Discovering new functions of the diabetes gene HNF1A in human pancreatic islets

The pancreatic islets of Langerhans are mighty micro-organs that dynamically maintain blood glucose in a homeostatic range. Within the pancreatic islets, beta cells secrete insulin and alpha cells secrete glucagon to antagonistically regulate glucose utilization, storage, and production throughout the body. Insulin functions to lower blood glucose levels in the fed state, while glucagon increases blood glucose levels during fasting. The mechanisms that govern insulin and glucagon secretion are complex, and dysregulation of islet cell function can lead to devastating metabolic consequences, including diabetes. While diabetes is classically defined as a disease of insulin insufficiency, growing evidence indicates that dysregulation of glucagon secretion also contributes to hyperglycemia in multiple diabetes subtypes. Namely, inappropriately high glucagon levels have been reported in patients with Type 2 (T2D) and monogenic forms of diabetes, including HNF1A-MODY. Mutations in HNF1A have been associated with both Maturity Onset Diabetes of the Young (MODY) and increased risk of T2D, and previous studies of monogenic diabetes have informed mechanistic understanding of and therapeutic targets for larger classes of diabetes. Rodent and human stem-cell models have greatly advanced our understanding of pancreatic development and disease. However, heterozygous hnf1a mice do not recapitulate the HNF1A haploinsufficiency phenotypes observed in HNF1A-MODY patients, and stem-cell derived beta-like cells lack certain properties of mature multicellular islets. Thus, there is still a need for models in which to study mechanisms underlying HNF1A-deficient diabetes. Similarly, the mechanisms underlying diabetic phenotypes after extended fasting remain poorly understood. "Starvation diabetes," a state characterized by low insulin levels and hyperglycemia upon refeeding after an extended fast, has been documented in humans and may represent adaptive changes in hormone output from islets to prevent dangerous hypoglycemia during nutrient deprivation. Circulating factors that act on islets to reduce insulin output during fasting and genetic factors that modulate starvation diabetes phenotypes have not yet been elucidated. In this thesis, we present primary human and mouse models that can be used to discover mechanisms underlying T2D, monogenic, and starvation diabetes. We describe methods for achieving reaggregated "pseudoislets" after dispersion of primary human islets, and how pseudoislets are amenable to efficient genetic manipulation while maintaining hallmark features of native islets. Combining the pseudoislet platform with modern genetic approaches, transplantation studies, hormone secretion assays, and chromatin-mapping methods, we identify novel functions of the transcription factor encoded by HNF1A (HNF1alpha) in mature pancreatic beta and alpha cells. Finally, we demonstrate genetic heterogeneity in susceptibility to diabetic phenotypes induced by starvation through characterization of six mouse strains. Overall, these studies reveal a direct mechanistic link between HNF1A loss and diabetic phenotypes in mature human islet cells and provide tools to investigate the mechanisms underlying beta and alpha cell dysfunction in multiple diabetes subtypes.

Keywords: Medicine.
Developmental Biology.
Advisor: Kim Seung K.
Gloyn Anna L.
Kingsley David M. (David Mark)
Nusse Roel