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Longitudinal, live imaging of islet autoimmune destruction in NOD mice

Gaetano Faleo¹, Midhat Abdulreda¹, R. Damaris Molano¹, Maite Lopez-Cabezas¹, Elsie Zahr-Akrawi¹, Judith Molina¹, Allison Bayer^1,2, Camillo Ricordi^1,2,3,4,5, Per-Olof Berggren^1,3,4, Alejandro Caicedo¹, Antonello Pileggi^1,2,3,5

¹Diabetes Research Institute, University of Miami; ²Microbiology & Immunology, University of Miami Miller School of Medicine; ³Surgery, University of Miami Miller School of Medicine, Miami, FL, United States; ⁴The Rolf Luft Research Center for Diabetes and Endocrinology, Karolinska Institutet, Stockholm, Sweden; ⁵Biomedical Engineering, University of Miami, Miami, FL, United States

NOD mice spontaneously develop Type-1 diabetes (T1D). Islet cells are difficult to access in vivo, because they are scattered within the exocrine tissue of the pancreas. Multiphoton/confocal fluorescent microscopy allows performing live imaging with cellular resolution. However, surgical exteriorization of the pancreas is required to study islet immunobiology, precluding longitudinal studies. We hypothesized that the anterior chamber of the eye (ACE) represents a useful in vivo tool to study islet immunobiology. The progression of autoimmunity was assessed by transplanting NOD.SCID islets into either spontaneously diabetic female NOD (recurrence model) or NOD.SCID mice (autoimmunity adoptive transfer model).

Recurrence of diabetes occurred with a median of 10 (range 5-12) and 8 (5-13) days in the ACE (n=6) and kidney capsule (KDN; n=12), respectively. Adoptive transfer of splenocytes from newly diabetic NOD mice induced diabetes within 30-35days. Longitudinal assessment of individual islet volume and granularity in the ACE of reconstituted NOD.SCID mice demonstrated islet swelling starting one week before onset of overt hyperglycemia followed by relatively quick volume reduction within a week. Live time-lapse studies performed after direct cytolabeling and cell viability dye injection in the ACE in selected animals allowed assessing the behavior of infiltrating B and T cells and cell death in the target tissue with single-cell resolution prior, during and after the onset of diabetes. Preliminary assessment of islet grafts explanted after diabetes onset from ACE and KDN showed infiltrating T and B lymphocyte populations similar to native pancreas by immunofluorescence staining.

In conclusion, islet transplantation into the ACE represents a valuable model to study islet immunity and offers unprecedented advantages compared to other transplantation sites, particularly the possibility to perform longitudinal in vivo studies on the very same islets with cellular resolution to characterize the effector phase kinetics of the infiltrating cells on the site of immune attack.

NIH-5U19AI050864-10