INVESTIGATING THE ROLE OF TRANSCRIPTION FACTORS IN THE DIFFERENTIATION AND DEVELOPMENT OF PANCREATIC β-CELLS USING HUMAN PLURIPOTENT STEM CELLS

Abstract

Diabetes is a chronic metabolic disorder characterized by loss or reduced insulin (INS)-secreting β-cells. Recent advances in stem cell research showed the ability of human pluripotent stem cells (hPSCs) to produce pancreatic β-cells that can be used to treat patients with diabetes. The need for transplantation therapy has urged researchers to use hPSCs to understand the process of pancreatic development and the regulatory network directing its differentiation into pancreatic β-cells. Although several studies were able to design efficient pancreatic differentiation protocols capable of producing high numbers of pancreatic progenitors (PPs) co-expressing PDX1 and NKX6.1 and ~30-50% INS-producing β-cells, the generation of functional, mono-hormonal β-cells remains an issue. Therefore, the aim of this study was to further understand the role of key transcription factors (TFs), RFX3/6 and FOXA2, in pancreatic development to mimic the ability to generate fully functional pancreatic β-cells in vitro. Some animal studies highlighted the importance of RFX3, RFX6, and FOXA2 TFs; however, their roles during early and late stages of pancreatic β-cell development as well as in INS secretion are not well-studied in human. Mapping the expression of RFX3 and RFX6 during the differentiation of hESCs into pancreatic β-cells showed that RFX6 was expressed at early stages of pancreatic differentiation, specifically at posterior foregut (PF) stage, and then became restricted to pancreatic β-cells where all INS+ cells were co-expressing RFX6. However, RFX3 was mostly expressed at late stages of pancreatic differentiation and was not restricted to a specific type of endocrine cell. PPs derived from FOXA2–/– hiPSCs showed a significant reduction in key pancreatic markers essential for pancreatic development. RNA-seq data revealed downregulation of genes associated with MODY, pancreatic development, Notch and WNT signaling pathways. Furthermore, absence of FOXA2 was associated with upregulation of several miRNAs controlling the expression of key TFs in pancreatic β-cell development. These findings indicate that RFX3, RFX6, and FOXA2 play essential roles in the development of human pancreatic β-cells. Also, the results suggest that FOXA2 play an essential role in regulating miRNAs targeting pancreatic endocrine TFs.

Date of Award	2022
Original language	American English
Awarding Institution	HBKU College of Health & Life Sciences