改良PIICS小鼠模型肠道菌群的多样性和结构变化  

The diversity and structural changes of intestinal microbiota in a modified persistent inflammatory-immunosuppressive-catabolic syndrome mouse model

作　　者：张亮[1] 罗板鑫 田中 龚龙波[1] ZHANG Liang;LUO Banxin;TIAN Zhong;GONG Longbo(Department of Gastrointestinal Surgery,Clinical School of Xuzhou Medical University,Xuzhou Central Hospital,Xuzhou,Jiangsu 221008,China;不详)

机构地区：[1]徐州市中心医院(徐州医科大学徐州临床学院)胃肠外科,江苏221008

出　　处：《中国微生态学杂志》2024年第12期1397-1405,共9页Chinese Journal of Microecology

基　　金：普通外科省级临床重点专科建设资金(1-2-2022TD001);徐州市卫生健康委科技项目(XWKYHT20220086);徐州市科技项目(KC23173)。

摘　　要：目的本研究改良持续性炎症-免疫抑制-分解代谢综合征(persistent inflammatory-immunosuppressive-catabolic syndrome,PⅡCS)小鼠模型使其肠道菌群更贴近PⅡCS患者肠道菌群的状况并初步阐述其肠道菌群的多样性和结构变化。方法28只雄性28周龄C57BL/6小鼠,体重30~35 g,随机分为假手术(SC)组(n=10)和PⅡCS组(n=18)。在现有PⅡCS小鼠模型的基础上,采用措施模拟可能对肠道菌群造成影响的临床过程,包括早期大剂量长期抗生素的应用和大量液体的补充,以及长期卧床。在第14天,使用ELISA法检测血液内的肿瘤坏死因子(tumor necrosis factor,TNF)-α等炎性因子和C-X-C基序化学因子配体(C-X-C motif chemokine ligand 1,CXCL1)等趋化因子;采用流式细胞术检测脾脏中CD8^(+)T细胞比例和CD11b^(+)的髓源性抑制细胞比例以反映免疫抑制情况,并以免疫组化和蛋白印记等方法分别检测肌肉纤维形态和蛋白表达以反映分解代谢的情况;使用16S rRNA基因测序技术并数据分析肠道菌群的多样性和结构变化。结果与SC组比较,PⅡCS组小鼠血液中白介素(interleukin,IL)-6(t=132.67,P<0.0010)、IL-10(t=65.76,P<0.0010)和TNF-α(t=23.77,P=0.0032)等炎症因子表达增加,而CXCL1(t=125.24,P<0.0010)、粒细胞-巨噬细胞集落刺激因子(granulocyte-macrophage colony-stimulating factor,GM-CSF)(t=111.24,P<0.0010)和巨噬细胞集落刺激因子(macrophage colony-stimulating factor,M-CSF)(t=154.22,P<0.0010)表达水平下降;CD8^(+)T细胞比例下降(t=37.11,P=0.0088),CD11b^(+)髓源性抑制细胞比例上升(t=45.12,P=0.0095);肌纤维形态变多边形,且截面积偏小(P<0.0010),肌肉蛋白MURF-1(t=123.20,P=0.0073)和FoxO-1(t=67.34,P=0.0091)表达下降;肠道菌群多样性下降(P<0.0010)。LEfSe分析显示PⅡCS组小鼠肠道菌群普拉梭菌属、瘤胃球菌科、杜氏杆菌属、丹毒丝菌纲丰度显著降低,变形菌门、变形菌纲、肠杆菌科、肠杆菌目、拜氏菌属、拟杆菌科、理研菌Objective In this study,the modified persistent inflammatory-immunosuppressive-catabolic syndrome(PIICS)mouse model made the intestinal microbiota more similar to the intestinal microbiota of PIICS patients and preliminarily elaborated the diversity and structural changes of the intestinal microbiota.Methods Twenty-eight male 28-weekold C57BL/6 mice weighing 30-35 g were randomly divided into the Sham Control(SC)group(n=10)and the PIICS group(n=18).Based on the existing PIICS mouse model,measures were used to simulate the clinical process that may affect the gut microbiota,including early application of high-dose long-term antibiotics,large amounts of fluids,and long-term bed rest.On day 14,inflammatory factors such as tumor necrosis factor(TNF)-αand chemokines such as C-X-C motif chemokine ligand 1(CXCL1)were detected by ELISA.Flow cytometry was used to detect the proportion of CD8^(+)T cells and the proportion of CD11b^(+)myeloid-derived suppressor cells in the spleen to reflect the immunosuppression,and immunohistochemistry and western imprinting was used to detect the morphology and protein expression of muscle fibers to reflect the catabolism.The diversity and structural changes of intestinal microbiota were analyzed by sequencing with 16S rRNA gene.Results Compared with the SC group,the expression of inflammatory factors such as interleukin(IL)-6(t=132.67,P<0.0010),IL-10(t=65.76,P<0.0010),and TNF-α(t=23.77,P=0.0032)in the blood of mice in the PIICS group increased.In contrast,CXCL1(t=125.24,P<0.0010),granulocyte-macrophage colony-stimulating factor(GMCSF)(t=111.24,P<0.0010),and macrophage colony-stimulating factor(M-CSF)(t=154.22,P<0.0010)decreased expression;The proportion of CD8^(+)T cells decreased(t=37.11,P=0.0088),and the proportion of CD11b^(+)myeloid-derived suppressor cells increased(t=45.12,P=0.0095).The morphology of muscle fibers became polygonal,and the cross-sectional area was small(P<0.0010),and the expressions of muscle protein MURF-1(t=123.20,P=0.0073)and FoxO-1(t=67.34,P=0.0091)decreased.

关 键 词：持续性炎症-免疫抑制-分解代谢综合征 肠道菌群 小鼠模型 

分 类 号：R459.7[医药卫生—急诊医学]