芹菜素抑制肿瘤细胞增殖机制研究进展

陈宁 叶丽红

·综述·

芹菜素抑制肿瘤细胞增殖机制研究进展

陈宁 叶丽红

本文通过参考近年来对芹菜素抑制肿瘤细胞增殖作用的研究的相关文献，于芹菜素对肿瘤细胞的增殖抑制作用机制做了归纳和探讨。研究发现，芹菜素对多种癌细胞都有抑制其生长和促使其凋亡的作用，其对各种癌细胞作用的机理研究已经深入到细胞和分子水平，包括对细胞凋亡和周期相关基因的调控，对细胞周期G0/G1,S及G2/M期的阻滞，对与凋亡相关的线粒体途径和非线粒体途径的各种酶和蛋白表达的影响，抑制肿瘤细胞血管生成以及增强肿瘤细胞对化疗药物的敏感性等。芹菜素抑制肿瘤细胞增殖机理值得我们深入探索，其作为一种自然界广泛存在的抗癌症化合物具有广阔的应用前景。

芹菜素； 癌细胞； 抑制增殖； 机制

芹菜素(apigenin，API)又称芹黄素，是一种天然的黄酮类化合物，广泛存在于自然界中,以植物黄色素的形式在茶叶、蔬菜、水果和香料中广泛分布，中药如地钱、西藏雪莲花、车前子、络石藤等中也有很高的含量，近年来通过对芹菜素的研究发现，芹菜素在抗肿瘤方面作用显著，对乳腺癌，前列腺癌，膀胱癌，卵巢癌，胃癌，肺癌，胰腺癌等均有一定的抑制作用，现就近年来芹菜素在抑制肿瘤细胞增殖机制方面的研究新进展作一综述。

1 调控凋亡基因

1.1 周期相关基因

杜俊瑶等[1]用芹菜素处理卵巢癌细胞CAOV3，发现存活素(survivin)mRNA的表达下调，p21 mRNA的表达上调，细胞周期在G2 /M 期停滞。Zhang等[2]发现芹菜素作用KYSE-510细胞后，p21waf1基因上调表达，与G2/M期细胞周期停滞有关的细胞周期素(cyclin)B1基因下调表达，p21waf1可能通过抑制细胞周期蛋白依赖性激酶[3](cyclin-dependent kinase ，CDK)1促进KYSE-510 细胞发生G2/M 期细胞周期停滞。而芹菜素作用OE33细胞后，GADD45β、14-3-3σ基因上调表达, cyclin B1基因下调表达，GADD45β能破坏cyclin B/CDK1 结合状态[4]，14-3-3σ则能通过Weel磷酸化CDK1，或使Cdc25c去磷酸化，促进cyclin B/CDK1向细胞核外的转运[5]，并控制细胞进入有丝分裂期[6]，从而引起OE33细胞发生G2/M期细胞周期停滞。APC是一种与大肠直肠癌发生有关的肿瘤抑制基因，它的功能失调对于芹菜素诱导细胞周期停滞有关键作用。Chung C S等[7]发现在人直肠癌细胞中，芹菜素使APC的表达上调，促使了癌细胞死亡。

1.2 凋亡相关基因

在对膀胱癌BIU-87细胞的研究中，姚善华等[8]研究发现芹菜素能通过下调survivin mRNA 和survivin 蛋白表达，上调半胱氨酸的天冬氨酸蛋白水解酶(cysteinyl aspartate specific proteinase，Caspase)-3蛋白表达，明显抑制细胞的生长，并促进细胞发生凋亡。Slusarz A等[9]发现芹菜素还可通过抑制Gli1 mRNA聚合，阻滞Hedgehog信号转导通路，抑制人前列腺癌细胞增殖。

1.3 能量摄取、血管形成相关基因

Melstrom等[10]等发现芹菜素可通过下调葡萄糖转运蛋白(GLUT-1)mRNA表达和蛋白质的表达显著抑制两种胰腺癌细胞系CD18和S2-013对葡萄糖的摄取，从而抑制胰腺癌细胞的生长；还可通过低氧诱导因子1α下调GLUT-1表达，以及阻碍血管内皮生长因子的表达，抑制在低氧状态下的胰腺癌细胞CD-18和S2-013的增殖。

2 调控细胞周期

2.1 G0/G1期阻滞

Gupta等[11]发现芹菜素可下调人前列腺癌细胞cyclin D1、cyclin D2和cyclin E以及通过p53途径上调CDK抑制剂WAF1/p21 and KIP1/p27表达, 从而抑制了CDK2、CDK4、CDK6活性，将细胞阻滞在G0/ G1期。Shukla S等[12]还发现经芹菜素处理的前列腺癌细胞和PC-3细胞中与p38和PI3K-Akt的磷酸化减少相关的cyclin D1的表达减少，cyclin D2，E以及结合的CDK 2, 4和6的明显抑制，使细胞阻滞于G0-G1期。

2.2 G1/S期阻滞

Zheng等[13]发现芹菜素通过P53途径的p21/WAF1表达增加使人宫颈癌细胞G1期阻滞。Bektic等[14]发现芹菜素可使cyclin D1表达减少而p21 /WAF1表达增加来使细胞发生G1/S期阻滞。Shukla S等[15]认为芹菜素通过使WAF1/p21、KIP1/p27、INK4a/p16 和INK4c/p18的表达增加，cyclinsD1、D2、E和CDK2、CDK4和CDK6表达下降，降低Rb在serine780的磷酸化，稳定p53 serine15的磷酸化，使芹菜素在前列腺癌移植裸鼠的肿瘤组织中阻滞细胞周期，从而抑制肿瘤的生长。

2.3 G2/M期阻滞

Kobayashi等[16]发现芹菜素可以通过P53依赖的途径诱导P21产生,进一步抑制CDK1使细胞周期阻滞于G2/M期从而发挥对前列腺癌细胞的生长抑制作用。Casagrande F等[17]研究发现芹菜素在黑色素瘤细胞中可以通过抑制CDK1(cdc2)的Tyr15残基的磷酸化而抑制CDK1(cdc2)活性,使肿瘤细胞被阻滞在G2/M期。Wang W等[18]认为芹菜素通过抑制cyclin B1结合的CDK1编码蛋白激酶的活性，减少cyclin B1及p34(cdc2)激酶的积聚将结肠癌细胞SW480，HT-29和Caco-2阻滞在G2/M期。Yin F等[19]则认为芹菜素抑制cyclin B1 和 CDK1 蛋白的表达，导致CDK1激酶活性的抑制，伴有CDK4，cyclin D1和cyclin A蛋白水平的下调，使MCF-7和MDA-MB-468细胞被阻滞在G2/M期。Ujiki MB等[20]发现芹菜素通过下调cyclin A、cyclin B表达，以及减少cdc2、cdc25磷酸化诱导胰腺癌细胞G2/M期阻滞。Choi EJ等[21]用芹菜素处理乳腺癌细胞SK-BR-3，细胞周期在G2/M期停滞，CDK1和cyclin A、B含量下降，p53的积累增加，p21水平提高。

3 调节参与凋亡的酶和蛋白的表达

3.1 线粒体途径

Choi EJ等[21]认为芹菜素通过抑制CDK1和积累P53，促进下游Bax和细胞色素C表达，进而增加p21(Cip1)水平，诱导乳腺癌细胞SK-BR-3凋亡。Gupta等[11]发现将人激素易感性前列腺癌细胞加入芹菜素共同培养后，观察到DNA呈片断化，Bax/Bcl-2比值增加，细胞发生凋亡。I-K1Wang等[22]认为芹菜素通过提高细胞内活性氧水平，改变线粒体跨膜电位促使细胞色素C释放到胞浆并激活Capase-9的作用，进而导致细胞内Caspase-3蛋白酶活化，引起细胞凋亡。Lu等[23]认为芹菜素通过上调细胞色素C，AIF和Endo G水平，调节Bax/Bcl-2比率，引起Caspase-9和Caspase-3活化，从而引发人肺癌A549细胞凋亡，而对于人肺癌H460细胞，芹菜素通过下调Bid，Bcl-2，procaspase-8；上调Bax，caspase-3，AIF，细胞色素C，GRP78和GADD153的表达；降低线粒体膜电位以及增加ROS和Ca2+来抑制增殖的[24]。Hussain等[25]研究芹菜素对淋巴瘤细胞系的影响，发现淋巴瘤细胞用芹菜素处理引起p-Bad蛋白去磷酸化，抗凋亡蛋白Bcl-2蛋白和细胞凋亡蛋白抑制剂(IAPs)下调，Bax/Bcl2比率增加，同时Bax蛋白构象改变，从细胞质移位到线粒体，引起线粒体膜电位下降，随后细胞色素C释放，激活Caspase-9和Caspase-3。

3.2 非线粒体途径

Tzong-Der Way等[26-27]研究发现芹菜素能够诱导HER2/neu过度表达的乳腺癌细胞发生凋亡，是通过减少HER2/neu蛋白的表达，从而抑制HER2/HER3-PI3K/Akt通路的信号转导，诱导细胞色素C及Caspase-3的激活而产生的。Lee等[28]认为芹菜素通过抑制肝细胞生长因子(HGF)诱导的AKT磷酸化和PI3K途径的beta4整合蛋白来抑制MDA-MB-231细胞增殖。Bektic等[14]发现芹菜素可使ERK1，ERK2的磷酸化减少，介导MAPK通路抑制前列腺间质细胞的增殖。Shukla S等[15]认为芹菜素抑制前列腺癌和PC-3细胞总RB蛋白和位于Ser780和Ser807/811位点RB蛋白的磷酸化，促进ERK1/2和JNK1/2的磷酸化，从而抑制了ELK-1的磷酸化及原癌基因c-fos的表达，抑制肿瘤细胞的存活。在其它研究中Shukla S等[29]发现芹菜素通过对胰岛素样生长因子(IGF)-I诱导的IGF-IR的抑制和对p-Akt的抑制导致GSK-3beta磷酸化和cyclin D1表达下降，p27/kip1表达增强，通过IGF/IGF-IR和PI3K/Akt途径诱导前列腺肿瘤细胞DU145凋亡。Liang Y C等[30]发现芹菜素可以通过抑制NF-κB抑制因子激酶活性，减少NF-κB抑制因子降解，下调NF-κB活性促使肿瘤细胞凋亡。Seo HS等[31]发现芹菜素能上调p53，phospho-p53和p21水平，抑制磷酸化c-Jun氨基端激酶-1(JNK-1)和磷酸化STAT3的表达和活性，降低胞质内的κB抑制因子α的磷酸化水平，抑制p65入核，阻断核因子-κB(NF-κB)通路的活化，诱导HER2过表达的乳腺癌细胞MCF-7 vec 和MCF-7 HER2的凋亡。

4 抑制肿瘤血管生成及增强对化疗药物的敏感性

LIU等[32]发现芹菜素通过抑制人肺癌A549细胞AKT和p70S6K1活性，致HIF-1和血管内皮生长因子表达下降，从而抑制肿瘤血管生成和细胞增殖。Choi等[33]发现芹菜素能下调Akt和ErbB2的表达来增加过度表达ErbB2的乳腺癌MDA-MB-453细胞对5-氟脲嘧啶的敏感性，促使细胞凋亡。Kachadourian等[34]报道，在A549细胞和人类前列腺癌PC-3细胞中，芹菜素可以显著降低谷胱甘肽水平，而谷胱甘肽水平的降低可能与细胞对抗肿瘤药物的敏感度增强相关。

5 结语与展望

芹菜素通过调控凋亡相关基因，调控细胞周期，调节参与细胞凋亡的蛋白和酶的表达，化疗药物的增敏等多种途径抑制肿瘤细胞增殖，发挥抗肿瘤作用，并且作为一种天然、低毒、高效的广泛存在于自然界的天然黄酮类化合物日益受到人们的重视，但是芹菜素具体抑制各种肿瘤细胞增殖的具体机理仍未探究清楚，值得深入研究，包括凋亡基因的表达与蛋白表达的一致性，芹菜素对其表达的量效关系，导致凋亡或周期阻滞的完整信号传导通路以及通路之间的选择性及相关性等。总之，芹菜素高效低毒，其抑制肿瘤细胞增殖的机制值得继续深入探究，并有望据此研发新型抗肿瘤药物。

[1]杜俊瑶,辛彦.芹菜素对人卵巢癌细胞系CAOV3细胞增殖周期的影响[J].中国实用妇科与产科杂志,2008,24(6):456-458.

[2]Zhang Qiang,Zhao Xin-Huai,Wang Zhu-Jun. Flavones and flavonols exert cytotoxic effects on a human oesophageal adenocarcinoma cell line (OE33) by causing G2/M arrest and inducing apoptosis[J]. Food and chemical toxicology, 2008 ,46(6):2042-2053.

[3]Vidal A,Koff A. Cell-cycle inhibitors: three families united by a common cause[J]. Gene, 2000, 247(1-2):1-15.

[4]Schwartz GK,Shah MA. Targeting the cell cycle: a new approach to cancer therapy[J]. Journal of clinical oncology, 2005, 23(36): 9408-9421.

[5]Haupt S,Berger M,Goldberg Z,et al. Apoptosis-the p53 network[J] . Journal of cell science, 2003, 116(20): 4077-4085.

[6]Chan T A, Hermeking H, Lengauer C,et al. 14-3-3 Sigma is required to prevent mitotic catastrophe after DNA damage[J]. Nature,1999, 401(6753): 616-620.

[7]Chung C S,Jiang Y,Cheng D. Impact of Adenomatous Polyposis Coli(APC) tumor supressor gene in human colon cancer cell lines on cell cycle arrest by apigenin[J]. Molecular carcinogenesis,2007,46(9): 773-782.

[8]姚善华,胡自力,邓新军. 芹菜素诱导膀胱癌细胞株BIU-87凋亡及机制的研究[J]. 重庆医科大学学报,2010,35(8):1194-1197.

[9]Slusarz Anna,Shenouda Nader S,Sakla Mary S,et al . Common botanical compounds inhibit the hedgehog signaling pathway in prostate cancer[J]. Cancer research,2010,70(8):3382-3390.

[10]Melstrom L G,Salabat M R,Ding Xian-Zhong,et al. Apigenin Inhibits the GLUT-1 Glucose Transporter and the Phosphoinositide 3-Kinase/Akt Pathway in Human Pancreatic Cancer Cells [J]. Pancreas,2008,37(4) : 426-431.

[11]Gupta S,Afaq F,Mukhtar H. Involvement of nuclear factor-kappa B, Bax and Bcl-2 in induction of cell cycle arrest and apoptosis by apigenin in human prostate carcinoma cells[J]. Oncogene. 2002, 21(23):3727-3238.

[12]Shukla S,Gupta S. Molecular targets for apigenin-induced cell cycle arrest and apoptosis in prostate cancer cell xenograft[J]. Molecular cancer therapeutics,2006,5(4):843-852.

[13]Zheng PW,Chiang LC,Lin CC. Apigenin induced apoptosis through p53-dependent pathway in human cervical carcinoma cells[J]. Life sciences,2005,76(12):1367-1379.

[14]Bektic Jasmin,Guggenberger Roman,Spengler Barbara,et al. The flavonoid apigenin inhibits the proliferation of prostatic strornal cells via the MAPK-pathway and cell-cycle arrest in G1/S [J] . Maturitas, 2006, 55(1): S37-S46.

[15]Shukla Sanjeev,Gupta Sanjay. Apigenin-induced cell cycle arrest is mediated by modulation of MAPK, PI3K-Akt, and loss of cyclin D1 associated retinoblastoma dephosphorylation in human prostate cancer cells[J]. Cell cycle, 2007, 6(9):1102-1114.

[16]Kobayashi T,Nakata T,Kuzumaki T. Effect of flavonoids on cell cycle progression in prostate cancer cells[J]. Cancer letters, 2002,176( 1):17-23.

[17]Casagrande F,Darbon JM. Effects of structurally related flavonoids on cell cycle progression of human melanoma cells: regulation of cyclin-dependent kinases CDK2 and CDK1 [J]. Biochemical pharmacology, 2001, 61(10): 1205-1215.

[18]Wang W,Heideman L,Chung C S.Cell cycle arrest at G2/M and growth inhibition by apigenin in human colon carcinoma cell lines[J]. Molecular carcinogenesis, 2000, 28(2):102-110.

[19]Yin F,Giuliano AE, Law RE,et al. Apigenin inhibits growth and induces G2/M arrest by modulating cyclin-CDK regulators and ERK MAP kinase activation in breast carcinoma cells [J]. Anticancer research, 2001, 21(1A): 413-420.

[20]Ujiki Michael B,Ding Xian-Zhong,Salabat M, et al. Apigenin inhibits pancreatic cancer cell proliferation through G2/M cell cycle arrest [J]. Molecular cancer,2006,5:76.

[21]Choi Eun Jeong,Gun-Hee Kim. Apigenin causes G2 /M arrest associated with the modulation of p21 and Cdc2 and activates p53-dependent apoptosis pathway in human breast cancer SK-BR-3 cells[J].Journal of nutritional biochemistry,2009,20(4): 285-290.

[22]Wang I K,Lin-Shiau S Y,Lin J K. Induction of apoptosis by apigenin and related flavonoids through cytochrome c release and activation of caspase-9 and caspase-3 in leukaemia HL-60 cells[J]. European journal of cancer ,1990,35(10):1517-1525.

[23]Lu Hsu-Peng,Chie Yu-Jie,Yang Ming-Sung,et al. Apigenin induces caspase-dependent apoptosis in human lung cancer A549 cells through Bax-and Bcl-2-triggered mitochondrial pathway[J].International journal of oncology ,2010,36( 6): 1477-1484．

[24]Lu Hsu-Feng,Chie Yu-Jie,Yang Ming-Sung,et al.Apigenin induces apoptosis in human lung cancer H460 cells through caspase-and mitochondria-dependent pathways[J].Human & experimental toxicology,2011,30(8):1053-1061．

[25]Hussain A R,Khan A S,Ahmed S O,et al. Apigenin induces apoptosis via downregulation of S-phase kinase-associated protein 2-mediated induction of p27Kip1 in primary effusion lymphoma cells [J].Cell proliferation,2010,43(2): 170-183.

[26]Way TD,Kao MC,Lin JK. Apigenin induces apoptosis through proteasomal degradation of HER2/neu in HER2/neu-overexpressing breast cancer cells via the phosphatidylinositol 3-kinase/Akt-dependent pathway[J]. Journal of biological chemistry,2004,279(6):4479-4489.

[27]Way TD,Kao MC,Lin JK. Degradation of HER2/neu by apigenin induces apoptosis through cytochrome c release and caspase-3 activation in HER2/neu-overexpressing breast cancer cells[J].Febs letters,2005,579(1):145-152.

[28]Lee WJ, Chen WK, Wang CJ,et al. Apigenin inhibits HGF-promoted invasive growth and metastasis involving blocking PI3K/Akt pathway and beta 4 integrin function in MDA-MB-231 breast cancer cells[J]. Toxicology and applied pharmacology,2008,226(2):178-191.

[29]Shukla S, Gupta S. Apigenin suppresses insulin-like growth factor I receptor signaling in human prostate cancer :an in vitro and in vivo study [J]. Molecular carcinogenesis,2009,48(3): 243-252.

[30]Liang Y C,Huang Y T,Tsai S H,et al. Suppression of inducible cyclooxygenase and inducible nitric oxide synthase by apigenin and related flavonoids in mouse macrophages[J]. Carcinogenesis, 1999,20(10): 1945-52.

[31]Seo Hye-Sook,Choi Han-Seok,Kim Soon-Re,et al.Apigenin induces apoptosis via extrinsic pathway,inducing p53 and inhibiting STAT3 and NF kappa B signaling in HER2-overexpressing breast cancer cells[J]. Molecular and cellular biochemistry,2012,366(1-2):319-334.

[32]LIU L Z,FANG J,ZHOU Q,et al.Apigenin inhibits expression of vascular endothelial growth factor and angiogenesis in human lung cancer cells: implication of chemoprevention of lung cancer [J].Molecular pharmacology,2005,68(3): 635-643．

[33]Choi E J,Kim G H. 5-Fluorouracil combined with apigenin enhances anticancer activity through induction of apoptosis in human breast cancer MDA-MB-453 cells[J]. Oncology reports,2009,22(6):1533-1537．

[34]Kachadourian Remy,Day Brian J. Flavonoid-induced glutathione depletion: potential implications for cancer treatment[J]. Free radical biology and medicine,2006,41(1): 65-76．

(本文编辑：蒲晓田)

Mechanismofapigenin’sinhibitionofcancercellproliferation:ReviewofProgress

CHENNing，YELi-hong.

NanjinguniversityoftraditionalChinesemedicine，Nanjing210046,China

YELi-hong，E-mail:cocolihongye@126.com

In this paper, the mechanism of apigenin’s inhibition of cancer cell proliferation was summarized and discussed with the reference of relevant literatures of inhibition in recent years. The research showed that apigenin had the effects of inhibition of proliferation and cell apotosis induction on various kinds of cancer cells and the mechanism was further concentrated on the cellular and molecular levels such as regulating cell gene related to cell cycle and apotosis，blocking cell cycle of G0/G1,S and G2/M stage，influencing the expression of protein and kidney related to mitochondrial or non-mitochondrial control of apotosis, inhibiting tumor angiogenesis and increasing the sensitivity of tumor cells to chemotherapy drugs. The mechanism of apigenin’s inhibition of cancer cell proliferation is worth our further exploration. Apigenin has broad application prospects as an anticancer compound widely distributed in nature.

Apigenin; Cancer cells; Inhibition of proliferation; Mechanism

科技部“十一五”国家科技支撑计划( 2006BAI11B08-01)；江苏省高校优势学科建设工程资助项目(PAPD)

210046 ，南京中医药大学第一临床医学院[陈宁(硕士研究生)、叶丽红]

陈宁(1989-)，女，2007级七年制在读硕士研究生。研究方向：中医学(中西医结合肿瘤内科)。E-mail：835569624@qq.com

叶丽红(1967-)，女，博士，教授，主任医师，硕士研究生导师。研究方向：内科杂病及肿瘤的中医药防治。E-mail: cocolihongye@126.com

R285.5

A

10.3969/j.issn.1674-1749.2014.02.023

2013-12-06)