丹参对巨噬细胞中Toll样受体2触发的炎症反应的影响。
Effect of Danshen on TLR2-triggered inflammation in macrophages.
作者信息
Ye Tingting, Xiong Difeifei, Chen Libing, Li Yufei, Gong Shuqing, Zhang Luquan, Li Bailing, Pan Jianyang, Qian Jing, Qu Haibin
机构信息
Pharmaceutical Informatics Institute, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, China.
Guizhou Baite Pharmaceutical co., Ltd., Guizhou, China.
出版信息
Phytomedicine. 2020 Apr 20;70:153228. doi: 10.1016/j.phymed.2020.153228.
BACKGROUND
Danshen (Salvia Miltiorrhiza Radix et Rhizoma) is a valued herbal plant widely used to treat cardiovascular diseases in Asian countries. In modern medicine, innate immunity-induced inflammation is considered a risk factor for cardiovascular diseases. However, little is known about the anti-inflammatory effects and molecular mechanism of Danshen.
PURPOSE
To evaluate the molecular mechanisms of Danshen on Toll-like receptor (TLR) 2-triggered inflammation in macrophages and identify its bioactive components.
METHODS
Pam3CSK4-stimulated bone marrow-derived macrophages (BMMs) were treated with Danshen water extract (DSE), and the levels of proinflammatory cytokines (interleukin (IL)-6, IL-12 and tumor necrosis factor (TNF)-α) were measured by both real-time quantitative PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). RNA sequencing (RNA-seq)-based bioinformatics analyses were applied to reveal the novel molecular mechanisms of DSE, followed by western blotting for verification. Additionally, HPLC-UV analysis along with bioassays was performed to identify the bioactive ingredients of DSE.
RESULTS
The results of RT-qPCR and ELISA showed that DSE significantly inhibited proinflammatory cytokine expression in a dose-dependent manner. Transcriptome analyses revealed that a wider panel of inflammatory cytokines responded to the regulatory effect of DSE, and that the TNF signaling pathway might have played a vital role. Western blotting data confirmed the involvement of extracellular signal-regulated protein kinases (ERK) and Jun N-terminal Kinase (JNK) related singling pathway. Among the seven components identified in DSE, Danshensu (DSS) and protocatechuic aldehyde (PA) were confirmed as bioactive ones with anti-inflammatory effects.
CONCLUSION
DSE showed a promising effect against TLR2-triggered inflammation associated with the inhibition of the TNF cascade down-streamed mitogen-activated protein kinase (MAPK) signaling pathway, in which IL-6 acts as the key effective molecule, and ERK and JNK phosphorylation was inhibited. Notably, DSS and PA were considered bioactive components with anti-inflammatory bioactivity.
背景
丹参是一种珍贵的草药，在亚洲国家被广泛用于治疗心血管疾病。在现代医学中，先天免疫诱导的炎症被认为是心血管疾病的一个危险因素。然而，关于丹参的抗炎作用及其分子机制知之甚少。
目的
评估丹参对巨噬细胞中Toll样受体（TLR）2触发的炎症的分子机制，并鉴定其生物活性成分。
方法
用丹参水提取物（DSE）处理Pam3CSK4刺激的骨髓来源巨噬细胞（BMM），通过实时定量PCR（RT-qPCR）和酶联免疫吸附测定（ELISA）测量促炎细胞因子（白细胞介素（IL）-6、IL-12和肿瘤坏死因子（TNF）-α）的水平。应用基于RNA测序（RNA-seq）的生物信息学分析来揭示DSE的新分子机制，随后进行蛋白质印迹法验证。此外，进行高效液相色谱-紫外分析以及生物测定以鉴定DSE的生物活性成分。
结果
RT-qPCR和ELISA结果表明，DSE以剂量依赖性方式显著抑制促炎细胞因子表达。转录组分析显示，更广泛的炎性细胞因子对DSE的调节作用有反应，并且TNF信号通路可能起了关键作用。蛋白质印迹数据证实细胞外信号调节蛋白激酶（ERK）和Jun N末端激酶（JNK）相关信号通路参与其中。在DSE中鉴定出的七种成分中，丹参素（DSS）和原儿茶醛（PA）被确认为具有抗炎作用的生物活性成分。
结论
DSE对TLR2触发的炎症显示出有前景的作用，这与抑制TNF级联下游的丝裂原活化蛋白激酶（MAPK）信号通路有关，其中IL-6是关键的有效分子，并且ERK和JNK磷酸化受到抑制。值得注意的是，DSS和PA被认为是具有抗炎生物活性的生物活性成分。
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