【摘要】 目的建立肺泡Ⅱ型上皮细胞(AECⅡ)与肺成纤维细胞(LF)共培养模型,检测胎鼠AECⅡ共培养和单独培养时Notch1、3受体表达的变化。方法应用Millipore插入式培养皿和Costar六孔板构建AECⅡ/LF共培养模型,将未接种LF的Millicell2PCF插入式细胞培养皿置于接种有AECⅡ的培养板为AECⅡ单独培养组。光镜、透射电镜观察共培养与单独培养条件下AECⅡ大体形态、超微结构;四甲基偶氮唑蓝(MTT)法检测其AECⅡ增殖活力;反转录PCR法检测其AECⅡ分泌表面活性蛋白C(SP-C)功能,并用荧光定量PCR法检测其AECⅡNotch1、3mRNA的表达。结果AECⅡ/LF共培养组较AECⅡ单独培养组稳定保持了其立方形结构和细胞表型并克隆样生长,增殖活力强。电镜下可见单独培养48hAECⅡ呈现AECⅠ样改变,而共培养AECⅡ维持其正常生理形态。与单独培养组比较,共培养组AECⅡ分泌SP-C明显增多(P<0.05),Notch1 mRNA表达显著升高(P<0.05),而Notch3 mRNA显著降低(P<0.05)。结论AECⅡ同型细胞培养时转分化为AECⅠ,不同型细胞(AECⅡ/LF)共培养抑制其转分化,此转分化过程受Notch信号分子调控。更多还原

【Abstract】 Objective To establish a co-culture model of alveolar epithelial type Ⅱ cell (AECⅡ) and lung fibroblast(LF) to detect the changes of of Notch1 and Notch3 receptors in heterocellular and homocellular cultures.Methods By utilizing Millipore snap-on plate and Costar six-well plate,AECⅡ/ LF co-culture system was established and put together the freshly isolated AECⅡs and 3rd passage LFs or AECⅡs alone for group research.The general configuration and ultramicrostructure of the 2 groups were observed by light microscope and transmission electron microscope separately.MTT method was used to detect their cell vitality and function of secreting SP-C(surfactant protein C)by reverse transcription PCR. Notch1, 3 mRNA were detected by fluorescent quantitative real-time PCR.Results AECⅡs of co-culture group showed a more stable configuration,more powerful vitality and secreted more SP-C. Notch1 mRNA in co-culture group was higher than homocellular culture while Notch3 mRNA is was lower.Conclusions AECⅡs of homocellular group may transdifferentiate into AECⅠs.When co-cultured with LF, AECⅡtransdifferentiation is suppressed, which is probably regulated by Notch signal.更多还原