文章摘要
选择性MAO-B抑制剂TM-5对胶质瘤U87细胞自噬和凋亡的影响
Effects of selective MAO-B inhibitor TM-5 on autophagy and apoptosis of glioma U87 cells
投稿时间:2022-11-21  
DOI:10.3969/j.issn.1000-0399.2023.10.004
中文关键词: TM-5  选择性MAO-B抑制剂  胶质瘤  细胞增殖  细胞凋亡  自噬
英文关键词: TM-5  Selective MAO-B inhibitor  Glioma  Cell proliferation  Apoptosis  Autophagy
基金项目:河南省医学科技攻关计划联合共建项目(编号:LHGJ20191452);河南省科技厅科技攻关项目(编号:202102310408)
作者单位E-mail
武海博 473000 河南南阳 南阳市中心医院神经内科  
梁燕 473000 河南南阳 南阳市中心医院神经内科  
沈雷 473000 河南南阳 南阳市中心医院神经内科  
范展 473000 河南南阳 南阳市中心医院神经内科  
傅国惠 473000 河南南阳 南阳市中心医院神经内科 fgh377818@sina.com 
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中文摘要:
      目的 探讨选择性单胺氧化酶B (MAO-B)抑制剂TM-5对胶质瘤U87细胞体外自噬及凋亡的影响。方法 正常培养胶质瘤细胞株U87,随机分为6组:对照组不做处理,5、10、25、50、100μmoL/L TM-5组分别予以5、10、25、50、100μmoL/L TM-5处理细胞,每组设3个复孔。采用噻唑蓝(MTT)法检测细胞活力;流式细胞术检测细胞凋亡;实时荧光定量聚合酶链反应(qRT-PCR)检测促凋亡蛋白Bcl-2相关蛋白X (Bax)、抑凋亡蛋白B淋巴细胞瘤-2(Bcl-2)的基因表达;蛋白质免疫印迹(WB)检测细胞凋亡相关蛋白Bax和切割后半胱氨酸蛋白酶3(cleaved caspase-3)以及自噬相关标志蛋白Beclin-1、自噬相关蛋白1轻链-Ⅱ(LC3-Ⅱ)和P62表达。结果 与对照组比较,10、25、50、100μmoL/L TM-5组U87细胞存活率降低(P<0.05),选取5、10、25μmoL/L TM-5组(细胞存活率>50%)进行后续实验。与对照组比较,10、25μmoL/L TM-5组相对细胞凋亡率、细胞中LC3-Ⅱ和Bax蛋白相对表达量升高,差异有统计学意义(P<0.05);5、10、25μmoL/L TM-5组U87细胞中促凋亡基因Bax mRNA相对表达水平、Beclin-1和cleaved caspase-3蛋白相对表达量升高,抑凋亡基因Bcl-2 mRNA、P62蛋白相对表达量降低,差异有统计学意义(P<0.05)。结论 TM-5可抑制胶质瘤U87细胞存活,可上调Beclin-1、LC3-Ⅱ、Bax和cleaved caspase-3,下调Bcl-2和P62表达,诱导细胞自噬和凋亡。
英文摘要:
      Objective To explore the effects of selective monoamine oxidase B(MAO-B) inhibitor TM-5 on autophagy and apoptosis of glioma U87 cells. Methods The normally cultured glioma cells U87 were randomly divided into control group(no treatment), 5, 10, 25, 50and 100 μmoL/L TM-5 groups(5, 10, 25, 50 and 100 μmoL/L of TM-5), with 3 experimental pores in each group. The cells activity was detected by MTT. Apoptosis was detected by flow cytometry. The expressions of pro-apoptosis protein Bax and anti-apoptosis protein Bcl-2genes were detected by real-time fluorescent quantitative polymerase chain reaction(qRT-PCR). The expressions of apoptosis-relatedproteins(Bax, cleaved caspase-3) and autophagy related markers(Beclin-1, LC3-II, P62) were detected by Western blot(WB). Results Compared with the control group, the survival rate of U87 cells decreased in 10, 25, 50 and 100 μmoL/L TM-5 groups(P<0.05). The follow-up experiments were performed under certain concentration(5, 10 and 25 μmoL/L, cells survival rate >50%). Compared with the control group, relative apoptosis rate of U87 cells and relative expression levels of LC3-Ⅱand Bax proteins increased in 10 and 25 μmoL/L TM-5 groups(P<0.05).Compared with control group, relative expression levels of pro-apoptosis gene Bax mRNA, Beclin-1 and cleaved caspase-3 proteins increased, while relative expression levels of anti-apoptosis gene Bcl-2mRNA and P62 protein decreased in 5, 10 and 25 μmoL/L TM-5 groups(P<0.05).Conclusion TM-5 can inhibit the survival of glioma U87 cells, up-regulate Beclin-1, LC3-Ⅱ, Bax and cleaved caspase-3, down-regulate Bcl-2 and P62, and induce autophagy and apoptosis.
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