良性气道狭窄小标本培养成纤维细胞方法中抗生素浓度的探讨  

An Investigation of Antibacterial Pretreatment Protocol for Primary Fibroblast Culture from Transbronchial Biopsies in Benign Tracheal Stenosis Patients

作　　者：王颖治[1] 陈愉[1] 李时悦[1] 

机构地区：[1]广州医科大学附属第一医院呼吸疾病国家重点实验室呼吸疾病国家临床研究中心,广东广州510120

出　　处：《中国呼吸与危重监护杂志》2015年第3期286-290,共5页Chinese Journal of Respiratory and Critical Care Medicine

基　　金：国家卫生和计划生育委员会公益性行业专项项目(编号:201402024);“十二五”国家科技支撑计划项目(编号:2012BAI05B01)

摘　　要：目的经支气管镜获取良性气道狭窄患者气道内肉芽组织，探讨小标本培养原代成纤维细胞的不同浓度抗生素处理方法。方法经支气管镜活检获得14例良性气道狭窄的患者15份气道内增生性肉芽组织，根据预处理方法不同分成3组：（1）常规抗生素浓度预处理组（组1），5份标本，给予1％～2％青霉素／链霉素（简称双抗）的磷酸盐缓冲液（PBS）预处理；（2）高浓度抗生素预处理组（组2），5份标本，给予6％双抗PBS预处理；（3）酒精联合高浓度抗生素预处理组（组3），5份标本，先用75％酒精漂洗3～4s，再用含6％双抗PBS漂洗干净。所有标本经不同的预处理后均用组织块法进行培养。结果（1）组1未能成功培养出成纤维细胞，并可见大量的细菌生长。组2和组3均能成功培养出成纤维细胞；（2）3种预处理方法培养成纤维细胞，细胞的增殖最远半径在不同的时间点的差异具有统计学意义（P〈0．01）。3种预处理方法培养成纤维细胞，在24、48及72h，3种方法间无差异（P〉0．05），在96h，3种方法间比较差异有统计学意义（P〈0．05）。结论经支气管镜获取气道肉芽组织的小标本，有效杀灭组织块细菌的预处理方法是培养原代成纤维细胞的重要因素。可采用增加抗生素的浓度和酒精漂洗进行预处理。Objective To investigate the antibacterial pretreatment protocol for primary fibroblast cell culture from transbronchial biopsies in patients with benign tracheal stenosis （BTS）. Methods Fifteen specimens of intratracheal hyperplastic granulation tissue were obtained from 14 BTS patients by transbronchial biopsies. The tissues were divided into 3 groups according to different antibacterial pretreatment with 5 specimens in each group. An usual concentration of antibiotics pretreatment group （group 1 ） was pretreated by washing with PBS contained 1% - 2% penicillin/streptomycin. A high concentration of antibiotics pretreatment group （ group 2） was pretreated by washing with PBS contained 6% penicillin/streptomycin. An alchohol and high concentration of antibiotics pretreatment group （ group 3） was pretreated by washing with 75% alcohol 3 - 4 seconds firstly, then by washing with 6% penicillin/ streptomycin. After different pretreatment, all tissues were cultured with tissue culture method in the same condition. Results The primary fibroblasts were successfully cultured from the tissue pretreated by method 2 and 3,but not cultured from the tissue pretreated by method 1 with large amount of bacteria. There were significant differences in the furthest radius of cell proliferation between different culture time points in three groups （P 〈 0.01 ）. The differences in the furthest radius of cell proliferation between three groups were not different at 24,48 or 72 h （ P 〉 O. 05 ）, but were significant between three groups at 96 h （ P 〈 O. 05 ）. Conclusion An pretreatment protocol with high concentration of antibiotics or 75% alcohol is feasible in human primary fibroblasts culture from small specimens obtained by transbronchial biopsy.

关 键 词：肉芽组织 成纤维细胞 原代细胞培养 抗生素 细菌 

分 类 号：R56[医药卫生—呼吸系统]