猪舍细颗粒物促进猪原代肺泡巨噬细胞向M1极化

doi:10.11843/j.issn.0366-6964.2021.06.026

摘要/Abstract

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（50 μg·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CD80和CD206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-α mRNA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CD80 mRNA表达水平（P<0.01），在后期显著降低CD206 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。

关键词: 细颗粒物, 猪肺泡巨噬细胞, 巨噬细胞极化

Abstract: 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 (50 μg·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-α and IL-10, and the relative expression of M1 and M2 markers CD80 and CD206 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-1β and TNF-α were 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 CD80 was significantly increased (P<0.01), and the mRNA expression level of CD206 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.

Key words: fine particulate matter, porcine alveolar macrophages, macrophage polarization

中图分类号:

R122.2
X513

沈家鲲, 唐倩, 崔洋洋, 金晓明, 李延森, 李春梅. 猪舍细颗粒物促进猪原代肺泡巨噬细胞向M1极化[J]. 畜牧兽医学报, 2021, 52(6): 1717-1726.

SHEN Jiakun, TANG Qian, CUI Yangyang, JIN Xiaoming, LI Yansen, LI Chunmei. Fine Particulate Matter from Pig House Promote M1 Polarization in Porcine Primary Alveolar Macrophage[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(6): 1717-1726.

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