摘要
本项研究的目的在于探讨蜂毒肽Melittin突变体多肽Melittin-K1对人肝癌耐药细胞BEL-7402/5-FU生长的影响,研究采用CCK-8法检测细胞活率,对比Melittin以及Melittin-K1抑制BEL-7402/5-FU细胞生长作用的差异。通过FITC荧光标记Melittin-K1,结合流式细胞术检测Melittin-K1与BEL-7402/5-FU细胞的结合作用。采用扫描电镜以及培养基上清中乳酸脱氢酶(lactate dehydrogenase,LDH)的含量检测评价Melittin-K1对BEL-7402/5-FU细胞膜的作用。通过构建BEL-7402/5-FU细胞裸鼠异种移植瘤模型并给予Melittin-K1注射来评价Melittin-K1的体内抑制耐药性肿瘤生长的作用。体外细胞研究结果表明,与Melittin相比,Melittin-K1抑制BEL-7402/5-FU细胞的生长的作用明显提高。Melittin-K1抑制BEL-7402/5-FU细胞的生长,呈现时间及剂量依赖性,在24,48,723个时间点的半数抑制浓度为1.00,0.81,0.62μmol/L。FITC标记的Melittin-K1能够直接与BEL-7402/5-FU细胞结合。扫描电镜以及LDH检测结果显示Melittin-K1具有破坏细胞膜完整性的作用。Melittin-K1体内抑制BEL-7402/5-FU细胞肿瘤生长作用明显并呈剂量依赖性。Melittin-K1在0.25,0.5,1 mg/kg 3个剂量组的抑瘤率为分23.95%、76.89%、79.16%。因此,Melittin-K1通过破坏细胞膜抑制BEL-7402/5-FU细胞生长,是一种极具应用前景的治疗耐药性肝癌的新药。
The aim of this study was to investigate the effect of Melittin-K1 on the growth of 5-fluorouracil resistant human hepatocellular carcinoma cell line BEL-7402/5-FU cells.CCK-8 method was used to detect the cell viability,and the difference of inhibitory effect of melittin and Melittin-K1 on BEL-7402/5-FU proliferation were compared.Further,the binding between FITC-labeled MelittinK1 and BEL-7402/5-FU cells was measured by detecting the fluorescence intensity via flow cytometry.The effect of Melittin-K1 on BEL-7402/5-FU cell membrane was evaluated by scanning electron microscope(SEM)and measuring the content of lactate dehydrogenase(LDH)in the culture medium supernatant.In order to further evaluate the inhibitory effect of Melittin-K1 in vivo,an xenograft model of BEL-7402/5-FU cells was established and different concentrations of Melittin-K1 were injected.The data obtained indicated that compared with Melittin,the inhibitory effect of Melittin-K1 on BEL-7402/5-FU cells’proliferation was obviously improved.Moreover,Melittin-K1 significantly inhibited the growth of BEL-7402/5-FU cells in a time and dose-dependent manners.The 50%inhibitory concentrations of Melittin-K1 on BEL-7402/5-FU cells were 1.00,0.81 and 0.62μmol/L at 24,48 and 72 h,respectively.The results of different concentrations of FITC labeled Melittin-K1 incubated with BEL-7402/5-FU cells indicated that FITC labeled Melittin-K1 could bind to BEL-7402/5-FU cells directly.The results of SEM scanning suggested that Melittin-K1 destroyed the integrity of cell membrane.The LDH secretion in supernatant after BEL-7402/5-FU cells treated with Melittin-K1 was increased.Moreover,compared with control group,Melittin-K1 remarkably inhibited the growth of BEL-7402/5-FU cells in vivo in a dosedependent manner.The tumor inhibition rates of Melittin-K1 were 23.95%,76.89%and 79.16%in the 0.25 mg/kg,0.5 mg/kg and 1 mg/kg group,respectively.In conclusion,Melittin-K1 inhibits BEL-7402/5-FU cell growth by destroying cell membrane and is a promising novel drug for the treatment of drug-resistant liver cancer.
作者
董健
赵诗迪
柯梦云
吕毅
DONG Jian;ZHAO Shi-di;KE Meng-yun;LV Yi(National Local Joint Engineering Research Center for Precision Surgery&Regenerative Medicine,Shaanxi Provincial Center for Regenerative Medicine and Surgical Engineering,First Affiliated Hospital of Xi'an Jiaotong University,Xi'an 710061,China;Department of Vascular surgery,First Affiliated Hospital of Xi'an Jiaotong University,Xi'an 710061,China)
出处
《药物生物技术》
CAS
2020年第5期457-461,共5页
Pharmaceutical Biotechnology
作者简介
董健(1989-),男,安徽芜湖人,住院医师,主要研究方向:肝癌的临床和基础研究,E-mail:dongjiandoctor@126.com;通信作者:吕毅(1963-),男,陕西宝鸡人,主任医师,主要研究方向:肝癌的临床和基础研究,E-mail:luyi169@126.com。
