Dataset Information

Profiling the microRNA Expression in Human iPS and iPS-derived Retinal Pigment Epithelium.

ABSTRACT: The purpose of this study is to characterize the microRNA (miRNA) expression profiles of induced pluripotent stem (iPS) cells and retinal pigment epithelium (RPE) derived from induced pluripotent stem cells (iPS-RPE). MiRNAs have been demonstrated to play critical roles in both maintaining pluripotency and facilitating differentiation. Gene expression networks accountable for maintenance and induction of pluripotency are linked and share components with those networks implicated in oncogenesis. Therefore, we hypothesize that miRNA expression profiling will distinguish iPS cells from their iPS-RPE progeny. To identify and analyze differentially expressed miRNAs, RPE was derived from iPS using a spontaneous differentiation method. MiRNA microarray analysis identified 155 probes that were statistically differentially expressed between iPS and iPS-RPE cells. Up-regulated miRNAs including miR-181c and miR-129-5p may play a role in promoting differentiation, while down-regulated miRNAs such as miR-367, miR-18b, and miR-20b are implicated in cell proliferation. Subsequent miRNA-target and network analysis revealed that these miRNAs are involved in cellular development, cell cycle progression, cell death, and survival. A systematic interrogation of temporal and spatial expression of iPS-RPE miRNAs and their associated target mRNAs will provide new insights into the molecular mechanisms of carcinogenesis, eye differentiation and development.

SUBMITTER: Wang HC

PROVIDER: S-EPMC4218680 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Profiling the microRNA Expression in Human iPS and iPS-derived Retinal Pigment Epithelium.

Wang Heuy-Ching HC Greene Whitney A WA Kaini Ramesh R RR Shen-Gunther Jane J Chen Hung-I H HI Cai Hong H Wang Yufeng Y

Cancer informatics 20141015 Suppl 5

The purpose of this study is to characterize the microRNA (miRNA) expression profiles of induced pluripotent stem (iPS) cells and retinal pigment epithelium (RPE) derived from induced pluripotent stem cells (iPS-RPE). MiRNAs have been demonstrated to play critical roles in both maintaining pluripotency and facilitating differentiation. Gene expression networks accountable for maintenance and induction of pluripotency are linked and share components with those networks implicated in oncogenesis. ...[more]

PMID: 25392691

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Project description:The therapeutic potential of pluripotent stem cells is great as they promise to usher in a new era of medicine where cells or organs may be prescribed to replace dysfunctional tissue. At the forefront are efforts in the eye to develop this technology as it lends itself to in vivo monitoring and sophisticated non-invasive imaging modalities. In the retina, retinal pigment epithelium (RPE) is the most promising replacement cell as it has a single layer, is relatively simple to transplant, and is associated with several eye diseases. However, after transplantation, the cells may transform and cause complications. This transformation may be partially due to incomplete maturation. With the goal of learning how to mature RPE, we compared induced pluripotent stem cell-derived RPE (iPSC-RPE) cells with adult human primary RPE (ahRPE) cells and the immortalized human ARPE-19 line. We cultured ARPE-19, iPSC-RPE, and ahRPE cells for one month, and evaluated morphology, RPE marker staining, and transepithelial electrical resistance (TEER) as quality control indicators. We then isolated RNA for bulk RNA-sequencing and DNA for genotyping. We genotyped ahRPE lines for the top age-related macular degeneration (AMD) and proliferative vitreoretinopathy (PVR) risk allele polymorphisms. Transcriptome data verified that both adult and iPSC-RPE exhibit similar RPE gene expression signatures, significantly higher than ARPE-19. In addition, in iPSC-RPE, genes relating to stem cell maintenance, retina development, and muscle contraction were significantly upregulated compared to ahRPE. We compared ahRPE to iPSC-RPE in a model of epithelial-mesenchymal transition (EMT) and observed an increased sensitivity of iPSC-RPE to producing contractile aggregates in vitro which resembles incident reports upon transplantation. P38 inhibition was capable of inhibiting iPSC-RPE-derived aggregates. In summary, we find that the transcriptomic signature of iPSC-RPE conveys an immature RPE state which may be ameliorated by targeting "immature" gene regulatory networks.

Project description:PURPOSE: The molecular mechanisms associated with human retinal pigment epithelium (RPE) development constitute the basis for cell replacement therapy for the treatment of retinal degenerative diseases. In the current study, gene expression analysis of the human fetal RPE during development was performed and was compared with the human native RPE. METHODS: Microdissection of the human RPE at three time points (13 weeks and 16 weeks of gestation and in mature adult eyes) was performed, and total RNA was isolated. Equal amounts of RNA were pooled from two or three independent donor eyes for each time point in each group. Gene expression was analyzed by hybridization to microarray chips. Validation was accomplished by comparing the microarray expression pro?les with quantitative real-time reverse transcriptase-polymerase chain reaction (qRT(2)-PCR) analysis of selected genes and by comparing selected expression pro?les with predicted pro?les based on previous studies. RESULTS: Of the 45,033 probe sets on the microarray, 30,736 were detected. A total of 3,498 differentially expressed genes could be clustered into eight patterns of expression that were statistically significant. Analysis of the expression patterns of genes coding for key functions (pigment synthesis, visual cycle, phagocytosis, adherens and tight junctions, and transcellular transport) indicated that the human RPE achieves a high degree of maturity during early pregnancy. Compared with 154 signature genes in the RPE, 148 candidate genes were identified in this study, including 53 downregulated genes and 5 upregulated genes. The qRT(2)-PCR results showed similar expression trends to those obtained by microarray analysis at the three time points. CONCLUSIONS: This study demonstrated gene expression profiles in the human RPE during normal development. These ?ndings indicate that the human RPE has different expression patterns than those of other animals. The results of this study may be helpful in furthering the understanding of the developmental processes occurring in humans and of the differentiation of RPE cells derived from human embryonic stem cells and from human induced pluripotent stem cells.

Dataset Information

Profiling the microRNA Expression in Human iPS and iPS-derived Retinal Pigment Epithelium.

Publications

Profiling the microRNA Expression in Human iPS and iPS-derived Retinal Pigment Epithelium.

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