重组铜绿假单胞菌分泌蛋白PA3611诱导支气管上皮细胞-间充质转化的初步观察

Pseudomonas aeruginosa recombinant protein PA3611 induces epithelial-mesenchymal transformation of bronchial epithelial cells

目的 探讨重组蛋白PA3611能否在体外使大鼠支气管上皮细胞发生上皮-间充质转化（EMT）.方法 通过DNA合成、基因扩增、载体构建、诱导表达、纯化等过程制备重组蛋白PA3611;利用CCK-8法检测PA3611刺激对支气管上皮细胞增殖的影响;镜下观察经PA3611作用后支气管上皮细胞的形态学变化;运用Western blot法和qPCR法检测支气管上皮细胞细胞E-钙粘蛋白（E-CAD）与 α-平滑肌肌动蛋白（α-SMA）的表达水平.结果 DNA测序证实成功制备重组蛋白PA3611;重组蛋白PA3611刺激能抑制支气管上皮细胞的增殖（P〈0.05）,诱导其细胞形态学发生EMT变化,抑制E-CAD表达及促进α-SMA的表达（P〈0.05）.结论 重组铜绿假单胞菌分泌蛋白PA3611可以诱导大鼠支气管上皮细胞发生EMT,其具体机制值得进一步深入探讨.

Objective To investigate whether Pseudomonas aeruginosa recombinant protein PA3611 can induce the in vitro epithelial-mesenebyma[ transformation （EMT） of rat bronchial epithelial cells. Methods A series of processes including DNA synthesis, gene amplification, vector construction, induction of expression and protein purification was used to synthesize the recombinant protein PA3611. CCK-8 kit was used to detect the proliferation of bronchial epithelial cells after stimulation with the protein PA3611. Morphological changes in bronchial epithelial cells were observed. Western blot and qPCR were performed to measure the expression of E-cadherin （E-CAD） and a-smooth muscle actin （a-SMA）. T test was used for statistical analysis. Results DNA sequencing verified the successful preparation of the recom- binant protein PA3611. PA3611 inhibited the proliferation （P〈0.05） and induced the EMT of bronchial epithelial cells. Moreover, it also inhibited the expression of E-CAD protein, but promoted the expression of a- SMA （P〈0.05）. Conclusion The recombinant protein PA36l 1 can induce the EMT of bronchial epithelial cells and the underlying mechanisms are worthy of further investigation.