(#49) Postnatal Development of Islet Macrophages

PRESENTED BY: Farhan Qureshi

Authors
First NameLast NameAffiliation/Institution
FarhanQureshiUniversity of Miami Department of Medicine, Division of Endocrinology
JuliaPanzerUniversity of Miami Department of Medicine, Division of Endocrinology
AlejandroCaicedoUniversity of Miami Department of Medicine, Division of Endocrinology
 

Purpose

The purpose of this study is to understand the physiological profile of the pancreatic islet resident macrophage at different stages during human postnatal development. The predominant leukocyte in the islet is the resident macrophage. This cell gauges the activation status of beta cells by detecting beta cell-derived ATP and adjusts its functions to contribute to islet tissue homeostasis. In the juvenile human pancreas, decreased levels of the ectonucleotidase NTPDase3 in beta cells may lead to increased levels of extracellular ATP in the islet. A large increase in ATP is an inflammatory signal and may lead to dysregulated immune responses that lead towards the beta cell-directed autoimmunity which emerges in early life. In the present study, we examine the functional phenotype of macrophages in pancreata from non-diabetic and type 1 diabetic (T1D) donors ranging from 3 months to 20 years old.
 

Methods

Living human pancreas slices were obtained from nPOD and incubated in HEPES solution (125 mmol/L NaCl, 5.9 mmol/L KCl, 2.56 mmol/L CaCl2, 1 mmol/L MgCl2, 25 mmol/L HEPES, 0.1% BSA, pH 7.4, and 10 mg/mL aprotinin). To visualize macrophages in situ, we used fluorescence conjugated antibodies for CD45 (1:50) and CD14 (1:50). To measure Ca2+ responses to stimuli, slices were incubated in the Ca2+ indicator dye Fluo-4 as well. Glucose was added to the buffered solution to give a basal glucose concentration of 3 mmol/L. All stimuli were bath applied. Confocal images were acquired in a Z-stack of 9-10 images every 3-5s using a Leica SP8 confocal laser-scanning microscope.
 

Summary of Results

In pancreata from young donors (3 months – 8 years), we observed that islet macrophages had a higher baseline activity and larger increases in intracellular Ca2+ in response to exogenous ATP. In contrast, macrophages from older individuals (12 – 20 years) had less activity at baseline and smaller responses to exogenous ATP.
 

Conclusions

The data from our study show that in young T1D and non-diabetic donors, macrophages are more active at baseline and respond strongly to ATP, as compared to macrophages from older donors. This may be due to increased availability of ATP signals in the juvenile pancreas. These results show that the islet resident macrophage undergoes functional changes throughout postnatal development. Moreover, they represent initial data that show the importance of measuring the functional maturation of immune cells and its relation to the development of T1D.