Transdifferentiation of Liver to Pancreas: Refining Therapeutic Liver Derived β-Cells for Type 1 Diabetes Mellitus

Abstract

The rising incidence in diabetes mellitus represents a major global health crisis and a growing burden on individual patients and society. Dysfunction of insulin producing β-cells is implicated in both type one (T1DM) and two (T2DM) diabetes mellitus. Currently available therapies including insulin injection or pharmaceuticals such as insulin secretagogues fall short of effective and sustainable treatment. Allotransplantation of β-cells can be used to cure T1DM in some patients although donor shortages impede widespread use. A potential alternative source of therapeutic β-cells is non-pancreatic tissue including the liver. Conversion of one differentiated cell type to another, termed ‘transdifferentiation’ presents a therapeutic opportunity for the generation of new β-cells from differentiated cells. Functional cures using transdifferentiated β-cells exist in animal models although protocols are inefficient and produce immature phenotypes. It has been proposed that an active Wnt/β-catenin signalling pathway is key to defining those hepatocytes able to undergo the transdifferentiation process to β-cells. Hepatocytes exhibit heterogeneity of functions known as ‘zonation’. One profound example of zonation is in ammonia detoxification. Two mutually exclusive systems exist and can be distinguished by their markers carbamoyl phosphate synthase 1 (CPS1) and glutamine synthetase (GS) respectively. CPS1, the key rate limiting enzyme of the urea cycle is expressed in cells of the periportal, intermediate and first few layers of perivenous zone. GS is expressed in a complementary pattern only in the last few layers of hepatocytes surrounding the central vein. GS expression is under the control of the Wnt/β-catenin pathway. It was hypothesised that by switching on the Wnt/β-catenin pathway in Wnt inactive CPS1 positive hepatocytes that the efficiency of the transdifferentiation process would be enhanced. Primary mouse hepatocytes were exposed to Wnt/β-catenin pathway activators and transduced with pancreatic transcription factors to test this hypothesis. I also utilised an in vitro model for the conversion of rat pancreatic B13 acinar cells to hepatocytes in order to test the role of the TGFβ family member activin A in the context of a dexamethasone induced transdifferentiated hepatocyte-like cell. Here I report the finding that manipulation of the Wnt/β-catenin signalling pathway confers limited maintenance of GS in a heterogenous population of hepatocytes. Furthermore, that ectopic expression of pancreatic transcription factors does not induce a switch in phenotype in adult mouse hepatocyte cultures. I demonstrate that addition of dexamethasone and activin A to pancreatic B13 cells promotes the perivenous hepatocyte phenotype (GS expression) while suppressing the periportal phenotype (CPSI). These data suggest that B13 cells might represent an alternative model for testing the utility of periportal versus perivenous hepatocyte-like cells as a source of cells for transdifferentiation to a β-cell phenotype.

Date of Award	27 Mar 2024
Original language	English
Awarding Institution	University of Bath
Supervisor	David Tosh (Supervisor) & Gernot Walko (Supervisor)