biostudies-arrayexpressKavitha Krishna SudhakarHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-16679Background: Age-related macular degeneration (AMD) is a leading cause of vision loss. Reticular pseudodrusen (RPD), deposits on the apical side of the retinal pigment epithelium (RPE), signify a distinctive and critical AMD phenotype. Yet, their molecular basis and relationship to the conventional drusen seen in AMD remain unclear. Results: We generated induced pluripotent stem cell-derived RPE cells from a clinically phenotyped cohort comprising only individuals with conventional drusen (AMD/RPD-) or drusen coexisting with RPD (AMD/RPD+). From these cells, we generated single-cell transcriptomics, proteomics, and functional data to identify differences between the two cohorts. We show that AMD/RPD+ RPE cells exhibit enrichment in extracellular matrix (ECM) remodelling, cytoskeletal, and hypoxia-responsive programs, whereas AMD/RPD- RPE cells display a relatively greater representation of mitochondrial and protein homeostasis pathways. Both subtypes engaged pathways classically linked to ageing, including ECM remodelling and mitochondrial function, but differed in the direction and extent of these changes. Expression and protein quantitative trait loci (QTLs) highlight shared genetic influences on mitochondrial and iron-handling pathways, while disease-interacting eQTLs and transcriptome-wide association study identify regulatory signals that are distinctive of the RPD subtype within AMD, including through regulation of ECM. Functionally, all iPSC-derived RPE formed drusen-like deposits in vitro: AMD/RPD- lines generated more basal deposits, whereas AMD/RPD+ cells exhibited greater structural instability under bisretinoid-induced stress.biostudies-arrayexpressNucleic Acid Extraction - The pooled samples were combined with the master mix containing the reverse transcriptase reaction and added to the first well of the third row of the 10x Genomics Chromium Test Chip. Partition oil was loaded into the first well of the first row, and gel beads were added to the first well in the second row. The remaining wells were filled with 50% glycerol and the bottom row was empty. As the chip runs, an emulsion occurs wherein each oil droplet contains one bead and one single cell termed GEM.Library Construction - Single cell RNA-sequencing libraries were prepped using the Chromium Single Cell 3’ Library & Gel bead Kit for 10X Genomics.Sample Collection - RPE cells were dissociated using trypsin-EDTA (0.25%) in phenol red (Life Technologies, 25200072) for 5 min at 37°C in a 5% CO2 incubator, the plate was tapped to release any non-RPE cells and washed with PBS-/- (Life Technologies, 14190-144). Trypsin-EDTA (0.25%) in phenol red (Life Technologies, 25200072) was added a second time and incubated for 10 min at 37°C in a 5% CO2 incubator. The trypsin was inactivated with RPE medium and cells were then harvested into a 15 mL conical tube, as a single-cell suspension. The cell suspension was centrifuged (5 minutes, 300 g, 4°C), then resuspended in 1 mL of 0.1% BSA-PBS. Cells were counted and assessed for viability with trypan blue, and pooled (eight samples maximum) at a concentration of 1000 live cells/μL (1 × 106 cells/mL).Sequencing - Libraries were sequenced on Illumina NovaSeq 6000.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - Raw UMI counts from remaining cells then underwent normalization and scaling using the SCTransform function as implemented in Seurat.MetabolomicsUnknownTranscriptomicsGenomicsProteomicsIllumina NovaSeq 6000RNA-seq of coding RNA from single cellsHomo sapiensKavitha Krishna SudhakarfalsePatient induced pluripotent stem cells identify specificities of a reticular pseudodrusen phenotype in age-related macular degenerationBackground: Age-related macular degeneration (AMD) is a leading cause of vision loss. Reticular pseudodrusen (RPD), deposits on the apical side of the retinal pigment epithelium (RPE), signify a distinctive and critical AMD phenotype. Yet, their molecular basis and relationship to the conventional drusen seen in AMD remain unclear. Results: We generated induced pluripotent stem cell-derived RPE cells from a clinically phenotyped cohort comprising only individuals with conventional drusen (AMD/RPD-) or drusen coexisting with RPD (AMD/RPD+). From these cells, we generated single-cell transcriptomics, proteomics, and functional data to identify differences between the two cohorts. We show that AMD/RPD+ RPE cells exhibit enrichment in extracellular matrix (ECM) remodelling, cytoskeletal, and hypoxia-responsive programs, whereas AMD/RPD- RPE cells display a relatively greater representation of mitochondrial and protein homeostasis pathways. Both subtypes engaged pathways classically linked to ageing, including ECM remodelling and mitochondrial function, but differed in the direction and extent of these changes. Expression and protein quantitative trait loci (QTLs) highlight shared genetic influences on mitochondrial and iron-handling pathways, while disease-interacting eQTLs and transcriptome-wide association study identify regulatory signals that are distinctive of the RPD subtype within AMD, including through regulation of ECM. Functionally, all iPSC-derived RPE formed drusen-like deposits in vitro: AMD/RPD- lines generated more basal deposits, whereas AMD/RPD+ cells exhibited greater structural instability under bisretinoid-induced stress.2026-04-09T00:00:00Z2026-04-09T10:19:37.889Z2026-02-25T00:42:45.208ZE-MTAB-16679ERP189575EFO_0002944EFO_0004170EFO_0005684EFO_0005518EFO_0003816EFO_0004184