α-苦瓜素经LRP1受体介导的JNK信号通路诱导肝细胞L02早期凋亡的机制研究
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篇名: α-苦瓜素经LRP1受体介导的JNK信号通路诱导肝细胞L02早期凋亡的机制研究
TITLE:
摘要: 目的:探讨α-苦瓜素诱导肝细胞L02(简称“L02细胞”)早期凋亡的信号通路。方法:制备并纯化α-苦瓜素。采用流式细胞术检测0(阴性对照)、160、80 μg/mL α-苦瓜素作用L02细胞2~8 h后的细胞凋亡率。小干扰RNA(siRNA)沉默低密度脂蛋白受体相关蛋白1(LRP1)得到LRP1-siRNA细胞,然后采用液相芯片分析术检测α-苦瓜素(160 μg/mL)分别作用L02细胞(正常组)和LRP1-siRNA细胞(沉默组)0、0.25、0.5、1、2 h后c-Jun氨基末端激酶(JNK)信号通路中促调蛋白磷酸化JNK(p-JNK)、磷酸化凋亡前体蛋白(p-Bad)、胱天蛋白酶9(Caspase-9)及抑调蛋白磷酸化蛋白53(p-p53)、磷酸化蛋白激酶B(p-Akt)、磷酸化B淋巴细胞瘤2(p-Bcl-2)、Caspase-8的表达情况,分析α-苦瓜素对L02细胞JNK信号通路的作用。结果:制备并纯化得到α-苦瓜素(纯度>97%);与阴性对照组比较,160 μg/mL α-苦瓜素作用8 h时,可显著诱导细胞早期凋亡(P<0.05),80 μg/mL α-苦瓜素诱导的早期凋亡不明显(P>0.05);与0 h比较,作用后0.25 h促调蛋白p-JNK、p-Bad和Caspase-9表达量显著增加(P<0.05),而抑调蛋白p-p53、p-Akt、p-Bcl-2和Caspase-8表达量无变化;与正常组相比,沉默组各时间点的p-JNK、p-Bad和Caspase-9的表达均显著减少(P<0.05)。结论:α-苦瓜素可能通过LRP1受体介导JNK信号通路诱导肝细胞凋亡,为揭示其肝毒性提供了参考。
ABSTRACT: OBJECTIVE: To explore the signaling pathway of α-momordicin inducing early apoptosis of hepatocyte L02 (called “the L02 cell” for short). METHODS: α-momordicin was prepared and purified. Flow cytometry was used to detect the apoptotic rate of the L02 cell after treated with 0 (negative control), 160 and 80 μg/mL of α-momordicin for 2-8 h. LRP1-siRNA cells were obtained by small interfering RNA (siRNA) silencing low density lipoprotein receptor-related protein 1 (LRP1). The expression of proapoptotic protein p-JNK, p-Bad, Caspase-9, inhibitor of apoptosis protein p-p53, p-Akt,p-Bcl-2 and active Caspase-8 in the L02 cell (normal group) and LRP1-siRNA (silence group) after treated with α-momordicin for 0, 0.25, 0.5, 1, 2 h were determined by liquid biochip analysis in c-Jun N-terminal kinase (JNK) signaling pathway. Effects of α-momordicin on JNK signaling pathway were analyzed. RESULTS: α-momordicin could be prepared and purified (purity>97%). Compared with negative control group, 160 μg/mL α-momordicin could significantly induced early apoptosis of the cell after treated for 8 h (P<0.05); early apoptosis of the cell induced by 80 μg/mL α-momordicin was not obvious (P>0.05). Compared with 0 h, the expression of p-JNK, p-Bad and Caspase-9 were increased significantly and even reached the peak value 0.25 h after medication (P<0.05), while the expression of p-p53, p-Akt, p-Bcl-2 and Caspase-8 had no change. Compared with normal group, the expression of p-JNK, p-Bad and Caspase-9 of silence group were decreased significantly at different time points (P<0.05). CONCLUSIONS: α-momordicin can induce the apoptosis of hepatocytes via LRP1 receptor mediated JNK signaling pathway, and will provide the reference for its hepatotoxicity.
期刊: 2018年第29卷第23期
作者: 邓念华,沈富兵,郑崛村,沈岱,王玲
AUTHORS: DENG Nianhua,SHEN Fubing,ZHENG Juecun,SHEN Dai,WANG Ling
关键字: α-苦瓜素;凋亡;肝细胞L02;c-Jun氨基末端激酶信号通路;低密度脂蛋白受体相关蛋白1
KEYWORDS: α-momordicin; Apoptosis; Hepatocyte L02; c-Jun N-terminal kinase signaling pathway; Low density lipoprotein receptor-related protein 1
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