微囊藻毒素致大鼠肝细胞氧化损伤机制探讨及B族维生素联合维生素C的保护作用
|
摘要
目的:
应用分子生物学的技术,探讨微囊藻毒素对体外培养的肝细胞的损伤机制,并且对抗氧化营养素及B族维生素对微囊藻毒素染毒所致肝细胞的损伤的保护作用进行研究,为寻找肝脏保护与肝癌预防的有效方法提供参考。
方法:
1、购买BRL(大鼠肝细胞)细胞株,待增殖情况稳定,开始试验。
2、MC-LR染毒肝细胞,24小时后MTT法检测细胞增殖情况。
3、应用流式细胞仪检测毒素对细胞凋亡情况的影响。
4、根据正交设计结果,BRL预先加入维生素作用,24小时后加入MC-LR染毒,检测超氧化物歧化酶(SOD)和谷胱甘肽过氧化酶(GSHPX)。
5、RT-PCR:结合正交设计实验,筛选出有效的保护组合,通过逆转录PCR技术,测定目的基因PCNA和UDG在mRNA水平上的改变。
结果:
1、与正常组细胞相比,MC-LR在浓度低于1μg/ml时,促进BRL增殖,浓度为0.5μg/ml时,细胞增长达到峰值;MC-LR浓度高于1.25μg/ml时,抑制细胞增殖。
2、MC-LR染毒BRL后,细胞凋亡发生变化,当浓度在0.001～0.5μg/ml范围,凋亡率逐渐增加,1～2.5μg/ml凋亡率逐渐减少。
3、与正常组细胞相比,不同浓度MC-LR作用下,PCNA表达量及UDG表达量差异有统计学意义。当MC-LR浓度在0.01～1.0μg/ml范围内,PCNA表达量随MC-LR浓度的增加而增加,在MC-LR浓度达到1.25μg/ml时,PCNA表达量开始下降;UDG表达的变化趋势与PCNA一致。
4、正交分析结果显示:与没有维生素保护的染毒细胞相比,VitB6、VitB12、VC作用后,细胞内SOD、GSH-PX含量增加,差异有统计学意义,并且VitB12与VC存在协同效应。
5、在高浓度MC-LR干预下,与没有维生素保护的染毒细胞相比,维生素B6、B12+VC作用后,PCNA表达量升高,差异有统计学意义。B6、B12、B12+VC作用后,UDG表达量增加,差异有统计学意义。
在低浓度MC-LR干预下,与没有维生素保护的染毒细胞相比,维生素B6、B12作用后,PCNA表达量增加,差异有统计学意义; B6、VC、B12、B12+VC作用后,UDG表达量减少,差异有统计学意义。
结论:
1、MC-LR在不同浓度时对肝细胞增殖和凋亡的作用是不同的,在低浓度时有促进细胞增生的作用,凋亡率逐渐增加;在高浓度时却表现为抑制细胞生长或细胞毒作用的表现,凋亡率逐渐降低。
2、MC-LR通过影响BRL的PCNA和UDG基因mRNA表达,启动BER修复途径,可能是BRL氧化损伤后的修复机制之一。
3.维生素VitB6、FA、VitB12、VC对MC-LR致BRL损伤有保护作用,其中,维生素一阶交互作用的最佳保护组合为:VitB12+ VC。
Objectives:
To study the mechanism of microcystin-induced oxidative damage in vitro with molecular biology technologies, and the effect of vitamin B group & antioxidant nutrients protecting liver cells from microcystin injury, which would help us find an effective way to protect liver and prevent liver cancer.
Methods:
1、Purchase BRL cells, initiate the testing after stable proliferation.
2、MC-LR treated liver cells, MTT was detected in cell proliferation after 24 hours.
3、Analyze toxin impact upon apoptosis by FCM.
4、According to results of orthogonal design, add vitamins for 24 hours in advances, then expose BRL to join MC-LR, measuring cells for ultra-oxide dismutase (SOD) and glutathione peroxidase (GSH-PX).
5、RT-PCR: With the combination of orthogonal experimental design, select the effective protection portfolio by PCR technology and measure the PCNA and UDG changes at mRNA level.
Results:
1、Compared with normal cells, MC-LR at the concentration of less than 1μg / ml accelerate the BRL proliferation, the cell growth reached its peak at the concentration of 0.5μg / ml; MC-LR at the concentration of higher than 1.25μg / ml inhibited the cell proliferation.
2、The apoptosis changes when MC-LR’s takes effect on BRL, the apoptosis rate gradually increase in the 0.001 ~ 0.5μg / ml while at 1 ~ 2.5μg / ml the rate gradually reduced.
3、Compared with normal cells, under different concentrations of MC-LR, PCNA expression and UDG expression had a significant difference. At the concentration of 0.01 ~ 1.0μg / ml, PCNA expression increased in accordance with the concentration increases of MC-LR. MC-LR in the concentration of 1.25μg / ml, PCNA expression began to decline. UDG follow with PCNA expression of change.
4、Orthogonal analysis showed that: compared to the cells without the protection of the vitamin, after VitB6, VitB12, VC take effects, content of SOD and GSH-PX changed, and VitB12 synergies with VC.
5、When without the protection of the vitamin, cells exposed in high concentrations of MC-LR intervention, PCNA expression increased and had a significant difference after vitamin B6, B12 +VC. After vitamin B6、B12、B12 +VC role, UDG expression increased and had a significant increase.
Without the protection of the vitamin cells exposed in low concentration MC-LR, PCNA expression increased and had a significant difference after vitamin B6, B12’s effect; after B6, VC, B12, B12+VC effect, UDG expression decreased and had a significant difference .
Conclusion:
1、The effect of MC-LR in different concentrations on liver cell proliferation and on apoptosis is different, in low concentration to promote the role of cell proliferation, apoptosis rate increased gradually; in the high concentration when expressed as inhibiting cell growth or the performance of cytotoxicity , the rate of apoptosis reduced gradually.
2、MC-LR changed the expression of PCNAmRNA and UDGmRNA, initiated BER repair channels, that may be the one of the repair mechanism for BRL oxidative damage.
3. VitB6, VitB12, VC play a protection role in the MC-LR of BRL injury, and vitamin first order interaction for the best combination of protection: VitB12 + VC
应用分子生物学的技术,探讨微囊藻毒素对体外培养的肝细胞的损伤机制,并且对抗氧化营养素及B族维生素对微囊藻毒素染毒所致肝细胞的损伤的保护作用进行研究,为寻找肝脏保护与肝癌预防的有效方法提供参考。
方法:
1、购买BRL(大鼠肝细胞)细胞株,待增殖情况稳定,开始试验。
2、MC-LR染毒肝细胞,24小时后MTT法检测细胞增殖情况。
3、应用流式细胞仪检测毒素对细胞凋亡情况的影响。
4、根据正交设计结果,BRL预先加入维生素作用,24小时后加入MC-LR染毒,检测超氧化物歧化酶(SOD)和谷胱甘肽过氧化酶(GSHPX)。
5、RT-PCR:结合正交设计实验,筛选出有效的保护组合,通过逆转录PCR技术,测定目的基因PCNA和UDG在mRNA水平上的改变。
结果:
1、与正常组细胞相比,MC-LR在浓度低于1μg/ml时,促进BRL增殖,浓度为0.5μg/ml时,细胞增长达到峰值;MC-LR浓度高于1.25μg/ml时,抑制细胞增殖。
2、MC-LR染毒BRL后,细胞凋亡发生变化,当浓度在0.001～0.5μg/ml范围,凋亡率逐渐增加,1～2.5μg/ml凋亡率逐渐减少。
3、与正常组细胞相比,不同浓度MC-LR作用下,PCNA表达量及UDG表达量差异有统计学意义。当MC-LR浓度在0.01～1.0μg/ml范围内,PCNA表达量随MC-LR浓度的增加而增加,在MC-LR浓度达到1.25μg/ml时,PCNA表达量开始下降;UDG表达的变化趋势与PCNA一致。
4、正交分析结果显示:与没有维生素保护的染毒细胞相比,VitB6、VitB12、VC作用后,细胞内SOD、GSH-PX含量增加,差异有统计学意义,并且VitB12与VC存在协同效应。
5、在高浓度MC-LR干预下,与没有维生素保护的染毒细胞相比,维生素B6、B12+VC作用后,PCNA表达量升高,差异有统计学意义。B6、B12、B12+VC作用后,UDG表达量增加,差异有统计学意义。
在低浓度MC-LR干预下,与没有维生素保护的染毒细胞相比,维生素B6、B12作用后,PCNA表达量增加,差异有统计学意义; B6、VC、B12、B12+VC作用后,UDG表达量减少,差异有统计学意义。
结论:
1、MC-LR在不同浓度时对肝细胞增殖和凋亡的作用是不同的,在低浓度时有促进细胞增生的作用,凋亡率逐渐增加;在高浓度时却表现为抑制细胞生长或细胞毒作用的表现,凋亡率逐渐降低。
2、MC-LR通过影响BRL的PCNA和UDG基因mRNA表达,启动BER修复途径,可能是BRL氧化损伤后的修复机制之一。
3.维生素VitB6、FA、VitB12、VC对MC-LR致BRL损伤有保护作用,其中,维生素一阶交互作用的最佳保护组合为:VitB12+ VC。
Objectives:
To study the mechanism of microcystin-induced oxidative damage in vitro with molecular biology technologies, and the effect of vitamin B group & antioxidant nutrients protecting liver cells from microcystin injury, which would help us find an effective way to protect liver and prevent liver cancer.
Methods:
1、Purchase BRL cells, initiate the testing after stable proliferation.
2、MC-LR treated liver cells, MTT was detected in cell proliferation after 24 hours.
3、Analyze toxin impact upon apoptosis by FCM.
4、According to results of orthogonal design, add vitamins for 24 hours in advances, then expose BRL to join MC-LR, measuring cells for ultra-oxide dismutase (SOD) and glutathione peroxidase (GSH-PX).
5、RT-PCR: With the combination of orthogonal experimental design, select the effective protection portfolio by PCR technology and measure the PCNA and UDG changes at mRNA level.
Results:
1、Compared with normal cells, MC-LR at the concentration of less than 1μg / ml accelerate the BRL proliferation, the cell growth reached its peak at the concentration of 0.5μg / ml; MC-LR at the concentration of higher than 1.25μg / ml inhibited the cell proliferation.
2、The apoptosis changes when MC-LR’s takes effect on BRL, the apoptosis rate gradually increase in the 0.001 ~ 0.5μg / ml while at 1 ~ 2.5μg / ml the rate gradually reduced.
3、Compared with normal cells, under different concentrations of MC-LR, PCNA expression and UDG expression had a significant difference. At the concentration of 0.01 ~ 1.0μg / ml, PCNA expression increased in accordance with the concentration increases of MC-LR. MC-LR in the concentration of 1.25μg / ml, PCNA expression began to decline. UDG follow with PCNA expression of change.
4、Orthogonal analysis showed that: compared to the cells without the protection of the vitamin, after VitB6, VitB12, VC take effects, content of SOD and GSH-PX changed, and VitB12 synergies with VC.
5、When without the protection of the vitamin, cells exposed in high concentrations of MC-LR intervention, PCNA expression increased and had a significant difference after vitamin B6, B12 +VC. After vitamin B6、B12、B12 +VC role, UDG expression increased and had a significant increase.
Without the protection of the vitamin cells exposed in low concentration MC-LR, PCNA expression increased and had a significant difference after vitamin B6, B12’s effect; after B6, VC, B12, B12+VC effect, UDG expression decreased and had a significant difference .
Conclusion:
1、The effect of MC-LR in different concentrations on liver cell proliferation and on apoptosis is different, in low concentration to promote the role of cell proliferation, apoptosis rate increased gradually; in the high concentration when expressed as inhibiting cell growth or the performance of cytotoxicity , the rate of apoptosis reduced gradually.
2、MC-LR changed the expression of PCNAmRNA and UDGmRNA, initiated BER repair channels, that may be the one of the repair mechanism for BRL oxidative damage.
3. VitB6, VitB12, VC play a protection role in the MC-LR of BRL injury, and vitamin first order interaction for the best combination of protection: VitB12 + VC
引文
[1] 孙昌盛,陈华等.同安居民饮用水藻类毒素染毒与原发性肝癌关系的研究.中华预防医学,1999,6(2):84.
[2] 朱长林,张文彬,孙鹏等.维生素C与维生素E的联合免疫调节及抗氧化作用.中国临床康复,2006,10(36):120-122
[3] 范晓岚,杨军,糜漫天等.β- 胡萝卜素的抗氧化作用与疾病预防,中国公共卫生2003,19(4):479-480
[4] 邵宗俊,井玲,李广生等.硒和维生素E缺乏诱导大鼠肝细胞凋亡.营养学报,2000,22(3):214-218
[5] Iwase K, Higaki J, Yoon HE, et al. Reduced thiamine (vitamin B1) levels following gastrectomy for gastric cancer. Gastric Cancer. 2002;5(2):77-82
[6] Ding WX, Shen HM, Zhu HG, et al .Studies on oxidative damage induced by cyanobacteria extract in primary cultured rat hepatocytes. Environ Res. 1998 Jul;78(1):12-8.
[7] Zegura B, Sedmak B, Filipic M. Microcystin-LR induces oxidative DNA damage in human hepatoma cell line HepG2. Toxicon. 2003 Jan;41(1):41-8.
[8] 陈华,孙昌盛,胡志坚等.饮水微囊藻毒素在大鼠肝癌发生期间对细胞增殖与凋亡的影响.癌变 畸变 突变,2002,14 (4) :214-217
[9] Guzman RE, Solter PF, Runnegar MT. Inhibition of nuclear protein phosphatase activity in mouse hepatocytes by the cyanobacterial toxin microcystin-LR. Toxicon. 2003 Jun;41(7):773-81.
[10] Lankoff A, Bialczyk J, Dziga D, et al. The repair of gamma-radiation-induced DNA damage is inhibited by microcystin-LR, the PP1 and PP2A phosphatase inhibitor. Mutagenesis. 2006 Jan;21(1):83-90.
[11] 朱守民.DNA 损伤修复基本方式的研究进展.国外医学分子生物学分册,2003;25(5):270-273
[12] 冯作化.医学分子生物学.北京:人民卫生出版社,2005.
[13] 马璇,张睿等.维生素 C、E 和 B2 协同抗大鼠体外,小鼠体内脂质过氧化及血脂的研究.大连医科大学学报.1995.17(1):12-13
[14] 胡志坚,李一伟等.微囊藻毒素促肝癌过程中肝细胞 PCNA、P21waf1 基因表达研究.中国公共卫生.2002.18 (5):538-540
[15] Lin Y, Desbois A, Jiang S, et al. Group B vitamins protect murine cerebellar granule cells from glutamate/NMDA toxicity. Neuroreport. 2004Oct 5;15(14):2241-4.
[16] 司徒镇强.细胞培养.北京:世界图书出版社,2007.
[17] Carmichael WW. The toxins of cyanobacteria. Sci Am. 1994 Jan;270(1):78-86.
[18] Pouria S, de Andrade A, Barbosa J, et al. Fatal microcystin intoxication in haemodialysis unit in Caruaru, Brazil. Lancet. 1998 Jul 4;352(9121):21-6.
[19] Srivastava JK, Gupta S. Tocotrienol-rich fraction of palm oil induces cell cycle arrest and apoptosis selectively in human prostate cancer cells. Biochem Biophys Res Commun. 2006 Jul 28;346(2):447-53.
[20] Sueoka E, Goto Y, Sueoka N, et al. Heterogeneous nuclear ribonucleoprotein B1 as a new marker of early detection for human lung cancers. Cancer Res. 1999 Apr 1;59(7):1404-7.
[21] 赵金明,朱惠刚.藻毒素对fos、jun基因表达及细胞周期的影响.中华预防医学杂志,2003, 37(1):23-25.
[22] Borner C. The Bcl-2 protein family: sensors and checkpoints for life-or-death decisions. Mol Immunol. 2003 Jan;39(11):615-47.
[23] Cross A,Mcdonnell JM, Korsmeyer SJ. Bcl-2 family members and the mitochondria in apoptosis.Genes Dev.1999Oct;13(15) :l899-911.
[24] 陈加平,傅文宇,王秀敏等.微囊藻毒素LR对BRL-3A凋亡相关蛋白表达的影响中国环境科学,2004,24(4):460-463
[25] Ding WX, Shen HM, Ong CN. Critical role of reactive oxygen species and mitochondrial permeability transition in microcystin-induced rapid apoptosis in rat hepatocytes. Hepatology. 2000 Sep;32(3):547-55.
[26] S.AL-Jassabi. Effect of Microcystin from Jordan on Ion Regulation and Antioxidant System in the Hepatocytes of Mice.Joumal of Biological Sciences,2004Apr;4(4):547-52.
[27] 殷丽红,屈卫东,朱惠刚.微囊藻毒素致肝癌研究进展.国外医学卫生学分册; 2005,3(2):32-34
[28] Guzman RE, Solter PF. Characterization of sublethal microcystin-LR exposure in mice. Vet Pathol. 2002 Jan;39(1):17-26.
[29] Ding Q, Reddy YV, Wang W, et al.Autophosphorylation of the catalytic subunit of the DNA-dependent protein kinase is required for efficient end processing during DNA double-strand break repair. Mol Cell Biol. 2003 Aug;23(16):5836-48.
[30] Douglas P, Moorhead GB, Ye R, et al. Protein phosphatases regulate DNA-dependent protein kinase activity. J Biol Chem. 2001 Jun 1;276(22):18992-8.
[31] MILLS GC. Hemoglobin catabolism. I. Glutathione peroxidase, an erythrocyte enzyme which protects hemoglobin from oxidative breakdown. J Biol Chem. 1957 Nov;229(1):189-97.
[32] Kovac D, Rubinic M, Krasevic M, et al. Proliferating cell nuclear antigen (PCNA) as a prognostic factor for colorectal cancer. Anticancer Res. 1995 Sep-Oct;15(5B):2301-2.
[33] Kasahara T, Kuwayama C, Hashiba M, et al.The gene expression of hepatic proteins responsible for DNA repair and cell proliferation in tamoxifen-induced hepatocarcinogenesis. Cancer Sci. 2003 Jul;94(7):582-8.
[34] Hall PA, Levison DA, Woods AL, et al. Proliferating cell nuclear antigen (PCNA) immunolocalization in paraffin sections: an index of cell proliferation with evidence of deregulated expression in some neoplasms. J Pathol. 1990 Dec;162(4):285-94.
[35] Hur K, Kim JR, Yoon BI, et al. Overexpression of cyclin D1 and cyclin E in 1,2-dimethylhydrazine dihydrochloride-induced rat colon carcinogenesis. J Vet Sci. 2000 Dec;1(2):121-6.
[36] Haupt S, Berger M, Goldberg Z, et al.Apoptosis - the p53 network. J Cell Sci. 2003 Oct 15;116(Pt 20):4077-85.
[37] Vairapandi M, Azam N, Balliet AG, et al. Characterization of MyD118, Gadd45, and proliferating cell nuclear antigen (PCNA) interacting domains. PCNA impedes MyD118 AND Gadd45-mediated negative growth control. J Biol Chem. 2000 Jun 2;275(22):16810-9.
[38] Scott M, Bonnefin P, Vieyra D, et al. UV-induced binding of ING1 to PCNA regulates the induction of apoptosis. J Cell Sci. 2001 Oct;114(Pt 19):3455-62.
[39] 施 玮,朱惠刚,晏晚蓉,微囊藻毒素MC-LR对肝细胞毒性机理研究, 中国公共卫生,2002,18(2):23-25
[40] 杨静,马力文.肿瘤与机体抗氧化系统.肿瘤防治杂志,2004,3(11):324-327
[41] Cheng T, Zhu Z, Masuda S, et al. Effects of multinutrient supplementation on antioxidant defense systems in healthy human beings. J Nutr Biochem. 2001 Jul;12(7):388-395.
[42] Riondel J, Wong HK, Glise D, et al. The effect of a water-dispersible beta-carotene formulation on the prevention of age-related lymphoid neoplasms in mice. Anticancer Res. 2002 Mar-Apr;22(2A):883-8.
[43] Wenger FA, Kilian M, Ridders J, et al. Influence of antioxidative vitamins A, C and E on lipid peroxidation in BOP-induced pancreatic cancer in Syrian hamsters. Prostaglandins Leukot Essent Fatty Acids. 2001 Sep;65(3):165-71.
[44] Sies H, Sharov VS, Klotz LO, et al. Glutathione peroxidase protects against peroxynitrite-mediated oxidations. A new function for selenoproteins as peroxynitrite reductase. J Biol Chem. 1997 Oct 31;272(44):27812-7.
[45] Avissar N, Finkelstein JN, Horowitz S, et al. Extracellular glutathione peroxidase in human lung epithelial lining fluid and in lung cells. Am J Physiol. 1996 Feb;270(2 Pt 1):L173-82.
[46] Wang HP, Qian SY, Schafer FQ, et al. Phospholipid hydroperoxide glutathione peroxidase protects against singlet oxygen-induced cell damage of photodynamic therapy. Free Radic Biol Med. 2001 Apr 15;30(8):825-35.
[47] Halliwell B. Why and how should we measure oxidative DNA damage in nutritional studies? How far have we come? Am J Clin Nutr. 2000 Nov;72(5):1082-7.
[48] 吴坤.营养与食品卫生学.北京:人民卫生出版社,2003.
[49] Hogenkamp HPC.B-12:1948-1998.In: Banerjee R, ed. Chemistry and biochemistry of B-12.New York: Wiley Interscience, 1999:3-8.
[50] Fenton WA, Rosenberg LE. Inherited disorders of cobalamin transport and metabolism. In: Scriver CR, Beaudet AL, Sly WS, Valle DR, eds. The metabolic and molecular bases of inherited disease. New York: McGraw-Hill, 1995:3129-49
[51] Coker M, de Klerk JB, Poll-The BT, et al. Plasma total odd-chain fatty acids in the monitoring of disorders of propionate, methylmalonate and biotin metabolism. J Inherit Metab Dis. 1996;19(6):743-51.
[52] Frenkel EP. Abnormal fatty acid metabolism in peripheral nerves of patients with pernicious anemia. J Clin Invest. 1973 May;52(5):1237-45.
[53] Mudd SH, Levy HL, Skouby F. Disorders of transsulfuration. In: Scriver CR, Beaudet AL, Sly WS, Valle DR, eds. The metabolic and molecular bases of inherited disease. New York: McGraw-Hill, 1995:1279-1327
[54] Mancia F, Keep NH, Nakagawa A, et al. How coenzyme B12 radicals are generated: the crystal structure of methylmalonyl-coenzyme A mutase at 2 A resolution. Structure. 1996 Mar 15;4(3):339-50.
[55] Drennan CL, Huang S, Drummond JT, et al. How a protein binds B12: A 3.0 A X-ray structure of B12-binding domains of methionine synthase. Science. 1994 Dec 9;266(5191):1669-74.
[56] Bowman, B.A.,Russell R.M..现代营养学,北京:化学工业出版社,2004.
[57] Chuang ISH, Ian HI, Koh TW, et a1. Human DNA-(Cytosine 5) methyltransferase-PCNA complex as a target for p21 AF1.Science, 1997, 277(26) : 1996- 2000
[1] Ding Z(丁震) , Chen X D (陈晓东) ,Lin P (林萍). The study of control and treatment of pollution on algae and algal toxin in drinking water. Practical Preventive Medicine ( in Chinese) (实用预防医学) , 2001, 8 (6) : 478-480.
[2] 陈伟,甘南琴,宋立荣. 微囊藻毒素在单波长紫外光照射下的光降解动态研究[J]. 化学学报,2004,62(2):142-147.
[3] Welker M , Steinberg C. Indirect photohysis of cyanotoxins: one possible mechanism for their low persistence. Wat.Res.1999, 33: 1159-1164.
[4] Feitz A , Walte D, Jones G, et al. Photocatalytic degradation of the blue green algal toxin microcystin-LR in anatural organicaqueous matrix. Environ. Sci. Technol. 1999, 33: 243-249.
[5] Zhang W H (张维昊) , Fang T (方涛) , Xu X Q (徐小清). Study on photodegradation of cyanobacterial toxin in blooms of Dian Chi Lake. China Environmental Sci.( in Chinese) (中国环境科学) , 2001, 21 (1) : 1～3.
[6] Park H D, Sasaki Y, Maruyama T, et al. Degradation of the cyanobacterial hepato toxin microcystin by a new bacterium isolated from a hypertrophic lake. Environ. Toxicol. , 2001, 16 (4) : 337～343.
[7] Takenak S, Watanabe M F. Microcystin- LR degradation by Pseudomona saeruginosa alkaline protease. Chemosphere, 1997, 34 (4) : 749～757.
[8] Bourne D G, Jones G J , Blakeley B L , et al. Enzymatic pathway for the bacterial degradation of the cyanobacterial cyclic peptide toxin microcystin-LR. Applied and Environ. Microbiology , 1996, 62 (11) : 4086-4094.
[9] Brourne D G, Riddles P, Jones G J , et al. Characterisation of a gene cluster involved in bacterial degradation of the cyanobacterial toxin microcystin LR. Environ. Toxicon, 2001, 16: 523-534.
[10] 李 祝,郭向前,赵以军. 水环境中微囊藻毒素的生物降解[J].生态科学,2005, 24(1):378-340.
[11]廖婉琴,梁旭方,王琳等.鲢鱼可溶性谷胱甘肽S.转移酶(s GST)基因c DNA全序列与5 调控区的克隆与分析[J].中国生物化学与分子生物学报,2006,22(6):470—476.
[12] 王立新,张玲,张余霞等.黑藻养殖水对MCs的抑制效应及其机制[J].植物生理与分子生物学学报, 2006,32(6):672-678.
[13] Gross EM.Allelopathic action by submersed macro—phytes on epiphytes and phytoplankton:Algicidal hydrolysable polyphenols from Myriophyllum spicatum [D].Kiel:Christian Albrechts University.1995.
[14] GROSS E M,MEYER H,SCHILLING G.Release and ecological impact of alOicidal hydrolysable polyphenols in Myriophyllum spicatum [J].Phytochemistly ,1996,4(1):133～138.
[15] NAKAI S,INOUE Y,HOSOMI M,et a1.Myrophyllum spicatum released alopathic polyphenoles inhibiting growth of blue — green algae Chlorella pyrenoidosa [J].Water Reseach,2000,34(I):3026-3032.
[16] R0SITAN0 J,NEWEBC0MBE G,NICHOLS0N B,et a1 .Ozonation of NOM and algal toxins in four treated waters[J].Water Res,2001,35(1):23-32.
[17]郑明岚.微囊藻毒素无害化处理的研究进展[J]. 卫生研究, 2007, 36(1):114-116.
[18] STEFAN J H,DANIEL R D,BETTINA C H.Effect of Ozena ion on the removal of cyan bacterial toxins during drinking water treatment[J].Environ Health Pers,2002,llO(11):l127-1132.
[19] Idil Arslan ,ISE il Akmehmet BalciogY Lu,Detlef W Bahnemann.Advanced chemical oxidation of reactive dyes in simulated dyehouse effluents by ferrioxalate—Fenton/UV—A and TiO2/UV—A processes[J].Dyes and Pigments,2000,47(3):207-218.
[20] Gajdek Piotr.Decomposition of microcystin-LR by Fenton oxidation[J].Toxicon ,2001,39(10):1 575—1 578.
[21] Erick R B,Dolores Martinea,Evaristo Martynez,et a1.Degradation of microcystin--LR toxin by Fenton and Photo-Fenton processes[J].Toxicon,2004,43(7):829—832.
[22] 冯小刚,卫 涛,袁春伟. 饮用水中微囊藻毒素污染及其光催化降解的研究进展[J]. 东南大学学报(自然科学版),2004, 34(5):452-456.
[23] 苑宝玲,陈一萍,郑雪琴等.光催化氧化协同高铁去除藻毒素的研究[J]. 环境科学, 2004,9(5):56-59.
[24] Pendleton P, Schumann R, Wong S H. Microcystin-LR adsorption by activated carbon. J. of Colloid and Interface Sci, 2001, 240 (1) : 1-8.
[25]刘成,高乃云等.氯对粉末活性炭吸附微囊藻毒素能力的影响[J].环 境 科 学, 2007 28(5):256-262.
[26] Morris R J , Williams D E, Lu H A , et al. The adsorption of microcystin-LR by natural clay particles. Toxicon,2000, 38: 303-308.
[27] Miller M J , Critchley M , Hutson J , et al. The adsorption of cyanobacterial hepao toxins from water onto soil during batch experiments. Wat. Res. 2001, 35 (6) : 1461-1468.
[28]刘振儒,赵纪强.壳聚糖去除MCs的工艺研究及机理探讨[J].青岛科技大学学报, 2006,26(5): 394-405.
[29] 纪荣平,李先宁,吕锡武等. 人工介质富集微生物对藻类和藻毒素降解试验研究[J].东南大学学报(自然科学版),2005, 35 (1): 486-491.
[30]王波,张光明,马伯志等.微囊藻毒素在超声场中的降解研究[J].环境科学, 2005, 26(6):101-104.
[31]张继彪,郑正,杨光俊等.辐照降解微囊藻毒素的研究[J].环境化学,2006,25(5):143-146.
[32] 陈铁晖 ,薛常镐 ,汪家梨等.姜黄素拮抗水华微囊藻毒素致动物肝氧化损伤[J].中国公共卫生,2006, 22(11):1402-1403.
[33] 许川 ,舒为群,邱志群等.绿茶对微囊藻毒素LR诱导肝细胞凋亡、Bcl-2表达及骨髓嗜多染红细胞微核的影响[J].癌变 畸变 突变,2007,19(1):29-32.
[2] 朱长林,张文彬,孙鹏等.维生素C与维生素E的联合免疫调节及抗氧化作用.中国临床康复,2006,10(36):120-122
[3] 范晓岚,杨军,糜漫天等.β- 胡萝卜素的抗氧化作用与疾病预防,中国公共卫生2003,19(4):479-480
[4] 邵宗俊,井玲,李广生等.硒和维生素E缺乏诱导大鼠肝细胞凋亡.营养学报,2000,22(3):214-218
[5] Iwase K, Higaki J, Yoon HE, et al. Reduced thiamine (vitamin B1) levels following gastrectomy for gastric cancer. Gastric Cancer. 2002;5(2):77-82
[6] Ding WX, Shen HM, Zhu HG, et al .Studies on oxidative damage induced by cyanobacteria extract in primary cultured rat hepatocytes. Environ Res. 1998 Jul;78(1):12-8.
[7] Zegura B, Sedmak B, Filipic M. Microcystin-LR induces oxidative DNA damage in human hepatoma cell line HepG2. Toxicon. 2003 Jan;41(1):41-8.
[8] 陈华,孙昌盛,胡志坚等.饮水微囊藻毒素在大鼠肝癌发生期间对细胞增殖与凋亡的影响.癌变 畸变 突变,2002,14 (4) :214-217
[9] Guzman RE, Solter PF, Runnegar MT. Inhibition of nuclear protein phosphatase activity in mouse hepatocytes by the cyanobacterial toxin microcystin-LR. Toxicon. 2003 Jun;41(7):773-81.
[10] Lankoff A, Bialczyk J, Dziga D, et al. The repair of gamma-radiation-induced DNA damage is inhibited by microcystin-LR, the PP1 and PP2A phosphatase inhibitor. Mutagenesis. 2006 Jan;21(1):83-90.
[11] 朱守民.DNA 损伤修复基本方式的研究进展.国外医学分子生物学分册,2003;25(5):270-273
[12] 冯作化.医学分子生物学.北京:人民卫生出版社,2005.
[13] 马璇,张睿等.维生素 C、E 和 B2 协同抗大鼠体外,小鼠体内脂质过氧化及血脂的研究.大连医科大学学报.1995.17(1):12-13
[14] 胡志坚,李一伟等.微囊藻毒素促肝癌过程中肝细胞 PCNA、P21waf1 基因表达研究.中国公共卫生.2002.18 (5):538-540
[15] Lin Y, Desbois A, Jiang S, et al. Group B vitamins protect murine cerebellar granule cells from glutamate/NMDA toxicity. Neuroreport. 2004Oct 5;15(14):2241-4.
[16] 司徒镇强.细胞培养.北京:世界图书出版社,2007.
[17] Carmichael WW. The toxins of cyanobacteria. Sci Am. 1994 Jan;270(1):78-86.
[18] Pouria S, de Andrade A, Barbosa J, et al. Fatal microcystin intoxication in haemodialysis unit in Caruaru, Brazil. Lancet. 1998 Jul 4;352(9121):21-6.
[19] Srivastava JK, Gupta S. Tocotrienol-rich fraction of palm oil induces cell cycle arrest and apoptosis selectively in human prostate cancer cells. Biochem Biophys Res Commun. 2006 Jul 28;346(2):447-53.
[20] Sueoka E, Goto Y, Sueoka N, et al. Heterogeneous nuclear ribonucleoprotein B1 as a new marker of early detection for human lung cancers. Cancer Res. 1999 Apr 1;59(7):1404-7.
[21] 赵金明,朱惠刚.藻毒素对fos、jun基因表达及细胞周期的影响.中华预防医学杂志,2003, 37(1):23-25.
[22] Borner C. The Bcl-2 protein family: sensors and checkpoints for life-or-death decisions. Mol Immunol. 2003 Jan;39(11):615-47.
[23] Cross A,Mcdonnell JM, Korsmeyer SJ. Bcl-2 family members and the mitochondria in apoptosis.Genes Dev.1999Oct;13(15) :l899-911.
[24] 陈加平,傅文宇,王秀敏等.微囊藻毒素LR对BRL-3A凋亡相关蛋白表达的影响中国环境科学,2004,24(4):460-463
[25] Ding WX, Shen HM, Ong CN. Critical role of reactive oxygen species and mitochondrial permeability transition in microcystin-induced rapid apoptosis in rat hepatocytes. Hepatology. 2000 Sep;32(3):547-55.
[26] S.AL-Jassabi. Effect of Microcystin from Jordan on Ion Regulation and Antioxidant System in the Hepatocytes of Mice.Joumal of Biological Sciences,2004Apr;4(4):547-52.
[27] 殷丽红,屈卫东,朱惠刚.微囊藻毒素致肝癌研究进展.国外医学卫生学分册; 2005,3(2):32-34
[28] Guzman RE, Solter PF. Characterization of sublethal microcystin-LR exposure in mice. Vet Pathol. 2002 Jan;39(1):17-26.
[29] Ding Q, Reddy YV, Wang W, et al.Autophosphorylation of the catalytic subunit of the DNA-dependent protein kinase is required for efficient end processing during DNA double-strand break repair. Mol Cell Biol. 2003 Aug;23(16):5836-48.
[30] Douglas P, Moorhead GB, Ye R, et al. Protein phosphatases regulate DNA-dependent protein kinase activity. J Biol Chem. 2001 Jun 1;276(22):18992-8.
[31] MILLS GC. Hemoglobin catabolism. I. Glutathione peroxidase, an erythrocyte enzyme which protects hemoglobin from oxidative breakdown. J Biol Chem. 1957 Nov;229(1):189-97.
[32] Kovac D, Rubinic M, Krasevic M, et al. Proliferating cell nuclear antigen (PCNA) as a prognostic factor for colorectal cancer. Anticancer Res. 1995 Sep-Oct;15(5B):2301-2.
[33] Kasahara T, Kuwayama C, Hashiba M, et al.The gene expression of hepatic proteins responsible for DNA repair and cell proliferation in tamoxifen-induced hepatocarcinogenesis. Cancer Sci. 2003 Jul;94(7):582-8.
[34] Hall PA, Levison DA, Woods AL, et al. Proliferating cell nuclear antigen (PCNA) immunolocalization in paraffin sections: an index of cell proliferation with evidence of deregulated expression in some neoplasms. J Pathol. 1990 Dec;162(4):285-94.
[35] Hur K, Kim JR, Yoon BI, et al. Overexpression of cyclin D1 and cyclin E in 1,2-dimethylhydrazine dihydrochloride-induced rat colon carcinogenesis. J Vet Sci. 2000 Dec;1(2):121-6.
[36] Haupt S, Berger M, Goldberg Z, et al.Apoptosis - the p53 network. J Cell Sci. 2003 Oct 15;116(Pt 20):4077-85.
[37] Vairapandi M, Azam N, Balliet AG, et al. Characterization of MyD118, Gadd45, and proliferating cell nuclear antigen (PCNA) interacting domains. PCNA impedes MyD118 AND Gadd45-mediated negative growth control. J Biol Chem. 2000 Jun 2;275(22):16810-9.
[38] Scott M, Bonnefin P, Vieyra D, et al. UV-induced binding of ING1 to PCNA regulates the induction of apoptosis. J Cell Sci. 2001 Oct;114(Pt 19):3455-62.
[39] 施 玮,朱惠刚,晏晚蓉,微囊藻毒素MC-LR对肝细胞毒性机理研究, 中国公共卫生,2002,18(2):23-25
[40] 杨静,马力文.肿瘤与机体抗氧化系统.肿瘤防治杂志,2004,3(11):324-327
[41] Cheng T, Zhu Z, Masuda S, et al. Effects of multinutrient supplementation on antioxidant defense systems in healthy human beings. J Nutr Biochem. 2001 Jul;12(7):388-395.
[42] Riondel J, Wong HK, Glise D, et al. The effect of a water-dispersible beta-carotene formulation on the prevention of age-related lymphoid neoplasms in mice. Anticancer Res. 2002 Mar-Apr;22(2A):883-8.
[43] Wenger FA, Kilian M, Ridders J, et al. Influence of antioxidative vitamins A, C and E on lipid peroxidation in BOP-induced pancreatic cancer in Syrian hamsters. Prostaglandins Leukot Essent Fatty Acids. 2001 Sep;65(3):165-71.
[44] Sies H, Sharov VS, Klotz LO, et al. Glutathione peroxidase protects against peroxynitrite-mediated oxidations. A new function for selenoproteins as peroxynitrite reductase. J Biol Chem. 1997 Oct 31;272(44):27812-7.
[45] Avissar N, Finkelstein JN, Horowitz S, et al. Extracellular glutathione peroxidase in human lung epithelial lining fluid and in lung cells. Am J Physiol. 1996 Feb;270(2 Pt 1):L173-82.
[46] Wang HP, Qian SY, Schafer FQ, et al. Phospholipid hydroperoxide glutathione peroxidase protects against singlet oxygen-induced cell damage of photodynamic therapy. Free Radic Biol Med. 2001 Apr 15;30(8):825-35.
[47] Halliwell B. Why and how should we measure oxidative DNA damage in nutritional studies? How far have we come? Am J Clin Nutr. 2000 Nov;72(5):1082-7.
[48] 吴坤.营养与食品卫生学.北京:人民卫生出版社,2003.
[49] Hogenkamp HPC.B-12:1948-1998.In: Banerjee R, ed. Chemistry and biochemistry of B-12.New York: Wiley Interscience, 1999:3-8.
[50] Fenton WA, Rosenberg LE. Inherited disorders of cobalamin transport and metabolism. In: Scriver CR, Beaudet AL, Sly WS, Valle DR, eds. The metabolic and molecular bases of inherited disease. New York: McGraw-Hill, 1995:3129-49
[51] Coker M, de Klerk JB, Poll-The BT, et al. Plasma total odd-chain fatty acids in the monitoring of disorders of propionate, methylmalonate and biotin metabolism. J Inherit Metab Dis. 1996;19(6):743-51.
[52] Frenkel EP. Abnormal fatty acid metabolism in peripheral nerves of patients with pernicious anemia. J Clin Invest. 1973 May;52(5):1237-45.
[53] Mudd SH, Levy HL, Skouby F. Disorders of transsulfuration. In: Scriver CR, Beaudet AL, Sly WS, Valle DR, eds. The metabolic and molecular bases of inherited disease. New York: McGraw-Hill, 1995:1279-1327
[54] Mancia F, Keep NH, Nakagawa A, et al. How coenzyme B12 radicals are generated: the crystal structure of methylmalonyl-coenzyme A mutase at 2 A resolution. Structure. 1996 Mar 15;4(3):339-50.
[55] Drennan CL, Huang S, Drummond JT, et al. How a protein binds B12: A 3.0 A X-ray structure of B12-binding domains of methionine synthase. Science. 1994 Dec 9;266(5191):1669-74.
[56] Bowman, B.A.,Russell R.M..现代营养学,北京:化学工业出版社,2004.
[57] Chuang ISH, Ian HI, Koh TW, et a1. Human DNA-(Cytosine 5) methyltransferase-PCNA complex as a target for p21 AF1.Science, 1997, 277(26) : 1996- 2000
[1] Ding Z(丁震) , Chen X D (陈晓东) ,Lin P (林萍). The study of control and treatment of pollution on algae and algal toxin in drinking water. Practical Preventive Medicine ( in Chinese) (实用预防医学) , 2001, 8 (6) : 478-480.
[2] 陈伟,甘南琴,宋立荣. 微囊藻毒素在单波长紫外光照射下的光降解动态研究[J]. 化学学报,2004,62(2):142-147.
[3] Welker M , Steinberg C. Indirect photohysis of cyanotoxins: one possible mechanism for their low persistence. Wat.Res.1999, 33: 1159-1164.
[4] Feitz A , Walte D, Jones G, et al. Photocatalytic degradation of the blue green algal toxin microcystin-LR in anatural organicaqueous matrix. Environ. Sci. Technol. 1999, 33: 243-249.
[5] Zhang W H (张维昊) , Fang T (方涛) , Xu X Q (徐小清). Study on photodegradation of cyanobacterial toxin in blooms of Dian Chi Lake. China Environmental Sci.( in Chinese) (中国环境科学) , 2001, 21 (1) : 1～3.
[6] Park H D, Sasaki Y, Maruyama T, et al. Degradation of the cyanobacterial hepato toxin microcystin by a new bacterium isolated from a hypertrophic lake. Environ. Toxicol. , 2001, 16 (4) : 337～343.
[7] Takenak S, Watanabe M F. Microcystin- LR degradation by Pseudomona saeruginosa alkaline protease. Chemosphere, 1997, 34 (4) : 749～757.
[8] Bourne D G, Jones G J , Blakeley B L , et al. Enzymatic pathway for the bacterial degradation of the cyanobacterial cyclic peptide toxin microcystin-LR. Applied and Environ. Microbiology , 1996, 62 (11) : 4086-4094.
[9] Brourne D G, Riddles P, Jones G J , et al. Characterisation of a gene cluster involved in bacterial degradation of the cyanobacterial toxin microcystin LR. Environ. Toxicon, 2001, 16: 523-534.
[10] 李 祝,郭向前,赵以军. 水环境中微囊藻毒素的生物降解[J].生态科学,2005, 24(1):378-340.
[11]廖婉琴,梁旭方,王琳等.鲢鱼可溶性谷胱甘肽S.转移酶(s GST)基因c DNA全序列与5 调控区的克隆与分析[J].中国生物化学与分子生物学报,2006,22(6):470—476.
[12] 王立新,张玲,张余霞等.黑藻养殖水对MCs的抑制效应及其机制[J].植物生理与分子生物学学报, 2006,32(6):672-678.
[13] Gross EM.Allelopathic action by submersed macro—phytes on epiphytes and phytoplankton:Algicidal hydrolysable polyphenols from Myriophyllum spicatum [D].Kiel:Christian Albrechts University.1995.
[14] GROSS E M,MEYER H,SCHILLING G.Release and ecological impact of alOicidal hydrolysable polyphenols in Myriophyllum spicatum [J].Phytochemistly ,1996,4(1):133～138.
[15] NAKAI S,INOUE Y,HOSOMI M,et a1.Myrophyllum spicatum released alopathic polyphenoles inhibiting growth of blue — green algae Chlorella pyrenoidosa [J].Water Reseach,2000,34(I):3026-3032.
[16] R0SITAN0 J,NEWEBC0MBE G,NICHOLS0N B,et a1 .Ozonation of NOM and algal toxins in four treated waters[J].Water Res,2001,35(1):23-32.
[17]郑明岚.微囊藻毒素无害化处理的研究进展[J]. 卫生研究, 2007, 36(1):114-116.
[18] STEFAN J H,DANIEL R D,BETTINA C H.Effect of Ozena ion on the removal of cyan bacterial toxins during drinking water treatment[J].Environ Health Pers,2002,llO(11):l127-1132.
[19] Idil Arslan ,ISE il Akmehmet BalciogY Lu,Detlef W Bahnemann.Advanced chemical oxidation of reactive dyes in simulated dyehouse effluents by ferrioxalate—Fenton/UV—A and TiO2/UV—A processes[J].Dyes and Pigments,2000,47(3):207-218.
[20] Gajdek Piotr.Decomposition of microcystin-LR by Fenton oxidation[J].Toxicon ,2001,39(10):1 575—1 578.
[21] Erick R B,Dolores Martinea,Evaristo Martynez,et a1.Degradation of microcystin--LR toxin by Fenton and Photo-Fenton processes[J].Toxicon,2004,43(7):829—832.
[22] 冯小刚,卫 涛,袁春伟. 饮用水中微囊藻毒素污染及其光催化降解的研究进展[J]. 东南大学学报(自然科学版),2004, 34(5):452-456.
[23] 苑宝玲,陈一萍,郑雪琴等.光催化氧化协同高铁去除藻毒素的研究[J]. 环境科学, 2004,9(5):56-59.
[24] Pendleton P, Schumann R, Wong S H. Microcystin-LR adsorption by activated carbon. J. of Colloid and Interface Sci, 2001, 240 (1) : 1-8.
[25]刘成,高乃云等.氯对粉末活性炭吸附微囊藻毒素能力的影响[J].环 境 科 学, 2007 28(5):256-262.
[26] Morris R J , Williams D E, Lu H A , et al. The adsorption of microcystin-LR by natural clay particles. Toxicon,2000, 38: 303-308.
[27] Miller M J , Critchley M , Hutson J , et al. The adsorption of cyanobacterial hepao toxins from water onto soil during batch experiments. Wat. Res. 2001, 35 (6) : 1461-1468.
[28]刘振儒,赵纪强.壳聚糖去除MCs的工艺研究及机理探讨[J].青岛科技大学学报, 2006,26(5): 394-405.
[29] 纪荣平,李先宁,吕锡武等. 人工介质富集微生物对藻类和藻毒素降解试验研究[J].东南大学学报(自然科学版),2005, 35 (1): 486-491.
[30]王波,张光明,马伯志等.微囊藻毒素在超声场中的降解研究[J].环境科学, 2005, 26(6):101-104.
[31]张继彪,郑正,杨光俊等.辐照降解微囊藻毒素的研究[J].环境化学,2006,25(5):143-146.
[32] 陈铁晖 ,薛常镐 ,汪家梨等.姜黄素拮抗水华微囊藻毒素致动物肝氧化损伤[J].中国公共卫生,2006, 22(11):1402-1403.
[33] 许川 ,舒为群,邱志群等.绿茶对微囊藻毒素LR诱导肝细胞凋亡、Bcl-2表达及骨髓嗜多染红细胞微核的影响[J].癌变 畸变 突变,2007,19(1):29-32.