李晞,王继红,肖亚雄
LI Xi,WANG Ji-hong,XIAO Ya-xiong(Laboratory of Biochemistry and Molecular Biology of Experimental Teaching Center

• 1:重庆医科大学基础医学实验教学中心生物化学与分子生物学实验室
• 2:重庆医科大学分子医学与肿瘤研究中心

摘要(Abstract)：

目的观察重楼提取液对人结肠癌SW480细胞增殖的影响,并探讨其作用机制。方法采用不同浓度(10、20、40、80μg/ml)的重楼提取液,分别作用于体外培养的SW480细胞12、24、36和48h,采用MTT法检测其对SW480细胞的抑制率;并以半数抑制浓度的重楼提取液与体外培养的SW480细胞作用24h后,显微镜下观察细胞形态,并通过流式细胞术分析其对SW480细胞细胞周期的影响,RT-PCR和Western blot法分别测定SW480细胞干细胞因子(SCF)mRNA和蛋白表达的变化。结果各浓度重楼提取液对SW480细胞均有不同程度的抑制作用,且存在剂量-效应和时间-效应关系,重楼提取液作用后的SW480细胞出现变性、坏死的形态改变;经重楼提取液处理24h后,SW480细胞在S期的分布比例下降,G0-G1期和G2/M期细胞分布增多(P<0.05);同时SCF表达增高(P<0.05)。结论重楼提取液可能通过抑制肿瘤细胞的蛋白质与DNA合成,抑制肿瘤细胞的有丝分裂,进而抑制SW480细胞增殖,且其作用机制可能与SCF无关。
Objective To observe the effect of Paridis extract on proliferation of human colon cancer SW480 cells and investigate the mechanism of the effect.Methods SW480 cells cultured in vitro were treated with various concentrations(10,20,40 and 80 μg/ml)of Paridis extract for 12,24,36 and 48 h separately.The inhibiting rate to SW480 cells was determined by MTT method.The SW480 cells cultured in vitro were treated with Paridis extract at median inhibitory concentration for 24 h,then observed for morphology and analyzed the effect on cell cycle by flow cytometry.The transcription of stem cell factor(SCF)mRNA in SW480 cells was determined by RT-PCR,and expression of SCF protein by Western blot.Results The Paridis extract at various concentrations showed inhibitory effect at different degrees on SW480 cells,in dose-and time-dependent modes.Morphological change of SW480 cells,such as denaturation and necrosis,were observed after treatment.The percentage of SW480 cells at S phage decreased 24 h after treatment with Paridis extract,while those at G0-G1 and G2 /M phases increased(P < 0.05).Meanwhile,the expression level of SCF increased(P < 0.05).Conclusion Paridis extract might inhibit the mitosis of tumor cells by inhibiting the synthesis of protein and DNA,thereby inhibit the proliferation of SW480 cells,and the inhibitory effect might be SCF-independent.

关键词(KeyWords)： 重楼;结肠肿瘤;细胞增殖;干细胞因子
Paridis;Colon tumor;Cell proliferation;Stem cell factor

Abstract:

Keywords:

基金项目(Foundation): 重庆市科委自然科学基金(CSTC,2008CE9142);; 重庆医科大学创新基金(CX200528)

作者(Author): 李晞,王继红,肖亚雄
LI Xi,WANG Ji-hong,XIAO Ya-xiong(Laboratory of Biochemistry and Molecular Biology of Experimental Teaching Center

DOI: 10.13200/j.cjb.2010.06.64.lix.007

参考文献(References)：

• [1]谭艺兰,许雪亮.人参皂甙Rg3诱导视网膜母细胞瘤HXO.RB44细胞凋亡[J].眼科研究,2007,25(9):674-676.
• [2]叶爱芳,尹丽慧,倪吴花,等.白藜芦醇对人白血病细胞增殖和细胞周期的影响[J].实用医学杂志,2009,25(l2):1955-1956.
• [3]黄浩,王航,罗荣城.冬虫夏草提取液抑制结肠癌细胞增殖的研究[J].中药材,2007,30(3):310-313.
• [4]许则丰,范钰,丁佳逸.姜黄素对结肠癌SW-480细胞抗失巢凋亡的影响[J].中国中医基础医学杂志,2006,12(3):187-189.
• [5]黄谟婉,马英.姜黄素对HeLa细胞增殖和凋亡的影响[J].中国生物制品学杂志,2008,21(12):1094-1097.
• [6]王艳霞.重楼抗肿瘤活性成分的体外实验研究[J].包头医学,2008,32(4):196-197.
• [7]金炜东,陈孝平,蔡红娇.重楼提取物对HepG2细胞的毒性作用[J].华中科技大学学报(医学版),2006,35(1):103-106.
• [8]肖毅良.菊藻丸抗肿瘤临床应用240例[J].中国中西医结合外科杂志,1997,3(2):132.
• [9]Bellone G,Silverstri S,Artusio E,et al.Growth stimulation of col-orectal carcinoma cells via the C-KIT receptor is inhibited byTGF-beta1[J].J Cell Physiol,1997,172(1):l-l1.
• [10]Bellone G,Carbone A,Sibona N,et a1.Aberrant activation0f C-KIT protects colon carcinoma cells against apoptosis an d en-hances their invasive potential[J].Cancer Res,2001,61(5):2200-2206.
• [11]Reed J,Ouban A,Frank K,et a1.Immunohistochemical staining for c-kit(CD117)is a rare event in human colorectal carcinoma[J].Clin Colorectal Cancer,2002,2(2):119-122.
• [12]马千里,马华,陈曦,等.C-KIT蛋白在结直肠癌中的表达及临床意义[J].第三军医大学学报,2006,28(8):831-833.
• [13]周满红,杜文胜,龙胜双.重楼总皂苷对脂多糖诱导大鼠腹腔巨噬细胞分泌TNF-α及IL-1β的影响[J].四川中医,2008,26(3):14-16.
• [14]周满红,贺华经,潘勇,等.重楼总皂苷对多发骨折一脂多糖两次打击模型大鼠血清细胞因子水平的影响[J].中国骨伤,2008,21(9):662-664.
• [15]Ren XD,Hu B,Colletti L,et a1.Stem cell factor and its receptor,c-kit,are important for hepatocyte proliferation in wild-type and tumor necrosis factor receptor-1knockout mice after70%hepate-ctomy[J].Surgery,2008,143(6):790-802.
• [16]周继陶,龚日祥,陈向征,等.人结直肠癌干细胞微球体培养及鉴定[J].四川大学学报(医学版),2009,40(5):918-922.