Single-Cell RNA Sequencing Reveals a Role for ROS Signaling in Nutrient-Induced beta-Cell Proliferation

In nondiabetic obese individuals, β-cell mass accretion maintains glucose homeostasis by balancing levels of circulating insulin and insulin resistance. Type 2 diabetes appears when these compensatory mechanisms fail. Deciphering the pathways controlling β-cell proliferation may help to identify therapeutic targets to expand β-cell mass and prevent or delay the onset of diabetes. Previously we demonstrated that the monounsaturated fatty-acid oleate potentiates glucose-induced rat β-cell proliferation. Aim: To identify the transcriptional mechanisms underlying oleate-induced rat β-cell proliferation. Methods: Rat islets were exposed to 16.7mM glucose (Glu) with or without 0.5 mM oleate (Glu+Ol) or palmitate (Glu+Pal) for 48h. Proliferation was assessed by flow cytometry (C-peptide staining and EdU incorporation). Single-cell cDNA libraries were generated using 10X Genomics technology and sequenced on the Illumina platform. Bioinformatic analyses were performed using Seurat package. Results: β-cell proliferation was potentiated by Ol, but not Pal. Following single-cell RNA sequencing, UMAP plots revealed several β-cell sub-populations, including proliferating β-cells. A number of differences in gene expression were identified between groups in non-proliferating sub-populations. Among these, gene sets involved in ER-stress and β-cell differentiation were significantly up- and down-regulated, respectively, in response to Pal, but not Ol. Gene set enrichment analyses comparing proliferative vs. non-proliferative β-cells revealed an up-regulation of reactive oxygen species (ROS) and oxidoreductase activity-related pathways in both Glu and Glu+Ol groups. The antioxidant N-acetyl cysteine or the peroxiredoxin inhibitor Conoidin A inhibited Glu+Ol-induced β-cell proliferation. Conclusion: Our results suggest the involvement of ROS signaling through a redox relay via peroxiredoxin activation in nutrient-induced β-cell proliferation. Disclosure A. Vivoli: None. J. Ghislain: None. A. Castell: None. A. Filali: None. R. M. Sladek: None. V. Poitout: None.