纳米二氧化钛经口暴露90天对大鼠粪便代谢组的影响

doi:10.19723/j.issn.1671-167X.2020.03.010

摘要/Abstract

摘要：

目的通过粪便代谢组学,探索纳米二氧化钛(titanium dioxide nanoparticles,TiO₂ NPs)经口染毒90 d对大鼠肠道及肠道菌群代谢的影响及其相关机制。方法 12只清洁级雄性Sprague Dawley(SD)大鼠按体质量随机分成两组,分别以0和50 mg/kg体质量的TiO₂ NPs持续灌胃90 d,对TiO₂ NPs的粒径、晶型、纯度、比表面积进行表征,并在第90天收集大鼠的新鲜粪便。经过冻干、亲水相萃取等前处理后,使用超高效液相色谱-轨道阱高分辨质谱仪联用系统(ultra performance liquid chromatography-Q-exactive orbitrap-high-resolution mass spectrometry system,UPLC-QEMS)对粪便代谢物进行非靶向测定,鉴定标注检测得到的代谢物,并进行代谢组学分析。结果与对照组相比,TiO₂ NPs染毒组大鼠体质量显著降低(P<0.05)。粪便代谢组学共发现22种代谢物浓度发生显著改变,其中黄嘌呤、甲基腺嘌呤、羟基吡啶、蛋氨酸亚砜等15种代谢物浓度显著上升,乙酰组胺、派可林酸、咪唑乳酸、缬氨酸等7种代谢物浓度显著下降。N-乙酰组胺、缬氨酸和蛋氨酸亚砜的改变倍数大于16倍。京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)通路分析发现,精氨酸生物合成通路和氨酰基-tRNA生物合成通路这两个代谢通路发生显著改变(错误发现率<0.05,通路受影响程度>0.10)。结论 TiO₂ NPs经口染毒90 d可扰乱肠道及肠道菌群代谢,并导致大鼠粪便中代谢物及代谢通路发生显著改变,提示TiO₂ NPs经口暴露对大鼠的毒性作用可能与肠道及肠道菌群代谢改变密切相关。

关键词: 纳米二氧化钛, 代谢组学, 粪便, 大鼠, Sprague-Dawley

Abstract:

Objective: To explore the effects and related mechanisms of oral exposure titanium dioxide nanoparticles (TiO₂ NPs) for 90 days on the intestinal and the gut microbiota of rats, through fecal metabolomics.Methods: Twelve 4-week-old clean-grade Sprague Dawley (SD) rats were randomly de-vided into 2 groups by body weight, treated with TiO₂ NPs at dose of 0 or 50 mg/kg body weight everyday respectively for 90 days. The solution of each infection was freshly prepared and shocked fully by ultrasonic. Characterization of the particle size, crystal form, purity, and specific surface area of TiO₂ NPs was conducted. And the fresh feces of the rats were collected on the 90th day. After lyophilized and hydrophilic phase extraction, ultra performance liquid chromatography-Q-exactive orbitrap-high-resolution mass spectrometry system (UPLC-QEMS) was utilized for non-targeted determination of fecal meta-bolites. The metabolites were identified and labeled through Compound Discoverer 3.0 software, and used for subsequent metabolomics analysis. Bioinformatics analysis was carried out including unsupervised principal component analysis and supervised orthogonal projection to latent structure discriminant analysis for the differential metabolites between the two groups. The differential metabolites were followed-up for Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis.Results: Compared with the control group, the body weight of the rats was significantly reduced (P<0.05) in the treatment group. A total of 22 metabolites in fecal metabolomics showed significant changes. Among them, xanthine, 1-methyladenine, 3-hydroxypyridine, methionine sulfoxide, pyridoxine, 1,5-isoquinolinediol, N-acetylornithine, N-acetyl-D-galactosamine, L-citrulline, L-methionine, leucine, DL-tryptophan, L-ornithine, 4-methyl-5-thiazoleethanol, and L-glutamic acid totaled 15 metabolites increased significantly. N-acetylhistamine, D-pipecolinic acid, imidazolelactic acid, L-valine, 2,3,4,6-tetramethylpyrazine, caprolactam, and histamine totaled 7 metabolites decreased significantly. N-acetylhistamine, L-valine and methionine sulfoxide were changed more than 16 times. Analysis of KEGG pathway revealed that the two metabolic pathways arginine biosynthesis and aminoacyl-tRNA biosynthesis were significantly changed (false discover rate < 0.05, pathway impact > 0.1).Conclusion: Oral exposure to TiO₂ NPs for 90 days could disrupt the metabolism of the intestine and gut microbiota, causing significant changes in metabolites and metabolic pathways which were related to inflammatory response, oxidative stress, glucose homeostasis, blood system and amino acid homeostasis in rat feces. It is suggested that the toxic effect of TiO₂ NPs on rats may be closely related to intestinal and gut microbiota metabolism.

Key words: Titanium dioxide nanoparticles, Metabolomics, Feces, Rats, Sprague-Dawley

中图分类号:

R994.4

韩硕,陈章健,周迪,郑湃,张家赫,贾光. 纳米二氧化钛经口暴露90天对大鼠粪便代谢组的影响[J]. 北京大学学报（医学版）, 2020, 52(3): 457-463.

Shuo HAN,Zhang-jian CHEN,Di ZHOU,Pai ZHENG,Jia-he ZHANG,Guang JIA. Effects of titanium dioxide nanoparticles on fecal metabolome in rats after oral administration for 90 days[J]. Journal of Peking University (Health Sciences), 2020, 52(3): 457-463.

图/表 6

图1

图2

图3

图4

表1

差异代谢物详细信息"

Super class	Metabolite name	Molecular formula	Relative molecular mass	HMDB ID	KEGG ID	Log change fold (log₂N)
Organoheterocyclic compounds	Xanthine	C₅H₄N₄O₂	152.110 9	HMDB0000292	C00385	2.857 1
Organoheterocyclic compounds	1-methyladenine	C₆H₇N₅	149.153 3	HMDB0011599	C02216	2.045 8
Organoheterocyclic compounds	3-hydroxypyridine	C₅H₅NO	95.099 3			2.191 9
Organoheterocyclic compounds	Pyridoxine	C₈H₁₁NO₃	169.177 8	HMDB0000239	C00314	2.149 2
Organic acids and derivatives	Methionine sulfoxide	C₅H₁₁NO₃S	165.210 0	HMDB0002005	C02989	7.581 8
Organoheterocyclic compounds	1,5-isoquinolinediol	C₉H₇NO₂	161.160 0			2.502 5
Organic acids and derivatives	N-acetylornithine	C₇H₁₄N₂O₃	174.197 7	HMDB0003357	C00437	0.564 7
Organic oxygen compounds	N-acetyl-D-galactosamine	C₈H₁₅NO₆	221.207 8	HMDB0000853	C05021	2.558 6
Organic acids and derivatives	L-citrulline	C₆H₁₃N₃O₃	175.185 7	HMDB0000904	C00327	1.895 0
Organic acids and derivatives	L-methionine	C₅H₁₁NO₂S	149.211 0	HMDB0000696	C00073	1.370 4
Organic acids and derivatives	Leucine	C₆H₁₃NO₂	131.172 9	HMDB0000687	C00123	0.754 9
Organoheterocyclic compounds	DL-tryptophan	C₁₁H₁₂N₂O₂	204.225 2	HMDB0013609	C00525	-0.140 4
Organic acids and derivatives	L-ornithine	C₅H₁₂N₂O₂	132.161 0	HMDB0000214	C00077	0.503 8
Organoheterocyclic compounds	4-methyl-5-thiazoleethanol	C₆H₉NOS	143.207 0	HMDB0032985	C04294	1.259 6
Organic acids and derivatives	L-glutamic acid	C₅H₉NO₄	147.129 3	HMDB0000148	C00025	0.714 0
Organic nitrogen compounds	Histamine	C₅H₉N₃	111.145 1	HMDB0000870	C00388	-1.004 3
Organoheterocyclic compounds	Caprolactam	C₆H₁₁NO	113.157 6	HMDB0062769	C06593	-8.288 4
Organoheterocyclic compounds	2,3,5,6-tetramethylpyrazine	C₈H₁₂N₂	136.194 3	HMDB0036584		-1.740 2
Organic acids and derivatives	L-valine	C₅H₁₁NO₂	117.146 3	HMDB0000883	C00183	-4.354 5
Organoheterocyclic compounds	Imidazolelactic acid	C₆H₈N₂O₃	156.139 3	HMDB0002320	C05132	-2.948 3
Organic acids and derivatives	D-pipecolinic acid	C₆H₁₁NO₂	129.157 0	HMDB0005960		-1.898 0
Organic acids and derivatives	N-acetylhistamine	C₇H₁₁N₃O	153.181 7	HMDB0013253	C05135	-8.991 5

表1

图5

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