average expression levels were consistently higher in ESC lines, and 84 signature genes whose average expression levels were consistently higher in EGC lines, irrespective of mouse strains. Similar analysis identified 250 signature genes whose average expression levels were consistently higher in a group of 129 cell lines, and 337 signature genes whose average expression levels were consistently higher in a group of C57BL/6 cell lines. Although none of the genes was exclusively expressed in either ESCs versus EGCs or 129 versus C57BL/6, in combination these signature genes provide a reliable separation and identification of each cell type. Differentiation-promoting conditions also revealed some minor differences between the cell lines. For example, in the presence of RA, EGCs showed a lower expression of muscle- and cardiac-related genes and a higher expression of gonad-related genes than ESCs. Taken together, the results provide a rich source of information about the similarities and differences between ESCs and EGCs as well as 129 lines and C57BL/6 lines. Such information will be crucial to our understanding of pluripotent stem cells. The results also underscore the importance of studying multiple cell lines from different strains when making comparisons based on gene expression analysis.

SUBMITTER: Sharova LV

PROVIDER: S-EPMC2000702 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundEver since their discovery, induced pluripotent stem cells (iPSCs) have been extensively differentiated into a large variety of cell types. However, a limited amount of work has been dedicated to differentiating iPSCs into osteoclasts. While several differentiation protocols have been published, it remains unclear which protocols or differentiation methods are preferrable regarding the differentiation of osteoclasts.MethodsIn this study we compare the osteoclastogenesis capacity of a peripheral blood mononuclear cell (PBMC)-derived iPSC line to a fibroblast-derived iPSC line in conjunction with either embryoid body-based or monolayer-based differentiation strategies. Both cell lines and differentiation protocols were investigated regarding their ability to generate osteoclasts and their inherent robustness and ease of use. The ability of both cell lines to remain undifferentiated while propagating using a feeder-free system was assessed using alkaline phosphatase staining. This was followed by evaluating mesodermal differentiation and the characterization of hematopoietic progenitor cells using flow cytometry. Finally, osteoclast yield and functionality based on resorptive activity, Cathepsin K and tartrate-resistant acid phosphatase (TRAP) expression were assessed. Results were validated using qRT-PCR throughout the differentiation stages.ResultsEmbryoid-body based differentiation yielded CD45+, CD14+, CD11b+ subpopulations which in turn differentiated into osteoclasts which demonstrated TRAP positivity, Cathepsin K expression and mineral resorptive capabilities. This was regardless of which iPSC line was used. Monolayer-based differentiation yielded lower quantities of hematopoietic cells that were mostly CD34+ and did not subsequently differentiate into osteoclasts.ConclusionsThe outcome of this study demonstrates the successful differentiation of osteoclasts from iPSCs in conjunction with the embryoid-based differentiation method, while the monolayer-based method did not yield osteoclasts. No differences were observed regarding osteoclast differentiation between the PBMC and fibroblast-derived iPSC lines.

Project description:BackgroundThe majority of parthenogenetic vertebrates derive from hybridization between sexually reproducing species, but the exact number of hybridization events ancestral to currently extant clonal lineages is difficult to determine. Usually, we do not know whether the parental species are able to contribute their genes to the parthenogenetic vertebrate lineages after the initial hybridization. In this paper, we address the hypothesis, whether some genotypes of seven phenotypically distinct parthenogenetic rock lizards (genus Darevskia) could have resulted from back-crosses of parthenogens with their presumed parental species. We also tried to identify, as precise as possible, the ancestral populations of all seven parthenogens.ResultsWe analysed partial mtDNA sequences and microsatellite genotypes of all seven parthenogens and their presumed ansectral species, sampled across the entire geographic range of parthenogenesis in this group. Our results confirm the previous designation of the parental species, but further specify the maternal populations that are likely ancestral to different parthenogenetic lineages. Contrary to the expectation of independent hybrid origins of the unisexual taxa, we found that genotypes at multiple loci were shared frequently between different parthenogenetic species. The highest proportions of shared genotypes were detected between (i) D. sapphirina and D. bendimahiensis and (ii) D. dahli and D. armeniaca, and less often between other parthenogens. In case (ii), genotypes at the remaining loci were notably distinct.ConclusionsWe suggest that both observations (i-ii) can be explained by two parthenogenetic forms tracing their origin to a single initial hybridization event. In case (ii), however, occasional gene exchange between the unisexual and the parental bisexual species could have taken place after the onset of parthenogenetic reproduction. Indeed, backcrossed polyploid hybrids are relatively frequent in Darevskia, although no direct evidence of recent gene flow has been previously documented. Our results further suggest that parthenogens are losing heterozygosity as a result of allelic conversion, hence their fitness is expected to decline over time as genetic diversity declines. Backcrosses with the parental species could be a rescue mechanism which might prevent this decline, and therefore increase the persistance of unisexual forms.

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Global gene expression profiling reveals similarities and differences among mouse pluripotent stem cells of different origins and strains.

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