两种牛蛙活性肽结构及体外抗肿瘤细胞增殖作用的研究
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摘要
试验旨在研究新发现的两种牛蛙活性肽对人肺癌细胞NCI-H446、人乳腺癌细胞MCF-7及小鼠白血病细胞K562 3种肿瘤细胞体外增殖的影响,为新的多肽抗肿瘤药筛选提供依据。本试验利用圆二色谱(CD)法检测两种新牛蛙活性肽的二级结构,通过MTS细胞毒试验测定不同浓度的两种新牛蛙活性肽对人肺癌细胞NCI-H446、人乳腺癌细胞MCF-7及小鼠白血病细胞K562 3种瘤细胞体外增殖的影响。圆二色谱法结果显示,Temporin-Lb的二级空间结构为聚脯氨酸Ⅱ型螺旋(PPⅡ)结构,Catesbeianin-1a的二级空间结构为无规则卷曲。MTS细胞毒试验结果显示,给予Catesbeianin-1a的3种瘤细胞培养24h后,瘤细胞形态无明显变化,正常增殖,而给予Temporin-Lb的3种瘤细胞培养24h后,瘤细胞发生皱缩、细胞变圆、脱落甚至死亡,增殖受到抑制,其中对TemporinLb在4~40μmol/L浓度范围内对小鼠白血病细胞K562的抑制效果最为明显。结果表明,新牛蛙活性肽Temporin-Lb具有一定的抑瘤作用,Catesbeianin-1a对瘤细胞的体外增殖没有明显影响。
The study was aimed to research the growth inhibitory effects of two new bioactive peptides Temporin-Lb and Catesbeianin-1afrom Rana catesbeiana on human lung cancer NCI-H446 cells,breast cancer MCF-7cells and mice leukemia K562 cells,and provide the basis for selecting the new peptide antitumor drugs.The second structures of two bioactive peptides were tested by circular dichroism spectrum(CD),and the effects of the two new bioactive peptides on human lung cancer NCI-H446 cells,breast cancer MCF-7cells and mice leukemia K562 cells were examined with MTS cytotoxicity assay.The circular dichroism spectrum results showed the secondary structure of Temporin-Lb was PPⅡ,and the secondary structure of Catesbeianin-1awas random coil.Using MTS cytotoxicity assay,it was found that given Catesbeianin-1a,the sharp of the three cancer cells above had little changed after culturing for 24 h,and the three cancer cells promoted normal.Given Temporin-Lb,the cell morphology of three cancer cells had changed after culturing for 24 h,the growth of the three cancer cells above had been inhibited,especially the bioactive peptide Temporin-Lb had a sharp antitumor effect to mice leukemia K562 cells between the concentra-tion of 4and 40μmol/L.Bioactive peptide Catesbeianin-1ahad no obvious effect on proliferation of the three cancer cells above.Bioactive peptide Temporin-Lb had a certain inhibitory effect to tumor cells.
The study was aimed to research the growth inhibitory effects of two new bioactive peptides Temporin-Lb and Catesbeianin-1afrom Rana catesbeiana on human lung cancer NCI-H446 cells,breast cancer MCF-7cells and mice leukemia K562 cells,and provide the basis for selecting the new peptide antitumor drugs.The second structures of two bioactive peptides were tested by circular dichroism spectrum(CD),and the effects of the two new bioactive peptides on human lung cancer NCI-H446 cells,breast cancer MCF-7cells and mice leukemia K562 cells were examined with MTS cytotoxicity assay.The circular dichroism spectrum results showed the secondary structure of Temporin-Lb was PPⅡ,and the secondary structure of Catesbeianin-1awas random coil.Using MTS cytotoxicity assay,it was found that given Catesbeianin-1a,the sharp of the three cancer cells above had little changed after culturing for 24 h,and the three cancer cells promoted normal.Given Temporin-Lb,the cell morphology of three cancer cells had changed after culturing for 24 h,the growth of the three cancer cells above had been inhibited,especially the bioactive peptide Temporin-Lb had a sharp antitumor effect to mice leukemia K562 cells between the concentra-tion of 4and 40μmol/L.Bioactive peptide Catesbeianin-1ahad no obvious effect on proliferation of the three cancer cells above.Bioactive peptide Temporin-Lb had a certain inhibitory effect to tumor cells.
引文
[1]Jemal A,Murray T,Samuels A,et al.Cancer stautistics 2003[J].CA Cancer J Clin,2003,53(1):5-26.
[2]Giordano A,Rustum Y M,Wenner C E.Cell cycle:Molecular targets for diagnosis and therapy:Tumor suppressor genes and cell cycle progression in cancer[J].J Cell Biochem,1998,70(1):1-7.
[3]杨纯正,刘淑仪.肿瘤耐药基因表达的临床意义及其逆转研究[J].中国肿瘤,2001,10(3):132-136.
[4]韩笑,李娜,杜培革.抗肿瘤多肽研究进展[J].中国生物工程杂志,2013,33(6):93-98.
[5]Hoskin D W,Ramamoorthy A.Studies on anticancer activities of antimicrobial peptides[J].Biochim Biophys Acta,2008,778(2):357-375.
[6]Gomes A,Giri B,Saha A,et al.Bioactive molecules from amphibian skin:Their biological activities with reference to therapeutic potentials for possible drug development[J].Indian J Exp Biol,2007,45(7):579-593.
[7]Zhao R L,Han J Y,Han W Y.Identification and cloning of two novel temporins from Lithobates catesbeianus[J].Progress in Biochemistry and Biophysics,2009,8:1064-1070.
[8]赵瑞利,韩俊友,韩文瑜,等.新牛蛙活性肽Catesbeianin-1a的抗菌抗肿瘤作用[J].中国兽医学报,2013,33(9):1407-1411.
[9]Gutierrez M E,Kummar S,Giaccone G.Next generation oncology drug development:Opportunities and challenges[J].Nat Rev Clin Oncol,2009,6(5):259-265.
[10]Dennison S R,Whittaker M,Harris F,et al.Anticancerα-helical peptides and structure function relationships underpinning their interactions with tumor cell membranes[J].Current Protein and Peptide Science,2006,7(6):487-499.
[11]Yang H,Wang X,Liu X,et al.Antioxidant peptidomics reveals novel skin antioxidant system[J].Mol Cell Proteomics 2009,8(3):571-583.
[12]Damien V,Bruston F,Nicolas P,et al.Antimicrobial peptides from hylid and ranin frogs originated from a150-million-year-old ancestral precursor with a conserved signal peptide but a hypermutable antimicrobial domain[J].Eur J Biochem 2003,270(9):2068-2081.
[13]张庆华,王青,尚田田,等.抗菌肽的活性机制及其分子设计研究进展[J].中国畜牧兽医,2014,41(5):163-167.
[14]Xu X,Li J,Han Y,et al.Two antimicrobial peptides from skin secretions of Rana grahami[J].Toxicon,2006,47(4):459-464.
[15]胡志帅,陈书明,晋大鹏.抗菌肽的结构特征、生物活性及应用[J].生物学杂志,2008,25(4):58-60.
[16]赵瑞利,韩文瑜,韩俊友,等.牛蛙皮肤抗菌肽的分离纯化与活性测定[J].中国生物制品学杂志,2008,21(8):694-697.
[17]Adzhubei A A,Sternberg M J,Makarov A A.Polyproline-Ⅱhelix in proteins:Structure and function[J].J Mol Biol,2013,425(12):2100-2132.
[2]Giordano A,Rustum Y M,Wenner C E.Cell cycle:Molecular targets for diagnosis and therapy:Tumor suppressor genes and cell cycle progression in cancer[J].J Cell Biochem,1998,70(1):1-7.
[3]杨纯正,刘淑仪.肿瘤耐药基因表达的临床意义及其逆转研究[J].中国肿瘤,2001,10(3):132-136.
[4]韩笑,李娜,杜培革.抗肿瘤多肽研究进展[J].中国生物工程杂志,2013,33(6):93-98.
[5]Hoskin D W,Ramamoorthy A.Studies on anticancer activities of antimicrobial peptides[J].Biochim Biophys Acta,2008,778(2):357-375.
[6]Gomes A,Giri B,Saha A,et al.Bioactive molecules from amphibian skin:Their biological activities with reference to therapeutic potentials for possible drug development[J].Indian J Exp Biol,2007,45(7):579-593.
[7]Zhao R L,Han J Y,Han W Y.Identification and cloning of two novel temporins from Lithobates catesbeianus[J].Progress in Biochemistry and Biophysics,2009,8:1064-1070.
[8]赵瑞利,韩俊友,韩文瑜,等.新牛蛙活性肽Catesbeianin-1a的抗菌抗肿瘤作用[J].中国兽医学报,2013,33(9):1407-1411.
[9]Gutierrez M E,Kummar S,Giaccone G.Next generation oncology drug development:Opportunities and challenges[J].Nat Rev Clin Oncol,2009,6(5):259-265.
[10]Dennison S R,Whittaker M,Harris F,et al.Anticancerα-helical peptides and structure function relationships underpinning their interactions with tumor cell membranes[J].Current Protein and Peptide Science,2006,7(6):487-499.
[11]Yang H,Wang X,Liu X,et al.Antioxidant peptidomics reveals novel skin antioxidant system[J].Mol Cell Proteomics 2009,8(3):571-583.
[12]Damien V,Bruston F,Nicolas P,et al.Antimicrobial peptides from hylid and ranin frogs originated from a150-million-year-old ancestral precursor with a conserved signal peptide but a hypermutable antimicrobial domain[J].Eur J Biochem 2003,270(9):2068-2081.
[13]张庆华,王青,尚田田,等.抗菌肽的活性机制及其分子设计研究进展[J].中国畜牧兽医,2014,41(5):163-167.
[14]Xu X,Li J,Han Y,et al.Two antimicrobial peptides from skin secretions of Rana grahami[J].Toxicon,2006,47(4):459-464.
[15]胡志帅,陈书明,晋大鹏.抗菌肽的结构特征、生物活性及应用[J].生物学杂志,2008,25(4):58-60.
[16]赵瑞利,韩文瑜,韩俊友,等.牛蛙皮肤抗菌肽的分离纯化与活性测定[J].中国生物制品学杂志,2008,21(8):694-697.
[17]Adzhubei A A,Sternberg M J,Makarov A A.Polyproline-Ⅱhelix in proteins:Structure and function[J].J Mol Biol,2013,425(12):2100-2132.