不同粒径聚苯乙烯微塑料和苯并(a)芘联合暴露影响炎性因子的表达

Co-exposure to different sized polystyrene microplastics and benzo[a]pyrene affected inflammation in zebrafish and bronchial-associated cells

研究发现,微塑料可吸附并富集持久性有机污染物,吸附污染物后的微塑料生物毒性增强.本文旨在探究不同粒径聚苯乙烯微塑料(polystyrene microplastics,PS-MPs)联合苯并(a)芘(benzo(a)pyrene,BaP)暴露,对成年斑马鱼鳃和支气管相关细胞炎性细胞因子表达的影响.以两种粒径的PS-MPs(0.5和5μm)分别与BaP联合暴露幼鱼7 d、成鱼14 d和细胞48 h.使用Real-time PCR测定不同暴露组炎性因子基因相对表达情况,体视镜观察微塑料颗粒进入幼鱼体内的情况,共聚焦显微镜观察微塑料颗粒进入细胞的情况.结果发现,粒径为0.5和5μm的PS-MPs均能进入幼鱼体内;0.5μm PS-MPs单独暴露时,TNF-α、IL-6和IL-8基因相对表达显著高于对照组;5μm PS-MPs单独暴露时,IFN-γ基因相对表达显著高于对照组.BaP和0.5μm PS-MPs联合暴露时,成鱼鳃组织IFN-γ、IL-6和IL-8基因的相对表达高于BaP单独暴露.细胞实验结果表明,仅0.5μm的PS-MPs能进入两种细胞胞质中.在人支气管上皮样细胞HBE中0.5μm PS-MPs组炎症因子TNF-α、IL-6和IL-8基因相对表达均高于对照组.在人支气管平滑肌细胞HBSMC中0.5μm PS-MPs组炎性细胞因子TNF-α、IL-8和IL-1β的相对表达显著高于对照组.HBE细胞中BaP分别与0.5μm/5μm微塑料联合暴露时,均未发现明显的联合作用.HBSMC细胞中BaP和0.5μm PS-MPs联合暴露时,对IFN-γ和IL-6炎症因子的表达有显著的联合作用;而BaP和5μm PS-MPs联合暴露时,有显著减弱BaP促炎作用的趋势,特别是对TNF-α和IL-8炎症因子表达的影响.由此推断,粒径可能是影响微塑料生物摄取和累积的主要因素之一,0.5μm微塑料因其易被细胞摄取,故联合有机污染物暴露可加重炎症反应.另外,因5μm微塑料未能进入细胞,其对炎症反应的影响仍需进一步探究.

In recent years,emerging research has revealed that microplastics(MPs)can pollute the air.Cellulose and plastic fibers have been found in the lungs of patients with lung cancer,indicating that MPs can"travel"through the atmosphere into the respiratory tract.Due to the hydrophobicity and relatively large surface area of MPs,the biotoxicity of MPs is greatly enhanced by their adsorption and enrichment in persistent organic pollutants(POPs).With the rapid development of the economy,emissions from coal burning and automobile exhaust have made polycyclic aromatic hydrocarbon(PAHs)one of the key pollutants in urban air that cannot be ignored.Epidemiological studies have shown that PAH exposure is associated with the development of respiratory diseases,such as bronchitis and asthma,in children.The aim of this study was to investigate the inflammatory effects of co-exposure to different size polystyrene MPs(PS-MPs)and benzo[a]pyrene(Ba P)on adult zebrafish gills and bronchial-related cells to provide basic information for assessment of the potential health risks of combined exposure to the human respiratory system.PS-MPs(with size of 0.5 and 5μm)and Ba P were co-administered to embryos for 7 d,adult zebrafish for 14 d,and HBE cells and HBMSCs for 48 h.The relative expression levels of inflammatory cytokine genes in the different exposure groups were determined by real-time polymerase chain reaction(PCR).The ingestion of microplastics into fish bodies and cells was observed by stereoscope and confocal microscopy,respectively.Result shows that microplastics of two sizes could be ingested by zebrafish embryos.When exposed only to0.5-μm PS-MPs,relative expression of the TNF-α,IL-6,and IL-8 genes was significantly increased compared with that in the control;relative expression of the IFN-γgene was largely increased in the gills of zebrafish in 5-μm PS-MP-exposed group.Moreover,relative expression of the IFN-γ,IL-6 and IL-8 genes in the group co-exposed to Ba P and 0.5-μm PS-MPs was higher than that in Ba P group.In addition,0.5-μm PS-MPs could be ingested by both cell lines.Upon exposure to 0.5-μm PS-MPs alone,relative expression of genes TNF-α,IL-6,and IL-8 was considerably increased compared with that in the control HBE cells.In HBSMCs,upon exposure to 0.5-μm PS-MPs alone,relative expression of the TNF-α,IL-8,and IL-1βgenes was substantially increased compared with that in the control group.Compared with that following exposure to Ba P alone,relative expression of the IFN-γand IL-6 genes in HBSMCs in 0.5-μm PS-MPs and Ba P co-exposure group was remarkably increased,indicating a possible combined effect;however,co-exposure of 5-μm PS-MPs and Ba P weakened the proinflammatory function of Ba P and especially influenced relative expression of the TNF-αand IL-8 genes.In addition,none of the co-exposure groups of HBE cells showed obvious combined effects.The corollary is that particle size may affect the ingestion of PS-MPs in cells.In addition,the size-dependent effects of PS-MPs influence the accumulation of organic pollutants and their biotoxicity.