痰热清对呼吸道合胞病毒感染人支气管上皮细胞分泌胸腺基质淋巴细胞生成素作用的体外研究.docVIP

痰热清对呼吸道合胞病毒感染人支气管上皮细胞分泌胸腺基质淋巴细胞生成素作用的体外研究 　　作者：蓝丹　檀卫平　陈环　吴葆菁　麦贤弟　黄花荣 作者单位：510120　广州，中山大学孙逸仙纪念医院儿科 　　【摘要】 目的　探讨痰热清对体外呼吸道合胞病毒(RSV)感染人支气管上皮细胞(NHBE)分泌胸腺基质淋巴细胞生成素(TSLP)的影响。方法　用Hep-2细胞扩增病毒，并进行毒力鉴定。建立RSV感染NHBE体外细胞模型。酶联免疫吸附(ELISA)法检测RSV感染NHBE后TSLP水平。将痰热清以不同给药方式(预防给药、直接灭活、治疗给药方式)干预RSV感染NHBE，ELISA法检测TSLP水平变化。结果　实时荧光定量RT-PCR方法鉴定细胞模型建立成功。(1)RSV对NHBE分泌TSLP的影响：相同感染时间内(12 h、24 h、48 h、72 h、96 h、120 h)，随着感染滴度增加(10 TCID50、50 TCID50、100 TCID50、500 TCID50、1000 TCID50)，各感染组TSLP分泌水平均呈上升趋势，与正常细胞对照组比较差异均有统计学意义(P均0.05)。相同感染滴度内(10 TCID50、50 TCID50、100 TCID50、500 TCID50、1000 TCID50)，随着感染时间增加(24 h、48 h、72 h、96 h、120 h)，各感染组TSLP分泌水平均呈上升趋势，与12 h组比较差异均有统计学意义(P均0.05)。(2)痰热清对RSV感染NHBE分泌TSLP的影响：在12 h、24 h、48 h、72 h给药时间内，痰热清、利巴韦林的不同给药方式与病毒对照组比较，均下调TSLP水平(P0.05)，以预防给药方式最为明显。预防给药方式下，痰热清组较利巴韦林组下调TSLP的作用更明显(P0.05)。结论　RSV能诱导NHBE分泌TSLP，痰热清可抑制RSV引起的TSLP分泌增加，以预防方式给药明显，痰热清可在病毒感染的控制中起到重要作用。 　　【关键词】 呼吸道合胞病毒;　上皮细胞;　支气管;　胸腺基质淋巴细胞生成素;　痰热清 　　【Abstract】　Objective　To study effects of traditional Chinese medicine Tanreqing injection on the secretion of thymic stromal lymphopoietin (TSLP) in human bronchial epithelial cell infected by respiratory syncytial virus (RSV) in vitro.Methods　1.Hep-2 cell lines were cultured in vitro and infected by RSV.Then the infectious titer of RSV was determined by TCID50 technique.We established a model of NHBE infected by RSV.2.NHBE were infected with series RSV titers and in series incubated hours.The level of TSLP in the culture supernatants was determined by enzyme-linked immunosorbent assay(ELISA).3.The effect of Tanreqing treatment on TSLP secretion was based on cell culture in vitro.The level of TSLP in three types of drug delivery(including precaution，direct deactivation and therapy) were determined by ELISA.Results　1.The mRNA of RSV was detected in total RNA of NHBE infected by RSV with QPCR method，which demonstrated that a model of RSV infection in vitro was established succesfully.2.The level of TSLP in NHBE cells supernatants infected by RSV significantly incre