M1902

MSCs were trypsined without Ethylenediaminetetraacetic acid (EDTA) (Gibco 15090046) and then mechanically lysed in a buffer containing mannitol (200 mmol/L; Sigma-Aldrich M1902), saccharose (70 mmol/L; Sigma-Aldrich S0389), EDTA (1 mmol/L; Sigma-Aldrich E9884), HEPES (10 mmol/L, pH = 7.4; Sigma-Aldrich H3375) and 1X cocktail of protease and phosphatase inhibitors (Roche 04693159001), by using a syringe with 25- and 27-gauge needles. MSC mitochondria were isolated by two differential centrifugations at 800 × g and 8,000 × g, respectively. All steps of the mitochondria isolation were carried out at 0°C and the mitochondria pellet was finally resuspended in ice-cold mannitol buffer.

Human proximal tubular epithelial cells (human kidney 2 [HK-2]) were procured from the BeNa Culture Collection (cat: BNCC339833, China). These cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM, cat: C11995500, Invitrogen, USA) supplemented with 10% fetal bovine serum (cat: 16-000-044, Invitrogen, USA), and 1% penicillin/streptomycin (cat: 1,510,122, Invitrogen, USA). The cells were maintained in a humidified incubator with 5% carbon dioxide at 37℃. HK-2 cells were treated with HG (30 mM glucose, cat: G-8769, Sigma-Aldrich, USA) for 12 h, 24 h, and 36 h. The control group was exposed to normal glucose (NG, 5.5 mM glucose + 24.5 mM mannitol, cat: M-1902, Sigma-Aldrich, USA).

Neutrophil isolation kit (17957, STEMCELL, Vancouver, Canada); Bovine serum albumin (A1933-25G, Sigma-Aldrich, St. Louis, MO, USA), FBS (iCell-0500, iCell Bioscience Inc, Shanghai, China); RPMI 1640 (SH30809.01, Hyclone, Logan, UT, USA); Fluorescein (FITC-dextran, 10 kDa; final concentration 5 μM, Sigma-Aldrich, St. Louis, MO, USA); fMLP (F3506, 1 mM, Sigma-Aldrich, St. Louis, MO, USA); Triton X-100 (9036-19-5, Sigma, St. Louis, MO, MO, USA); Giemsa (G5637, Sigma, St. Louis, MO, USA); D-glucose (G7021, Sigma-Aldrich, St. Louis, MO, USA); mannitol (10 mM, an osmotic control reagent, M1902, Sigma-Aldrich, St. Louis, MO, USA); Fibronectin (F2006, 2 μg mL⁻¹, Sigma-Aldrich, St. Louis, MO, USA); Commercial magnetic Neutrophil isolation kit (17957, STEMCELL Technologies, Inc., Toronto, Canada); Precursors and initiators of Polydimethylsiloxane (PDMS) (Dow Corning, Midland, MI, USA); Glass slide (80350-0001, Jiangsu Shitai Xinchuang Scientific Instrument Co., Ltd., Nanjing, China); Microscope (DMi8, Leica, Wezler, Hesse, Germany); Plasma cleaner (PDC-002, Harrick Scientific Products, Inc., New York, NY, USA).