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Investigating type 1 diabetes pathogenesis with the live pancreas tissue slice platform

Abstract Text

Mollie Huber1, Denise Drotar2, Helmut Hiller3, Maria Beery3, Paul Joseph3, Irina Kusmartseva3 , Stephan Speier2 , Todd Brusko1, Maigan Brusko1, Mark Atkinson3 , Edward Phelps4, and Clayton Mathews1

1Dept. of Pathology, University of Florida; 2Paul Langerhans Institute, Technische Universität Dresden; 3nPOD Laboratory, Dept. of Pathology, University of Florida; 4 J. Crayton Pruitt Family Dept. of Biomedical Engineering, University of Florida

Type 1 diabetes (T1D) results from the autoimmune destruction of pancreatic beta cells. Previously it has been difficult to study the islet microenvironment and corresponding islet-immune cell interactions in live, intact pancreatic tissue, particularly from human samples. The development and application of the slice methods allow for the in-depth study of T1D pathogenesis in the context of the genuine and live islet environment.

In these studies, mouse pancreas tissue slices were prepared from NOD-Rag1-/- and NOD-Rag1-/--AI4α/β TCR transgenic (AI4) mice. Due to the consistent development of T1D in this model, the effects of insulitis at various stages of disease progression on islet functionality and tissue condition can be observed. These observations were achieved through calcium flux recordings as well as immune cell staining and tracking performed using confocal microscopy. Furthermore, studies were conducted in live human pancreas tissue slices prepared from control donors, autoantibody-positive donors, and donors with recent-onset T1D. When CD3+ cells were stained in slices from an autoantibody positive donor, an islet was found with apparent focal insulitis. Using HLA-dextramer reagents we further identified autoreactive T cells within the insulitis. To our knowledge, these are the first ever images of live, endogenous human immune cells attacking insulin-producing beta cells in situ.

Through the application of the pancreatic slice model, the effects of insulitis on beta cell function, particularly at early stages of disease, can be studied in detail. T cells can be introduced in to live human pancreatic tissue slices and their interactions and functional impacts recorded. Differences in immune cell behavior can be studied in tissues from individuals at varying degrees of disease risk. Further studies using live pancreas slices will help to discern many of the processes and effects of islet-immune cell interactions.