Shi Xiaoyan, Huang Hanwen, Zhou Min, Liu Yarong, Wu Hongfei, Dai Min
College of Pharmacy, Anhui University of Chinese Medicine, Hefei, China.
Anhui Key Laboratory for Research and Development of Traditional Chinese Medicine, Hefei, China.
Front Pharmacol. 2021 Nov 22;12:765482. doi: 10.3389/fphar.2021.765482. eCollection 2021.
Paeonol (Pae) is a natural phenolic compound isolated from Cortex Moutan, which exhibits anti-atherosclerosis (AS) effects. Our previous work demonstrated that gut microbiota plays an important role during AS treatment as it affects the efficacy of Pae. However, the mechanism of Pae in protecting against vascular fibrosis as related to gut microbiota has yet to be elucidated. To investigate the antifibrosis effect of Pae on AS mice and demonstrate the underlying gut microbiota-dependent mechanism. ApoE mice were fed with high-fat diet (HFD) to replicate the AS model. H&E and Masson staining were used to observe the plaque formation and collagen deposition. Short-chain fatty acid (SCFA) production was analyzed through LC-MS/MS. The frequency of immune cells in spleen was phenotyped by flow cytometry. The mRNA expression of aortic inflammatory cytokines was detected by qRT-PCR. The protein expression of LOX and fibrosis-related indicators were examined by western blot. Pae restricted the development of AS and collagen deposition. Notably, the antifibrosis effect of Pae was achieved by regulating the gut microbiota. LC-MS/MS data indicated that the level of SCFAs was increased in caecum contents. Additionally, Pae administration selectively upregulated the frequency of regulatory T (Treg) cells as well as downregulated the ratio of T helper type 17 (Th17) cells in the spleen of AS mice, improving the Treg/Th17 balance. In addition, as expected, Pae intervention can significantly downregulate the levels of proinflammatory cytokines IL-1β, IL-6, TNF-α, and IL-17 in the aorta, and upregulate the levels of anti-inflammatory factor IL-10, a marker of Treg cells. Finally, Pae's intervention in the gut microbiota resulted in the restoration of the balance of Treg/Th17, which indirectly downregulated the protein expression level of LOX and fibrosis-related indicators (MMP-2/9 and collagen I/III). Pae attenuated vascular fibrosis in a gut microbiota-dependent manner. The underlying protective mechanism was associated with the improved Treg/Th17 balance in spleen mediated through the increased microbiota-derived SCFA production. Collectively, our results demonstrated the role of Pae as a potential gut microbiota modulator to prevent and treat AS.
丹皮酚（Pae）是一种从牡丹皮中分离出的天然酚类化合物，具有抗动脉粥样硬化（AS）作用。我们之前的研究表明，肠道微生物群在AS治疗过程中发挥重要作用，因为它会影响Pae的疗效。然而，Pae预防血管纤维化与肠道微生物群相关的机制尚待阐明。为了研究Pae对AS小鼠的抗纤维化作用，并阐明潜在的肠道微生物群依赖性机制。给载脂蛋白E（ApoE）小鼠喂食高脂饮食（HFD）以复制AS模型。采用苏木精-伊红（H&E）染色和Masson染色观察斑块形成和胶原沉积。通过液相色谱-串联质谱（LC-MS/MS）分析短链脂肪酸（SCFA）的产生。通过流式细胞术对脾脏中免疫细胞的频率进行表型分析。通过实时定量聚合酶链反应（qRT-PCR）检测主动脉炎性细胞因子的mRNA表达。通过蛋白质印迹法检测赖氨氧化酶（LOX）和纤维化相关指标的蛋白表达。Pae抑制了AS的发展和胶原沉积。值得注意的是，Pae的抗纤维化作用是通过调节肠道微生物群实现的。LC-MS/MS数据表明，盲肠内容物中SCFAs水平升高。此外，给予Pae选择性上调了AS小鼠脾脏中调节性T（Treg）细胞的频率，并下调了辅助性T细胞17（Th17）细胞的比例，改善了Treg/Th17平衡。此外，正如预期的那样，Pae干预可显著下调主动脉中促炎细胞因子白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）和白细胞介素-17（IL-17）的水平，并上调抗炎因子白细胞介素-10（Treg细胞的标志物）的水平。最后，Pae对肠道微生物群的干预导致Treg/Th17平衡的恢复，这间接下调了LOX和纤维化相关指标（基质金属蛋白酶-2/9和I/III型胶原）的蛋白表达水平。Pae以肠道微生物群依赖性方式减轻血管纤维化。潜在的保护机制与通过微生物群衍生的SCFA产生增加介导的脾脏中Treg/Th17平衡改善有关。总的来说，我们的结果证明了Pae作为一种潜在的肠道微生物群调节剂在预防和治疗AS中的作用。
