毛冬青三萜皂苷对动脉粥样硬化大鼠粪便和尿液代谢组学的影响

白荣钰; 易欢; 邱静文; 陈丰连; 王莹; 陈冰莹; 李瑜; 张蕾

doi:10.14148/j.issn.1672-0482.2022.0424

毛冬青三萜皂苷对动脉粥样硬化大鼠粪便和尿液代谢组学的影响

doi: 10.14148/j.issn.1672-0482.2022.0424

白荣钰^1,,
易欢¹,
邱静文¹,
陈丰连¹,
王莹¹,
陈冰莹¹,
李瑜²,
张蕾^1, ,

1.
广州中医药大学中药学院, 广东广州 510006
2.
广州中医药大学护理学院, 广东广州 510006

基金项目:

广东省教育厅创新强校工程项目 2016KTSCX019

广东省科技计划项目 2016A020226031

广州市科技局一般科学研究项目 201607010334

详细信息

作者简介:
白荣钰, 女, 硕士研究生, E-mail: bairongyu2021@163.com

通讯作者:
张蕾, 女, 教授，主要从事中药药效物质基础及其作用机制研究, E-mail: zhangleic431@163.com

中图分类号: R285.5
计量
- 文章访问数: 171
- HTML全文浏览量: 46
- PDF下载量: 12
- 被引次数: 0
出版历程
- 收稿日期: 2021-11-18
- 发布日期: 2022-05-10

Effects of Ilex Pubescens Triterpenoid Saponins on Fecal and Urine Metabolomics in Atherosclerotic Rats

1.
School of Pharmaceutical Science, Guangzhou University of Chinese Medicine, Guangzhou 510006, China
2.
School of Nursing, Guangzhou University of Chinese Medicine, Guangzhou 510006, China

摘要

摘要: 目的探讨毛冬青三萜皂苷(IPTS)对动脉粥样硬化(AS)模型大鼠粪便和尿液代谢组学的影响。方法采用高脂饮食联合腹腔注射维生素D₃诱导大鼠AS模型, 实验分为对照组、模型组、IPTS组。检测大鼠血清中血脂四项水平, 应用核磁共振(NMR)技术对粪样和尿样进行代谢组学分析。结果 IPTS干预后, 大鼠血清TG含量显著下降(P < 0.05), HDL-C含量显著升高(P < 0.05)。代谢组学分析结果表明, 模型组大鼠中上调的差异代谢产物主要包括氨基酸类(亮氨酸、酮亮氨酸、胍基乙酸酯)和肠道菌群相关代谢物(胆碱、甜菜碱、氧化三甲胺等), 下调的代谢产物包括三羧酸循环产物(琥珀酸盐、富马酸盐和戊二酸)、核苷酸类(腺嘌呤、黄嘌呤等)以及氨基酸类(色氨酸、犬尿氨酸等), 经IPTS干预后, 部分三羧酸循环产物、氨基酸和核苷酸代谢产物以及肠道菌群代谢物得到了回调。结论 IPTS对部分粪便和尿液差异代谢物有调节改善作用, 对AS大鼠的代谢紊乱一定的预防和改善作用。
- 毛冬青 /
- 三萜皂苷 /
- 动脉粥样硬化 /
- 代谢组学 /
- 粪便 /
- 尿液
Abstract: OBJECTIVE To investigate the effect of Ilex pubescens triterpene saponins (IPTS) on fecal and urine metabolomics in atherosclerotic (AS) rats. METHODS AS model rats were induced by high-fat diet combined with intraperitoneal injection of vitamin D₃. The rats were divided into control group, model group and IPTS group. The contents of total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) in the serum were tested. The fecal and urine samples of the rats were taken for metabolomic analysis using nuclear magnetic resonance (NMR) technology. RESULTS After administration of IPTS, the content of TG significantly decreased (P < 0.05), and the content of HDL-C significantly increased (P < 0.05) in rat serum. The result of metabonomics analysis showed that the differential metabolites up-regulated in the model group mainly included amino acids (leucine, ketooleucine, guanidoacetate) and intestinal flora-related metabolites (choline, betaine, trimethylamine oxide, etc.). The down-regulated metabolites included the tricarboxylic acid cycle products (succinate, fumarate and glutarate), nucleotides (adenine, scutellaria, etc.) and amino acids (tryptophan, kynurenine, etc.), which showed that the energy metabolism, amino acid metabolism, nucleotide metabolism and intestinal flora-host co-metabolism of the model group rats were disturbed. After administration of IPTS, some products of TCA cycle, nucleotide metabolites, amino acid metabolites, and metabolites of gut microbiota were corrected, which indicated IPTS could ameliorate the metabolic disorders caused by atherosclerosis. CONCLUSION IPTS can partly regulate the differential metabolites of fecal and urine samples in AS rats and have a good therapeutic effect on AS rats.
- Ilex pubescens /
- triterpene saponins /
- atherosclerosis /
- metabolomics /
- feces /
- urine

HTML全文

图 1 各组大鼠血清中TC、TG、LDL-C、HDL-C含量

注: 与对照组比较, ^##P < 0.01;与模型组比较, ^*P < 0.05。x±s, n=10。

Figure 1. Serum contents of TC, TG, LDL-C and HDL-C in rats

下载: 全尺寸图片幻灯片

图 2 对照组与模型组粪样¹H-NMR谱图的PCA得分图(A)、OPLS-DA得分图(B)、PLS-DA得分图(C)以及PLS排列实验验证图(D)

Figure 2. PCA scores plots (A), OPLS-DA scores plots (B), PLS-DA scores plots (C), and PLS permutation test plots (D) for the ¹H-NMR data of fecal samples between control and model groups

下载: 全尺寸图片幻灯片

图 3 对照组与模型组尿样¹H-NMR谱图的PCA得分图(A)、OPLS-DA得分图(B)、PLS-DA得分图(C)以及PLS排列实验验证图(D)

Figure 3. PCA scores plots (A), OPLS-DA scores plots (B), PLS-DA scores plots (C), and PLS permutation test plots (D) for the ¹H-NMR data of urine samples between control and model groups

下载: 全尺寸图片幻灯片

图 4 IPTS组与模型组的粪样¹H-NMR谱图的PCA得分图(A)、OPLS-DA得分图(B)、PLS-DA得分图(C) 以及PLS排列实验验证图(D)

Figure 4. PCA scores plots (A), OPLS-DA scores plots (B), PLS-DA scores plots (C), and PLS permutation test plots (D) for the ¹H-NMR data of fecal samples between IPTS and model groups

下载: 全尺寸图片幻灯片

图 5 IPTS组与模型组的尿样¹H-NMR谱图的PCA得分图(A)、OPLS-DA得分图(B)、PLS-DA得分图(C) 以及PLS排列实验验证图(D)

Figure 5. PCA scores plots (A), OPLS-DA scores plots (B), PLS-DA scores plots (C), and PLS permutation test plots (D) for the ¹H-NMR data of urine samples between IPTS and model groups

下载: 全尺寸图片幻灯片

图 6 造模后的粪样(A)和尿样(B)代谢通路图

Figure 6. Metabolomics pathway analysis for fecal(A) and urine(B) samples after modeling

下载: 全尺寸图片幻灯片

图 7 IPTS干预后的粪样(A)和尿样(B)代谢通路图

Figure 7. Metabolomics pathway analysis for fecal(A) and urine(B) samples after the intervention of IPTS

下载: 全尺寸图片幻灯片

图 8 造模后差异代谢物参与的代谢通路图

注：↑.上调; ↓.下调; 黑色字体.未检测到的代谢物; 绿色字体.检测到的差异代谢物

Figure 8. Involved metabolomics pathway analysis of differential metabolites after modeling

下载: 全尺寸图片幻灯片

图 9 IPTS干预后差异代谢物参与的代谢通路图

注：↑.上调; ↓.下调; 黑色字体.未检测到的代谢物; 绿色字体.与造模后检测到的差异代谢物相同; 棕色字体.得到纠正的代谢物; 黄色字体.给药后引起新变化的代谢物

Figure 9. Involved metabolomics pathway analysis of differential metabolites after the intervention of IPTS

下载: 全尺寸图片幻灯片

表 1 IPTS浓度测定标准曲线

Table 1. Standard curve line of concentration determination of IPTS

浓度/(mg·mL^-1)	吸光度	浓度/(mg·mL^-1)	吸光度
0.000 68	0.193	0.003 40	1.203
0.001 36	0.452	0.005 10	1.837
0.002 72	0.960	0.006 80	2.459

下载: 导出CSV

表 2 大鼠粪样和尿样中差异代谢物汇总

Table 2. The summary of the differential metabolites between the feces and urine samples in rats

代谢通路	差异代谢物	模型组vs.对照组		IPTS组vs.模型组
代谢通路	差异代谢物	粪便	尿液	粪便	尿液
核苷酸代谢(Nucleotide metabolism)	腺嘌呤(Adenine)	↓^*
	次黄嘌呤(Hypoxanthine)	↓^***			↓^**
	黄嘌呤(Xanthine)	↓^***			↑^*
	黄嘌呤核苷(Xanthosine)	↓^***
	鸟嘌呤(Guanine)				↓^*
	胞嘧啶(Cytosine)				↑^*
肠道菌群宿主共代谢(Gut microbiota and host co-metabolism)	丁酸盐(Butyrate)	↓^**		↑^*
	胆碱(Choline)	↑^**
	丙酸盐(Propionate)	↓^**
	三甲胺(TMA)	↓^*
	氧化三甲胺(TMAO)		↑^***		↓^*
	戊酸盐(Valerate)	↓^**		↑^*
	甲酸盐(Formate)		↑^*	↓^*
	乙酸盐(Acetate)			↓^**	↓^**
	丙酮(Acetone)		↑^**
	甜菜碱(Betaine)		↑^***
	肉碱(Carnitine)		↓^***		↓^*
	肌酸(Creatine)		↓^**
	马尿酸盐(Hippurate)		↓^***		↑^*
	乙酸苯酯(Phenylacetate)		↑^***	↓^*
氨基酸代谢(Amino acids metabolism)	丙氨酸(Alanine)	↑^**
	瓜氨酸(Citrulline)	↓^*
	亮氨酸(Leucine)	↑^***
	胍基乙酸酯(Guanidoacetate)	↑^**
	4-氨基丁酸酯(4-Aminobutyrate)		↓^**		↑^**
	吲哚-3-乙酸(Indole-3-acetate)		↓^**		↑^*
	酮亮氨酸(Ketoleucine)		↑^***
	犬尿氨酸(Kynurenine)		↓^***		↑^*
	色氨酸(Tryptophan)		↓^***
	缬氨酸(Valine)		↓^**
	β-丙氨酸(β-Alanine)		↑^***
	二甲基甘氨酸(Dimethylglycine)		↑^**
	甘氨酸(Glycine)		↑^***
能量代谢(Energy metabolism)	富马酸盐(Fumarate)	↓^***		↑^***
	戊二酸(Glutarate)	↓^**
	琥珀酸盐(Succinate)	↓^**	↓^**
	牛磺胆酸(Taurocholic acid)	↑^*
	2-氧戊二酸(2-Oxoglutarate)		↓^**		↑^***
	柠檬酸盐(Citrate)				↑^**
	乳酸(Lactate)	↑^*
注: 2组间比较，^P < 0.05, ^{ }P < 0.01, ^{ * *}P < 0.001。

下载: 导出CSV

参考文献(18)

[1]	MOORE KJ, KOPLEV S, FISHER EA, et al. Macrophage trafficking, inflammatory resolution, and genomics in atherosclerosis: JACC macrophage in CVD series (part 2)[J]. J Am Coll Cardiol, 2018, 72(18): 2181-2197. doi: 10.1016/j.jacc.2018.08.2147
[2]	梅丽, 牛瑞娟, 蒋玲, 等. 毛冬青化学成分及药理活性研究进展[J]. 生物化工, 2018, 4(2): 129-131. doi: 10.3969/j.issn.2096-0387.2018.02.037 MEI L, NIU RJ, JIANG L, et al. Advances in the study on the chemical constituents and pharmacological activities of Ilex pubescens hook. et arn[J]. Biol Chem Eng, 2018, 4(2): 129-131. doi: 10.3969/j.issn.2096-0387.2018.02.037
[3]	ZHOU Y, ZENG KW, ZHANG JY, et al. Triterpene saponins from the roots of Ilex pubescens[J]. Fitoterapia, 2014, 97: 98-104. doi: 10.1016/j.fitote.2014.05.020
[4]	李美芬. 毛冬青中乌苏烷型三萜皂苷的药物代谢研究[D]. 广州: 广州中医药大学, 2011. LI MF. Study on drug metabolism of ursane-type saponins from Radix ilicis[D]. Guangzhou: Guangzhou University of Chinese Medicine, 2011.
[5]	钟海森, 覃骊兰, 杜正彩, 等. 壮药毛冬青的应用概况[J]. 河北中医, 2018, 40(5): 793-797. doi: 10.3969/j.issn.1002-2619.2018.05.036 ZHONG HS, QIN LL, DU ZC, et al. Overview of the application of the Zhuang medicine Ilex pubescens[J]. Hebei J Tradit Chin Med, 2018, 40(5): 793-797. doi: 10.3969/j.issn.1002-2619.2018.05.036
[6]	白荣钰, 易欢, 陈丰连, 等. 毛冬青三萜皂苷对动脉粥样硬化大鼠肠道菌群的影响[J]. 中草药, 2021, 52(20): 6245-6253. doi: 10.7501/j.issn.0253-2670.2021.20.014 BAI RY, YI H, CHEN FL, et al. Effect of triterpenoid saponins from Ilex pubescens on intestinal flora in atherosclerotic rats[J]. Chin Tradit Herb Drugs, 2021, 52(20): 6245-6253. doi: 10.7501/j.issn.0253-2670.2021.20.014
[7]	陈钰泉, 刘玉婷, 邱杰, 等. 不同比色法测定植物源总三萜皂苷含量的对比[J]. 黑龙江农业科学, 2018(3): 108-112. https://www.cnki.com.cn/Article/CJFDTOTAL-HLJN201803027.htm CHEN YQ, LIU YT, QIU J, et al. Comparation of different colorimetry for determination of the total herbal triterpenoid saponins[J]. Heilongjiang Agric Sci, 2018(3): 108-112. https://www.cnki.com.cn/Article/CJFDTOTAL-HLJN201803027.htm
[8]	XIAO CN, HAO FH, QIN XR, et al. An optimized buffer system for NMR-based urinary metabonomics with effective pH control, chemical shift consistency and dilution minimization[J]. Analyst, 2009, 134(5): 916-925. doi: 10.1039/b818802e
[9]	WOJTOWICZ W, ZABEK A, DEJA S, et al. Serum and urine ¹H NMR-based metabolomics in the diagnosis of selected thyroid diseases[J]. Sci Rep, 2017, 7(1): 9108. doi: 10.1038/s41598-017-09203-3
[10]	刘卫红, 张琪, 颜贤忠, 等. 高脂血症及动脉粥样硬化痰瘀演变的代谢组学研究[J]. 中医杂志, 2008, 49(8): 738-741. https://www.cnki.com.cn/Article/CJFDTOTAL-ZZYZ200808043.htm LIU WH, ZHANG Q, YAN XZ, et al. Metabonomics study on phlegm and blood stasis evolution of hyperlipidemia and atherosclerosis[J]. J Tradit Chin Med, 2008, 49(8): 738-741. https://www.cnki.com.cn/Article/CJFDTOTAL-ZZYZ200808043.htm
[11]	KUSHIYAMA A, NAKATSU Y, MATSUNAGA Y, et al. Role of uric acid metabolism-related inflammation in the pathogenesis of metabolic syndrome components such as atherosclerosis and nonalcoholic steatohepatitis[J]. Mediators Inflamm, 2016, 2016: 8603164.
[12]	CORRY DB, ESLAMI P, YAMAMOTO K, et al. Uric acid stimulates vascular smooth muscle cell proliferation and oxidative stress via the vascular renin-angiotensin system[J]. J Hypertens, 2008, 26(2): 269-275. doi: 10.1097/HJH.0b013e3282f240bf
[13]	庞博, 越晧, 王恩鹏, 等. 动脉粥样硬化患者尿液的代谢组学研究[J]. 分析化学, 2015, 43(11): 1766-1771. doi: 10.11895/j.issn.0253-3820.150434 PANG B, YUE H, WANG EP, et al. A metabonomics study of atherosclerosis by rapid resolution liquid chromatography quadrupole time-of-flight mass spectrometry[J]. Chin J Anal Chem, 2015, 43(11): 1766-1771. doi: 10.11895/j.issn.0253-3820.150434
[14]	PEDERSEN ER, MIDTTUN O, UELAND PM, et al. Systemic markers of interferon-γ-mediated immune activation and long-term prognosis in patients with stable coronary artery disease[J]. Arterioscler Thromb Vasc Biol, 2011, 31(3): 698-704. doi: 10.1161/ATVBAHA.110.219329
[15]	BROWN JM, HAZEN SL. The gut microbial endocrine organ: Bacterially derived signals driving cardiometabolic diseases[J]. Annu Rev Med, 2015, 66: 343-359. doi: 10.1146/annurev-med-060513-093205
[16]	WANG ZN, ROBERTS AB, BUFFA JA, et al. Non-lethal inhibition of gut microbial trimethylamine production for the treatment of atherosclerosis[J]. Cell, 2015, 163(7): 1585-1595. doi: 10.1016/j.cell.2015.11.055
[17]	KOETH RA, LEVISON BS, CULLEY MK, et al. γ-Butyrobetaine is a proatherogenic intermediate in gut microbial metabolism of L-carnitine to TMAO[J]. Cell Metab, 2014, 20(5): 799-812. doi: 10.1016/j.cmet.2014.10.006
[18]	AGUILAR EC, DA SILVA JF, NAVIA-PELAEZ JM, et al. Sodium butyrate modulates adipocyte expansion, adipogenesis, and insulin receptor signaling by upregulation of PPAR-γ in obese Apo E knockout mice[J]. Nutrition, 2018, 47: 75-82. doi: 10.1016/j.nut.2017.10.007