IGFBP-3 Expression In Alveolar Epithelial Cells Injury Induceded By Cigarette Smoke Extract

Objective To investigate the expression of insulin-like growth factor binding protein-3(IGFBP-3) in cigarette smoke extract (CSE) induced alveolar epithelial cell and analyze the potential protective effect of N-acetylcysteine(NAC) and Sulforaphane (SFN).Methods Rat type Ⅱ alveolar epithelial cells (RLE-6TN cell line) was maintained in vitro.5%,10% and 20% CSE were prepared and used to treat the cell line for 24 h. MTT assay was employed to evaluate cell viability after treatment with CSE or 1,5,10,20,40,80 and 160 μM SFN, The potential protective effect of SFN (5 μM) or NAC (1 mM) was also observed by MTT assay. Moreover, flow cytometry was performed to analyze the cell cycle changes in RLE-6TN cells after 5% CSE treatment and SFN or NAC co-treatment. Finally, real-time fluorescence quantitative RT-PCR (RT-qPCR) and western blot were performed to analyze mRNA and protein level of IGFBP-3 and IGFBP-5.Results CSE could induce significant inhibition of RLE-6TN cells in a dose-dependent manner. Both 5μM SFN and 1 mM NAC could reduce cell injury induced by CSE. As the result of cell cycle analysis,5% CSE could cause G0/G1 arrest of RLE-6TN cells, more interestingly, SFN or NAC antagonized against the G0/G1 arrest induced by CSE. With the treatment of CSE, Increased IGFBP-3 mRNA level was observed, however,5μM SFN and 1 mM NAC could decrease it.Corresponding to the result of mRNA, western blot showed an increased expression of IGFBP-3 after 5% CSE exposure and decreased IGFBP-3 was observed after SFN or NAC co-treatment, on the contrary 5% CSE could decrease IGFBP-5mRNA level and this effect could be antagonized by SFN and NAC. However, there was no significant expression changes in IGFBP-5 protein level.Conclusions CSE may induce inhibition of cell cycle of alveolar epithelial cells by up-regulating IGFBP-3, SFN and NAC could antagonize against the injury by down-regulating the expression of IGFBP-3.