摘要
目的建立免疫力低下大鼠感染耐药株鲍曼不动杆菌肺炎模型,为临床防治细菌性肺炎提供动物模型支持。方法筛选鲍曼不动杆菌耐药株后,将48只SD大鼠随机分为正常组、环磷酰胺对照组(腹腔注射45 mg/kg环磷酰胺)、细菌感染组(气管滴注1.5×108 CFU鲍曼不动杆菌悬液)、细菌感染+免疫抑制组(腹腔注射45 mg/kg环磷酰胺+气管滴注1.5×108 CFU鲍曼不动杆菌悬液);采用流式细胞术检测感染前和感染后3、7 d外周血CD4+、CD8+、NK细胞比例;使用肺功能仪检测造模后3、7 d最大吸气流量(PIF)、最大呼气流量(PEF)、潮气量(Vt)、第2秒用力呼气容积/用力肺活量(FEV200/FVC);ELISA法检测肺泡灌洗液中IL-6、TNF-α及IL-10水平;HE染色观察各组大鼠肺组织形态;细菌培养计数观察肺组织细菌负荷。结果环磷酰胺对照组、细菌感染组和细菌感染+免疫抑制组大鼠造模后均出现自主活动减少,细菌感染组和细菌感染+免疫抑制组大鼠可闻及肺部啰音;与正常组比较,环磷酰胺对照组大鼠外周血中CD4+及NK细胞中CD11b比例减少,CD8+比例增加,细菌感染组大鼠PIF、PEF、Vt及FEV200/FVC降低,肺泡灌洗液中IL-6、TNF-α水平增加,IL-10水平降低,肺组织细菌负荷增多,并伴有轻度的肺泡壁增宽和炎症细胞浸润(P<0.05,P<0.01);与环磷酰胺对照组和细菌感染组比较,细菌感染+免疫抑制组大鼠外周血中CD4+比例减少,CD8+比例增加,PIF、PEF、Vt及FEV200/FVC减少,肺泡灌洗液中IL-6、TNF-α水平增加,IL-10水平降低,肺组织细菌负荷增多,肺泡壁增宽和炎症细胞浸润程度加重,且细菌感染+免疫抑制组大鼠外周血NK细胞中CD11b比例显著低于细菌感染组(P<0.05,P<0.01)。结论本实验采用鲍曼不动杆菌、鲍曼不动杆菌联合环磷酰胺免疫抑制剂构建鲍曼不动杆菌感染的细菌性肺炎模型,均能成功构建鲍曼不动杆菌感染性肺炎模型,其中鲍曼不动杆菌联合环磷酰胺免疫抑制剂构建肺炎模型较为严重,可能与环磷酰胺降低机体细胞免疫功能,加重肺部细菌感染有关。
Objective To establish an animal model of pneumonia for research on clinical prevention and treatment of bacterial pneumonia by infecting immunocompromised rats with drug-resistant Acinetobacter baumannii(Ab)strains.Methods Drug-resistant Ab strains were selected.Forty-eight SD rats were randomly divided into four groups:normal group,cyclophosphamide control group(intraperitoneal injection of 45 mg/kg cyclophosphamide),bacterial infection group(intratracheal instillation of 1.5×108 CFU Ab suspension),and bacterial infection+immunosuppression group(intraperitoneal injection of 45 mg/kg cyclophosphamide+intratracheal instillation of 1.5×108 CFU Ab suspension).Flow cytometry analysis was used to detect the proportion of CD4+,CD8+and NK cells in rat peripheral blood before as well as 3 d and 7 d after infection.A lung function meter was used to detect peak inspiratory flow(PIF),peak expiratory flow(PEF),tidal volume(Vt)and forced expiratory volume in the second second/forced vital capacity(FEV200/FVC)at 3 d and 7 d after modeling.ELISA was used to detect the levels of IL-6,TNF-αand IL-10 in the alveolar lavage fluid.HE staining was used to observe the morphology of rat lung tissues in each group.Bacterial loads in rat lung tissues were counted by bacterial culturing.Results A decrease in voluntary activity was observed in rats in the cyclophosphamide control group,bacterial infection group and bacterial infection+immunosuppression group after modeling.Lung rales could be heard in the bacterial infection group and bacterial infection+immunosuppression group.Compared with the normal group,the cyclophosphamide control group showed decreased proportion of CD4+and CD11b+NK cells and increased CD8+cells in peripheral blood;the bacterial infection group showed decreased PIF,PEF,Vt and FEV200/FVC,increased IL-6 and TNF-αlevels and decreased IL-10 level in the alveolar lavage fluid,and higher bacterial load in lung tissues with mild widening of alveolar walls and inflammatory cell infiltration(P<0.05,P<0.01).Compared with the cyclophosphamide control group and the bacterial infection group,the bacterial infection+immunosuppression group showed a lower proportion of CD4+cells and a higher proportion of CD8+cells in rat peripheral blood,decreased PIF,PEF,Vt and FEV200/FVC,increased IL-6 and TNF-αlevels and decreased IL-10 level in alveolar lavage fluid,higher bacterial load in lung tissues,and aggravated widening of alveolar walls and inflammatory cell infiltration(P<0.05,P<0.01).The proportion of CD11b+NK cells in peripheral blood of rats in the bacterial infection+immunosuppression group was significantly lower than that in the bacterial infection group(P<0.05,P<0.01).Conclusions A bacterial pneumonia model was successfully constructed by infecting rats with Ab alone or in combination with cyclophosphamide immunosuppression.In the model constructed with Ab and cyclophosphamide immunosuppression,the rats had more severe pneumonia,which might be related to the reduced cellular immune function and the aggravated bacterial infection in rat lung tissues by cyclophosphamide.
作者
唐梓宁
刘学武
兰前清
唐海亮
肖洒
袁湘中
彭冬冬
Tang Zining;Liu Xuewu;Lan Qianqing;Tang Hailiang;Xiao Sa;Yuan Xiangzhong;Peng Dongdong(Hunan Key Laboratory of Pharmacodynamics and Safety Evaluation of New Drugs,Hunan Prima Drug Research Center Co.Ltd.,Changsha 410329,China)
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2023年第12期908-917,共10页
Chinese Journal of Microbiology and Immunology
基金
湖南省企业科技创新创业团队支持计划(2021)。
关键词
鲍曼不动杆菌
环磷酰胺
感染性肺炎
疾病模型构建
Acinetobacter baumannii
Cyclophosphamide
Infectious pneumonia
Disease model establishment
作者简介
通信作者:彭冬冬,Email:pengdongdong@hnse.org,电话:0731-83285167。
