摘要
背景:温热中药半夏拥有可人工广泛种植、易提纯等优势,其提取物可抑制慢性髓系白血病、急性髓系白血病、急性T淋巴白血病细胞增殖,但是作用机制尚不明确。目的:探讨中药半夏提取物对3种白血病细胞凋亡的作用机制。方法:慢性髓系白血病细胞株(K562)、急性髓系白血病M3细胞株(HL-60)、急性T淋巴白血病细胞株(C8166)在半夏提取物5种浓度梯度中分别作用24,48,72 h,采用CCK-8法评估3种白血病细胞增殖情况并从中筛选出有明显抑制作用的中(300 mg/L)、高(500 mg/L)质量浓度。然后进一步采用流式细胞术检测3种白血病细胞早期凋亡发生率,采用RT-PCR与Western blot检测Bax、Bcl-2、Caspase-3 mRNA及蛋白水平。结果与结论:①5种质量浓度的半夏提取物作用3种白血病细胞具有细胞增殖抑制作用,中(300 mg/L)、高(500 mg/L)质量浓度半夏提取物增殖抑制率较高,与低质量浓度组(100 mg/L)、对照组比较差异有显著性意义(P<0.01);②中、高质量浓度半夏提取物可诱导3种白血病细胞早期凋亡;③中、高质量浓度半夏提取物可以下调3种白血病细胞的Bcl-2 mRNA表达,上调Bax及Caspase-3 mRNA表达(P<0.05;P<0.01);④中、高质量浓度半夏提取物作用72 h后,K562、C8166细胞的Bcl-2蛋白条带明显减弱,而HL-60细胞无减弱变化;Bax及Caspase-3蛋白条带明显增强;⑤结果表明,半夏提取物能有效抑制髓系白血病、T淋巴细胞白血病细胞的增殖,促进其凋亡,其作用机制可能与其调节Bax/Bcl-2、Caspase-3蛋白表达有关。
BACKGROUND:Pinellia ternata can be cultivated artificially and is easy to purify.Its extract can inhibit the cell proliferation of chronic myeloid leukemia,acute myelogenous leukemia M3,and T-cell acute lymphoblastic leukemia,but the underlying mechanism remains unclear.OBJECTIVE:To explore the mechanism of action of pinellia ternata extract on apoptosis of three kinds of leukemia cells.METHODS:Chronic myeloid leukemia cell lines(K562),acute myeloid leukemia–M3 cell line(HL-60),acute T lymphocyte leukemia cell line(C8166)were incubated in five kinds of concentration in pinellia ternata extracts in vitro for 24,48,and 72 hours,respectively.Cell counting kit-8 assay was used to assess the influence of three kinds of leukemia cell proliferation to screen the optimal inhibitory effect of moderate(300 mg/L)and high(500 mg/L)concentration.Flow cytometry was used to determine the incidence of early apoptosis of three kinds of leukemia cells.The mRNA and protein expression levels of Bax,Bcl-2 and Caspase-3 were detected by RT-PCR and western blot assay.RESULTS AND CONCLUSION:Cell counting kit-8 assay results showed that five different concentrations of pinellia extracts could inhibit the proliferation of three kinds of leukemia cells.The inhibition rates of medium concentration(300 mg/L)and high concentration(500 mg/L)were significantly higher than those of low concentration(100 mg/L)and control group(P<0.01).Flow cytometry(Annexin PE/7AAD and Annexin V/PI)double staining showed that moderate and high concentrations of pinellia extract could induce early apoptosis of three kinds of leukemia cells.RT-PCR results revealed moderate and high concentrations of pinellia extract could down-regulate the mRNA expression level of Bcl-2,and up-regulate the mRNA expression levels of Bax,Bcl-2,Caspase-3(P<0.05,P<0.01).Western blot assay results showed that the Bcl-2 protein bands of K562 and C8166 cells were significantly weakened after 72 hours of incubation with moderate and high concentrations of pinellia tubulin extract,while n
作者
冯嘉昆
刘伟
李正发
赵晓晨
李增政
杨同华
赵仁彬
胡芃
裴强
关心
Feng Jiakun;Liu Wei;Li Zhengfa;Zhao Xiaochen;Li Zengzheng;Yang Tonghua;Zhao Renbin;Hu Peng;Pei Qiang;Guan Xin(School of Medicine,Kunming University of Science and Technology,Kunming 650504,Yunnan Province,China;the First People’s Hospital of Yunnan Province(Affiliated Hospital of Kunming University of Science and Technology),Kunming 650100,Yunnan Province,China;Yunnan University of Traditional Chinese Medicine,Kunming 650032,Yunnan Province,China)
出处
《中国组织工程研究》
CAS
北大核心
2020年第31期5023-5029,共7页
Chinese Journal of Tissue Engineering Research
基金
云南省应用基础研究项目(2017EF468(-242)),项目负责人:李正发
云南省临床医学中心开放课题(2019LCZXKF-XY01),项目负责人:李正发。
