Hacat cell line

Human keratinocyte cell line (HaCaT cell line) was purchased from the China Center for Type Culture Collection (Wuhan, China). The HaCaT cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, HyClone, United States) containing 10% fetal bovine serum (FBS) and 1% penicillin and streptomycin in a humidified incubator with 5% CO₂ at 37°C.

Particulate matter in the air was sampled by an air sampler (Thermo Anderson G-2.5, Thermo Scientific, USA) in Lanzhou, Gansu province for 3 months. After the collection was completed, PM2.5 was obtained by filtering through an ultra-fine glass fiber net. After that, PM2.5 was sterilized, then freeze-dried and concentrated and stored at −80°C for later use. HaCaT cell line was purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). LBP was purchased from Natural Field Bio-Technique Co., Ltd (Shaanxi, China). DMEM medium was obtained from Hyclone (Beijing, China). Fetal Bovine Serum (FBS) was purchased from Si-Ji-Qing Biotechnology (Hangzhou, China). 2’,7’-Dichlorofluorescin diacetate was purchased from Sigma-Aldrich (St. Louis, MO, USA). Anti-LC3B and anti-P62 antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA). Anti-GRP78 and anti-CHOP antibodies were purchased from Beyotime (Shanghai, China). Ad-GFP-LC3B adenovirus, Annexin V-FITC/PI apoptosis detection kit, Mitochondrial membrane potential detection kit (JC-1) and the Ca²⁺ probe Fluo-4 AM were also purchased form Beyotime (Shanghai, China).

hBMSCs were purchased from American Type Culture Collection (ATCC, Manassas, USA). mBMSCs were obtained from Cyagen Biosciences Company (Guangzhou, China). These cells were cultured with a Mesenchymal Stem Cell Growth Kit for Bone Marrow-derived MSCs (ATCC PCS-500-041) (supplemented with rh FGF basic: 125 pg/mL, rh IGF-1: 15 ng/mL, 7% fetal bovine serum (FBS) and L-Alanyl-L-Glutamine: 2.4 mM). Hypoxic preconditioning was induced in an oxygen control incubator (Smartor 118, Zhejiang, China) filled with 5% CO₂ and 90–94% N₂. The oxygen concentrations and hypoxic incubation times were indicated as shown. Cells cultured under normoxic conditions (21% O₂) served as a control.
The human keratinocyte (HaCaT) cell line was purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China) (originally from American Type Culture Collection (Manassas, USA)) and was cultured in Minimum Essential Medium (MEM; Procell Life Science & Technology, PM150410) containing 10% FBS (Gibco, A3160802) and 1% penicillin and streptomycin (Gibco). HaCaT cells were treated with high glucose (HG) (25 mM glucose) and 1% O₂ hypoxia.