能量可控陡脉冲联合H-ras干扰性RNA治疗卵巢癌裸鼠皮下移植瘤的实验研究

Experimental study of energy controllable steep pulses and RNA interference targeting H-ras gene on inhibiting growth of xenografted tumor of ovarian cancer cells

目的 利用陡脉冲电场的可逆性电穿孔电转基因,观察陡脉冲治疗仪联合基因治疗对肿瘤生长的影响。方 法 构建抗H- ras基因表达的小发夹状RNA重组质粒载体(pshRNA H- ras);建立卵巢癌荷瘤裸鼠模型,光、电镜观察陡脉 冲对肿瘤组织的病理学改变,荧光报告质粒观察陡脉冲电场的可逆性电穿孔;实验分为4组:对照组、陡脉冲治疗组、陡脉 冲加基因单次干扰治疗组和加基因重复干扰治疗组,记录治疗后21d各荷瘤鼠肿瘤的体积和体质量。结果 陡脉冲对肿 瘤组织有明显的破坏作用,肿瘤边缘残存有活性的癌细胞;各治疗组肿瘤体积与体质量明显低于对照组,差异具有极显著 性(P<0.01),其中,陡脉冲加基因重复干扰治疗组肿瘤体积和重量显著低于仅用陡脉冲治疗组(P<0.01)。结论 陡脉 冲是一种新型有效的治癌工具,联合基因治疗能更有效地抑制肿瘤生长。

Objective To evaluate the effects of energy controllable steep pulses(ECSP) and the small hairpin RNA recombinant plasmid targeting H-ras gene (pshRNA/H-ras) on the growth of xenografted ovarian tumor cells. Methods The recombinant plasmids targeting H-ras gene coding sequence (pshRNA/H-ras) were constructed successfully. ECSP were applied to ovarian tumor-bearing nude mice. After exposed to ECSP, tissues were obtained and present for routine HE and transmission electron microscope (TEM) observation. The reversible electroporation were observed by fluorescent reporter plasmids. Tumor modeling were achieved in 20 nude mice and divided into four groups: ECSP group, ECSP and pshRNA/H-ras 1 (injecting one time), ECSP and pshRNA/H-ras 4 (injecting 4 times), and unexposed group. The volume and weight of tumors in each group were recorded at predetermined times. Results ECSP show effectively destructive action as compared with unexposed group. Irreversible cell changes could be seen under TEM. There were survived cells close to the necrotized regions. The volume and weight of tumors in each experimental group were significantly smaller than unexposed group (P<0.01). In ECSP and pshRNA/H-ras 4 group, ECSP was far more effective than ECSP group in retarding tumor growth (P<0.01). Conclusion ECSP is a new and effective way for solid tumor treatment and can inhibit the growth of tumors with the combination of gene method.