Study on Molecular Mechanism of Benzo (ɑ) pyrene on CMA by HSP90ɑ and HIF-1ɑ

Abstract

OBJECTIVE: The effects of benzo (α) pyrene (BaP) on molecular chaperone autophagy (CMA) through heat shock protein 90 (HSP90) and hypoxia inducible factor-1 (HIF-1) were studied by RNA interference and subcutaneous tumor formation technique in nude mice. METHODS: 40 nude mice inoculated with the shHSP90ɑ A549 cell line under the armpits of the forelimbs were divided into 4 groups, 1.80 mg/kg/d BaP-corn oil solution was intragastrically administered for 60 days (except the Control group), and the growth of nude mice and transplanted tumors was recorded curve. The size and morphological changes of tumors were observed by small animal imaging technique and HE staining method. qPCR, Western blot and Immunohistochemistry were used to detect the expression of HSP90ɑ, HSC70 and Lamp-2A. A549 cells were treated with 0.1μmol/L, 1μmol/L and 10μmol/L BaP for 24h, EPO, HIF-1ɑ concentration and HIF-1ɑ protein expression were detected by Elisa and Western blot; A549 cells were treated with 10μmol/L BaP and added HIF-1ɑ inhibitor 24h, qPCR, Western blot and Immunofluorescence method were used to detect the expression of HSP90ɑ, HSC70 and Lamp-2A. RESULTS: BaP can reduce silence HSP90ɑ the weight of nude mice and transplanted tumors (P<0.01); BaP can reduce silence HSP90ɑ the expression of HSP90ɑ, HSC70, Lamp-2A mRNA and protein in transplanted tumor tissues (P<0.05); BaP has no significant difference in the organization structure of silence HSP90ɑ and non-silence HSP90ɑ; BaP can reduce the total number of bioluminescence photons of silence HSP90ɑ transplanted tumors (P<0.01). 10μmol/L BaP can increase the concentration of EPO and HIF-1ɑ and the expression of HIF-1ɑ protein in A549 cells (P<0.05); and after adding HIF-1ɑ inhibitors treat A549 cells, HSP90ɑ, HSC70, Lamp-2A mRNA and protein expression were decreased (P<0.05), and the fluorescence intensity of HSP90ɑ was decreased. CONCLUSIONS: BaP can promote the growth of transplanted tumor in nude mice, while the growth of transplanted tumor in nude mice is inhibited shHSP90ɑ. BaP promotes the occurrence of CMA by promoting the expression of HSP90ɑ and HIF-1ɑ, which are important regulatory genes for BaP to activate CMA.