摘要
目的
白血病是人类好发的恶性肿瘤,是由于造血干细胞增殖分化异常而导致的恶性增殖性疾病。迄今白血病的治疗仍主要采用传统的大剂量联合化疗,但此疗法副作用大,复发率高,而且对正常机体免疫与造血系统有极大的损害。人参总皂苷(TSPG)是人参“补气生血”或促进造血的有效成分。我们既往研究表明:TSPG对造血系统恶性肿瘤的增殖有抑制作用。但TSPG对白血病增殖抑制及诱导分化的机理尚不清楚。本研究采用体外细胞培养技术观察TSPG对K562细胞增殖分化的影响,旨在阐述人参对血细胞发生的调控机制,为研制开发天然中药分化诱导剂提供理论与实验依据。
方法
1、体外培养白血病细胞株K562。
2、采用MTT法、细胞形态观察探讨TSPG对K562细胞增殖的抑
Objective
Leukemia is a very familiar malignancy in human being. Its pathogeny is due to bone marrow hematopoietic stem cells proliferation and differentiation abnormally. So far, traditional chemotherapy and radiotherapy are still the main therapies, but they have many side effects and the ratio of relapse is very high. What’s more, it greatly damages the normal immunity and hematopoiesis. Total saponins of Panaxginseng (TSPG) is one of the main effective ingredient extracted from Panax Ginseng which plays a vital role in“invigorating qi”or promoting haematopoietic function. Our previous study suggested that TSPG can inhibit the proliferation of leukemia cells. But the molecular biological mechanism for TSPG inhibiting proliferation and inducing differentiation of K562 cells is not clear. By using techniques of cell culture ex-vivo, we can observe the effect of TSPG on proliferation and differentiation of K562 cells. Our purpose is to clarify the modern biological mechanism for TSPG regulating haematopoietic function, and to provide the theoretical basis and the experimental evidence of its clinic application.
Methods
1, The leukemia cell line (K562) was cultured out of body.
引文
[1] El-Bayonmy K. Sinha R. Mechanisms of mammary cancer chemoprevention by organoselenium compounds[J]. Mutal Res, 2004,551(1-2):181-197
[2] 李丽艳,路新国.人参皂甙的最新研究进展[J].四川省卫生管理干部学院学报,2004,23(4):293-295.
[3] Yun T K, Lee Y S, Lee Y H, et al. Anticarcinogenic effect of Panax ginseng C.A. Meyer and identification of active compounds[J]. J Korean Med Sci, 2001,16(suppl):S6-18.
[4] Shibata S. Chemistry and cancer preventing activities of ginseng saponins and some related triterpenoid compounds[J]. J Korean Med Sci, 2001,16(suppl):28-37.
[5] 陈婷梅,王亚平。人参总皂甙对 K562 细胞增殖抑制与诱导分化的实验研究[J]. 重庆医科大学学报,199,24(2):115-116.
[6] 陈婷梅,王亚平,陈地龙,等.人参总皂甙诱导 K562 细胞凋亡的实验研究[J].中草药,2003,34(3):235-237.
[7] 王勇,王亚平,祝彼得等. 人参总皂甙对小鼠造血细胞增殖的影响[J].解剖学杂志,1995,18(2):153-156.
[8] 王亚平,王勇. 人参总皂甙对小鼠造血细胞增殖的影响及其调节机理研究[J]. 中药药理与临床研究进展,1996,P273-283.
[9] 王勇,王亚平,祝彼得. 人参总皂甙体外刺激小鼠造血作用机制的研究[J].中华血液学杂志,1995,16(12):648-649.
[10] 陈地龙,王亚平,陈婷梅. 人参总皂甙协同 EPO 和 GM-CSF 对 K562 细胞诱导分化的实验研究[J].第三军医大学学报,2003,25(22):2018-2021.
[11] Lazzio CB, Lazzio BB. Human chronic myelogenous leukemia cellline with positive Philadelphia chromosome[J].Blood,1975,45(6):321-324.
[12] Tabilio A, Pelicci P G, Vinci G, et al. Myeloid and megakaryocytic properties of K562 cell lines [J]. Cancer Res, 1983,43:4569-4573.
[13] Sachs L. The molecular regulation of normal and leukemia blood cells[J]. Pro R Scotland, 1987,231:289.
[14] 陈婷梅,祝彼得. 抗白血病中药及天然药物的研究现状[J].中国中西医结合杂志,1995,15(5):317-319.
[15] Chong SKF, Oberholzer VG. Ginseng is there a use in clinical medicine[J]. Postgr Med J 1988,64(6):841
[16] 陆平成,张旭,周坤福,等. 人参皂甙联合 TNF 对肿瘤的作用[J].南京中医药大学学报,1995,11(4):27-29.
[17] 窦德强,陈英杰.人参的化学成分及药理活性的研究进展与展望[J].沈阳药科大学学报,1999,16(2):151-156.
[18] 李静,王亚平. 人参总皂甙对骨髓巨噬细胞的影响及与造血调控的关系[J].解剖学杂志,2004,27(2):109-113.
[19] 王璐,王亚平. 人参总皂甙对人淋巴细胞表达早期造血生长因子的影响[J]. 解剖学杂志,2004,27(2):157-161.
[20] 王璐,王亚平. 人参总皂甙诱导人造血基质细胞表达 IL-3 的实验研究[J]. 解剖学杂志,2004,35(1):49-54
[21] Charles J. S, Mammalian G1 cyclines[J]. Cell, 1993, 73:1059-1065.
[22] 王建林,张太峰,李海艳.细胞周期及其对肿瘤防治的意义[J].中国兽医科技,1994,24(1):46-47.
[23] Johnoson DG, S. Rajagopalan, V. Prakash et al. Annu Rev Pharmacol Toxicol[M], 1999,39:295-312.
[24] Carson DA, Ribeiro J. Apoptosis and disease [J].Lancet, 1993,341:1231.
[25] Suzuk M, Shinnar AE, Nacker LA, et al. Laurintero from red alga L. Nidificayamada [J]. Bull Chem Soc Japan, 1997,52(3):352-355.
[26] 马靖,庞大本,池旭生.药物诱导细胞凋亡时 DNA 检测技术的探讨[J].癌症,2000,19(3):11-13.
[27] 姜泊.细胞凋亡基础与临床[M].北京:人民军医出版社,1999.11-12.
[28] Gorczyca W, Gong J, Ardelt B, et al. The cell cycle related differencesin susceptibility of HL260 cells to apoptosis induced byvarious antitumor agents[J]. Cancers,1998,53(13):3186-3192.
[29] 孙 宪 寿 . 白 血 病 细 胞 诱 导 分 化 的 研 究 进 展 [J]. 基 础 医 学 与 临床,1995,15(3):10.
[1] Mulcahy L. The erythropoietin receptor [J]. Semin Oncol,2001,28(2 suppl 8):19
[2] Miura Y, Miura O, Ihle J N, et al. Activation of the mitogen activated protein kinase pathway by the erythropoietin receptor[J]. J Biol Chem,1994,269:29962-29969.
[3] Ren H Y, Komatsu N, Shimizu R, et al. Erythropoietin induces tyrosine phosphorylation and activation of phospholipase Cγ1 in a human erythropoietin-dependent cell line[J]. J Biol Chem, 1994,269:19633-19638.
[4] Anagnostou A, lee E S, Kessimian N, et al. Erythropoietin has a mitogenic and positive chemotactic effect on endothelial cells [J]. Proc Natl Acad Sci USA. 1990,87:5978-5982.
[5] Batra S, Perelman N, Luck L R, et al. Pediatric tumor cells express erythropoietin and a functional erythropoietin receptor that promotes angiogenesis and tumor cell survival[J]. Nature,2003,83(10):1477-1487.
[6] Arcasoy M O, Amin K, Vollmer R T, et al. Erythropoietin and erythropoietin receptor expression in human prostate cancer[J]. Mod Pathol,2004,10:1038.
[7] 金锋,樊华,路平,等. EpoR 表达与乳腺癌的侵润相关性研究[J]. 肿瘤研究与临床,2003,15(1):8-9.
[8] 樊华,杉田完尔,瞿明,等.红细胞生成素受体介导的白血病细胞系增殖信号传导的研究[J]. 中华血液学杂志,2000,21:295.
[9] Touw L, Van Gurp R, Schipper P, et al. Blood[M]. 1992,79:2867.
[10] Kittler EL, McGrath H, Temeles D, et al. Biologic significance of constitutive and subliminal growth factor production by bone marrowstroma[J]. Blood.1992,79(12):3168-3178.
[11] Sutherland J A, Turner A R, Mannoni P, et al. Differentiation of K562 leukemia cells along erythroid, macrophage, and megakaryocyte lineages[J]. J Biol Response mod,1986,5:250-262.
[12] 王传清,袁弥满。氯高铁血红素对血红蛋白合成及红细胞生成素受体的诱导作用[J]. 上海医科大学学报,1998,25(5):357-359.
[13] Stopka T, Zivny JH, Stopkova P,et al. Human hematopoietic progenitors express erythropoietin[J]. Blood,1998,91(10):3766-3772.
[14] Jeannesson P, Lahlil R, Chenais B, et al. Anthracyclines as tumor cell differentiating agents: effects on the regulation of erythroid gene expression[J]. Leuk Lymphoma, 1997,26(5-6):575-587
[15] 陈婷梅,王亚平。人参总皂甙对 K562 细胞增殖抑制与诱导分化的实验研究[J]. 重庆医科大学学报,1999;24(2):115-116.
[16] J.萨姆布鲁克,E.F.弗里奇,T.曼尼阿蒂斯,分子克隆实验指南(第三版)[[M].科学出版社.
[17] 赵家增,王鸿利,韩忠朝.血液实验学[M].上海科学技术出版社,1997,126.
[18] Weimar IS, Mirauda N, Muller EJ, et al. Hepatocyte growth factor/scatter factor (HGF/SF) is produced by human bone marrow stromal cells peomotes proliferation, adhesion and surivial of human hematopoietic progenitor cells(CD+34)[J]. Experimantal Hematology, 1998,26:885.
[19] Bazan JF. Structural design and molecular evolution of a cytokine receptor superfamily[J]. Proc Natl Acad Sci USA,1990,87(18):6934-6939.
[20] Dusanter-Fourt I, Casadevall N, Lacombe C, et al. Erythropoietin induces the tyrosine phosphorylation of its own receptor in human erythropoietin-responsive cells[J].J Biol Chem,1992,267(15): 10670- 10675.
[21] Sawyer ST. et al. Erythropoietin cell biology[J]. Hematol Oncol Clin North Am 1994 Oct;8(5):895-898.
[22] 樊华,杉田完尔,瞿明,等.白血病细胞株红细胞生成素受体的表达及机能的研究[J].白血病,2000;9(2):98-99.
[23] 张克强,唐佩弦. 红细胞生成素功能及其介导的信号转导[J].国外医学输血及血液学分册,2001,24(1)36-39.
[24] Neumann D, Wikstrom L, Watowich SS, et al. Intermediates in degradation of the erythropoietin receptor accumulate and are degraded in lysosomes[J]. J Biol Chem, 1993,268(18):13639-13649.
1、Ostor W, Frisch J, Nicolay U, et al. Interleukin-3 biologic effects and clinical impact. Cancer 1991, 67(2):2712
2、Reddy EP, Korapati A, Chaturvedi P, et al. Il-3 signaling and the role of Src kinases, JAKs and STATs : a covert liaison unveiled. Oncogene 2000, 19: 2532-2547
3、Martinez M,Margarita PhD; Huston,et al. Biology of common [beta] receptor-signaling cytokines: IL-3, IL-5, and GM-CSF.The Journal of Allergy and Clinical Immunology 2003, 112(4): 653-665
4、Carr PD, Gustin SE, Church AP, et al. Structure of the complete extracellular domain of the common β subunit of the human GM-CSF, IL-3, and IL-5 receptors reveals a novel dimmer configuration. Cell 2001, 104:291-300
5、Gustin SE, Church AP, Ford SC, et al. Expression, crystallization and derivatization of the complete extracellular domain of theβc subunit of the human GM-CSF, IL-3, and IL-5 receptors. Eur. J. Biochem 2001, 268:2905-2911
6、Ausushi M, Alice L-FM, Toshiya O, et al. Receptors for granulocyte-macrophage colony-stimulating factor, interleukin(IL)-3, and IL-5. Blood 1993,82(1): 1960-1974
7、Stefania1 M, Roberta1 R, Isabella1 P. Expression of interleukin 3 and granulocyte-macrophage colony-stimulating factor receptor common chain [bgr]c, [bgr] IT in normal haematopoiesis[colon] lineage specificity and proliferation-independent induction. British Journal of Haematology 2000, 111(2): 441-451
8、Miyajirna A, Ito Y, Kinoshita T. Cytokine signaling for proliferation, surival and death in hematopoietic cells. Hematol 1999,69(3): 137-146
9、Barry SC, Bagley CJ, Phillips J, et al. Two contiguous residues in human interleukin-3, Asp21 and Glu22, selectively interact with the alpha- andbeta-chains of its receptor and participate in function. J Biol Chem 1994; 269:8488-8492
10、Woodeock JM, Bagley CJ, Zacharakis B, et al. A single tyrosine residue in the membrane-proximal domainof the granulocyte-macrophage colony-stimulating factor, interleukin-3, and IL-5 receptor common β-chain is necessary and sufficient for hign affinity binding and signaling by all three ligands. J Biol Chem 1996;271(42):25999-26006
11、Rane SG, Reddy EP. Janus kinases: Components of multiple signaling pathways. Oncogene 2000,19:5662-5679
12、Ward AC, Touw J, Yoshimure A. The JAK-STAT pathway in normal and perturbed hematopoiesis. Blood 2000, 95:19
13、Helen W, Pamela J R, David C L. Differentiation-linked changes in granulocyte-macrophage colony-stimulating factor receptor mediated signalling in the HL-60 promyelocytic cell line. British Journal of Haematology 1998, 101(1): 82-89
14、Santos SCR, Dumon S, Mayeux P, et al. Cooperation between STAT5 and phosphatidylinositol 3-kinase in the IL-3-dependent surival of a bone marrow derived cell line. Oncogene 2000,19:1164-1172
15、W Low, G Olmos-Centenera, c Madsen, et al. Role of Bax in apoptosis of IL-3-dependent cells. Oncogene 2001,20:4476-4483
16、Briggs SD, Bryant SS, Jove R, et al. The Ras GTPame-activating protein (GAP) is an SH3 domain-binding protein and substrate for the Src-related tyrosine kinase. J Biol Chem 1995,270(24): 14718-14724
17、 Luis DP, Maribel GG, Carmen P, et al. Interleukin-3 induced phosphorylation of BAD through the protein kinase Akt. Science 1997, 278(5338):687-689
18、He P, Tang ZhY. Cytokine receptor superfamily and its role in signal transduction. Prog Biochem Biophys 1998,25(3):201-204
19、Santos SCR, Lacronique V, Bouchaerrt I, et al. Constitutively activeSTAT5 variants induce growth and surival of hematopoietic cells through a PI-3K/Akt dependent pathway. Oncogene 2000,19,1164-1172