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目的探讨肿瘤增殖相关基因Ki67反义肽核酸(PNAs)、反义寡核酸(ASODNs)对人肾癌细胞增殖及凋亡的调控。寻找肾癌反义治疗的合适药物。方法将PNAs转染人肾癌786-0细胞系,采用免疫组化、Westernblot技术检测Ki67表达,细胞生长曲线、3H-thymidine掺入试验检测肾癌细胞增殖,TUNEL法检测癌细胞凋亡。并与相同浓度的ASODNs进行对比。结果PNAs处理组(10μmol/L)786-0细胞Ki67表达阳性率(%)(16.9±0.7)降低,Ki67蛋白(%)(42.1±2.2)降低,与ASODNs处理组(28.6±0.4)(83.6±1.4)比较差异有显著性(P<0.01,P<0.01)。PNAs处理组3H-thymidine掺入率(%)(20.7±1.5)减少,与ASODNs处理组(58.6±1.4)比较差异有显著性(P<0.01)。PNAs处理组凋亡细胞阳性率(%)(28.7±2.3)增加,与ASODNs处理组(13.8±1.0)比较差异有显著性(P<0.01)。结论PNAs可在反义及反基因二个环节发挥抗肿瘤作用,与ASODNs相比,PNAs有更强的阻抑人肾癌Ki67基因表达、增殖及促进凋亡作用,是一种有前途的基因治疗药物。
Objective To investigate the effects of Ki67 antisense oligodeoxynucleotides (PNAs) and antisense oligodeoxynucleotides (ASODNs) on the proliferation and apoptosis of human renal cell carcinoma cells. Looking for antineoplastic treatment of renal cancer appropriate drugs. Methods PNAs were transfected into human renal cell carcinoma 786-0 cell line. Immunohistochemistry and Western blotting were used to detect the expression of Ki67 and cell growth curve. 3H-thymidine incorporation assay was used to detect the proliferation of renal cancer cells. TUNEL assay was used to detect the apoptosis of cancer cells. And compared with the same concentration of ASODNs. Results The Ki67 positive rate (16.9 ± 0.7) and Ki67 protein (42.1 ± 2.2) in 786-0 cells of PNAs treatment group were significantly lower than that of ASODNs treatment group (28.6 ± 0.4) (83.6 ± 0.4) ± 1.4), the difference was significant (P <0.01, P <0.01). 3H-thymidine incorporation (%) (20.7 ± 1.5) decreased in PNAs-treated group compared with that in ASODNs-treated group (58.6 ± 1.4) (P <0.01). The positive rate of apoptotic cells in PNAs treatment group (28.7 ± 2.3) was significantly higher than that in ASODNs treatment group (13.8 ± 1.0) (P <0.01). Conclusions PNAs can exert anti-tumor effect in both antisense and anti-gene. Compared with ASODNs, PNAs have a stronger inhibitory effect on Ki67 gene expression, proliferation and apoptosis in human renal cell carcinoma, which is a promising gene medicine.