猪舍细颗粒物促进猪原代肺泡巨噬细胞向M1极化  

Fine Particulate Matter from Pig House Promote M1 Polarization in Porcine Primary Alveolar Macrophage

作　　者：沈家鲲 唐倩 崔洋洋 金晓明 李延森[1] 李春梅[1] SHEN Jiakun;TANG Qian;CUI Yangyang;JIN Xiaoming;LI Yansen;LI Chunmei(Research Center for Livestock Environmental Control and Smart Production,College of Animal Science and Technology,Nanjing Agricultural University,Nanjing 210095,China)

机构地区：[1]南京农业大学动物科技学院家畜环境控制与智慧生产研究中心,南京210095

出　　处：《畜牧兽医学报》2021年第6期1717-1726,共10页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(31772648,32072781);国家重点研发计划项目(2016YFD0500505)。

摘　　要：旨在研究猪舍细颗粒物(PM_(2.5))对猪原代肺泡巨噬细胞(PAMs)精氨酸代谢、炎症因子表达和极化标志物表达的影响。利用支气管肺泡灌洗方法,体外分离PAMs,细胞纯化后,利用Diff-quik染色和F4/80标记鉴定PAMs,并测定细胞活力。将纯化培养的PAMs分为对照组和PM_(2.5)处理组(50μg·mL^(-1)),继续培养4、8和12 h,测定细胞活力,收集细胞上清测定一氧化氮(NO)的含量,裂解PAMs测定精氨酸酶的活性,并采用RT-PCR检测PAMs炎症因子IL-1β、TNF-α和IL-10以及M1、M2型巨噬细胞标志物CD 80和CD 206的表达水平。结果显示,通过支气管肺泡灌洗方法成功分离培养了PAMs,并发现PAMs的细胞活力随着培养时间的延长逐渐降低(P<0.05)。PM_(2.5)处理PAMs,显著提高了细胞上清中NO含量(P<0.05),显著降低了细胞精氨酸酶的活性(P<0.01),并显著提高了细胞炎症因子IL-1β和TNF-αmRNA表达水平(P<0.01),降低了IL-10 mRNA表达水平(P<0.05)。另外,PM_(2.5)处理PAMs早期,显著提高了CD 80 mRNA表达水平(P<0.01),在后期显著降低CD 206 mRNA表达水平(P<0.01)。综上表明,猪舍PM_(2.5)促进了PAMs向M1表型极化,促进了炎症的发展。This experiment aimed to study the effects of fine particulate matter(PM_(2.5))from pig house on the arginine metabolism,expression of inflammatory cytokines and macrophage polarization markers in primary porcine alveolar macrophages(PAMs).Bronchoalveolar lavage was conducted to isolate PAMs.After PAMs were purified,Diff-quik staining were used to identify PAMs and F4/80 labeling were used to identity PAMs,and cell viability of PAMs was determined.The PAMs were further divided into the control and the PM_(2.5)(50μg·mL^(-1))groups,continuing cultured for 4,8,or 12 h to determine cell viability,the cellular supernatant was collected to detect the concentration of nitric oxide(NO),PAMs were lysed to detect the activity of arginase,and RT-PCR was used to detect the relative expression of cytokines IL-1β,TNF-αand IL-10,and the relative expression of M1 and M2 markers CD 80 and CD 206 in PAMs.The results showed that PAMs were successfully isolated by bronchoalveolar lavage and purified,the cell viability of PAMs gradually decreased when the culture time was increased(P<0.05).After PM_(2.5) treatment,the concentration of NO in the cellular supernatant was significantly increased(P<0.05)and the activity of arginase significantly decreased(P<0.01).The mRNA expression of inflammatory cytokines IL-1βand TNF-αwere significantly increased(P<0.01),and the mRNA expression of IL-10 was decreased(P<0.05).In the early stage of PM_(2.5) treatment,the mRNA expression of CD 80 was significantly increased(P<0.01),and the mRNA expression level of CD 206 was significantly decreased in the later stage(P<0.01).These results suggested that PM_(2.5) from pig house promotes PAMs to the M1 polarization and promotes the development of inflammation.

关 键 词：细颗粒物 猪肺泡巨噬细胞 巨噬细胞极化 

分 类 号：R122.2[医药卫生—环境卫生学] X513[医药卫生—公共卫生与预防医学]