Survival and potential of insulin-deficient beta cells in type 1 diabetes - SUMMARY / ABSTRACT
Recent evidence put forth by our group and others suggests that the pancreas of patients with recent-onset type
1 diabetes (T1D) contains a significant number of “empty” beta cells that have lost the ability to secrete mature
insulin but retained other hallmark features such as proinsulin expression. The mechanisms that underlie
formation of such cells and their fate remain poorly understood. We hypothesize that empty beta cells, which
are invisible to standard insulin immunostaining and potentially also to the immune system, play important roles
in the pathophysiology of T1D while holding therapeutic potential for reversion of diabetes. We propose in depth
experiments to characterize empty beta cells, using MACSima ultra high-content immunofluorescence imaging
to assess in situ expression of 78 immune and pancreas cell markers, including vascular and lymphatic
annotation with signatures of inflammation, extravasation/trafficking, and immune cell residency in autoantibody
positive (AAb+) and T1D donors as compared to donors with type 2 diabetes (T2D) and non-diabetic controls
available through the Network for Pancreatic Organ donors with Diabetes (nPOD) repository. We will compare
insulin containing versus insulin negative islets, within and across donors, to identify empty beta cells and
determine how they correlate with islet, acinar, and immune cell phenotypes. These data will serve as a template
for a serial section to undergo laser capture microdissection (LCM) of insulin containing islets, insulin negative
islets, and acinar tissue regions, which will be subjected to bulk RNAseq as well as our novel method for
quantifying beta cells based on DNA methylation patterns. In addition, we will use a novel mouse model (beta
cell-specific, tamoxifen-inducible Adar1-mutant) and cultured human pancreas slices to functionally interrogate
molecular pathways underlying the formation of empty beta cells. Specifically, we propose to leverage these two
model systems to test therapeutic candidates— including an incretin mimetic (GLP1), an endoplasmic reticulum
(ER) stress inhibitor (ISRIB), and multiple biologics targeting specific immune subsets— for their ability to
modulate empty beta cells, their insulin content and insulin secretion. We will then correlate these functional data
with molecular and cellular features of hormone negative islet cells via MACSima, RNAseq, and DNA
methylation. These studies are expected to yield insights into a fundamental yet little understood process taking
place in human T1D. We anticipate that empty beta cells can be re-functionalized, paving the way for therapeutic
development to restore endogenous beta cell function in T1D. Ultimately, when combined with effective
interventions to constrain autoimmunity, it is our hope that the metabolic modalities explored here could provide
a means to reduce insulin requirements or even achieve insulin independence after T1D onset, dramatically
improving longevity and quality of life for these patients.