SOM230促进HepG2细胞原位移植瘤坏死的机制

目的 观察一种新合成的生长抑素类似物SOM230对肝癌细胞株HepG2体外、体内生长的影响,探索非细胞毒性药物导致肝癌坏死的可能机制. 方法 采用四甲基偶氮唑盐法、末端脱氧核苷酸转移酶介导的脱氧三磷酸尿苷缺口末端标记(TUNEL)及流式细胞技术检测SOM230对体外培养的肝癌细胞HepG2生长及凋亡的作用.用肝癌细胞株HepG2建立裸鼠肝癌原位移植瘤模型,分为SOM230组(100 μg·kg~1·d~(-1)皮下注射)及对照组(等量等渗盐水皮下注射),连续用药8周,测定移植瘤坏死体积.Western blot测定肿瘤组织中生长抑素受体2(SSTR2)的表达水平;免疫组织化学显示肿瘤组织中血管内皮生长因子(VEGF)、SsTR2的表达部位;酶联免疫吸附法检测移植瘤组织中肿瘤坏死因子α(TNF α)含量.两组间均数比较采用t检验,多组间均数比较采用单因素方差分析. 结果 SOM230在体外对HepG2细胞增殖有一定的抑制作用,IC_(50)为2.73×10~6mol/L,对其凋亡无明显影响(TUNEL实验F=0.16,P>0.05;Annexin-V实验F=0.13,P>0.05).SOM230组的移植瘤坏死体积为73.4%±7.0%,明显高于对照组的30.2%±14.0%(t=-8.02,P<0.01).两组移植瘤的血窦中均有SSTR2阳性染色,表达水平差异无统计学意义;s0M230组肝癌移植瘤组织中的VEGF表达为阴性,但移植瘤内TNF α水平在两组的差异无统计学意义SOM230组与对照组,(10.3±8.0)ng/ml与(9.2±5.8)ng/ml,t=-0.24,P>0.05)].结论 SOM230通过SSTR2介导,显著下调肝癌组织VEGF的表达水平,仅阻断生长旺盛肿瘤的血供,导致肝癌移植瘤明显坏死.

Induction of necrosis in the hepatocellular carcinoma HepG2 xenografts treated with SOM230

Objective To investigate the effects of SOM230, a new somatostatin analogue, on the proliferation of hepatocellular carcinoma (HCC) cell line HepG2 in vitro and in vivo, and explore the mecha-nism underline the necrosis of tumors. Methods MTr, TdT-mediated dUTP nick end labeling assay (TUNEL) and flow cytometric assay were used to measure the effects of SOM230 on the proliferation and apoptosis of HCC HepG2 cells. Nude mice bearing HCC xenografts of the HepG2 cell line were treated with SOM230 (100μg·kg~(-1)·d~(-1)) subcutanenusly injection) and saline as a control for eight weeks. The mass and percentage of necrotic volume of the HCC xenografts in nude mice were determined. Western blot was used to detect SSTR2 in HCC xenografts. Immunohistochemical method was used to detect the expression sites of SSTR2 and VEGF in HCC xenografts. ELISA was used to detect the levels of TNF α. Results No proliferation and apoptosis of HepG2 cells were induced by SOM230 in vitro (F=0.16, P>0.05). The percentage of necrotic volume in SOM230 were significantly higher than that of control group (73.4%±7.0% vs 30.2%±14.0%, t=-8.02, P<0.01). SSTR2 was expressed in blood sinus of HCC xenografts in nude mice. There was no significance difference in the level of SSTR2 expression between SOM230 group and saline treated group. VEGF expression in xenografts was down-regulated by SOM230 treatment. SOM230 treatment did not affect the level of TNF α in HCC xenograftst=-0.24, P>0.05]. Conclusions SOM230 can induce massive necrosis of HCC xenografts only after the blockage of blood flow through down-regulation of VEGF mediated by SSTR2.