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目的:研究携带阿尔法干扰素(hαIFN)基因的内皮祖细胞(EPCs)对肿瘤免疫治疗的效果。方法:用健康捐献者的骨髓体外诱导培养EPCs,携带hαIFN基因的腺病毒Ad5-hαIFN转染EPCs制备EPCs-hαIFN,ELISA法检测EPCs-hαIFN培养上清液中hαIFN浓度。体外EPCs-hαIFN与HepG-2肝癌细胞按照1∶5的浓度混合培养,观察肿瘤细胞增殖情况。制作裸鼠肝癌模型,静脉注射EPCs-hαIFN,观察对肿瘤的抑制作用和对荷瘤动物生存期的影响。结果:使用人骨髓可以培养出CD133、KDR和CD34阳性的EPCs。携带hαIFN基因的EPCs-hαIFN培养液中可以测得hαIFN,最高浓度可以达1400pg/ml以上,在相当长的一段时间内细胞培养液的hαIFN浓度保持稳定。EPCs-hαIFN与HepG-2细胞按照1∶5的浓度混合培养4天,HepG-2细胞的存活率为(70.50±3.33)%(P<0.05)。通过静脉注射EPCs-hαIFN可以抑制肿瘤的生长[肿瘤重量:EPCs-hαIFN组(0.71±0.23)g,空白对照组(3.27±1.28)g,P<0.05],而皮下注射hαIFN和静脉注射EPCs-EGFP对肿瘤没有抑制作用。尾静脉注射EPCs-hαIFN可延长荷瘤裸鼠的生存时间(中位生存时间:EPCs-hαIFN组为39天,空白对照组为33天,P<0.05)。结论:肿瘤靶向性的EPCs-hαIFN可以通过分泌hαIFN抑制肿瘤的进展。
Objective: To study the effect of endothelial progenitor cells (EPCs) carrying alpha-interferon (IFN-α) gene on tumor immunotherapy. Methods: EPCs-hαIFN were transfected into EPCs by transfection of EPCs with adenovirus Ad5-hαIFN carrying hαIFN gene in bone marrow of healthy donors in vitro. The concentration of hαIFN in EPCs-hαIFN culture supernatants was detected by ELISA. In vitro EPCs-hαIFN and HepG-2 hepatocellular carcinoma cells were mixed and cultured according to the concentration of 1: 5 to observe the proliferation of tumor cells. The nude mouse model of hepatocellular carcinoma was established. EPCs-hαIFN was injected intravenously to observe the inhibitory effect on tumor and the effect on the survival of tumor-bearing animals. Results: CD133, KDR and CD34-positive EPCs could be cultured using human bone marrow. The hαIFN can be measured in EPCs-hαIFN culture medium carrying hαIFN gene, and the highest concentration can reach 1400 pg / ml. The hαIFN concentration of cell culture medium remained stable for a long period of time. EPCs-hαIFN and HepG-2 cells were mixed and cultured at the concentration of 1: 5 for 4 days. The survival rate of HepG-2 cells was (70.50 ± 3.33)% (P <0.05). EPCs-hαIFN group (0.71 ± 0.23g) and blank control group (3.27 ± 1.28g g, P <0.05) were also inhibited by intravenous injection of EPCs-hαIFN, while subcutaneous injection of hαIFN and EPCs- EGFP does not inhibit the tumor. The tail vein injection of EPCs-hαIFN prolonged the survival time of tumor-bearing nude mice (median survival time: 39 days in EPCs-hαIFN group and 33 days in blank control group, P <0.05). Conclusion: Tumor-targeted EPCs-hαIFN can inhibit tumor progression by secreting hαIFN.