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目的:使用流体力学方法向人肝癌裸鼠原位移植瘤转染含突变型IκBα的重组逆转录病毒质粒pBABE-puro-IκBα super repressor(pBABE-puro-IκBαSR),观察其对移植瘤生长的作用及其相关机制。方法:使用人肝癌细胞株MHCC97-H、MHCC97-L,建立裸鼠人肝癌原位移植瘤模型,将荷瘤鼠分为MHCC97-H组(对照组)、MHCC97-H+IκBαSR组(转染组)、MHCC97-L组(对照组)和MHCC97-L+IκBαSR组(转染组),1周后,使用流体力学方法通过尾静脉向对照组注射pBABE-puro空载体质粒,向转染组注射pBABE-puro-IκBαSR质粒。每周1次,共3次。观察荷瘤鼠的生存率,检测原位移植瘤块的大小、重量、以及转移瘤数目,使用全自动生化检测仪检测血清中丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)水平。采用免疫组化SP法分析核因子(NF)-κBp65蛋白表达,使用Westernblot检测总蛋白中IκBα蛋白含量。结果:pBABE-puro-IκBαSR质粒显著提高荷瘤小鼠的生存率,转染组肿瘤的体积、重量显著小于对照组,抑瘤率分别为29.3%和33.0%(均P<0.05);转染组荷瘤小鼠ALT和AST显著低于对照组(均P<0.05);NF-κBp65在对照组胞浆中及在细胞核中均呈强阳性表达,在转染组胞浆中呈浅棕色,为弱阳性表达,胞核中极少或没有阳性表达,差异具有统计学意义(P<0.01)。Western blot结果显示,转染组IκBα表达水平高于对照组(P<0.05)。结论:通过流体力学方法向人肝癌原位移植瘤转染突变型IκBαSR质粒,可以抑制移植瘤的生长。其机制可能是IκBαSR抑制了NF-κB的活化。
OBJECTIVE: To transfect human orthotopic transplanted hepatocellular carcinoma xenografts into human orthotopically transplanted human hepatocellular carcinoma xenografts with the recombinant retrovirus (pBABE-puro-IκBα super repressor) (pBABE-puro-IκBαSR), and observe its effect on the growth of xenografts. And its related mechanisms. Methods: Human hepatocellular carcinoma cell lines MHCC97-H and MHCC97-L were used to establish an orthotopic xenograft model of human hepatocellular carcinoma in nude mice. The mice were divided into MHCC97-H group (control group) and MHCC97-H+IκBαSR group (transfection group). Group), MHCC97-L group (control group) and MHCC97-L+IκBαSR group (transfected group), 1 week later, the pBABE-puro empty vector plasmid was injected into the control group via the tail vein using a hydrodynamic method and transfected into the transfection group. The pBABE-puro-IκBαSR plasmid was injected. Once a week, a total of 3 times. The survival rate of tumor-bearing mice was observed, and the size, weight, and number of metastases were measured. The alanine aminotransferase (ALT) and aspartate aminotransferase in serum were detected using an automatic biochemical analyzer. (AST) level. The expression of nuclear factor-κBp65 protein was analyzed by immunohistochemical SP method, and the protein content of IκBα was detected by Western blot. RESULTS: The pBABE-puro-IκBαSR plasmid significantly increased the survival rate of tumor-bearing mice. The volume and weight of tumors in the transfection group were significantly lower than those in the control group. The tumor inhibition rates were 29.3% and 33.0%, respectively (all P<0.05); The ALT and AST in the tumor-bearing mice were significantly lower than those in the control group (all P<0.05). NF-κB p65 was strongly expressed in the cytoplasm and in the nucleus of the control group, and was light brown in the cytoplasm of the transfection group. For weakly positive expression, there was little or no positive expression in nucleus, and the difference was statistically significant (P<0.01). Western blot results showed that the expression level of IκBα was higher in the transfection group than in the control group (P<0.05). CONCLUSION: Transfection of the mutant IκBαSR plasmid into human hepatocellular carcinoma orthotopic transplanted tumors by hydrodynamics can inhibit the growth of the transplanted tumor. The mechanism may be that IκBαSR inhibits the activation of NF-κB.