蜂毒肽突变体Melittin-K1抑制人肝癌细胞BEL-7402生长的作用
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摘要
目的:为了提高蜂毒肽Melittin的抗肿瘤活性,我们设计出新型多肽并将其命名为Melittin-K1。并对多肽Melittin-K1抑制人肝癌细胞BEL-7402生长的作用进行研究。方法:采用CCK-8法检测细胞活力。扫描电镜拍照结合培养基上清中乳酸脱氢酶(lactic dehydrogenase,LDH)的表达水平来评价细胞膜的损伤水平。构建BEL-7402细胞裸鼠异种移植瘤模型来评价Melittin-K1、Melittin体内对肝癌生长的影响以及毒性。结果:与Melittin相比,Melittin-K1抑制BEL-7402细胞生长的作用更显著;与L0_2细胞相比,BEL-7402对Melittin-K1的杀伤作用更敏感; Melittin-K1体外抑制BEL-7402细胞生长呈时间、剂量依赖性。扫描电镜以及LDH检测结果均显示Melittin-K1处理组细胞膜的破损情况严重。BEL-7402细胞裸鼠皮下移植瘤模型结果表明,Melittin-K1体内显著抑制肿瘤生长,其抗肿瘤作用明显高于同剂量Melittin。MelittinK1和Melittin体内毒性小,主要表现在,与模型组相比,体重曲线、肝功指标均未发生明显改变。结论:Melittin-K1能够明显抑制肝癌细胞BEL-7402的增殖,是一种极具潜力的抗肝癌药物。
Objective: In order to improve the anti-tumor activity of Melittin,we designed a novel peptide named Melittin-K1. In this study,we investigated the effect of Melittin-K1 on the growth of human hepatocellular carcinoma cell line BEL-7402. Methods: CCK-8 assay was used to detect the cell viability. The damage level of cell membranes after Melittin-K1 treatment was evaluated by observing scanning electron microscopy( SEM) image and detecting the level of lactate dehydrogenase( LDH) in the supernatant of cell culture medium. The effect of Melittin-K1 and Melittin on the growth of BEL-7402 cells and toxicity in vivo was evaluated by establishing the model of subcutaneous transplanted tumors in nude mice. Results: Melittin-K1 inhibited proliferation of BEL-7402 cells more significantly than Melittin. BEL-7402 cells were more sensitive to Melittin-K1 than L0_2 cells. Melittin-K1 suppressed the growth of BEL-7402 cells in a time-and dose-dependent manner. The results of scanning electron microscopy and LDH secretion in the supernatant of cell culture medium showed that the damage level of cell membrane was serious in Melittin-K1 treatment group. Subcutaneous transplantation of BEL-7402 cells in nude mice showed that Melittin-K1 significantly inhibited the growth of tumors in vivo,and its anti-tumor effect was significantly higher than that of Melittin at the same dose. Melittin-K1 and Melittin exhibited less toxic in mice. The body weight curve and liver function parameters were similar to that in the model group. Conclusion: Melittin-K1 significantly inhibits the proliferation of BEL-7402 cells and is a potential anti-tumor drug.
Objective: In order to improve the anti-tumor activity of Melittin,we designed a novel peptide named Melittin-K1. In this study,we investigated the effect of Melittin-K1 on the growth of human hepatocellular carcinoma cell line BEL-7402. Methods: CCK-8 assay was used to detect the cell viability. The damage level of cell membranes after Melittin-K1 treatment was evaluated by observing scanning electron microscopy( SEM) image and detecting the level of lactate dehydrogenase( LDH) in the supernatant of cell culture medium. The effect of Melittin-K1 and Melittin on the growth of BEL-7402 cells and toxicity in vivo was evaluated by establishing the model of subcutaneous transplanted tumors in nude mice. Results: Melittin-K1 inhibited proliferation of BEL-7402 cells more significantly than Melittin. BEL-7402 cells were more sensitive to Melittin-K1 than L0_2 cells. Melittin-K1 suppressed the growth of BEL-7402 cells in a time-and dose-dependent manner. The results of scanning electron microscopy and LDH secretion in the supernatant of cell culture medium showed that the damage level of cell membrane was serious in Melittin-K1 treatment group. Subcutaneous transplantation of BEL-7402 cells in nude mice showed that Melittin-K1 significantly inhibited the growth of tumors in vivo,and its anti-tumor effect was significantly higher than that of Melittin at the same dose. Melittin-K1 and Melittin exhibited less toxic in mice. The body weight curve and liver function parameters were similar to that in the model group. Conclusion: Melittin-K1 significantly inhibits the proliferation of BEL-7402 cells and is a potential anti-tumor drug.
引文
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[2]SHAO Y,GUAN JZ.Progress in treatment of primary liver cancer[J].Infectious Disease Information,2016,29(4):248-252.[邵艳,管静芝.原发性肝癌治疗进展[J].传染病信息,2016,29(4):248-252.]
[3]CHEN Q,LIU X,JIANG XF,et al.Research progress of pharmacological effects of melittin[J].Journal of Jilin Medical College,2018,39(1):65-68.[陈琪,刘欣,姜小凡,等.蜂毒肽药理作用的研究进展[J].吉林医药学院学报,2018,39(1):65-68.]
[4]WANG ZC,CHEN Q,LIU X,et al.Research progress of the antitumor mechanisms of Melittin[J].Journal of Jilin Medical College,2018,39(6):462-464.[王智传,陈琪,刘欣,等.蜂毒肽抗肿瘤作用机制研究进展[J].吉林医药学院学报,2018,39(6):462-464.]
[5]Wu X,Zhao B,Cheng Y,et al.Melittin induces PTCH1 expression by down-regulating Me CP2 in human hepatocellular carcinoma SMMC-7721 cells[J].Toxicol Appl Pharmacol,2015,288(1):74-83.
[6]Zhang SF,Chen Z.Melittin exerts an antitumor effect on nonsmall cell lung cancer cells[J].Mol Med Rep,2017,16(3):3581-3586.
[7]Kong GM,Tao WH,Diao YL,et al.Melittin induces human gastric cancer cell apoptosis via activation of mitochondrial pathway[J].World J Gastroenterol,2016,22(11):3186-3195.
[8]Park MH,Choi MS,Kwak DH,et al.Anti-cancer effect of bee venom in prostate cancer cells through activation of caspase pathway via inactivation of NF-kappaB[J].Prostate,2011,71(8):801-812.
[9]Matar P,Alaniz L,Rozados V,et al.Immunotherapy for liver tumors:Present status and future prospects[J].J Biomed Sci,2009(16):30.
[10]Riedl S,Zweytick D,Lohner K,et al.Membrane-active host defense peptides-challenges and perspectives for the development of novel anticancer drugs[J].2011,164(8):766-781.
[11]GAO J,XIE C,Lu WY.Progress in research of antitumor activity of antimicrobial peptides[J].World Clinical Drugs,2012,33(10):620-626.[高洁,谢操,陆伟跃.抗菌肽的抗肿瘤作用研究新进展[J].世界临床药物,2012,33(10):620-626.]
[12]Li C,Liu H,Yang Y,et al.N-myristoylation of antimicrobial peptide CM4 enhances its anticancer activity by interacting with cell membrane and targeting mitochondria in breast cancer cells[J].Front Pharmacol,2018(9):1297.
[13]Deslouches B,Di YP.Antimicrobial peptides with selective antitumor mechanisms:Prospect for anticancer applications[J].Oncotarget,2017,8(28):46635-46651.
[14]Rady I,Siddiqui IA,Rady M,et al.Melittin,a major peptide component of bee venom,and its conjugates in cancer therapy[J].Cancer Lett,2017(402):16-31.
[15]HUANG YB,ZHAI NC,GAO G,et al.Effects of net charge on biological activities of alpha-helical anticancer peptides[J].Chemical Journal of Chinese Universities,2012,33(6):1252-1258.[黄宜兵,翟乃翠,高贵,等.净电荷对螺旋型抗癌肽生物活性的影响[J].高等学校化学学报,2012,33(6):1252-1258.]
[16]Ke MY,Dong J,Wang Y,et al.MEL-pep,an analog of melittin,disrupts cell membranes and reverses 5-Fluorouracil resistance in human hepatocellular carcinoma cells[J].Int J Biochem Cell Biol,2018(101):39-48.