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After a centrifugation at 100 g for 1 min, cells were recovered and washed with 2 ml HBSS with Ca 2+ and Mg 2+ medium (Wako Pure Chemical Industries) containing 14 mM glucose (Sigma-Aldrich) for 3 times, and then preincubated at 37 C for 1 hr in a shaker at 100 rpm (Taitec Bio-shaker BR-40LF, Saitama, Japan) before used for experiments