首页|CH25H调控细胞自噬和凋亡对喉鳞状细胞癌进展影响

CH25H调控细胞自噬和凋亡对喉鳞状细胞癌进展影响

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目的 探讨胆固醇25-羟化酶(CH25H)通过调控细胞自噬和凋亡对喉鳞状细胞癌(LSCC)进展的影响.方法 通过转染CH25H过表达质粒,构建CH25H稳定过表达的人LSCC细胞系AMC-HN-8细胞(AMC-HN-8/CH25H),转染空白质粒为转染对照(AMC-HN-8/Mock),同时设置未转染空白对照(AMC-HN-8).采用实时荧光定量PCR检测各组细胞CH25H mRNA表达水平,Western bolt检测CH25H、凋亡相关蛋白[B淋巴细胞瘤-2(Bcl-2)、Bcl-2关联X蛋白(Bax)、裂解的半胱氨酸蛋白酶-3(C-caspase-3)]、自噬相关蛋白[微管相关蛋白1A/1B-轻链3(LC3)Ⅱ、Beclin-1、p62]以及磷脂酰肌醇-3-激酶(PI3K)/蛋白激酶B(Akt)通路蛋白[PI3K、磷酸化PI3K(p-PI3K)、Akt、磷酸化Akt(p-Akt)]的表达水平.流式细胞术检测各组细胞凋亡率,免疫荧光检测各组细胞LC3 Ⅱ表达.构建裸鼠LSCC移植瘤动物模型,采用随机数字表法将裸鼠分为AMC-HN-8组、AMC-HN-8/Mock组、AMC-HN-8/CH25H组,每组8只,分别接种对应细胞,检测CH25H对LSCC细胞生长以及凋亡相关蛋白和自噬相关蛋白的影响.结果 相比AMC-HN-8和AMC-HN-8/Mock组细胞,AMC-HN-8/CH25H组细胞CH25H表达上调(P<0.05),凋亡率显著增加(P<0.05),CH25H表达促进了 Bax和C-caspase-3表达,抑制了 Bcl-2的表达.同时CH25H表达促进了 LC3 Ⅱ和Beclin-1的表达,抑制了 p62的表达,表明过表达CH25H可促进LSCC细胞的凋亡和自噬.此外,CH25H抑制了 PI3K/Akt致癌信号通路的激活.在移植瘤裸鼠模型中,过表达CH25H可抑制体内LSCC肿瘤生长.结论 CH25H在LSCC细胞中通过PI3K/Akt信号通路调控细胞自噬和凋亡进而影响肿瘤进展.
CH25H affects laryngeal squamous carcinoma progression through regulating autophagy and apoptosis
Objective To investigate the effect of cholesterol 25-hydroxylase(CH25H)on the progression of laryngeal squamous cell carcinoma(LSCC)and its relation with autophagy and apoptosis of tumor cells.Methods Human LSCC AMC-HN-8 cells were tansfected with CH25-plasmid or blank plasmid to construct AMC-HN-8/CH25H cell line and transfection control cell line AMC-HN-8/Mock,and untransfected AMC-HN-8 cells served as blank control.The expression level of CH25H mRNA was detected by real-time fluorescence quantitative PCR.The expression levels of CH25H protein,apoptosis-related proteins(Bcl-2,Bax and C-caspase-3),autophagy-related proteins(LC3 Ⅱ,Beclin-1 and p62),and PI3K/Akt pathway proteins(p-PI3K,PI3K,p-Akt,and Akt)were detected by Western blot.Flow cytometry was used to detect the cell apoptosis rate,and immunofluorescence was used to detect the expression of LC3 Ⅱ.The nude mice were transplanted with AMC-HN-8,AMC-HN-8/Mock,and AMC-HN-8/CH25H cells(n=8 in each group),respectively.The tumor volume was measured,and the apoptosis protein and autophagy protein levels were detected in each group.Results Compared with AMC-HN-8 and AMC-HN-8/Mock cells,the expression of CH25H in AMC-HN-8/CH25H cells was upregulated(P<0.05),the apoptosis rate was significantly increased(P<0.05);the expression of Bax and C-caspase-3 was enhanced,and Bcl-2 expression was inhibited.Meanwhile,the expression of LC3 Ⅱ and Beclin-1 was increased,and the expression of p62 was inhibited,indicating that overexpression of CH25H promoted apoptosis and autophagy in laryngeal squamous cell carcinoma cells.Furthermore,CH25H inhibited the activation of the PI3K/Akt oncogenic signaling pathway.Overexpression of CH25H inhibited LSCC tumor growth in xenografted nude mice.Conclusion CH25H regulates autophagy and apoptosis through the PI3K/Akt signaling pathway in laryngeal squamous cell carcinoma cells,thereby affecting tumor progression.

Cholesterol 25-hydroxylaseLaryngeal squamous cell carcinomaAutophagyApoptosisPhospha-tidylinositol 3-kinase/protein kinase B

熊华才、许雨欣、邬振华、胡丹飞、李群、项振飞

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315000 宁波市医疗中心李惠利医院放疗科

宁波大学医学部

315000 宁波市医疗中心李惠利医院耳鼻喉科

胆固醇25-羟化酶 喉鳞癌 自噬 凋亡 磷脂酰肌醇-3-激酶/蛋白激酶B

浙江省医药卫生科技计划宁波市自然科学基金

2022KY2952021J289

2024

浙江医学
浙江省医学会

浙江医学

CSTPCD
影响因子:0.428
ISSN:1006-2785
年,卷(期):2024.46(8)
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