The Role of Circular RNA Derived from Endothelial Cells in Promoting the Formation of Colonies in Procr+ Islet Cell

Diabetes mellitus is a chronic metabolic disorder characterized by impaired insulin secretion or insulin resistance, and current therapeutic approaches are unable to achieve complete remission. Recent studies have demonstrated that Procr islet cells within murine islets can form colonies and differentiate into islet organoids through co-culture with endothelial cells, subsequently reversing diabetes upon transplantation. However, the underlying mechanism by which endothelial cells promote the growth of Procr islet cells remain incompletely understood. This study aims to investigate circular RNA (circRNA) within the extracellular vesicles (EVs) of endothelial cells as a potential breakthrough. This investigation seeks to elucidate the role of circCARD6, derived from endothelial cells, in promoting the colony formation of Procr cells. I hypothesize that circCARD6 is selectively encapsulated into EVs, subsequently transferred into Procr cells, and functions as a microRNA (miRNA) sponge to modulate the expression of downstream genes, thereby enhancing cellular proliferation and differentiation. To validate this hypothesis, I employed co-culture of Procr cells and endothelial cells (ECs), followed by circRNA sequencing (RNA-seq) to identify differentially expressed circRNAs. I will employ fluorescence in situ hybridization (FISH) using BSJ-specific probes to confirm the transfer of circCARD6 from EC-derived EVs to Procr cells. Furthermore, I will use biotin-labeled miRNA pull-down assays and functional over-expression experiments to verify the sponge activity of circCARD6 on miR-31 and miR-29b-3p, as well as its impact on downstream protein expression and colony formation capacity. I expect proving that circCARD6 serves as a critical EV-derived circRNA that promotes colony formation and differentiation of Procr cells by acting as a miRNA sponge. This study provides novel insights into the mechanism of circRNA-mediated intercellular communication and identifies circCARD6 as a potential molecular target for enhancing islet regeneration. Our research findings may contribute to the development of novel therapeutic strategies for diabetes management.