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目的探讨3株不同来源的铜绿假单胞菌分泌性蛋白的抗肿瘤活性。方法将3株不同来源的铜绿假单胞菌经LB培养基对其过夜静置培养,离心获得上清液,采用硫酸铵盐析沉淀蛋白质,再经PBS透析除盐。然后将蛋白作用于人肝癌细胞Hep-G2、宫颈癌细胞HeLa、肺癌细胞A549和人永生化表皮细胞HaCaT,CCK-8检测其对细胞的毒性作用,吉姆萨染色观察凋亡细胞形态变化。结果 SDS-PAGE证实成功获得不同分子量的分泌蛋白,CCK-8结果显示混合蛋白对3种不同肿瘤细胞的生长均有不同程度的抑制作用,且其抑制作用存在一定的时间和浓度依赖性,但对人正常细胞无明显抑制作用。吉姆萨染色初步显示细胞形态为凋亡状态。结论初步证实该3株铜绿假单胞菌产生的分泌性蛋白具有不同程度的抗肿瘤作用,为进一步分离纯化具有抗肿瘤活性的单一蛋白质及研究其抗肿瘤机制提供依据。
Objective To investigate the antitumor activity of three secreted proteins of Pseudomonas aeruginosa from three different sources. Methods Three strains of Pseudomonas aeruginosa from different sources were cultured overnight in LB medium and centrifuged to obtain the supernatant. The proteins were precipitated by ammonium sulfate and desalted by dialysis with PBS. Then, the effect of the protein on Hep-G2, HeLa, A549 and HaCaT and CCK-8 cells was detected by MTT assay. The morphological changes of apoptotic cells were observed by Giemsa staining. Results SDS-PAGE confirmed the successful production of secreted proteins with different molecular weights. The results of CCK-8 showed that the mixed protein had different degrees of inhibition on the growth of three different tumor cells, and the inhibitory effect was time and concentration-dependent No significant inhibition of normal human cells. Giemsa staining initially showed that the cell morphology was apoptotic. Conclusions The secreted proteins produced by the three strains of Pseudomonas aeruginosa are preliminary confirmed to have antitumor activity in varying degrees and provide the basis for further isolation and purification of a single protein with anti-tumor activity and to study its anti-tumor mechanism.