不同灵芝菌株菌丝体乙醇提取物体外抑瘤活性的比较和机理
A COMPARISON STUDY OF THE IN VITROANTICANCER ACTIVITY AND MECHANISM OF ETHANOL EXTRACTS FROM MYCELIA OF DIFFERENT GANODERMA LUCIDUM STRAINS
杨晓彤 周月琴 李绪全 冯慧琴 糜可 杨庆尧
摘 要：以人急性早幼粒白血病细胞HL-60细胞株作为模型,用MTT法测定生长抑制率,流式细胞Annexin V实验和DNA含量法研究抑瘤机制,比较了8个灵芝Ganodermalucidum菌株发酵菌丝体乙醇提取物的体外抗肿瘤活性和抑瘤机制.筛选结果得到了能产生高抑瘤活性乙醇提取物的灵芝菌株L5.其对HL-60细胞的抑制率为91.4±0.9%(72小时,125μg/mL);作用48小时后,13.3%的细胞发生早期凋亡,G0/G1期细胞比例比对照组增加15.9%,而S期细胞比例则下降了8.4%,G2/M期细胞数减少7.6%.本研究证明了灵芝菌丝体乙醇提取物能有效抑制HL-60肿瘤细胞的体外生长,抑制率与菌株相关,其抑瘤机制与细胞Go/G1期阻滞和诱导凋亡有关.
关键词：细胞周期,凋亡,HL-60细胞
分类号：Q936 文献标识码：A
文章编号：1672-6472(2005)02-0251-0258
基金项目：上海市教育委员会科学项目资助(项目号04DB20)
作者简介：杨晓彤,通讯作者:email:xtyang@shnu.edu.cn
作者单位：杨晓彤（上海师范大学微生物与免疫学研究所,上海,200234）
周月琴（上海师范大学微生物与免疫学研究所,上海,200234）
李绪全（上海师范大学微生物与免疫学研究所,上海,200234）
冯慧琴（上海师范大学微生物与免疫学研究所,上海,200234）
糜可（上海师范大学微生物与免疫学研究所,上海,200234）
杨庆尧（上海师范大学微生物与免疫学研究所,上海,200234）
参考文献：
[1]Cai H, Wang FS, Wang LF, Bai Y, Xu L, Yang JS, Zhang YM, 2001. Method of determination of content of biological activity constituents ganoderic acid in Ganoderma lucidum. Chinese Journal of Veterinary Science, 21(4): 381～383 (in Chinese)
[2]Chen RY, Yu DQ, 1990. Progress of studies on chemical constituents of Ganoderma lucidum. Acta Pharmaceutica Sinica,25 (12): 940～953 (in Chinese)
[3]Fang QH, Zhong JJ, 2002. Two-stage culture process for improved production of ganoderic acid by liquid fermentation of higher fungus Ganoderma lucidum. Biotechnol. Progr, 18:51～54
[4]Gao JJ, Min BS, Ahn EM, Nakamura N, Lee HK, Hattori M, 2002. New titerpene aldehydes, lucialdehydes A-C, from Ganoderma lucidum and their cytotoxicity against murine and human tumor cells. Chem Pharm Bull, 50 (6): 867～940
[5]Gonzalez AG, Leon F, Rivera A, Padron JI, Gonzalez-Plata J, Zuluaga JC, Quintana J, Estevez F, Bermejo J, 2002. New lanostanoids from the fungus Ganoderma concinna. J Nat Prod, 5:417～421
[6]Gu J, 2002. Extraction of ganoderic acid from Ganoderma lucidum by using ultrasonic assistance. Journal of Nantong Institute of Technology (Natural Science), 1(3):13～16 (in Chinese)
[7]Hirotani M, Furuya T, 1990. Changes of the triterpene patterns during formation of the fruiting body in Ganoderma lucidum.Phytochemistry, 29:3767～3771
[8]Hu H, Ahn NS, Yang XL, Lee YS, Kang KS, 2002. Ganoderma lucidum extract induces cell cycle arrest and apoptosis in MCF-7 human breast cancer cell. Iht J Cancer, 102:250～253
[9]Jong SC, Birmingham JM, 1992. Medicinal benefits of the mushroom Ganoderma. Adv Appl Microbiol, 37:101～34
[10]Kimura Y, Taniguchi M, Baba K, 2002. Antitumor and antimetastic effects on liver triterpene fractions of Ganoderma lucidum: mechanism of action and isolation of an active substance. Anticancer Res, 22 (6A): 3309～3318
[11]Kohda H, Tokumoro W, Sakamoto K, Fuiji M, Hirai Y, Yamasaki K, Komada Y, Nakamura H, Ishihara S, Uchida M, 1985. The biologically active constituents of Ganoderma lucidum (Fr.) Karst. histamine release-inhibitory triterpenes. Chem Pharm Bull,33(4): 1367～1374
[12]Li PZ, Xu R, Xia JH, Zhang KC, 2000. Production of teracyclic triterpene acids by submerged fermentation of Ganoderma lucidum.Industrial Microbiology, 20 (1): 15～17 (in Chinese)
[13]Lin SB, Li CH, Lee SS, Kan LS, 2003. Triterpene-enriched extracts from Ganoderma lucidum inhibit growth of hepatoma cells via suppressing protein kinase C, activating mitogen-activated protein kinases and G2-phase cell cycle arrest. Life Sciences, 72:2381～2390
[14]Lin ZB, Zhang HN, 2004. Anti-tumor and immunoregulatory activities of Ganoderma lucidum and its possible mechanisms. Acta Pharm Sin, 25 (11): 1387～1395
[15]Luo J, Lin ZB, 2002. Advances of pharmacological effects of triterpenes from Ganoderma lucidum. Acta Pharm Sin, 37 (7): 574～578(in Chinese)
[16]Ma L, Wu F, Chen RY, 2003. Analysis of triterpene constituents from Ganoderma lucidum. Acta Pharm Sin, 38 (1): 50～52 (in Chinese)
[17]Min BS, Gao JJ, Nakamura N, Hattori M, 2000. Triterpenes from the spores of Ganoderma lucidum and their cytotoxicity against meth-A and LLC tumor cells. Chem Pharm Bull, 48 (7): 1026～1033
[18]Mosmann T, 1983. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods, 65:55～63
[19]Nishitoba T, Sato H, Kasai T, Kawagishi H, Sakamura S, 1985. New bitter C27 and C30 terpenoids from the fungus Ganoderma lucidum (Reishi). Agric Biol Chem, 49:1793～1798
[20]Ranynal P, Pollard HB, 1994. Annexins: the problem of assessing the biological role for a gene family of multifunctional calcium and phospholipid-binding protein. Biochemica et Biophysica Acta, 1197: 63～93.
[21]Sliva D, 2003. Ganoderma lucidum (Reishi) in cancer treatment. Integr Cancer Ther, 2(4): 358～364
[22]Toth JO, Luu B, Ourisson GL, 1983. Acides ganoderiques T a Z: triterpenes cytotoxiques de Ganoderma lucidum (Polyporaceae).Terahedr Lett, 24:1081～1084
[23]Vindelov LL, 1977. Flow microfluorometric analysis of nuclear DNA in cells from solid tumors and cell suspension.Virchows-Arch-B-Cell-Pathology, 24 (3): 227～242
[24]Wang WX, Gu ZL, 2001. Colorimetric detection of total triterpene content in Crataegus pinnatifida. China Wilding Resources,20 (5): 47～48 (in Chinese)
[25]Wu TS, Shi LS, Kuo SC, 2001. Cytotoxicity ofGanoderma lucidum triterpenes. JNatProd, 64:1121～1122
[26]Xiang SP, Tang HC, Chen G, Shi YS, 2001. Studied on colorimetric determination of oleanolic acid in Chinese quince. Natural Product Research and Development, 13 (4): 23～26 (in Chinese)
[27]Zhu HS, Yang XL, Wang LB, Zhao DX, Chen L, 2000. Effects of extracts from sporoderm-broken spores of Ganoderma lucidum on Hela cells. Cell Biol Toxicol, 16:201～206
[28]陈若芸,于德泉,1990.灵芝三萜化学成分研究进展.药学学报,25(12):940～953
[29]罗俊,林志彬,2002.灵芝三萜化合物药理作用研究进展.药学学报,37(7):574～578
[30]顾剑,2002.利用超声辅助从培养的灵芝中提取灵芝酸.南通工学院学报,1 (3):13～16
[31]王文祥,顾振纶,2001.比色法测定山渣总三萜酸的含量.中国野生植物资源,20(5):14～48
[32]项昭保,唐春红,2001.木瓜中齐墩果酸测定方法的研究.天然产物研究与开发,13(4):23～26
[33]蔡辉,王芳生,王丽凤,白燕,许莉,杨峻山,张津梅,2001.赤芝中灵芝酸类生物活性成分含量的测定方法.中国兽医学报,21(4):381～383
[34]马林,吴丰,陈若芸,2003.灵芝三萜成分分析.药学学报,1:50～52
[35]李平作,徐柔,夏结红,章克昌,2000.灵芝深层发酵生产四环三萜酸的研究.工业微生物,30(1):15～17
菌物学报
MYCOSYSTEMA
2005 Vol.24 No.2 P.251-258
