轻微肝性脑病患者经颈静脉肝内门体分流术后不同预后组肠道菌群的变化

李萌浩; 李凯; 汤世豪; 王钲钰; 郭文刚; 殷占新; 韩国宏

doi:10.3969/j.issn.1001-5256.2021.02.016

轻微肝性脑病患者经颈静脉肝内门体分流术后不同预后组肠道菌群的变化

DOI: 10.3969/j.issn.1001-5256.2021.02.016

空军军医大学西京消化病医院消化介入科，西安 710032

基金项目:

国家自然科学基金 (81420108020)

利益冲突声明：本研究不存在研究者、伦理委员会成员、受试者监护人以及与公开研究成果有关的利益冲突，特此声明。

作者贡献声明：韩国宏教授负责课题设计；李萌浩、汤世豪、李凯、王钲钰、郭文刚和殷占新参与收集数据；李萌浩进行资料分析和撰写论文；李凯副主任医师和韩国宏教授指导撰写文章，修改论文并最后定稿。

详细信息

作者简介:
李萌浩(1997—)，男，博士，主要从事肠道菌群与肝硬化门静脉高压并发症方面的研究

通信作者:
韩国宏，hangh@fmmu.edu.cn

中图分类号: R575.2
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出版历程
- 收稿日期: 2020-09-13
- 录用日期: 2020-09-30
- 出版日期: 2021-02-20

Changes in gut microbiota after transjugular intrahepatic portosystemic shunt in cirrhotic patients with mild hepatic encephalopathy in different prognosis groups

Department of Liver Disease and Digestive Interventional Radiology, Xijing Hospital of Digestive Diseases, Air Force Medical University, Xi'an 710032, China

摘要

摘要: 目的对比分析肝硬化轻微肝性脑病(MHE)患者经颈静脉肝内门体分流术(TIPS)后不同预后组肠道菌群结构与术前的差异。方法选取2016年7月—2017年7月西京医院消化病院住院并接受TIPS治疗的28例MHE患者，分别于术前1~3 d和术后1个月收集粪便样本及采样时临床资料信息，根据术后预后不同分为3组:无HE组(n=8)、MHE组(n=12)和显性脑病(OHE)组(n=8)。对粪便样本采取16S rRNA高通量测序技术进行测序得到菌群相对丰度并在属水平使用SPSS和R语言对各组间菌群物种多样性，术后变化和变化差异展开分析。计数资料组间比较采用χ²检验，计量资料3组间比较采用Kruskal-Wallis H检验，采用Bonferroni法进行多个样本的多重比较，同组患者术前术后的比较采用Wilcoxo符号秩检验。微生物组Beta分析基于Bray Curtis距离矩阵进行主坐标分析(PCoA)，使用Adonis法(PerMANOVA)对比组间差异。结果基于Bray Curtis距离矩阵的PCoA分析显示，仅MHE组术前和术后的beta多样性明显改变(F=2.71，P=0.049)。术后无HE组原生菌群小杆菌属、粪球菌属、瘤胃菌科某属、解黄酮菌属和狭义梭菌属丰度较术前明显升高(Z值分别为2.521、2.1、2.1、2.1和1.96，P值均＜0.05)；MHE组术后有害菌群颗粒链菌属(Z=2.521, P=0.012)、肠球菌属(Z=2.51, P=0.012)、链球菌属(Z=2.432, P=0.015)和罗氏菌属(Z=2.001, P=0.045)丰度较术前明显下降，但韦荣球菌属(Z=2.353, P=0.019)和巨球菌属(Z=1.955, P=0.05)丰度却明显上升; OHE组术后仅观察到韦荣球菌属丰度较术前升高(Z=2.38，P= 0.017)。3组间菌群变化量(术后丰度/术前丰度)比较差异有统计学意义[无HE组vs MHE组vs OHE组：2.00(1.11~91.61) vs 1.21(0.26~6.79) vs 0.09(0.01~0.92)，χ²=6.249，P=0.043]。结论 TIPS术后不同预后患者肠道菌群的变化有明显的差异，原生菌群丰度的升高可能对缓解HE病情有一定影响。
- 肝硬化 /
- 肝性脑病 /
- 门体分流术, 经颈静脉肝内 /
- 胃肠道微生物组
Abstract: Objective To investigate the changes in gut microbiota after transjugular intrahepatic portosystemic shunt (TIPS) in cirrhotic patients with mild hepatic encephalopathy (MHE) in different prognosis groups. Methods A total of 28 MHE cirrhotic patients who were hospitalized and underwent TIPS in Xijing Hospital of Digestive Diseases from July 2016 to July 2017 were enrolled. Fecal samples and related clinical data were collected on days 1-3 before surgery and at 1 month after surgery. According to the prognosis after surgery, the patients were divided into none-hepatic encephalopathy (HE) group with 8 patients, MHE group with 12 patients, and overt hepatic encephalopathy (OHE) group with 8 patients. Fecal samples were analyzed by 16S rRNA sequencing to obtain the relative abundance of gut microbiota, and SPSS and R packages were used to analyze the biodiversity, postoperative changes, and differences in such changes of gut microbiota at the genus level between groups. The chi-square test was used for comparison of categorical data between groups; the Kruskal-Wallis H test was used for comparison of continuous data between three groups; the Bonferroni method was used for multiple comparisons of multiple samples; the Wilcoxon signed-rank test was used for comparison before and after surgery within each group. For microbiome beta-diversity analyses, a principal coordinate analysis (PCoA) was performed based on Bray-Curtis distance matrix, and the Adonis method (PerMANOVA) was used for comparison between groups. Results PCoA based on Bray-Curtis distance matrix showed that only the MHE group had a significant change in beta diversity after surgery (F=2.71, P=0.049). After surgery, the non-HE group had significant increases in the abundance of the native flora Dialister, Coprococcus, Ruminococcaceae_uncultured, Flavonifractor, and Clostridium_sensu_stricto_1 (Z=2.521, 2.1, 2.1, 2.1, and 1.96, all P < 0.05); the MHE group had significant reductions in the abundance of the harmful flora Granulicatella(Z=2.521, P=0.012), Enterococcus(Z=2.51, P=0.012), Streptococcus(Z=2.432, P=0.015), and Rothia(Z=2.001, P=0.045) and significant increases in the abundance of Veillonella(Z=2.353, P=0.019) and Megasphaera(Z=1.955, P=0.05); the OHE group only had a significant increase in the abundance of Veillonella after surgery (Z=2.38, P=0.017). There was a significant difference in the change in gut microbiota (postoperative abundance/preoperative abundance) between the non-HE group, the MHE group, and the OHE group [2.00 (1.11-91.61) vs 1.21 (0.26-6.79) vs 0.09 (0.01-0.92), χ²=6.249, P=0.043]. Conclusion There is a significant difference in the change in gut microbiota after TIPS between patients with different prognoses, and the increase in the abundance of native flora may have a certain influence on the remission of MHE.
- Liver Cirrhosis /
- Hepatic Encephalopathy /
- Portasystemic Shunt, Transjugular Intrahepatic /
- Gastrointestinal Microbiome

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图 1 MHE组患者术前术后菌群beta多样性对比分析

注：基于Bray Curtis距离矩阵的主坐标分析，MHE组术前术后beta多样性有统计学差异(F=2.71，P=0.049)。

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图 2 不同预后组TIPS术前术后肠道菌群变化情况

下载: 全尺寸图片幻灯片

图 3 不同预后组TIPS术后粪球菌属变化量比较

注：3组间菌群变化量有明显差异(χ²=6.249，P=0.043)，球菌属变化量在无HE组和MHE组中均明显高于OHE组(P值分别为0.021、0.050)，而无HE组与MHE组差异无统计学意义(P＞0.05)。

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表 1 不同预后组术前基线特征比较

指标	无HE组(n=8)	MHE组(n=12)	OHE组(n=8)	χ²值	P值
年龄(岁)	49.5(45.8~52.8)	54.5(49.8~59.5)	46.5(44.3~52.8)	5.28	0.07
性别[男，例(%)]	3(37.5)	6(50.0)	4(50.0)	3.08	0.61
BMI(kg/m²)	18.7(18.3~20.7)	21.6(18.6~24.1)	21.4(19.6~23.8)	4.43	0.11
肝硬化病因[HBV，例(%)]	4(50.0)	7(58.3)	7(87.5)	7.39	0.35
MELD评分	10.9(7.8~13.2)	13.4(10.0~17.5)	13.4(12.7~14.9)	4.82	0.10
Child-Pugh分级[例(%)]				2.44	0.49
A(5~6)	4(50.0)	3(12.5)	2(25.0)
B(7~9)	4(50.0)	6(50.0)	5(62.5)
C(10~13)	0(0)	3(12.5)	1(12.5)
INR	1.4(1.1~1.6)	1.4(1.2~1.9)	1.6(1.5~1.7)	4.92	0.10
血红蛋白(g/dl)	80.0(75.5~83.8)	87.0(79.3~94.3)	80.0(59.0~92.0)	1.69	0.43
总蛋白(mg/dl)	68.2(53.1~73.9)	68.0(55.7~78.9)	66.2(54.7~68.0)	0.83	0.66
Alb(g/dl)	36.5(30.6~39.3)	34.4(28.1~37.2)	32.7(31.4~35.5)	1.10	0.58
TBil(μmol/L)	15.2(10.6~20.6)	23.6(14.7~49.6)	28.1(20.4~36.1)	4.86	0.09
AST(U/L)	21.5(13.0~40.3)	21.0(11.3~30.0)	28.5(20.3~34.8)	1.77	0.41
ALT(U/L)	30.5(20.8~42.8)	27.5(21.0~42.5)	31.0(21.5~37.5)	0.01	0.99
ALP(U/L)	97.0(73.8~117.0)	65.5(49.8~92.8)	79.0(59.8~112.3)	3.69	0.16
γ-氨基丁酸(U/L)	32.0(21.0~45.8)	23.5(14.0~56.5)	31.0(20.0~49.0)	0.63	0.73
肌酐(mg/dl)	77.5(66.5~93.8)	85.5(73.0~105.0)	72.0(68.3~95.0)	1.46	0.48
静脉血氨(μg/dl)	51.0(21.0~60.0)	37.5(23.8~60.5)	36.0(27.5~47.5)	0.29	0.86
注：MELD，终末期肝病模型。

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表 2 不同预后组术前alpha多样性比较

多样性指数	无HE组(n=8)	MHE组(n=12)	OHE组(n=8)	χ²值	P值
Ace指数	186.70(137.96~232.29)	214.69(144.20~354.14)	186.68(130.79~241.34)	0.96	0.62
Chao1指数	194.64(133.60~215.46)	203.40(138.58~352.64)	183.79(118.53~208.50)	1.03	0.60
Shannon指数	2.65(2.02~3.12)	2.95(2.56~3.95)	2.67(2.04~3.50)	1.71	0.43
Simpson指数	0.16(0.09~0.23)	0.09(0.05~0.16)	0.13(0.07~0.26)	2.46	0.29

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