实验研究
donghuiwei
2020, 44(11): 0-0.
摘要: 目的 通过环磷酰胺注射建立免疫抑制大鼠模型，探究免疫抑制大鼠肺部菌群变化，为探究免疫低下患者肺部感染菌群的结构与来源提供依据。方法 选用140～160 g雄性SD大鼠，随机分为对照组（A）和免疫抑制组（B）,每组10只。每天按60 mg/kg体重给予免疫抑制组大鼠腹腔注射浓度为10 mg/mL的环磷酰胺溶液,对照组大鼠仅腹腔注射等体积的生理盐水溶剂。连续注射3天后，分别对两组大鼠心脏采血测血细胞计数，取肺组织进行16s rDNA高通量测序和生物信息学分析。结果 B组大鼠中性粒细胞、淋巴细胞、单核细胞等白细胞数量明显低于A组。A组各门水平上优势菌门有厚壁菌门（Firmicutes）、变形菌门（Proteobacteria）、拟杆菌门（Bacteroidetes）、放线杆菌门（Actinobacteria）；在属水平上优势菌属有乳酸杆菌（Lactobacillus）、拟杆菌（Bacteroides）、未定型瘤胃球菌（unidentified_Ruminococcaceae）等。B组在门和属水平上与A组没有明显区别。对样本进行Alpha多样性指数（Alpha Diversity）组间差异分析，ACE、Chao、Shannon、Simpson四个指数间差异并不显著(pACE=0.569，pChao=0.599，pShannon=0.370，pSimpson=0.460)。PCoA分析结果表明A组与B组的各样品分布相似度较高。结论 通过高剂量环磷酰胺腹腔注射，成功制作免疫抑制大鼠模型。阐明免疫抑制大鼠和正常大鼠肺部菌群结构，并发现两者在丰度和多样性上差别并不显著。这提示，免疫低下患者患肺部感染的致病菌可能并非源自肺部机会致病菌的异常增殖，而是来源于外界环境吸入。
关键词: 环磷酰胺；免疫抑制；16S rDNA；肺部菌群；菌群结构
中图分类号：R378.99
Abstract: Objective The immunosuppressive rat model was established by cyclophosphamide injection to explore the changes of pulmonary microflora in immunosuppressive rats so as to provide a basis for exploring the source of pulmonary infection in immunocompromised patients. Methods Male SD rats (140g-160g) were randomly divided into control group (A) ,(n = 10) and immunosuppressive group (B), (n = 10). Rats in the immunosuppressive group were given intraperitoneal injection of cyclophosphamide at the concentration of 10 mg/kg per day according to the body weight of 60 mg/mL, while the rats in the control group were only injected intraperitoneally with the same volume of normal saline solvent. After 3 days of continuous injection, blood samples were collected from the hearts of the two groups for blood cell count, and lung tissues were taken for 16s rDNA high-throughput sequencing and bioinformatics analysis. Results The number of neutrophils, lymphocytes and monocytes in group B was significantly lower than that in group A. In the group A, the dominant phyla were Firmicutes, Proteobacteria, Bacteroidetes and Actinobacteria. At the genus level, the dominant bacteria are Lactobacillus, Bacteroides, unidentified_Ruminococcaceae and so on. There are no significant difference of the dominat phyla and genus level between group A and B. The difference analysis of Alpha Diversity index among groups showed that there was no significant difference among ACE, Chao, Shannon and Simpson (pACE=0.569,pChao=0.599,pShannon=0.370,pSimpson=0.460). The results of PCoA analysis showed that the similarity of sample distribution between group A and group B was very high. Conclusion The immunosuppressive rat model was successfully established by intraperitoneal injection of high-dose cyclophosphamide. The structure of pulmonary microflora in immunosuppressive rats and normal rats was clarified, and it was found that there was no significant difference in abundance and diversity between them. This suggests that the pathogens of pulmonary infection in immunocompromised patients are not caused by the amplification of opportunistic pathogens in the lungs, but probably by inhalation in the external environment.
Key words: Cyclophosphamide; Immunosuppression; 16s rDNA; Pulmonary flora; Flora structure
