黑水虻抗菌肽分离纯化及活性组分对HCT-8细胞的增殖抑制作用  

Isolation and purification of antibacterial peptides from H.illucens larvae and the inhibitory effects of active components on HCT-8 cell proliferation

作　　者：冯群 田忠 高嘉敏 李笑驹 夏嫱 FENG Qun;TIAN Zhong;GAO Jia-min;LI Xiao-ju;XIA Qiang(Zhuhai Campus of Zunyi Medical University,Innovation Base of Immunology for Graduate Education in Guizhou Province,Zhuhai 519041,Guangdong,China)

机构地区：[1]遵义医科大学珠海校区/贵州省免疫学研究生教育创新基地,珠海519041 [2]遵义医科大学第三附属医院

出　　处：《医学研究生学报》2022年第1期17-22,共6页Journal of Medical Postgraduates

基　　金：国家自然科学基金(32060127);贵州省教育厅创新群体重大研究项目(黔教合KY字[2016]037)。

摘　　要：目的黑水虻抗菌肽抗癌效应的研究鲜见。文章旨在探讨黑水虻抗菌肽活性组分对敏感肿瘤细胞株(HCT-8)体外增殖的影响,探索其可能的抑癌机制。方法采用RP-HPLC分离纯化黑水虻抗菌肽粗提物,抑菌实验筛选活性组分;酶标仪测定抗菌肽活性组分对豚鼠正常红细胞的溶血性;CCK-8法测定抗菌肽活性组分对敏感肿瘤细胞株(HCT-8)和正常人胚肾细胞(HEK293)的增殖抑制作用;流式细胞仪检测抗菌肽活性组分对HCT-8细胞周期的影响。结果分离纯化并通过抑菌实验筛选获得高活性组分抗菌肽HI-3,其对正常红细胞不溶血。抗菌肽HI-3对HCT-8细胞的增殖抑制作用具有明显的浓度依赖性,抑制率随处理浓度增加而增大,浓度在400μg/mL时抑制率最大,为60.11%±0.75%,显著高于其他处理组细胞的抑制率(P<0.01),且对正常HEK293细胞无明显影响。随着HI-3处理浓度增加,G0/G1期的HCT-8细胞明显增加,G2/M期的HCT-8细胞明显下降,变化呈浓度依赖性,而S期的变化呈现先增加后下降的变化趋势。结论黑水虻抗菌肽活性组分HI-3可显著抑制HCT-8细胞的增殖,且对正常细胞无不良影响;可能是通过阻滞HCT-8细胞周期于G0/G1期或S期,从而影响细胞增殖,具体作用机制有待进一步研究。Objective Research on the anti-cancer effect of antibacterial peptides of black soldier fly is rare.This article is aimed to explore the effect of the active components of antibacterial peptides of black soldier fly on the proliferation of sensitive tumor cell lines(HCT-8)in vitro and explore its possible anti-cancer mechanism.Methods The crude extract of antimicrobial peptide was separated and purified by RP-HPLC.The active component was screened out by antibacterial experiment,and the hemolysis of antibacterial peptide active component on guinea pig erythrocytes was determined by microplate reader.The inhibition of proliferation of antimicrobial peptide active component on sensitive tumor cell lines(HCT-8)and normal human embryonic kidney cells(HEK293)was detected by CCK-8 assays.The effect of antimicrobial peptide active component on the cells cycle of HCT-8 was detected by flow cytometry.Results(1)HI-3,a highly active component of antimicrobial peptide,was purified and screened through antibacterial experiments,which did not hemolyze normal red blood cells.(2)The inhibitory effect of antimicrobial peptide HI-3 on the proliferation of HCT-8 cells was obviously concentration-dependent,and the inhibition rate increased with the increasing of the treatment concentration.When the concentration was 400μg/mL,the inhibition rate was the largest(60.11±0.75%),which was significantly higher than the inhibition rate of other treatment groups(P<0.01),but had no significant effect on normal HEK293 cells.(3)With the increase of HI-3 treatment concentration,HCT-8 cells in G0/G1 phase was significantly increased,HCT-8 cells in G2/M phase was significantly decreased.The changes were concentration-dependent,while the changes in S phase showed an increase first and then a decrease,indicating antimicrobial peptide HI-3 could block the HCT-8 cells cycle in G0/G1 and S phase at low concentrations and block the cells cycle of HCT-8 in G0/G1 phases at high concentrations.Conclusion In this study,HI-3,the active component of the ant

关 键 词：黑水虻 抗菌肽 分离纯化 结肠癌细胞 增殖 机制 

分 类 号：R735.3[医药卫生—肿瘤]