CAJ | 학술논문

目的　观察香烟烟雾提取物(CSE)和酪氨酸激酶抑制剂(tyrophostin 25)对气道上皮细胞(AEC) 酪氨酸磷酸化程度、细胞Src蛋白(c-Src)及β-连环素（β-cat）表达的影响，探讨细胞酪氨酸磷酸化程度及β-cat在吸烟致AEC损伤和修复中的作用及调控机制。方法　体外培养猪AEC，将培养细胞分成3组：N组：加无血清培养基；C组：在无血清培养基上分别加CSE；CT组：加tyrophostin 25及CSE。每组随机检测300个AEC中β-cat免疫细胞化学染色的分布规律；利用图像分析技术，每组随机检测30个AEC的酪氨酸磷酸化程度、c-Src、β-cat mRNA的变化。结果　C组加入20%的CSE作用4 h和24 h后，AEC蛋白质酪氨酸磷酸化程度分别是N组的1.51倍和3.44倍(P<0.000 1)，胞膜上β-cat阳性表达率分别为26.7%和38.7%，较N组（分别为83.3%和81.3%）明显下降（P<0.001 0）。与N组比较C组胞质中c-Src的浓度分别降低了47.9%和51.4%(P<0.000 1)；CT组酪氨酸磷酸化程度及胞膜上β-cat表达的变化接近N组。而C组和CT组β-cat mRNA含量均较N组有明显下降（P<0.000 1）。结论　上皮细胞的蛋白质酪氨酸磷酸化程度的变化可能介导了香烟对气道上皮细胞的损伤，而β-cat蛋白质可逆性磷酸化对粘附连接的结构和功能的维持发挥重要作用。
목적　관찰향연연무제취물(CSE)화락안산격매억제제(tyrophostin 25)대기도상피세포(AEC) 락안산린산화정도、세포Src단백(c-Src)급β-련배소（β-cat）표체적영향，탐토세포락안산린산화정도급β-cat재흡연치AEC손상화수복중적작용급조공궤제。 방법　체외배양저AEC，장배양세포분성3조：N조：가무혈청배양기；C조：재무혈청배양기상분별가CSE；CT조：가tyrophostin 25급CSE。매조수궤검측300개AEC중β-cat면역세포화학염색적분포규률；이용도상분석기술，매조수궤검측30개AEC적락안산린산화정도、c-Src、β-cat mRNA적변화。결과　C조가입20%적CSE작용4 h화24 h후，AEC단백질락안산린산화정도분별시N조적1.51배화3.44배(P<0.000 1)，포막상β-cat양성표체솔분별위26.7%화38.7%，교N조（분별위83.3%화81.3%）명현하강（P<0.001 0）。여N조비교C조포질중c-Src적농도분별강저료47.9%화51.4%(P<0.000 1)；CT조락안산린산화정도급포막상β-cat표체적변화접근N조。이C조화CT조β-cat mRNA함량균교N조유명현하강（P<0.000 1）。결론　상피세포적단백질락안산린산화정도적변화가능개도료향연대기도상피세포적손상，이β-cat단백질가역성린산화대점부련접적결구화공능적유지발휘중요작용。
Objective　To explore the possible mechanism of injury by smoking and its repairing, the effects of cigarette smoke extract on the expression of β-catenin（β-cat），c-Src and level of phosphoric tyrosine in airway epithelial cells (AEC). Methods　Pig AECs were cultured in vitro. After CSE and tyrphostin 25 exposure, (1) the expression of β-cat was determined by immunocytochemical stain in 300 randomly selected AECs of each group, (2) the level of phosphoric tyrosine, content of c-Src and β-cat mRNA in AECs were evaluated by immunocytochemical stain, in situ hybridization and image analysis in 30 randomly selected AECs of each group. Results　After exposure to CSE for 4 hours and 24 hours, the levels of phosphoric tyrosine were 1.51 and 3.44 times of that of the control groups of the same time (P<0.0001) respectively; the ratio of positive stain of β-cat on membrane was markedly lower than that in the control groups (78.3% vs 83.3%, 77.7% vs 81.3%，P<0.0001）, and the cytoplasmic content of c-Src decreased 47.9% and 51.4% (P<0.0001) compared with the control groups of the same time, respectively. While dealt with CSE and tyrphostin 25 together, the level of phosphoric tyrosine, the expression of β-cat and c-Src resembled those in normal control. Contends of β-cat mRNA decreased in the both two situations (P<0.0010). Conclusion　CSE decreased the membranous expression of β-cat and increased the level of tyrosine phosporylation of AECs. The change of β-cat expression might mediate the airway epithelial injuries caused by smoking, and tyrphostin 25 might prevent such injuries through inhibiting the tyrosine kinase.