Fig. 6: Comparative analysis of 6 months in vitro and in vivo SC-β cells and improving in vitro maturation by ARID1B gene knockdown.
a, Schematic showing comparison of changes associated with 6 month in vitro culture and 6 month in vivo transplants. b, Number of genes, by gene expression or ATAC promoter accessibility, or motif chromatin accessibility, downregulated or upregulated in 6 months in vitro and in vivo SC-β cells. Number of features were determined by differential gene, promoter accessibility or motif accessibility analysis comparing 2 week SC-β cells with 6 month SC-β cells in vitro or 6 month SC-β cells in vivo (24,491 cells from 5 independent biological samples; week 2 representative SC-islets, month 6 SC-islets, and 3 samples of month 6 in vivo SC-islets). c, Volcano plots showing differential motif accessibility analysis (left) and differential gene expression analysis (right) comparing 6 months SC-β cells from in vitro and in vivo SC-islets. Statistical significance was assessed by two-sided Wilcoxon rank sum test for RNA expression and two-sided logistic regression for motif chromatin accessibility. d, Schematic of SC-islets transfected with shRNA lentivirus for ARID1B gene knockdown. e, qPCR plots of SC-islets with ARID1B shRNA showing mean ± s.e.m. (n = 4 biologically independent samples) of expression of β-cell-associated genes, INS (P = 0.0014), DLK1 (P = 4.3 × 10−5) and IAPP (P = 3.0 × 10−6). Statistical significance was assessed by unpaired two-sided t-test. f, Protein quantification plot showing mean ± s.e.m. (n = 4 biologically independent samples) of human insulin content (P = 7.6 × 10−4) by ELISA (left), and pro-insulin/insulin ratio (P = 3.6 × 10−4) by ELISA (right). Statistical significance was assessed by unpaired two-sided t-test. g, Volcano plot from single-nucleus multi-omics comparing motif chromatin accessibility of SC-β cells from control and ARID1B shRNA condition (21,969 cells from 2 independent biological samples, 1 of each condition from the same differentiation batch; integration of all samples). Statistical significance was assessed by two-sided logistic regression. SC, stem cell derived; Txp, transplant; EC, enterochromaffin cells.
