- B [6] (TAC) [DOI] /j.issn

Transcription

1 [ ] (TAC) 12 C57BL/6 (N ) ( ) 6 N 22d 16S rdna N [ ] [ ] R543.1 [ ] A [ ] (216) [DOI] /j.issn Intestinal flora changes in a mouse model of transverse aortic constriction ZHOU Xian-jie 1, GONG Li-jing 2, LI Jun 3, LIU Qing-chun 1*, ZHU De-sheng 3* 1 Joint Graduate Training Base of the Armed Police General Hospital, Jinzhou Medical University, Beijing 139, China 2 Sport Science Research Center, Beijing Sport University, Beijing 184, China 3 Laboratory Animal Center, Peking University, Beijing 1871, China * Corresponding author. LIU Qing-chun, lqc@vip.163.com; ZHU De-sheng, deshengz@ pku.edu.cn This work was supported by the Special Projects of Major Scientific Instrument and Equipment of China (211YQ3114) [Abstract] Objective To investigate intestinal microbiota changes of model mice with transverse aortic constriction (TAC) to find a new target point for preventing and treating cardiovascular disease. Methods Twelve C57BL/6 mice were randomly divided into TAC model group (group ) and control group (group N) of 6 mice each. The group received minimally invasive surgery for ligating the aorta to make its constriction to the appropriate degree, while the group N received same operation but no constriction. Faecal samples were collected 22 days after the treatment. Intestinal flora were determined by 16S rdna pyrosequencing and bioinformatics clustering analysis were performed with software of Quantitative Insights into Microbial Ecology. Results A higher abundance of Parabacteroides and a lower abundances of Lactobacillaceae, Lactobacillus and Cocleatum were present in TAC mice compared with the controls. Conclusion Intestinal flora changes would take place in the TAC mice. Intestinal flora may be a potential target for prevention and treatment of cardiovascular diseases, but further validation should be performed to verify the relationship between those intestinal flora changes and disease progression in animal models. [Key words] transverse aortic constriction; intestinal flora; diseases models, animal (TAC) [1] [ ] (211YQ3114) [ ] [ ] 139 ( ) 184 ( ) 1871 () [ ] E -mai l lqc@v ip. 163.com deshengz@ pku.edu.cn [2] [3] [4-5] - B [6] [7] [8] 16S rrna 1

2 Med J Chin PLA, Vol. 41, No. 1, October 1, S rdna rrna DNA 16S rdna ~1 C57BL/6 2~25g [ SCXK( )212-1] [ SYXK( )211-3] 12h/12h (N ) ( ) [9] [1] 37.5~1.cm 2~3mm DNA 22.1g DNA DNA ( ) Qubit DNA 1.4 PCR 16S rdna16s rdna 4PCR PCR 341F 5'- CCTAYG G GRBGCA SCAG -3' 86R 5'-GGACTACNNGGGTATCTA AT-3' 98 1min 98 1s 5 3s 72 3s min PCR Thermo Scientific GeneJET 2 (New England Biolabs NEB Next Ultra DNA Library Prep Kit for Illumina ) Qubit Illumina HiSeq 1.5 (operational taxonomic units OTUs) OTUs- 1.6 SPSS 2. t OTUs - (QIIME) [11] P< N g g g g (P.5) 2.2 QIIME N N OTUs P ( 1) Number of species Number of sequencing Fig.1 Dilution curve between random sequencing number and the corresponding species s rdna ( ) 2 N N1 N2 N3 N4 N5 N

3 Number of species 2 Fig.2 Sequence number with annotation information in the kingdom, phylum, class, order, family, genus and species level between N and group Relative abundance 3 N Fig.3 Top 1 of the relative abundance of species at the phylum level in the N and group N N Group Group Species Genus Family Order Class Phylum Kingdom 1 ( 3) 3 ( ) ( ) ( ) (P.5) Others Acidobacteria Defembacteres Crenarchaeota Cyanobacteria Actinobacteria Tenericutes Verrucomicrobia Proteobacteria Firmicutes Bacteroidetes 15 (Lactobacillaceae).147%.561% (P=.37) 15 (Lactobacillus) (Parabacteroides).147%.561%.75%.535%(P=.37) 15 (Cocleatum).5%.46%(P=.19) ( 4) N Lactobacillaceae Lactobacillus Parabacteroides Cocleatum N group group 4 Fig.4 Significant differences of bacteria in the group of N and 3 [1] [12] 16S rdna N OTUs OUT N N N N N [13-14] [15-16]

4 Med J Chin PLA, Vol. 41, No. 1, October 1, [17] [18] N [19] Hata [2] ( ) [21] N [22] N [1] Tarnavski O, McMullen JR, Schinke M, et al. Mouse cardiac surgery: comprehensive techniques for the generation of mouse models of human diseases and their application for genomic studies[ J]. Physiol Genomics, 24, 16(3): [2] Diwan A, Dorn GW. Decompensation of cardiac hypertrophy: cellular mechanisms and novel therapeutic targets[ J]. Physiology (Bethesda), 27, 22(1): [3] Ta n g W H, Wa n g Z, Fa n Y, e t a l. P r o g n o s t i c v a l u e o f elevated levels of intestinal microbe-generated metabolite trimethylamine-n-oxide in patients with heart failure: refining the gut hypothesis[ J]. J Am Coll Cardiol, 214, 64(18): [4] Organ CL, Otsuka H, Bhushan S, et al. Choline diet and its gut microbe-derived metabolite, trimethylamine N-oxide, exacerbate pressure overload-induced heart failure[ J]. Circ Heart Fail, 216, 9(1): 1-1. [5] Sheng W, Li LB, Lu JY. Role of TGF-, MMPs and TIMPs in persistent atrial fibrillation associated with rheumatic mitral stenosis[ J]. Med J Chin PLA, 214, 39(8): [,,. TGF- MMPs TIMPs [ J]., 214, 39(8): ] [6] Seldin MM, Meng Y, Qi H, et al. Trimethylamine N-ox ide promotes vascular inflammation through signaling of mitogenactivated protein kinase and nuclear factor- B[ J]. J Am Heart Assoc, 216, 5(2): [7] Ho n o u r J W. Hi s to r i c a l p e r s p e c t i v e : g u t d y s b i o s i s a n d hypertension[ J]. Physiol Genomics, 215, 47(1): [8] Niu M, Shao TB, Chen RC, et al. Analysis of intestinal flora distribution and detection of bacteria toxin gene in ulcerative colitis patients[ J]. J Zhengzhou Univ (Med Sci), 215, 5(4): [,,,. [ J]. ( ), 215, 5(4): ] [9] Faerber G, Barreto-Perreia F, Schoepe M, et al. Induction of heart failure by minimally invasive aortic constriction in mice: reduced peroxisome proliferator-activated receptor coactivator levels and mitochondrial dysfunction[ J]. J Thorac Cardiovasc Surg, 211, 141(2): [1] Chen Y, Guo H, Xu D, et al. Left ventricular failure produces profound lung remodeling and pulmonary hypertension in mice: heart failure causes severe lung disease[ J]. Hypertension, 212, 59(6): [11] Moreno-Indias I, Torres M, Montserrat JM, et al. Intermittent hypoxia alters gut microbiota diversity in a mouse model of sleep apnoea[ J]. Eur Respir J, 215, 45(4): [12] Xin W, Li XY, Lu XC, et al. Cardiac function improved by sarcoplasmic reticulum Ca 2+ -ATPase overexpression in a heart failure model induced by chronic myocardial ischemia[ J]. Med J Chin PLA, 211, 36(4): [,,,. Ca 2+ -ATP [ J]., 211, 36(4): ] [13] Awadel-K ar iem FM, Patel P, K apoor J, et al. First repor t of Parabacteroides goldsteinii bacteraemia in a patient with complicated intra-abdominal infection[ J]. Anaerobe, 21, 16(3): [14] Song Y, Liu C, Lee J, et al. "Bacteroides goldsteinii sp. nov." isolated from clinical specimens of human intestinal origin[ J]. J Clin Microbiol, 25, 43(9): [15] B o e n te R F, Fe r re i ra LQ, Fa l c ão L S, e t al. D e tection o f resistance genes and susceptibility patterns in Bacteroides and Parabacteroides strains[ J]. Anaerobe, 21, 16(3): [16] Boyanova L, Kolarov R, Mitov I. Recent evolution of antibiotic resistance in the anaerobes as compared to previous decades[ J]. Anaerobe, 215, 31(1): 4-1. [17] Dziarski R, Park SY, Kashyap DR, et al. Pglyrp-regulated gut microflora Prevotella falsenii, Parabacteroides distasonis and Bacteroides eggerthii enhance and Alistipes finegoldii attenuates colitis in mice[ J]. PLoS One, 216, 11(1): 1-24.

5 [18] Li J, Shang HB, Liu QC, et al. Minimally invasive transverse aortic constriction procedure for constructionof heart failure animal model in mice[ J]. Shandong Med J, 214, 54(12): 5-7, 112. [,,,. [ J]., 214, 54(12): 5-7, 112.] [19] Ivey KL, Hodgson JM, Kerr DA, et al. The effect of yoghurt and its probiotics on blood pressure and serum lipid profile: a randomised controlled trial[ J]. Nutr Metab Cardiovasc Dis, 215, 25(1): [2] Hata Y, Yamamoto M, Ohni M, et al. A placebo-controlled study of the effect of sour milk on blood pressure in hypertensive subjects[ J]. Am J Clin Nutr, 1996, 64(5): [21] S h a r a f e d t i n o v K K, P l o t n i k o v a OA, A l e x e e v a R I, e t a l. Hy p o c a l o r i c d i e t s u p p l e m e n ted w i t h p ro b i o t i c c h e e s e improves body mass index and blood pressure indices of obese hypertensive patients--a randomized double-blind placebocontrolled pilot study[ J]. Nutr J, 213, 12(1): [22] Chen L, Tai WC, Brar MS, et al. Tumor grafting induces changes of gut microbiota in athymic nude mice in the presence and absence of medicinal Gynostemma saponins[ J]. PLoS One, 215, 1(5): ( ) ( )