小檗碱对人胶质瘤T98G细胞迁移和侵袭的抑制作用及其机制

doi:10.13481/j.1671-587X.20240107

摘要/Abstract

摘要：

目的探讨小檗碱（BBR）对人胶质瘤T98G细胞脂肪酸的调控作用及细胞增殖、迁移和侵袭的影响，阐明其潜在的作用机制。方法对数生长期T98G细胞分为对照组和不同浓度（25、50及100 mg·L^－1）BBR组，采用细胞划痕愈合实验检测各组细胞迁移率，Transwell小室实验检测各组细胞侵袭率。对数生长期T98G细胞分为对照组和100 mg·L^－1 BBR组，质谱法检测2组细胞中脂肪酸含量。对数生长期T98G细胞分为对照组和不同浓度（50、100及150 mg·L^－1）BBR组，采用Western blotting法检测各组细胞中磷脂酰肌醇3-激酶（PI3K）、磷酸化PI3K（p-PI3K）、蛋白激酶B（AKT）、磷酸化AKT（p-AKT）、固醇调节元件结合蛋白1（SREBP-1）和脂肪酸合成酶（FASN）蛋白表达水平。应用基因沉默技术抑制FASN表达，将对数生长期T98G细胞分为对照组、shFASN1组和shFASN2组，采用Western blotting法检测各组细胞中FASN蛋白表达水平，克隆形成实验检测各组细胞克隆形成情况，细胞划痕愈合实验检测各组细胞迁移率。结果与对照组比较，不同浓度BBR组细胞迁移率和侵袭率呈浓度依赖性降低（P<0.01），100 mg·L^－1 BBR组细胞中各类脂肪酸含量明显降低（P<0.01）。与对照组比较，150 mg·L^－1 BBR组细胞中p-PI3K、p-AKT、SREBP-1和FASN蛋白表达水平均明显降低（P<0.05或P<0.01），100和150 mg·L^－1 BBR组细胞中SREBP-1蛋白表达水平明显降低（P<0.01）。抑制FASN表达后，与对照组比较，shFASN1组和shFASN2组细胞中FASN蛋白表达水平均明显降低（P<0.01），且shFASN2组细胞中FASN蛋白表达水平低于shFASN1组（P<0.05）；与对照组比较，shFASN1组和shFASN2组细胞中克隆形成数明显减少，细胞迁移率明显降低（P<0.01），且shFASN2组细胞迁移率明显低于shFASN1组（P<0.05）。结论 BBR通过降低细胞中PI3K/AKT/SREBP-1/FASN通路相关蛋白的表达干扰胶质瘤T98G细胞中脂肪酸合成，进而降低其迁移和侵袭能力。

关键词: 小檗碱, 胶质瘤, 脂肪酸, 细胞迁移, 细胞侵袭

Abstract:

Objective To discuss the regulatory effect of berberine （BBR） on fatty acids in the human glioma T98G cells and its effect on the cell proliferation， migration， and invasion， and to clarify its potential mechanism. Methods The T98G cells at logarithmic growth phase were divided into control group and different concentrations （25， 50， and 100 mg·L^－1） of BBR groups. Cell wound healing assay was used to detect the migration rates of the cells in various groups； Transwell chamber assay was used to detect the invasion rates of the cells in various groups.The T98G cells at logarithmic growth phase were divided into control group and 100 mg·L^－1BBR group，and Mass spectrometry was used to detect the fatty acid contents in the cells in two groups. The T98G cells at logarithmic growth phase were divided into control group and different concentrations （50， 100， and 150 mg·L^－1） of BBR groups； Western blotting method was used to detect the expression levels of phosphatidylinositol 3-kinase （PI3K）， phosphorylated PI3K （p-PI3K）， protein kinase B （AKT）， phosphorylated AKT （p-AKT）， sterol regulatory element-binding protein 1 （SREBP-1），and fatty acid synthase （FASN） in the cells in various groups.The expression of FASN was suppressed by gene silencing technology， and the T98G cells at logarithmic growth phase were divided into control group， shFASN1 group， and shFASN2 group. Western blotting method was used to detect the expression levels of FASN protein in the cells in various groups； clone formation assay was used to detect the clone formation of the cells in various groups；cell wound healing assay was used to detect the migration rates of the cells in various groups. Results Compared with control group， the migration rates and invasion rates of the cells in different concentrations of BBR groups were decreased in a concentration-dependent manner （P<0.01）， and the fatty acid content in the cells in 100 mg·L^－1 BBR group was significantly decreased （P<0.01）.Compared with control group，the expression levels of p-PI3K，p-AKT， SREBP-1， and FASN proteins in the cells in 150 mg·L^－1 BBR group were significantly decreased （P<0.05 or P<0.01）， and the expression level of SREBP-1 protein in the cells in 100 and 150 mg·L^－1 BBR groups were significantly decreased （P<0.01）. After suppression of FASN expression， compared with control group， the expression levels of FASN protein in the cells in shFASN1 and shFASN2 groups were significantly decreased （P<0.01）， and the expression level of FASN protein in the cells in shFASN2 group was lower than that in shFASN1 group （P<0.05）； compared with control group， the numbers of clone formation and migration rates of the cells in shFASN1 and shFASN2 groups were significantly decreased （P<0.01）， and the migration rate of the cells in shFASN2 group was significantly lower than that in shFASN1 group （P<0.05）. Conclusion BBR interferes with fatty acid synthesis in the glioma T98G cells by reducing the expression of the PI3K/AKT/SREBP-1/FASN pathway related proteins， and decrease their migration and invasion capabilities.

Key words: Berberine, Glioma, Fatty acid, Cell migration, Cell invasion

中图分类号:

R285.5

孙玉学,刘自强,吴豪,赵黎明,高涛,黄海燕,栗超跃. 小檗碱对人胶质瘤T98G细胞迁移和侵袭的抑制作用及其机制[J]. 吉林大学学报(医学版), 2024, 50(1): 50-57.

Yuxue SUN,Ziqiang LIU,Hao WU,Liming ZHAO,Tao GAO,Haiyan HUANG,Chaoyue LI. Inhibitory effect of berberine on migration and invasion of human glioma T98G cells and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2024, 50(1): 50-57.

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参考文献 24

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Group	Saturated fatty acid	Mono-unsaturated fatty acid	Polyunsaturated fatty acid	Total fatty acid
Control	100.03±5.33	103.26±3.18	103.25±3.17	102.43±2.43
100 mg·L^－1 BBR	60.46±2.35^*	64.43±1.32^*	70.49±2.01^*	65.23±2.13^*

Group	p-PI3K protein	PI3K protein	p-AKT protein	AKT protein	SREBP-1 protein	FASN protein
Control	100.05±3.03	100.03±5.11	100.02±6.12	100.12±2.51	100.11±5.32	100.23±7.92
BBR(mg·L^－1) 50	97.23±8.33	123.93±7.58	88.73±7.23	128.65±3.45	92.12±9.15	92.33±6.43
100	74.22±6.18^*	116.25±8.59	58.32±11.02^**	130.04±6.52	88.32±7.95	85.78±8.11^*
150	67.95±15.32^**	112.21±5.23	9.45±6.56^**	132.29±4.67	77.28±13.12^**	73.43±9.13^**
^P<0.05，^*P<0.01 compared with control group.

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