本文選題：高三尖杉酯堿 + K562細胞�。� 參考：《南京醫(yī)科大學》2014年碩士論文

【摘要】：目的觀察經高三尖杉酯堿(Homoharringtonine, HHT)處理后的不同時間點K562細胞凋亡率的變化。方法分別采用MTT、流式細胞術和western blot等方法檢測HHT處理后的不同時間點K562細胞的增殖活力、凋亡率及Bcl-2蛋白和Caspase-3蛋白的表達。結果①HHT (終濃度為lOng/ml)作用后, K562細胞增殖活力逐漸下降,但作用后期細胞增殖活力又逐漸升高,第1天到第8天的細胞增殖活力分別為(93.6±1.2)%、(81.3±0.9)%、(69.8+0.7)%、(57.2±2.1)%、(43.8+1.1)%、(54.1+1.4)%、(64.5±1.3)%和(70.6+1.8)%。與之相對應的是K562細胞凋亡率在HHT作用早期逐步上升而在作用后期逐漸下降。②HHT處理后,K562細胞的激活型caspase-3蛋白(17KD及19KD)表達從第1天至第7天逐漸增高,第8天開始下降(p0.05)。③HHT處理后1-8天K562細胞Bcl-2陽性表達呈現先低后高的特點：前7天逐漸下降,第8天開始上升,差異有統(tǒng)計學意義(p0.05)結論在體外處理K562細胞的過程中,HHT的細胞凋亡誘導作用在體外培養(yǎng)早期較強,但在作用后期凋亡誘導作用明顯減弱。目的 觀察單用高三尖杉酯堿(Homoharring-tonine,HHT)及HHT聯(lián)合自噬抑制劑3-MA對K562細胞增殖、凋亡以及自噬的影響。方法10ng/ml的HHT及1.5mmol/L的3-MA作用K562細胞后,檢測細胞Beclinl基因表達(RT-PCR法)、LC3 Ⅱ/Ⅰ及caspase-3凋亡蛋白表達(Western blot印跡法)和細胞自噬(透射電子顯微鏡)。結果 ①HHT處理后,K562細胞的Beclin 1基因和LC3 Ⅱ/Ⅰ表達水平在作用的早期(分別為d1-d5和d1-d6)逐步下降,但HHT作用后期(分別為d6-d8和d7-d8)兩者的表達水平開始上升；②caspase-3蛋白表達水平在HHT作用早期(d1-d7)逐步升高,d8開始下降。③HHT聯(lián)用3-MA組的Beclin 1基因及LC3II/I表達水平在d1-d8逐漸下降,無后期升高現象發(fā)生。④HHT聯(lián)用3-MA組的caspas e-3蛋白表達水平在d1--d8內持續(xù)逐漸升高。⑤電鏡下,HHT作用的第8天K562細胞自噬體顯著增多,而HHT聯(lián)合3-MA作用第8天無自噬體出現。結論HHT可誘導K562細胞凋亡,但HHT持續(xù)后,K562細胞可發(fā)生自噬而使HHT的細胞凋亡誘導作用減弱；聯(lián)用自噬抑制劑可增強HHT的細胞凋亡誘導作用。
[Abstract]:Objective to observe the apoptosis rate of K562 cells treated with homoharringtonine (HHT) at different time points. Methods MTT, flow cytometry and western blot were used to detect the proliferation activity, apoptosis rate and the expression of Bcl-2 protein and Caspase-3 protein in K562 cells treated with HHT at different time points. Results the proliferative activity of K562 cells decreased gradually after treatment with 1HHT (final concentration was lOng / ml), but the proliferative activity of K562 cells increased gradually in the late stage of treatment. The proliferative activities of K562 cells were 93.6 鹵1.29.80.73 鹵0.99.70.78 from day 1 to day 8, respectively. The proliferative activity of K562 cells was 54.2 鹵2.11g / ml and 64.5 鹵1.3g% and 70.6 / 1.8U / d, respectively. In contrast, the apoptotic rate of K562 cells increased gradually at the early stage of HHT treatment, but decreased gradually at the late stage of treatment. The expression of activated caspase-3 protein (17KD and 19KD) in K562 cells increased gradually from day 1 to day 7 after treatment with .2HHT. On the 8th day, the expression of Bcl-2 in K562 cells decreased at first and then increased after treatment with HHT. The expression of Bcl-2 decreased gradually at the first 7 days, and then increased on the 8th day. Conclusion the apoptosis induction of HHT in K562 cells was stronger at the early stage of culture in vitro, but the apoptotic effect was significantly weakened in the late stage of treatment of K562 cells. Objective to observe the effects of homoharringtonine (HHT) and HHT combined with autophagy inhibitor 3-MA on the proliferation, apoptosis and autophagy of K562 cells. Methods after K562 cells were treated with HHT of 10ng/ml and 3-MA of 1.5mmol/L, the expression of Beclinl gene was detected by RT-PCR and the expression of caspase-3 apoptosis protein was detected by Western blot and autophagy (transmission electron microscope). Results after 1HHT treatment, the expression of Beclin 1 gene and LC3 鈪，

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