川楝素通过mTORC1信号通路调控自噬增强阿糖胞苷抗急性髓系白血病的作用及机制 点击下载
| 论文标题： | 川楝素通过mTORC1信号通路调控自噬增强阿糖胞苷抗急性髓系白血病的作用及机制 |
| 英文标题： | |
| 中文摘要： | 目的 基于哺乳动物雷帕霉素靶蛋白复合物1（mTORC1）信号通路探讨川楝素增强阿糖胞苷抗急性髓系白血病（AML）的作用及潜在机制。方法以雌性NSG免疫缺陷小鼠为对象，以单次γ射线照射+单次尾静脉注射U937细胞悬液的方式构建AML模型，探讨阿糖胞苷（2.5mg/kg）、川楝素（1.0mg/kg）、阿糖胞苷+川楝素（2.5mg/kg+1.0mg/kg）对小鼠体重、生存期、外周血常规指标、组织病理形态学及自噬相关蛋白表达的影响；以U937细胞为对象，探讨阿糖胞苷（1μmol/L）、川楝素（160nmol/L）、阿糖胞苷+川楝素（1μmol/L+160nmol/L）对细胞中自噬相关蛋白、mTORC1信号通路相关蛋白及mRNA表达的影响；验证川楝素与选择性自噬接头蛋白1（又称p62）的靶向关系，并进行基因本体（GO）、京都基因和基因组数据库（KEGG）富集分析和分子对接；构建p62敲减的U937稳转株，验证川楝素增敏作用的潜在机制。结果动物实验证实，川楝素联合阿糖胞苷可显著改善AML小鼠的体重、生存期和外周血常规指标（P＜0.05），改善其骨髓、肝脏、脾脏组织白血病细胞浸润、局灶性坏死等病理改变，并有回调自噬相关蛋白表达的作用。细胞实验证实，川楝素能逆转阿糖胞苷引起的自噬流激活（表现为微管相关蛋白1轻链3B、Beclin-1表达下调，p62蛋白表达上调），并可通过激活mTORC1信号通路，进而抑制下游Unc-51样自噬激活激酶1（ULK1）复合物（包括ULK1、2，自噬相关蛋白13，黏着斑激酶家族相互作用蛋白200kDa）的表达（P＜0.05）。川楝素的增敏作用与抑制自噬、调控mTORC1信号通路有关；其可通过疏水作用与p62蛋白稳定结合。机制验证研究表明，敲减p62后，川楝素无法再激活mTORC1信号通路或抑制自噬。结论川楝素可能通过直接靶向p62蛋白，激活mTORC1信号通路，进而抑制阿糖胞苷诱导的AML细胞保护性自噬，从而增强该细胞对阿糖胞苷的化疗敏感性。 |
| 英文摘要： | OBJECTIVE To investigate the effect and potential mechanism of toosendanin in enhancing the antileukemic activity of cytosine arabinoside against acute myeloid leukemia （AML） based on the mammalian target of rapamycin complex 1 （mTORC1） signaling pathway.METHODS Female NSG immunodeficient mice were used to establish an AML model via a single γ-ray irradiation followed by a single tail-vein injection of a U937 cell suspension. The effects of cytosine arabinoside （2.5 mg/kg）， toosendanin （1.0 mg/kg）， and cytosine arabinoside+toosendanin （2.5 mg/kg+1.0 mg/kg） on body weight， survival time， peripheral blood routine indicators， histopathological morphology， and the expressions of autophagy-related proteins were explored. Using U937 cells as the subject， the effects of cytosine arabinoside （1 μmol/L）， toosendanin （160 nmol/L）， and cytosine arabinoside+toosendanin （1 μmol/L+160 nmol/L） on the expressions of autophagy-related proteins， mTORC1 signaling pathway-related proteins and mRNAs were investigated. The targeting relationship between toosendanin and the selective autophagy ligand 1 （also known as p62） was validated， and the Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analyses， as well as molecular docking， were performed. A p62 -knockdown U937 stable cell line was constructed， and the underlying mechanism of toosendanin-induced sensitization was verified.RESULTS Animal experiments confirmed that the combination of toosendanin and cytosine arabinoside significantly improved body weight， survival time and peripheral blood routine indicators in AML mice （ P ＜0.05）， alleviated pathological changes such as leukemic cell infiltration and focal necrosis in bone marrow， liver， and spleen tissues， and modulated the expressions of autophagy-related proteins. Cell experiments confirmed that toosendanin could reverse cytosine arabinoside-induced activation of the autophagy cascade （manifested by downregulation of the protein expressions of microtubule-associated protein 1 light chain 3B and Beclin-1， upregulation the protein expression of p62）， and might inhibit the expression of the downstream Unc-51-like autophagy activating kinase 1 （ULK1） complex （including ULK1， ULK2， autophagy-related protein 13， and FAK family interacting protein of 200 kDa） by activating the mTORC1 signaling pathway （ P ＜0.05）. The sensitizing effect of toosendanin was associated with the inhibition of autophagy and the regulation of the mTORC1 signaling pathway； it could stably bind to p62 via hydrophobic interactions. Mechanism verification experiments indicated that upon p62 knockdown， toosendanin was no longer able to activate the mTORC1 signaling pathway or inhibit autophagy.CONCLUSIONS Toosendanin may enhance chemo-sensitivity of AML cells to cytosine arabinoside by directly targeting the p62 protein， which subsequently activates the mTORC1 signaling pathway， and thereby suppresses cytosine arabinoside-induced protective autophagy. |
| 期刊： | 2026年第37卷第17期 |
| 作者： | 欧阳贤凤；康建华；郭瑛；黄燕；江炜；晏建国 |
| 英文作者： | OUYANG Xianfeng，KANG Jianhua，GUO Ying，HUANG Yan，JIANG Wei，YAN Jianguo |
| 关键字： | 川楝素；阿糖胞苷；急性髓系白血病；保护性自噬；p62；mTORC1信号通路 |
| KEYWORDS： | cytosine arabinoside;acute myeloid leukaemia;protective autophagy;p62;mTORC1 signaling pathway |
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