Additional file 1 of Transplantation of retinal pigment epithelium and photoreceptors generated concomitantly via small molecule-mediated differentiation rescues visual function in rodent models of retinal degeneration
Creators: name: Harshini Surendran
name: Nandakumar, Swapna
name: K, Vijay Bhaskar Reddy
name: Stoddard, Jonathan
name: K, Varsha Mohan
name: Upadhyay, Pramod K.
name: McGill, Trevor J.
name: Rajarshi Pal
Publisher: name: figshare
Year: 2021
Subjects: subject: Biophysics
subject: Medicine
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Neuroscience
subject: Physiology
subject: Immunology
subject: FOS: Clinical medicine
subject: FOS: Clinical medicine
subject: 69999 Biological Sciences not elsewhere classified
subject: Developmental Biology

description: Additional file 1: Supplementary methods, tables, and figures. Table S1. List of primers. Table S2. List of antibodies. Figure S1: A) Status of neuro retinal markers in RPE progenitors at day 35- PAX6, Nestin, CRX. B) neural markers GFAPΔ, SOX2. C) stem cell marker OCT4 at RPE day 70, D) proliferation marker Ki67 in day 70 RPE co expressed with MITF and FOXG1. E) Real time PCR data of non-rosette population selected for RPE at days 20, 35, 50, 70 quantified as fold change and expressed as heat map (Left panel).) NGS results provides confirmation on same set of genes in iPSC, retinal progenitors, RPE progenitors, RPE, PRP samples shows clustering and differential expression (Right panel). Figure S2: A) Expression of RPE markers in PRP; MITF at day 35, ZO1 at day 35 and day 50. B) Status of neural markers in SOX1, Nestin, FOXG1 and GFAPΔ in day 35 PRP cultures. C) Status of stem cell marker OCT4, proliferation marker Ki67 in day 70 PRP. D) Real time PCR data on rosette selected population destined to form PRP at days 20, 35, 50, 60, 75 quantified as fold change and expressed as heat map. E) KCl (+/-) induced intracellular Calcium imaging of day 75 PRP showed positive response. Figure S3: A) Bright field images showing RPE differentiation cultures of an in-house generated iPSC line. B) Expression of RPE markers MITF, PMEL17 and Tyrosinase at day 75. Figure S4: A) Primary data score, data quality and read alignment summary. B) Gene Ontology highlighted in retinal progenitors’ stage represented as fold enrichment. C) Differentially up and down regulated genes expressed in different pathways and signaling channels in retinal progenitors, RPE, PRP samples. D) Three sample Venn diagrams showing differentially up and down regulated genes in retinal progenitors, RPE, and PRP. Figure S5: OMICS data analysis: Log transformed normalized count from RNAseq data shows clustering and differential expression of iPSC, retinal progenitors, RPE progenitors, RPE, PRP samples represented as heat map. A) Retinal ganglion genes; B) Central Nervous System (CNS) related genes; C) embryonic germ layer- specific genes and D, E) adult RPE and fetal RPE signature genes. F) Significantly dysregulated pathway genes. Heat map color: Red to green through black. Figure S6: A) Study design table for RPE animal studies. B) Quantification of histological rescue presented as counted nuclei thick (left) and cones per image (right). C) High dose transplanted animals from P60 and P90 were stained for KI67 and HNA. D) Study design table for PRP animal studies. E) Depth perception behavioral study response in treated animals at different time points.