自噬
PI3K/AKT/mTOR通路
巨噬细胞极化
蛋白激酶B
污渍
CD86
巨噬细胞
免疫印迹
肿瘤坏死因子α
分子生物学
M2巨噬细胞
分泌物
细胞生物学
医学
生物
信号转导
免疫学
细胞凋亡
内科学
生物化学
体外
免疫系统
基因
T细胞
作者
Yingyu Xie,Lijun Tian,Zihan Fang,Aimei Zhong,Ao Zhang,Shixin Xu,Yanan Wang,Junping Zhang
摘要
Objective To investigate the efficacy of Bushen Kangshuai (BS-KS) tablet on autophagy and polarization in mouse macrophage RAW 264.7. Meyhods Macrophage autophagy was induced by oxidized low-density lipoprotein (100 μg/mL). To detect the levels of autophagy, macrophage were transfected with double fluorescence LC3 autophagy adenovirus, then the numbers of autophagosomes and autophagic lysosomes were asessed by confocal microscopy. The autophagy related proteins expression of PI3K, Akt, phospho-mAkt (p-Akt) and mTOR, phospho-mTOR ([p-TOR), p62, microtubule-associated protein 1 (LC3-Ⅱ)were determined by western blotting. The macrophage polarization model was induced by lipopolysaccharide (1 μg/mL). The mRNA levels of iNOS, CD86 (M1 macrophages marker molecules), and CD206, Arg-1 (M2 macrophages marker molecules) were detected by real-time quantitative PCR. The concentration of cytokines TNF-α and IL-10 was determined by enzyme-linked immunosorbent assay. The protein expression of nuclear proteins PPAR-γ, NF-κB, and cytoplasmic protein IKB α was determined by western blotting. Results The expression of the autophagy-related protein LC3-Ⅱ was increased and the expression of p62 was decreased in the BS-KS intervention group. The protein expression of PI3K, p-Akt, and p-mTOR was also reduced. BS-KS also inhibited the mRNA expression of iNOS and CD86 on M2 macrophage, but promoted the expression of CD206 and Arg-1 on M2 macrophage. With respect to the regulation of inflammatory factors, BS-KS could inhibit the secretion of pro-inflammatory TNF-α and promote the secretion of anti-inflammatory IL-10. It also inhibited the protein expression of IKB-α and NF-κB, and promoted the expression of nuclear protein PPAR-γ. Conclusion We believe that BS-KS promotes macrophage autophagy by increasing the level of autophagy protein and inhibiting the PI3K/Akt/mTOR signaling pathway. Furthermore, BS-KS seems to inhibit macrophage M1 polarization and promote M2 polarization via the PPAR gamma /NF-κB signaling pathway, thus playing an inhibitory role in atherosclerosis.
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