基于16S核糖体RNA高通量测序探讨加味四君子汤对严重烫伤兔肠道菌群多样性的影响  

Investigating the effects of Modified Sijunzi Decoction on the diversity of intestinal microflora of severe scald rabbits based on 16S ribosomal RNA high-throughput sequencing

作　　者：罗锦花[1] 詹剑华[1] 廖文强 程兴 黄凯[1] Luo Jinhua;Zhan Jianhua;Liao Wenqiang;Cheng Xing;Huang Kai(Burn and Wound Repair Center,the First Affiliated Hospital of Nanchang University,Nanchang 330006,China)

机构地区：[1]南昌大学第一附属医院烧伤与创面修复中心,南昌330006

出　　处：《中华烧伤与创面修复杂志》2022年第3期227-235,共9页Chinese Journal of Burns And Wounds

基　　金：江西省科技计划(20171BBG70061、2020BBG73020)。

摘　　要：目的基于16S核糖体RNA(16S rRNA)高通量测序,探讨加味四君子汤对严重烫伤兔肠道菌群多样性的影响。方法采用实验研究方法。选取90只6~8个月龄雌雄不限日本大耳兔,按照随机数字表法分为正常对照组、单纯烫伤组、烫伤+低剂量组、烫伤+中剂量组、烫伤+高剂量组,每组18只。正常对照组兔自由饮食、饮水,单纯烫伤组、烫伤+低剂量组、烫伤+中剂量组、烫伤+高剂量组兔制作30%体表总面积Ⅲ度烫伤模型后分别给予生理盐水、0.2 g/mL加味四君子汤、1.0 g/mL加味四君子汤、5.0 g/mL加味四君子汤灌胃,持续给药7 d。于分组处理第1、3、7天,采用酶联免疫吸附测定法检测各组兔回肠黏膜组织中的肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-10水平,各组每个时间点的样本数为6。根据前述实验结果,另外选取9只兔分为正常对照组、单纯烫伤组、烫伤+中剂量组,每组3只,各组兔分组及处理方法同前。于分组处理第7天,采用高通量测序技术对3组兔回肠黏膜菌群的16S rRNA V3、V4区进行测序;采用QIIME软件统计菌群优质序列数目;使用RDP Classifier软件分析菌群的门、纲、目、科、属水平;采用Illumina MiSeq测序技术对菌群行α多样性(Ace、Chao1、Simpson、Shannon指数)和β多样性分析。对数据行析因设计方差分析、SNK检验及Bonferroni校正。结果与正常对照组相比,单纯烫伤组、烫伤+低剂量组、烫伤+高剂量组分组处理第1、3、7天和烫伤+中剂量组分组处理第1、3天兔回肠黏膜组织中的TNF-α水平均明显升高(P<0.01),单纯烫伤组、烫伤+低剂量组、烫伤+中剂量组、烫伤+高剂量组分组处理第1、3、7天兔回肠黏膜组织中的IL-1β水平均明显升高(P<0.05或P<0.01),单纯烫伤组、烫伤+低剂量组、烫伤+中剂量组、烫伤+高剂量组分组处理第1、3、7天兔回肠黏膜组织中的IL-10水平均显著降低(P<0.01)。与单纯烫伤�Objective To investigate the effects of Modified Sijunzi Decoction on the diversity of intestinal microflora of in severe scald rabbits based on 16S ribosomal RNA(16S rRNA)high-throughput sequencing.Methods The experimental research method was adopted.Ninety Japanese big-ear rabbits regardless gender,aged 6 to 8 months,were randomly divided into normal control group,scald alone group,scald+low-dose group,scald+medium-dose group,and scald+high-dose group,with 18 rabbits in each group.The rabbits in normal control group were free to eat and drink,and the rabbits in scald alone group,scald+low-dose group,scald+medium-dose group,and scald+high-dose group were intragastrically administered normal saline,0.2 g/mL Modified Sijunzi Decoction,1.0 g/mL Modified Sijunzi Decoction,and 5.0 g/mL Modified Sijunzi Decoction,respectively for 7 days after sustaining full-thickness scalding of 30%total body surface area.On the 1^(st),3^(rd),and 7^(th)day after grouping,the levels of tumor necrosis factorα(TNF-α),interleukin 1β(IL-1β),and IL-10 in ileal mucosa tissue of rabbits in each group were determined by enzyme-linked immunosorbent assay,and the number of samples in each group at each time point was 6.According to the above experimental results,another 9 rabbits were selected and divided into normal control group,scald alone group and scald+medium-dose group,with 3 rabbits in each group.The grouping and treatment methods of rabbits in each group were the same as before.On the 7^(th) day after grouping,the V3,V4 region of 16S rRNA of ileum mucosa of rabbits in three groups were sequenced by high-throughput sequencing technology.The number of quality bacteria was counted by QIME software.The classifications of phylum,class,order,family and genus of microflora were analyzed by RDP Classifier software.Theαdiversity(Ace,Chao1,Simpson,and Shannon indexes)andβdiversity were analyzed by Illumina MiSeq sequencing technology,and the number of experiment samples in each group was 3.Data were statistically analyzed with analysis for

关 键 词：植物制剂 高通量核苷酸序列分析 加味四君子汤 严重烫伤 肠道屏障 肠道菌群 16S核糖体RNA 

分 类 号：R285.5[医药卫生—中药学]