Homo sapiens
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ABSTRACT:
PROVIDER: PRJNA110837 | ENA |
REPOSITORIES: ENA
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Project description:Human fibroblasts can be induced into pluripotent stem cells (iPS cells), but the reprogramming efficiency is quite low. Here, we screened a panel of candidate factors in the presence of OCT4, SOX2, KLF4 and c-MYC in an effort to improve the reprogramming efficiency from human adult fibroblasts. We found that p53 siRNA and UTF1 enhanced the efficiency of iPS cell generation up to 100-fold, even when the oncogene c-MYC was removed from the combinations. We further demonstrated that by using a novel combination of the four factors OCT4, SOX2, KLF4 and UTF1, iPS cells could be generated at a frequency at least 10 times higher than using the original four reprogramming factors without c-MYC. The iPS cells generated in this work have a similar gene expression profile and differentiation potential as human embryonic stem (hES) cells. In conclusion, two novel supporting factors that increase the efficiency of direct reprogramming have been identified, and a more-efficient method for the generation of human iPS cells has been developed in the absence of the oncogene c-MYC. Keywords: cell type comparison
2008-11-07 | GSE12922 | GEO
Project description:Human fibroblasts can be induced into pluripotent stem cells (iPS cells), but the reprogramming efficiency is quite low. Here, we screened a panel of candidate factors in the presence of OCT4, SOX2, KLF4 and c-MYC in an effort to improve the reprogramming efficiency from human adult fibroblasts. We found that p53 siRNA and UTF1 enhanced the efficiency of iPS cell generation up to 100-fold, even when the oncogene c-MYC was removed from the combinations. We further demonstrated that by using a novel combination of the four factors OCT4, SOX2, KLF4 and UTF1, iPS cells could be generated at a frequency at least 10 times higher than using the original four reprogramming factors without c-MYC. The iPS cells generated in this work have a similar gene expression profile and differentiation potential as human embryonic stem (hES) cells. In conclusion, two novel supporting factors that increase the efficiency of direct reprogramming have been identified, and a more-efficient method for the generation of human iPS cells has been developed in the absence of the oncogene c-MYC. Keywords: cell type comparison Total RNA from hFSF, hAFF, hES cells (H1, H7) and 7 established iPS cell lines were labeled with Cy5, hybridized to a human Oligo Microarray (Phalanx Human Whole Genome OneArray™, Phalanx Biotech) according to the manufacturer's protocol. Three technical repetitions were performed. After hybridization, arrays were scanned using GenePix 4000B scanner (Molecular Devices) and processed using the GenePix Pro 6.0 software (Molecular Devices). After removing control probes, a 14/33 presence call (SNR>=5 and foreground-background>0) was used to filter probes for the 33 microarrays, resulting in 12311 probes for further quantile normalization.
2008-11-06 | E-GEOD-12922 | biostudies-arrayexpress
Project description:Ectopic expression of four transcription factors including Oct4, Sox2, Klf4 and c-Myc in differentiated fibroblast cells could reset the cell fate of fibroblast cells to pluripotent state. Subsequently, fully pluripotency of these so-called induced pluripotent stem cells (iPSCs) has been demonstrated as viable mice could be generated autonomously from iPS cells through tetraploid blastocyst complementation. Moreover, the generation of human and patient-specific iPS cells have raised the possibility of utilizing iPS cells clinically. However, the utilization of c-Myc in iPS cells induction greatly increased the incidence of tumorigenecity in the iPS-chimeric mice and also might hinder the clinical application of human iPS cells in the future. Fortunately, c-Myc has been recently found dispensable for iPS induction even though the iPS induction efficiency is greatly reduced in the absence of c-Myc. However, it remains unknown if these three factors-induced iPS cells are fully pluripotent. In the present study, we have successfully demonstrated that 3-factor iPS cells could also be fully pluripotent as viable mice could be generated from 3-factor iPS cells autonomously via tetraploid complementation and moreover, our data indicated that the pluripotency regulatory mechanism in 3-factor iPS cells might be distinct from 4-factor iPS cells.
2010-10-19 | GSE24046 | GEO
Project description:Ectopic expression of four transcription factors including Oct4, Sox2, Klf4 and c-Myc in differentiated fibroblast cells could reset the cell fate of fibroblast cells to pluripotent state. Subsequently, fully pluripotency of these so-called induced pluripotent stem cells (iPSCs) has been demonstrated as viable mice could be generated autonomously from iPS cells through tetraploid blastocyst complementation. Moreover, the generation of human and patient-specific iPS cells have raised the possibility of utilizing iPS cells clinically. However, the utilization of c-Myc in iPS cells induction greatly increased the incidence of tumorigenecity in the iPS-chimeric mice and also might hinder the clinical application of human iPS cells in the future. Fortunately, c-Myc has been recently found dispensable for iPS induction even though the iPS induction efficiency is greatly reduced in the absence of c-Myc. However, it remains unknown if these three factors-induced iPS cells are fully pluripotent. In the present study, we have successfully demonstrated that 3-factor iPS cells could also be fully pluripotent as viable mice could be generated from 3-factor iPS cells autonomously via tetraploid complementation and moreover, our data indicated that the pluripotency regulatory mechanism in 3-factor iPS cells might be distinct from 4-factor iPS cells. We compared the gene expression profile of iPS cells with and without the tetraploid embryo complementation competence. Three biological repeats were included for each line.
2010-10-19 | E-GEOD-24046 | biostudies-arrayexpress
Project description:p53 knockdown by shRNA markedly increased efficiency of human iPS cell generation
2009-03-01 | GSE13334 | GEO
Project description:Introducing four transcription factors, Oct4, Sox2, Klf4 and c-Myc can reprogram both human and mouse somatic cells. first, the oncogenic properties of reprogramming factors such as Klf4 and c-Myc, second, the viral integrations of transcription factors in the genome of somatic cells which may also hinder a clinical setting and finally, the low efficiency of the generation of induced pluripotent stem cells (iPS). Some recent studies overcame the first two problems, the oncogenic properties and genomic integration of the reprogramming factors by either replacing Klf4 and/or c-Myc with small molecules or using different iPS protocols such as integration-free transcription factor inductions as well as introducing proteins. However, the slow reprogramming process and low efficiency of the iPS generation still need to be improved for either clinical application or studying the reprogramming mechanism. Different approaches including adding soluble Wnt3a, and generating iPS cells in hypoxic condition also showed increased efficiency in both 4 factors (Oct4/Sox2/Klf4/c-Myc) or less factors combination without further genetic manipulation. In the current study, we show that basic fibroblast growth factor (bFGF) dramatically increased the efficiency of iPS generation and shortened the reprogramming timing of mouse embryonic fibroblast (MEF) cells in both 4 factors (4F Oct4/Sox2/Klf4/c-Myc) and 3 factors (3F Oct4/Sox2/Klf4) combination without further exogenous genetic manipulation. RNA samples to be analyzed on microarrays were prepared using Qiagen RNeasy columns with on-column DNA digestion. 300 ng of total RNA per sample was used as input into a linear amplification protocol (Ambion), which involved synthesis of T7-linked double-stranded cDNA and 12 hrs of in-vitro transcription incorporating biotin-labelled nucleotides. Purified and labelled cRNA was then hybridized for 18 hrs onto MouseRef-8 v2 expression BeadChips (Illumina) according to the manufacturer's instructions. After washing, as recommended, chips were stained with streptavidin-Cy3 (GE Healthcare) and scanned using the iScan reader (Illumina) and accompanying software. Samples were hybridized as biological replicates.
2016-10-13 | GSE18888 | GEO
Project description:The expression of four transcription factors (OCT3/4, SOX2, KLF4, and c-MYC) can reprogram mouse as well as human somatic cells to induced pluripotent stem (iPS) cells. Expression of the c-MYC, also known as an oncogene, might induce carcinogenesis and thus, iPS cells produced with the use of c-MYC transduction cannot be used for human therapeutic applications. Furthermore, reprogramming efficiency was significantly reduced in the absence of c-MYC transduction. Here, we generated iPS cells from mesenchymal stromal cells (MSCs) derived from human third molars (wisdom teeth) by retroviral transduction of OCT3/4, SOX2, and KLF4 without c-MYC. Interestingly, clonally expanded MSCs, named 10F-15, could be used for iPS cell generation with 100-fold higher efficiency compared to that of other clonally expanded MSCs and human dermal fibroblasts. These iPS cells resembled human embryonic stem (ES) cells in many aspects, including morphology, ES markers expression, global gene expression, epigenetic states, and the ability to differentiate into the three germ layers in vitro and in vivo. Because human third molars are discarded as clinical waste, our data indicate that MSCs isolated from human third molars are a valuable cell source for the generation of iPS cells.
2010-06-26 | GSE16963 | GEO
Project description:Human somatic fibroblasts can be reprogrammed to induced pluripotent stem (iPS) cells by exogenic expression of the Yamanaka factors (OCT4, SOX2, KLF4 and MYC) after about 1 month. To gain some insight into the early processes operative in fibroblast reprogramming, we profiled genome-wide transcription levels using Illumina microarrays in the starting donor cells-human foreskin fibroblast (HFF1) cells and at three time points after OSKM transduction (24 h, 48 h, 72 h), as well as two iPS cell lines (iPS2, iPS4) and hES cell lines (H1, H9). We show that within the context of the viral transduction reprogramming protocol, the donor cell response to viral transfection perturbs redox homeostasis, which induces oxidative damage on the donor cells' protein and DNA. This leads to activation of p53, senescence, and apoptosis, greatly reducing the efficiency of reprogramming.
2011-10-25 | GSE28688 | GEO
Project description:Human somatic fibroblasts can be reprogrammed to induced pluripotent stem (iPS) cells by exogenic expression of the Yamanaka factors (OCT4, SOX2, KLF4 and MYC) after about 1 month. To gain some insight into the early processes operative in fibroblast reprogramming, we profiled genome-wide transcription levels using Illumina microarrays in the starting donor cells-human foreskin fibroblast (HFF1) cells and at three time points after OSKM transduction (24 h, 48 h, 72 h), as well as two iPS cell lines (iPS2, iPS4) and hES cell lines (H1, H9). We show that within the context of the viral transduction reprogramming protocol, the donor cell response to viral transfection perturbs redox homeostasis, which induces oxidative damage on the donor cells' protein and DNA. This leads to activation of p53, senescence, and apoptosis, greatly reducing the efficiency of reprogramming. Total RNA obtained from HFF1 (human foreskin fibroblast) cells, OSKM-transduced HFF1 cells after 24h, 48h, 72h, undifferentiated hESCs, iPSCs.
2011-10-24 | E-GEOD-28688 | biostudies-arrayexpress
Project description:p53 knockdown by shRNA markedly increased efficiency of human iPS cell generation Gene expression patterns were compared between human ES cells and dermal fibroblasts. Gene expression patterns were also compared between p53 shRNA-treated fibroblasts and control fibroblasts.
2009-02-28 | E-GEOD-13334 | biostudies-arrayexpress