蜂胶对细菌脂多糖刺激下奶牛乳腺上皮细胞炎症相关基因mRNA转录水平和紧密连接蛋白的影响

doi:10.11843/j.issn.0366-6964.2020.05.026

Abstract

Abstract: This study aimed to investigate the effects of ethanol extract of Chinese propolis (EECP) on transcript levels of inflammation-related genes and tight junctional permeability in bovine mammary epithelial cells stimulated by bacterial endotoxin (lipopolysaccharide,LPS). The total content of phenolic acids and flavonoids in the EECP was determined by Folin-phenol method and ALNO₃ colorimetry, respectively. Moreover, the inflammatory model of bovine mammary epithelial cells (MAC-T) was induced by bacterial lipopolysaccharide and the effect of EECP on the relative cell proliferation rate was detected via CCK-8 method. Real-time quantitative PCR (RT-qPCR) was used to evaluate LPS-induced bovine mammary epithelial cells inflammatory factors (IL-6, IL-8, TNF-α and IL-1β) relative mRNA transcription levels. To determine the effect of EECP on LPS-induced MAC-T cell, inflammation tight junction permeability. We used RT-qPCR to detect the relative mRNA transcription levels of tight junction proteins (occludin, ZO-1), and further used immunofluorescence to localize tight junction membrane proteins. The results showed that the content of total phenolic acids and total flavonoids in EECP were 106.35 mg GAE·g^-1 and 320.85 mg RE·g^-1, respectively. CCK-8 results showed that the safe concentration of EECP was 0-15 μg·mL ^-1, and can effectively increase the viability of MAC-T cells under LPS stimulation. LPS stimulation significantly increased the mRNA transcription levels of cellular inflammation-related factors IL-6, IL-8, TNF-α and IL-1β (P<0.001). However, the mRNA transcription of IL-6, IL-8, TNF-α and IL-1β were significantly reduced under pretreatment with 2.5-15.0 μg·mL^-1 EECP. Similarly, the LPS model group significantly inhibited (P<0.01) the mRNA transcription of tight junction proteins (ZO-1 and occludin), while the mRNA transcription of tight junction proteins was increased by EECP pretreatment (P<0.05). Immunofluorescence staining experiments also confirmed that EECP could alleviate LPS-induced mammary epithelial barrier dysfunction by up-regulating the expression of tight junction proteins occludin and ZO-1. The results confirmed that EECP has a good protective effect on bacterial lipopolysaccharide-induced inflammation of bovine mammary epithelial cells, which provides a basis for the use of Chinese propolis to prevent mastitis in dairy cows.

Key words: Chinese propolis, anti-inflammatory, tight junction, bovine mammary epithelial cells

CLC Number:

OU Aiqun, WANG Kai, WU Liming, LI Jianghong, PENG Wenjun. Effects of Propolis on Transcript Levels of Inflammation-related Genes and Tight Junction Proteins of Bovine Mammary Epithelial Cells Stimulated by Bacterial Lipopolysaccharide[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(5): 1149-1157.

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Effects of Propolis on Transcript Levels of Inflammation-related Genes and Tight Junction Proteins of Bovine Mammary Epithelial Cells Stimulated by Bacterial Lipopolysaccharide

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