蝎毒多肽提取物抑制肝癌H22细胞化疗期间再增殖实验研究

Inhibitive effect of polypeptide extract from scorpion venom on repopulation in H22 tumor cell during chemotherapy

目的：观察蝎毒多肽提取物（polypeptide extract from scorpion venom，PESV）对H22肝癌荷瘤小鼠化疗期间再增殖的抑制作用并探讨可能的机制。方法：96只Balb／c小鼠皮下接种小鼠肝癌H22细胞，随机分为模型组、PESV高、低剂量组和衙瘤对照组。以5．Fu对荷瘤小鼠进行化疗建立再增殖模型。按不同的用药方法对4组小鼠进行干预，每周测量肿瘤体积2次。各组每7d处死6只小鼠，实验共进行35d。以免疫组织化学方法检测各组肿瘤组织CD105，PCNA，VEGF，PDGF蛋白水半表达。以CD105标记微血管，计算微血管密度（MVD）。结果：荷瘤对照组肿瘤体积在13—24d增加迅速，荷瘤对照组小鼠在第27天全部死亡，模型组肿瘤体积在17d前增长较快，在第17～22天增加较慢，之后体积增加又较快，在第31天全部死亡。PESV高、低剂量组肿瘤体积全程缓慢增加，体积在17d以后显著低于模型组，PESV高低剂量组之间仅在第17天存在差异（P〈0．05）。免疫组织化学检测湿示，模型组肿瘤组织第31天PCNA表达水平高于第21，28天（P〈0．01），PESV高、低剂量组表达水平显著低于模型组（P〈0．01），PESV高、低剂量组仅在第17天存在显著差异（P〈0．05）。免疫组织化学检测显示，与PESV高、低剂量组相比，模型组CD105-MVD在第21，28天（P〈0．05），第35天（P〈0．01）存在显著差异，PESV高低剂量组之间无差别。模型组VEGF表达第35天高于第21，28天（P〈0．05），PESV高、低剂量VEGF表达在第21，28，35天表达水平均显著低于模型组（P〈0．01），PESV高低剂量组无差别。模型组肿瘤PDGF表达水平逐渐下降，PESV高，低组存第21天表达水平最低，之后逐渐增高，PESV高低剂量组之间在第35天存在显著差异（P〈0．01）。相关性分析显示，肿瘤组织VEGF表达水平与肿瘤组织CD105-MVD呈正相关（r=0．669）。结论：PESV可抑制H22肿瘤在化疔期间再增殖作用，其机制可能是抑制血管生成和使肿瘤血管正常化。

Objective： To observe the inhabitive effect and mechanism of polypeptide extract from scorpion venom （PESV）on repopulation in H22 tumor cell during chemotherapy. Method： H22 tumor cells were injected into 96 mice subcutaneously, then mice were divided into 4 groups radomly ： Model, low-dose-PESV, high-dose-PESV, and control. Reppulation model was established by 5- Fu treating mice with H22. Four groups was treated differectly, 6 mice of each group was sacrificed every 7 days, measured tumor volume twice one week. The expression of vascular endothelial growth factor（ VEGF ） , proliferating cell nuclear antigen（PCNA） , CD105 microvessel density（CD105-MVD） and platelet derived growth factor （PDGF） in H22 tumor issue was observed by using immunohistochemistry and grey analysis, the relation of VEGF and MVD was affirmed by correlation analysis. Result： In control group tumor volume of H22 increased quickly in 13-24 day, and all mice died before 27 day. In model tumor volume increased quickly before 17, in 17-22 day slowly, after 22 day quickly again, and all the mice died before 31 day. In low and high dose PESV, tumor volume added slowly, and only in 17 day there was significant difference between these two groups. Immunohistochemistry showed , PCNA expression of model group in 31 day was higher than in 21,28 day, the expression level of high and low PESV group was lower than model group all the time, only in 17 day there was significant difference between high and low PESV group. Immunohistochemitry showed, compared with high and low dose PESV group, CD105-MVD of model group was higher in 21, 28 day（P 〈0. 05） and in 35 day （P 〈 0. 01 ） , and no difference was found between high and low dose PESV. VEGF expression of model group in 35 day was higher than in 21, 28 day（P 〈0. 01 ） , and model group higher than high and low dose PESV in 21, 28, 35 day. The expression of PDGF in model decreased gradually, in high and low dose PESV, the expression was lowest in 21 day. In day 35 high dose PESV higher than low dose PESV. There was positive correlation（ r = 0. 669）between VEGF expression and CD105-MVD. Conclusion： PESV can inhabit repopulation of H22 tumor cell during chemotherapy, and the mechanism maybe is through antingiogenesis and nomalizing tumor vessels.