Project description:The integrity of the epithelium is maintained by a complex but regulated interplay of processes that allow seamless orchestration of a proliferative state into a stably differentiated state. In this study, using stem cell derived Retinal Pigment Epithelium (RPE) cells as a model; we have investigated the molecular mechanisms that affect attainment of the epithelial phenotype. We identify a novel role for the proto-oncogene FOXM1 in determining epithelial fate of RPE by directly regulating cell proliferation and by indirectly regulating expression of signalling factors BMP7 and Wnt5B; both of which are intimately required for eventual acquisition of the epithelial phenotype. This study uncovers the role of FOXM1 in a non-oncogenic, native-like setting and shows that human ES derived RPE can serve as a useful model system to address biological questions not restricted to visual function.
Project description:To evaluate the effect of oxidative stress on transcript localization in the retinal pigment epithelium (RPE), we performed poly-A RNA sequencing on nuclear and cytoplasmic fractions from induced pluripotent stem cell-derived retinal pigment epithelium (iPSC-RPE) cells exposed to hydrogen peroxide, as well as untreated controls.
Project description:The retinal pigment epithelium (RPE) provides vital support to photoreceptor cells and its dysfunction is associated with the onset and progression of age-related macular degeneration (AMD). Surgical provision of RPE cells may ameliorate AMD and thus it would be valuable to develop sources of patient-matched RPE cells for this application of regenerative medicine. We describe here the generation of functional RPE-like cells from fibroblasts that represent an important step toward that goal. We identified candidate master transcriptional regulators of RPEs using a novel computational method and then used these regulators to guide exploration of the transcriptional regulatory circuitry of RPE cells and to reprogram human fibroblasts into RPE-like cells. The RPE-like cells share key features with RPEs derived from healthy individuals, including morphology, gene expression and function, and thus represent a step toward the goal of generating patient-matched RPE cells for treatment of macular degeneration. Expression analysis was performed on induced retinal pigment epithelium-like cells.
Project description:Development of efficient and reproducible conditions for directed differentiation of pluripotent stem cells into specific cell types is important not only to understand early human development but also to enable more practical applications, such as in vitro models of disease, drug discovery, and cell therapies. The differentiation of stem cells to retinal pigment epithelium (RPE) in particular holds promise as a source of cells for therapeutic replacement in age-related macular degeneration. Here we show development of a robust and efficient method to derive RPE with high reproducibility in an adherent, monolayer system using sequential inhibition and activation of the Activin and BMP signalling pathways. We use whole genome transcript analysis to characterize cells at different stages of differentiation to gain further understanding of the developmental dynamics and fate specification of RPE.
Project description:Illumina Infinium HumanMethylation450 BeadChip data from genomic DNA of retinal pigment epithelium from Age-related Macular Degeneration patients or age-matched controls.
Project description:Transcriptional comparison of adult human primary Retinal Pigment Epithelium, human pluripotent stem cell-derived Retinal Pigment Epithelium, and ARPE19 cells
Project description:To identify disease-specific transcriptional programs in retinal pigment epithelium (RPE) cells, fibroblasts from 43 patients with geographic atrophy (GA) were reprogrammed into induced pluripotent stem cells (iPSCs) before being differentiated into RPE and compared to those from 36 healthy individuals. 127,659 RPE cells were profiled via single cell RNA-sequencing (scRNA-seq) and cell classification identified 7 cellular states related to RPE maturation.