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目的探讨神经纤毛蛋白1(Neuropilin 1,NRP1)基因小干扰RNA(siRNA)质粒对裸鼠胃癌移植瘤生长的抑制作用。方法设计靶向人NRP1基因的siRNA序列,合成一对互补的寡核苷酸,退火后克隆至质粒pGenesil-1.1上,构建重组质粒siRNA-NRP1;将人胃癌细胞SGC7901注入裸鼠右臀部皮下,构建移植瘤模型,成瘤后将重组质粒siRNA-NRP1注入瘤内,观察裸鼠临床表现,并测定裸鼠移植瘤的体积及重量;病理切片观察肿瘤组织及血管生长情况;免疫组化方法观察裸鼠移植瘤中NRP1的表达及微血管密度。结果酶切分析及测序鉴定证实目的寡核苷酸片段已克隆至质粒pGenesil-1.1中;siRNA-NRP1组裸鼠移植瘤的体积及重量均小于对照组(P<0.01);病理切片显示siRNA-NRP1组肿瘤细胞坏死较对照组明显;免疫组化显示siRNA-NRP1组裸鼠移植瘤中NRP1的表达及微血管密度均低于对照组(P<0.01)。结论通过siRNA-NRP1抑制裸鼠胃癌移植瘤中NRP1的表达,可影响裸鼠移植瘤中血管的生成,最终抑制肿瘤的生长。
Objective To investigate the inhibitory effect of neuropilin 1 (NRP1) gene small interfering RNA (siRNA) plasmid on the growth of transplanted gastric cancer in nude mice. Methods siRNA targeting NRP1 gene was designed and synthesized. A pair of complementary oligonucleotides were designed and annealed, then cloned into plasmid pGenesil-1.1 to construct recombinant plasmid siRNA-NRP1. Human gastric cancer cell line SGC7901 was injected subcutaneously into the right hip of nude mice. The model of xenograft tumor was constructed. After tumorigenesis, the recombinant plasmid siRNA-NRP1 was injected into the tumor to observe the clinical manifestations of the nude mice. The volume and weight of the transplanted tumor in nude mice were determined. The pathological changes of tumor tissue and blood vessel were observed. Expression of NRP1 in Nude Mouse Xenografts and Microvessel Density. Results Enzyme digestion analysis and sequencing confirmed that the target oligonucleotide fragment was cloned into plasmid pGenesil-1.1. The size and weight of siRNA-NRP1 group were smaller than that of the control group (P <0.01) NRP1 group tumor necrosis than the control group was obvious; immunohistochemistry showed that siRNA-NRP1 group nude mice transplanted tumor NRP1 expression and microvessel density were lower than the control group (P <0.01). Conclusion Inhibition of NRP1 expression by siRNA-NRP1 in transplanted gastric cancer in nude mice can affect the formation of blood vessels in xenografts of nude mice and eventually inhibit the growth of the tumor.