NB1RGB (human skin fibroblast) (Riken BRC) and IMR90 (human lung fibroblast) (JCRB Cell Bank) cells were maintained in MEMα (Wako, Tokyo, Japna) supplemented with 10% FBS, 100 U/ml penicillin, and 100 μg/ml streptomycin. Cell cultures were passaged (1:4, every 3 days) and maintained at 37°C under 5% CO2. We defined two types of each cell line according to the number of days in culture: NB1RGB: young cells: from 8 to 20 days and aging cells: from 60 to 70 days; IMR90: young cells: from 6 to 12 days and aging cells: from 30 to 40 days34 (link)
NB1RGB cells were transiently transfected using lipofection (Lipofectamine 3000, Invitrogen, Carlsbad, CA, USA) with plasmids expressing SOD2 (pcDNA6.2-SOD2, cloned from pENTR221-SOD2 (DNAFORM ID 100008212) using Gateway cloning system (Invitrogen, Carlsbad, CA, USA).)
NB1RGB cells were transiently transfected using lipofection (Lipofectamine 3000, Invitrogen, Carlsbad, CA, USA) with plasmids expressing SOD2 (pcDNA6.2-SOD2, cloned from pENTR221-SOD2 (DNAFORM ID 100008212) using Gateway cloning system (Invitrogen, Carlsbad, CA, USA).)
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