杨梅酮的体外抗氧化及抗肿瘤活性
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摘要
为研究杨梅酮的抗氧化和抗膀胱癌活性,进行了抗氧化试验和环境扫描电子显微镜(ESEM)观察.结果表明,杨梅酮能有效抑制AAPH诱导的红细胞溶血,且效果呈浓度依赖性,80μmol/L杨梅酮的溶血抑制率达到(93.67±0.01)%.研究表明,杨梅酮可通过增强细胞内SOD、GPx的酶活来调控ROS水平,减少MDA产生,避免脂质过氧化.抗肿瘤活性研究发现,杨梅酮对膀胱癌细胞EJ和T24以及正常膀胱细胞SV-HUC-1的抑制效果也有剂量相关性,IC_(50)分别为(39.8±1.0)、(49.0±0.9)、(75.9±1.8)μmol/L.进一步细胞吸收量分析表明,可能是EJ对杨梅酮有更强的细胞吸收导致了杨梅酮对EJ的IC_(50)比T24更低(更敏感的细胞毒性).采用流式细胞术分析杨梅酮对EJ细胞周期DNA分布的影响,发现杨梅酮主要通过诱导EJ细胞产生凋亡及S期阻滞抑制细胞增殖.
Antioxidant assay and ESEM observation were carried out to investigate the antioxidant and anti-bladder cancer activities of myricetin. The results show that myricetin can effectively inhibit the AAPH-induced hemolysis of erythrocytes in a dose-dependent manner. The hemolysis inhibition rate of myricetin increases to(93.67±0.01)%at a concentration of 80 μmol/L. A further experiment confirms that SOD and GPx antioxidant enzyme activities are enhanced to control ROS and MDA generation when erythrocytes are preconditioned by myricetin. Moreover, MTT tests indicate that myricetin could suppress the EJ and T24 bladder cancer cells proliferation in a dose-related manner with the half-maximal inhibitory concentration(IC_(50)) of(39.8±1.0) μmol/L and(49.0±0.9) μmol/L, as well as a lower cytotoxicity on normal bladder cell SV-HUC-1 with the IC_(50) of(75.9±1.8) μmol/L. In addition, myricetin exhibits higher antitumor activity in EJ cell than that in T24 cells due to higher intracellular uptake amount of myricetin in EJ cells. Further flow cytometric studies of EJ cell cycle distribution show that myricetin inhibits the proliferation of EJ cancer cells mainly through induction of apoptosis and S cycle arrest.
Antioxidant assay and ESEM observation were carried out to investigate the antioxidant and anti-bladder cancer activities of myricetin. The results show that myricetin can effectively inhibit the AAPH-induced hemolysis of erythrocytes in a dose-dependent manner. The hemolysis inhibition rate of myricetin increases to(93.67±0.01)%at a concentration of 80 μmol/L. A further experiment confirms that SOD and GPx antioxidant enzyme activities are enhanced to control ROS and MDA generation when erythrocytes are preconditioned by myricetin. Moreover, MTT tests indicate that myricetin could suppress the EJ and T24 bladder cancer cells proliferation in a dose-related manner with the half-maximal inhibitory concentration(IC_(50)) of(39.8±1.0) μmol/L and(49.0±0.9) μmol/L, as well as a lower cytotoxicity on normal bladder cell SV-HUC-1 with the IC_(50) of(75.9±1.8) μmol/L. In addition, myricetin exhibits higher antitumor activity in EJ cell than that in T24 cells due to higher intracellular uptake amount of myricetin in EJ cells. Further flow cytometric studies of EJ cell cycle distribution show that myricetin inhibits the proliferation of EJ cancer cells mainly through induction of apoptosis and S cycle arrest.
引文
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[2] 高蔚娜,蒲玲玲,韦京豫,等.中国四地区成人蔬菜和水果中杨梅黄酮、槲皮素、坎二菲醇、芹菜配基和玉米黄酮摄入量的比较 [J].营养学报,2014,36(6):559- 564,628.GAO Wei-na,PU Ling-ling,WEI Jing-yu,et al.Comparison of dietary intakes of myricetin,quercetin,kaempferol,apigenin and luteolin from vegetables and fruits in Chinese adults from four different regions [J].Acta Nutrimenta Sinica,2014,36(6):559- 564,628.
[3] 刘合生,戚向阳,曹少谦,等.杨梅乙醇提取物对小鼠酒精性肝损伤的保护作用 [J].中国食品学报,2014,14(8):34- 40.LIU He-sheng,QI Xiang-yang,CAO Shao-qian,et al.The protective effect of ethanol extract of Chinese bayberry fruit on alcohol-induced liver injury in mice [J].Journal of Chinese Institute of Food Science & Technology,2014,14(8):34- 40.
[4] KO C H,SHEN S C,LEE T J F,et al.Myricetin inhibits matrix metalloproteinase 2 protein expression and enzyme activity in colorectal carcinoma cells [J].Molecular Cancer Therapeutics,2005,4(2):281- 290.
[5] ONO K,NAKANE H,FUKUSHIMA M,et al.Differential inhibitory effects of various flavonoids on the activities of reverse transcriptase and cellular DNA and RNA polyme-rases [J].European Journal of Biochemistry,1990,53(5):469- 476.
[6] DAJAS F,RIVERA F,BLASINA F,et al.Cell culture protection and in vivo neuroprotective capacity of flavonoids [J].Neurotoxicity Research,2003,5(6):425- 432.
[7] ONG K C,KHOO H E.Biological effects of myricetin [J].General Pharmacology,1997,29(2):121- 126.
[8] GHASEMZADEH A,JAAFAR H Z E,ASHKANI S,et al.Variation in secondary metabolite production as well as antioxidant and antibacterial activities of Zingiber zerumbet (L.)at different stages of growth [J].BMC Complementary and Alternative Medicine,2016,16:104(1-10).
[9] SEN S,DE B,DEVANNA N,et al.Total phenolic,total flavonoid content,and antioxidant capacity of the leaves of Meyna spinosa Roxb.,an Indian medicinal plant [J].Chinese Journal of Natural Medicines,2013,11(2):149- 157.
[10] ZHAO L,CHEN J,SU J,et al.In vitro antioxidant and antiproliferative activities of 5-hydroxymethylfurfural [J].Journal of Agricultural & Food Chemistry,2013,61(44):10604- 10611.
[11] TANG X,DE ROOIJ M,DE JONG L.Volume change measurements of rice by environmental scanning electron microscopy and stereoscopy [J].Scanning,2007,29(5):197- 207.
[12] BERTO A,RIBEIRO A B,DE SOUZA N E,et al.Bioactive compounds and scavenging capacity of pulp,peel and seed extracts of the Amazonian fruit Quararibea cordata against ROS and RNS [J].Food Research International,2015,77:236- 243.
[13] LEE J S,HONG E K.Hericium erinaceus enhances doxorubicin-induced apoptosis in human hepatocellular carcinoma cells [J].Cancer Letters,2010,297(2):144- 154.
[14] HUANG Y Y,LUO Y,ZHENG W J,et al.Rational design of cancer-targeted BSA protein nanoparticles as radiosensitizer to overcome cancer radioresistance [J].ACS Applied Materials & Interfaces,2014,6(21):19217- 19228.
[15] CHEN J P,LI L,SU J Y,et al.Enhancing effect of natural borneol on the cellular uptake of demethoxycurcumin and their combined induction of G2/M arrest in HepG2 cells via ROS generation [J].Journal of Functional Foods,2015,17(10):103- 114.
[16] HORTOLA P.Secondary-electron SEM bioimaging of human erythrocytes in bloodstains on high-carbon steel substrate without specimen preparation [J].Micron,2008,39(1):53- 55.
[17] BARZEGAR A.Antioxidant activity of polyphenolic myri-cetin in vitro cell-free and cell-based systems [J].Molecular Biology Research Communications,2016,5(2):87- 95.
[18] EL-BESHBISHY H A,BAHASHWAN S A,ALY H A A,et al.Abrogation of cisplatin-induced nephrotoxicity in mice by alpha lipoic acid through ameliorating oxidative stress and enhancing gene expression of antioxidant enzymes [J].European Journal of Pharmacology,2011,668(1/2):278- 284.
[19] CHEN J P,LI L,SU J Y,et al.Proteomic analysis of G2/M arrest triggered by natural borneol/curcumin in HepG2 cells,the importance of the reactive oxygen species-p53 pathway [J].Journal of Agricultural and Food Chemistry,2015,63(28):6440- 6449.
[20] BATISH D R,SINGH H P,SETIA N,et al.2-Benzoxazolinone(BOA)induced oxidative stress,lipid peroxidation and changes in some antioxidant enzyme activities in mung bean (Phaseolus aureus) [J].Plant Physiology and Biochemistry,2006,44(11/12):819- 827.
[21] SAUER H,WARTENBERG M,HESCHELER J.Reactive oxygen species as intracellular messengers during cell growth and differentiation [J].Cellular Physiology Biochemistry & Pharmacology,2001,11(4):173- 186.
[22] DONOVAN M,COTTER T G.Control of mitochondrial integrity by Bcl-2 family members and caspase-independent cell death [J].Biochimica et Biophysica Acta,2004,1644(2/3):133- 147.
[23] CHEN J P,LI L,SU J Y,et al.Natural borneol enhances bisdemethoxycurcumin-induced cell cycle arrest in the G2/M phase through up-regulation of intracellular ROS in HepG2 cells [J].Food & Function,2015,6(3):740- 748.
[24] SUN F,ZHENG X Y,YE J,et al.Potential anticancer activity of myricetin in human T24 bladder cancer cells both in vitro and in vivo [J].Nutrition & Cancer,2012,64(4):599- 606.
[25] 韦伟.杨梅素体外诱导膀胱癌细胞株BIU- 87凋亡机制的研究[D].重庆:重庆医科大学,2011.
[26] WU P,LIU S,SU J,et al.Apoptosis triggered by isoquercitrin in bladder cancer cells by activating the AMPK-activated protein kinase pathway [J].Food & Function,2017,8(10):3707(1-39).
[27] 赵雪巍,刘培玉,刘丹,等.黄酮类化合物的构效关系研究进展 [J].中草药,2015,46(21):3264- 3271.ZHAO Xue-Wei,LIU Pei-yu,LIU Dan,et al.Research progress in structure-activity relationship of flavoniods [J].Chinese Traditional & Herbal Drugs,2015,46(21):3264- 3271.