2mml glutamine

Islets were washed briefly with VERSENE solution (ThermoFisher
15040066), and dissociated using TrypLE Select Enzyme solution (ThermoFisher
12563011) at 37°C, with occasional trituration. For human islets, the
dissociated cells were purified into β cell and non-β cell
fractions using mouse anti-human NTPDase3 antibodies (Ectonucleotidases
antibodies hN3-B3S) and CELLection Pan Mouse IgG kit (ThermoFisher
11531D).^{35 (link)} Cells
were plated on sterilized glass shards and cultured overnight at 37°C
before experiments. For mouse cells, culture media was RPMI-1640
supplemented with 10% (v/v) fetal bovine serum (ThermoFisher A31605), 1%
(v/v) penicillin-streptomycin (Fisher Scientific 15070063), and 2 mM
L-glutamine (Fisher Scientific 25030081). For human cells, the media was
adjusted to contain 8 mM glucose.

Male wild-type C57BL/6 mice were purchased from Jackson Laboratory (Indianapolis, IN). The mice were maintained and bred in the Division of Laboratory Animal Resources at the University of Cincinnati Medical Center. All the animal experiments conformed to the Guidelines for the Care and Use of Laboratory Animals prepared by the National Academy of Sciences, published by the National Institutes of Health, and approved by the University of Cincinnati Animal Care and Use Committee (Animal Welfare Assurance Number: A3295-01). The mouse macrophage cell line RAW264.7 was purchased from American Type Culture Collection (ATCC), Rockville, MD. The macrophages were maintained in Dulbecco’s modified Eagle’s medium (DMEM; Sigma) containing 15% of fetal bovine serum (FBS; Sigma), 2mm L-glutamine (Gibco, USA), 100 u/ml penicillin and 100 u/ml streptomycin (Sigma). The macrophages were grown at 37°C with 5% CO₂ in fully humidified air. Culture medium was changed every 1–2 days. Subsequent passages were performed with a 0.025% Trypsin (Sigma) containing 0.02% EDTA for 10 min at 37 °C. The fourth passage macrophages were used for experiments in this study.