黄连水提物对人皮肤肿瘤A431细胞凋亡、增殖的影响  

Effects of water extract of berberine on apoptosis and proliferation of human skin tumor A431 cells

作　　者：高耀星[1] 于建设[1] 都日娜 杨丽敏 赵鹏伟[3] 孙鹏[3] GAO Yaoxing;YU Jianshe;DU Rina;YANG Limin;ZHAO Pengwei;SUN Peng(Department of Anesthesiology, Affiliated Hospital of Inner Mongolia Medical University, Inner Mongolia Autonomous Region, Hohhot 010010,China;Department of Dermatology, International Mongolian Medical Hospital of Inner Mongolia Autonomous Region, Inner Mongolia Autonomous Region, Hohhot 010010,China;Inner Mongolia Medical University, Inner Mongolia Autonomous Region, Hohhot 010010,China)

机构地区：[1]内蒙古医科大学附属医院麻醉科,内蒙古呼和浩特010010 [2]内蒙古自治区国际蒙医医院皮肤科,内蒙古呼和浩特010010 [3]内蒙古医科大学,内蒙古呼和浩特010010

出　　处：《中国医药导报》2019年第16期21-24,49,F0004,共6页China Medical Herald

基　　金：国家知识产权局专利受理号:21510574607.5;内蒙古自治区自然科学基金项目(2014MS0366)

摘　　要：目的探讨黄连水提物对人皮肤肿瘤A431细胞凋亡、增殖的影响。方法采用低温提取工艺,从药材中提取出水溶性的小分子混合物,用紫外色谱分析仪检测混合物成分;采用MTT法检测药物对人皮肤肿瘤A431细胞的抑制作用,药物设4个浓度,即6.3、12.5、25、50μg/μL,并将其作为药物组,未加药物的为对照组,观察24、48、72 h抑瘤率;然后用Tunel检测细胞凋亡现象;通过MTT筛选后,用25μg/μL黄连水提物作用肿瘤A431细胞48 h后,用Western blot检测Caspase-3、Caspase-8、Caspase-9、Bcl-2凋亡途径的影响。结果紫外光谱分析黄连水提物主要成分为核酸、生物碱等,MTT显示25μg/μL黄连水提取物在48 h对A431细胞的抑制率已达到最佳(P <0.05)。显微镜下,对照组无变化。25μg/μL黄连水提物孵育48 h后,细胞明显的数量减少、变圆、漂浮;Tunel显示25μg/μL黄连水提物核定位处绿光明显增多,提示细胞大量凋亡。本研究检测显示,25μg/μL黄连水提取物孵育48 h后,肿瘤A-431细胞中Caspase-3、Caspase-8、Caspase-9表达量显著升高,而Bcl-2的表达量显著降低。结论本研究初步表明黄连水提小分子混合物可促进A-431细胞凋亡,抑制细胞增殖。其机制包括细胞外凋亡通路和细胞内凋亡通路,导致肿瘤细胞中Caspase-3、Caspase-8、Caspase-9上调及抑制凋亡蛋白Bcl-2的下调。这进一步明确了黄连水提物在治疗皮肤肿瘤中的作用机制,为皮肤肿瘤的治疗提供的方法。Objective To investigate the effect of water extract of berberine on apoptosis and proliferation of human skin tumor A431 cells. Methods The low temperature extraction process was used to extract the water-soluble small molecule mixture from the medicinal materials. MTT assay was used to detect the inhibitory effect of the drug on human skin tumor A431 cells. Four concentrations of 6.3, 12.5, 25, 50 μg/μL were set as the drug group, and those without the drug were set as the control group. The tumor inhibition rates at 24, 48 and 72 h were observed. Tunel was used to detect apoptosis. After being screened by MTT, the apoptosis pathways of caspase-3, caspase-8, caspase-9 and Bcl-2 were detected by Western blot after treating tumor A431 cells with 25 μg/μL water extract for 48 h. Results The main components were nucleic acid, alkaloid, etc. MTT showed that the inhibition rate of 25 μg/μL water extract of berberine on A431 cells reached the best at 48 h (P < 0.05). Under the microscope, there was no change in the control group. After incubated with 25 μg/μL water extract of berberine for 48 h, the number of cells significantly decreased, became round and floated. Tunel showed 25 μg/μL water extract of berberine significantly increased green light at the approved location, indicating a large number of apoptosis. In this study, after incubated with 25 μg/μL water extract of berberine for 48 h, and the expressions of caspase-3, caspase-8 and caspase-9 in tumor A431 cells were significantly increased, while the expression of Bcl-2 was significantly decreased. Conclusion This study preliminarily showed that the small molecule mixture of water extract from rhizoma coptiliae could promote apoptosis of A431 cells and inhibit cell proliferation. The mechanisms include the extracellular apoptosis pathway and the intracellular apoptosis pathway, leading to the up-regulation of caspase-3, caspase-8 and caspase-9 in tumor cells and the down-regulation of the apoptotic protein Bcl-2. This study further clarified the m

关 键 词：黄连水提物 皮肤肿瘤 A431细胞 细胞凋亡 BCL-2 CASPASE-3 CASPASE-8 CASPASE-9 

分 类 号：R739.5[医药卫生—肿瘤]