Recombinant human CC16 inhibits senescence in human bronchial epithelial cells induced by cigarette smoke extract

Abstract

Objective To investigate the effect and mechanism of recombinant human CC16 protein (rhCC16) on cigarette smoke extract (CSE)-induced senescence of human bronchial epithelial cells (HBECs). Methods CCK-8 assay was used to evaluate the cell viability in HBECs after treated with 0%, 1%, 2.5%, 5%, 7.5% and 10% CSE and/or 0, 10, 100, 250 and 500 ng/mL rhCC16.β-galactosidase staining was used to observe the activity of the enzyme in control, CSE, and CSE+rhCC16 treated HBECs.The mRNA levels of P16 and P21 in above cell groups were detected by real-time fluorescence quantitative PCR (qPCR), and the protein levels of P16, P21, p-P53, P38, p-P38, ERK1/2 and p-ERK1/2 were detected with Western blot in control group, CSE and CSE+rhCC16 groups. Results Compared with the control group, 5% CSE stimulation for 72 h or the treatment of 500 ng/mL or lower dose of rhCC16 had no significant effect on cell viability of HBECs.Stimulation 5% CSE for 72 h resulted in significantly higher positive rate to β-galactosidase staining (P < 0.05), elevated mRNA and protein levels of P16 and P21(P < 0.05), and enhanced protein levels of p-P53, p-P38 and p-ERK1/2(P < 0.05) when compared with the control group.Compared with the simple CSE stimulation, 250 ng/mL rhCC16 treatment decreased positive rate to β-galactosidase staining (P < 0.05), reduced mRNA and protein levels of P16 and P21(P < 0.05) and protein levels of p-P53, p-P38 and p-ERK1/2(P < 0.05). Conclusion rhCC16 inhibits CSE-induced cellular senescence of HBECs, which may due to its suppression in P38 MAPK and ERK1/2 signaling pathway.

Keywords

• cellular senescence
• cigarette smoke extract
• cc16 protein
• p38 mapk/erk1/2