非抗凝N-去硫酸肝素对胃癌转移抑制作用的实验研究

Inhibition of N-desulfated heparin on metastasis of orthotopic implantation of human gastric carcinoma

目的研究N-去硫酸肝素对人胃癌组织原位移植非肥胖性糖尿病(non-obesity diabetes,NOD)重度联合免疫缺陷(SCID)小鼠转移模型的肿瘤转移抑制、血管生成和血管内皮生长因子(VEGF)表达的影响。方法建立人胃癌组织原位移植NOD SCID小鼠胃痛转移模型,20只小鼠均分成2组。移植后1周,分别静脉注射0．9%氯化钠溶液(0．9%氯化钠溶液组)与10 mg/kg N-去硫酸肝素(N-去硫酸肝素组),每周2次,共3周。第6周处死动物,观察肿瘤转移情况,免疫组化法检测肿瘤组织微血管密度、VEGF表达,荧光定量PCR检测肿瘤组织VEGF mRNA表达。结果0．9%氯化钠溶液组10只小鼠9只有肿瘤转移,N-去硫酸肝素组10只小鼠只有2只转移,两组间差异有统计学意义(P＜0．05)。未发现出血等不良反应。0．9%氯化钠溶液组平均微血管密度为9．1±3．4,N-去硫酸肝素组为4．7±1．8,两组间差异有统计学意义(t=3．617,P＜0．05)。0．9%氯化钠溶液组有9只VEGF阳性表达,明显高于N-去硫酸肝素组的2只(P＜0．05)。荧光定量PCR测定显示,N-去硫酸肝素组VEGF mRNA(Ct：19．56±1．53)表达较0．9%氯化钠溶液组(Ct=16．38±1．68)低,两组间差异有统计学意义(t=4．425,P＜0．05)。结论N-去硫酸肝素通过抑制肿瘤组织VEGF表达和血管生成,抑制肿瘤转移。N-去硫酸肝素无明显出血等不良反应。

Objective To investigate the effect of N-desulfated heparin on tumor metastasis, tumor angiogenesis, and vascular endothelial growth factor（VEGF） of orthotopic implantated human gastric carcinoma in non-obesity diabetes（NOD） severe combined immune deficiency（SCID） mice. Methods Human gastric cancer SGC-7901 tissues were orthotopically implanted into the stomach of the NOD SCID mice. Twenty mice were randomly divided into two groups which received either intravenous injections of 0. 9% NaCl solution（0.9% NaCl solution group） or 10 mg/kg N-desulfated heparin（N-desulfated heparin group） twice weekly for three weeks. Mice were sacrificed six weeks after implantation. Tissues from stomach and other organs were obtained for histopathological evaluation. The intratumoral microvessel density（MVD） and VEGF protein expression in tumor were evaluated immunochemically. Real time PCR technique was used to detect VEGF mRNA expression. Results The tumor metastasis rates were 9/10 in 0.9% NaCl solution group and 2/10 in N-desulfated heparin group（P〈0.05）. MVD was 9.1±3.4 in 0.9% NaCl solution group and 4. 7±1. 8 in N-desulfated heparin group （t = 3. 617, P 〈 0.05）. VEGF protein expression was 90% in 0.9% NaCl solution group and 20% in N-desulfated heparin group（ P 〈 0.05）. VEGF mRNA expression was lower in N-desulfated heparin group （Ct： 19.56±1.53） than that in 0.9% NaCl solution group（Ct： 16.38±1.68, t = 4. 425, P〈0.05）. Conclusions N-desulfated heparin can inhibit the metastasis of gastric cancer through inhibiting tumor VEGF expression, and tumor angiogenesis with no obvious anticoagulant activity.