摘要
Background:Benzo[a]pyrene(B[a]P),a carcinogen pollutant produced by combustion processes,is present in the western diet with grilled meats.Chronic exposure of B[a]P in hepatocellular carcinoma(HCC)cells promotes metastasis rather than primary proliferation,implying an unknown mechanism of B[a]P-induced malignancy.Given that exosomes carry bioactive molecules to distant sites,we investigated whether and how exosomes mediate cancer-stroma communications for a toxicologically associated microenvironment.Method:Exosomes were isolated from B[a]P stimulated BEL7404 HCC cells(7404-100Bap Exo)at an environmental relevant dose(100 nmol/L).Lung preeducation animal model was prepared via injection of exosomes and cytokines.The inflammatory genes of educated lungs were evaluated using quantitative reverse transcription PCR array.HCC LM3 cells transfected with firefly luciferase were next injected to monitor tumor burdens and organotropic metastasis.Profile of B[a]P-exposed exosomes were determined by ceRNA microarray.Interactions between circular RNA(circRNA)and microRNAs(miRNAs)were detected using RNA pull-down in target lung fibroblasts.Fluorescence in situ hybridization and RNA immunoprecipitation assay was used to evaluate the“on-off”interaction of circRNA-miRNA pairs.We further developed an adenoassociated virus inhalation model to examine mRNA expression specific in lung,thereby exploring the mRNA targets of B[a]P induced circRNA-miRNA cascade.Results:Lung fibroblasts exert activation phenotypes,including focal adhesion and motility were altered by 7404-100Bap Exo.In the exosome-educated in vivo model,fibrosis factors and pro-inflammatory molecules of are up-regulated when injected with exosomes.Compared to non-exposed 7404 cells,circ_0011496 was up-regulated following B[a]P treatment and wasmainly packaged into 7404-100Bap Exo.Exosomal circ_0011496 were delivered and competitively bound to miR-486-5p in recipient fibroblasts.The down-regulation of miR-486-5p converted fibroblast to cancer-associated fibroblast via regulating the downstream of Twinfilin-1(TWF1)and matrix metalloproteinase-9(MMP9)cascade.Additionally,increased TWF1,specifically in exosomal circ_0011496 educated lungs,could promote cancer-stroma crosstalk via activating vascular endothelial growth factor(VEGF).These modulated fibroblasts promoted endothelial cells angiogenesis and recruited primary HCC cells invasion,as a consequence of a pre-metastatic niche formation.Conclusion:We demonstrated that B[a]P-induced tumor exosomes can deliver circ_0011496 to activate miR-486-5p/TWF1/MMP9 cascade in the lung fibroblasts,generating a feedback loop that promoted HCC metastasis.
基金
National Nature Science Foundation,Grant/Award Numbers:82173543,81902939
Innovative Research Team of High-level Local Universities in Shanghai,Grant/Award Number:SHSMU-ZLCX20211602
Key laboratory of the Ministry of Education Foundation,Grant/Award Number:2022-MEKLLC-MS-003
Sanming Project of Medicine in Shenzhen,Grant/Award Number:SZSM202311019
Shanghai Key Discipline of Public Health,Grant/Award Number:GWVI-11.1-20
Shanghai Science and Technology Development Funds,Grant/Award Number:23QA1405700。
作者简介
Wei Mu,https://orcid.org/0000-0003-2828-8096,contributed equally to this work;Correspondence:Yang Ge,School of Public Health,Shanghai Jiao Tong University School of Medicine,No.227,South Chongqing Rd,Shanghai 200025,P.R.China.Email:geyang19861026@icloud.com;Correspondence:Weiping Jia,Shanghai Clinical Center for Diabetes,Shanghai Sixth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine,No.600,Yishan Rd,Shanghai 200233,P.R.China.Email:wpjia@sjtu.edu.cn;Pengfei Gu,contributed equally to this work
