Effects of nobiletin on proliferation and apoptosis of laryngeal squamous cell carcinoma cells by regulating the FAK/AKT signaling pathway

doi:10.11958/20250433

Abstract

Abstract:

Objective To investigate the effect of nobiletin on the proliferation and apoptosis of laryngeal squamous cell carcinoma (LSCC) cells and its mechanism related to the focal adhesion kinase (FAK)/protein kinase B (AKT) signaling pathway. Methods TU177 cells of LSCC were randomly assigned into the control group, the L-nobiletin group (10 mg/L) group, the M-nobiletin group (20 mg/L), the H-nobiletin group (40 mg/L) and the H-nobiletin+FAK activator ZINC40099027 group (40 mg/L nobiletin+2.5 g/L ZINC40099027). CCK-8 method was used to detect cell viability. Flow cytometry was used to detect cell apoptosis. Scratch experiment was used to detect cell migration. Transwell method was used to detect cell invasion. Western blot assay was used to detect apoptosis and FAK/AKT pathway related proteins. Results Compared with the control group, the cell viability, scratch healing rate, number of invasive cells, the protein expressions of Bcl-2, p-FAK/FAK and p-AKT/AKT were decreased in a dose-dependent manner in the L-nobiletin group, the M-nobiletin group and the H-nobiletin group, while the cell apoptosis rate and protein expressions of Bax and caspase-3 were increased in a dose-dependent manner (P<0.05). The OD value, scratch healing rate, number of invasive cells, Bcl-2, p-FAK/FAK and p-AKT/AKT protein expression levels were higher in the H-nobiletin+ZINC40099027 group than those of the H-nobiletin group, and cell apoptosis rate, Bax and caspase-3 protein expression were lower compared to the H-nobiletin group (P<0.05). Conclusion Nobiletin may inhibit the proliferation and promote the apoptosis of LSCC cells by suppressing the FAK/AKT signaling pathway.

Key words: laryngeal neoplasms, carcinoma, squamous cell, nobiletin, focal adhesion protein-tyrosine kinases, proto-oncogene proteins c-akt, cell proliferation, apoptosis

CLC Number:

R739.65

Figures/Tables 9

References 25

[1]	CAVALIERE M, BISOGNO A, SCARPA A, et al. Biomarkers of laryngeal squamous cell carcinoma:a review[J]. Ann Diagn Pathol, 2021,54:151787. doi:10.1016/j.anndiagpath.2021.151787.
[2]	BRAY F, LAVERSANNE M, SUNG H, et al. Global cancer statistics 2022:GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin, 2024, 74(3):229-263. doi:10.3322/caac.21834.
[3]	NOCINI R, MOLTENI G, MATTIUZZI C, et al. Updates on larynx cancer epidemiology[J]. Chin J Cancer Res, 2020, 32(1):18-25. doi:10.21147/j.issn.1000-9604.2020.01.03.
[4]	BARSOUK A, ALURU J S, RAWLA P, et al. Epidemiology,risk factors,and prevention of head and neck squamous cell carcinoma[J]. Med Sci (Basel), 2023, 11(2):42-46. doi:10.3390/medsci11020042.
[5]	JOHNSON D E, BURTNESS B, LEEMANS C R, et al. Head and neck squamous cell carcinoma[J]. Nat Rev Dis Primers, 2020, 6(1):92. doi:10.1038/s41572-020-00224-3.
[6]	杨雪竹, 张浩, 崔西玉, 等. 川陈皮素抑制胃癌SGC-7901细胞侵袭能力的机制探讨[J]. 现代肿瘤医学, 2020, 28(18):3099-3104.
	YANG X Z, ZHANG H, CUI X Y, et al. Study on the mechanism of nobiletin inhibiting the invasiveness of SGC-7901 gastric cancer cells[J]. J Mod Oncol, 2020, 28(18):3099-3104. doi:10.3969/j.issn.1672-4992.2020.18.001.
[7]	张阳, 姜华茂. 川陈皮素对人前列腺癌DU145细胞生长的抑制作用及其机制[J]. 吉林大学学报(医学版), 2020, 46(6):1260-1266.
	ZHANG Y, JIANG H M. Inhibitory effect of nobiletin on growth of human prostate cancer DU145 cells and its mechanism[J]. J Jilin Univ(Med Ed), 2020, 46(6):1260-1266. doi:10.13481/j.1671-587x.20200624.
[8]	ROSAS-MARTÍNEZ M, GONZÁLEZ ROSALES J A, GUTIÉRREZ-VENEGAS G. Nobiletin regulates invasion,migration and metastasis in hypopharyngeal squamous cell carcinoma[J]. Cell Mol Biol (Noisy-le-grand), 2023, 69(7):1-6. doi:10.14715/cmb/2023.69.7.1.
[9]	WANG R, WANG S, LI Z, et al. PLEKHH2 binds β-arrestin1 through its FERM domain,activates FAK/PI3K/AKT phosphorylation,and promotes the malignant phenotype of non-small cell lung cancer[J]. Cell Death Dis, 2022, 13(10):858. doi:10.1038/s41419-022-05307-5.
[10]	XU Z, LI H, LIN C, et al. Knockdown of RABL3 suppresses the proliferation and invasion of oral squamous cell carcinoma through inactivating the FAK/AKT pathway[J]. J Bioenerg Biomembr, 2021, 53(2):203-211. doi:10.1007/s10863-021-09871-x.
[11]	陈晓, 谢涵. 川陈皮素调节TLR4/TRIF/IRF3信号通路对宫颈癌细胞增殖、凋亡和侵袭的影响[J]. 中国优生与遗传杂志, 2022, 30(8):1305-1311.
	CHEN X, XIE H. Effects of Nobiletin on the proliferation,apoptosis and invasion of cervical cancer cells by regulating TLR4/TRIF/IRF3 signaling pathway[J]. Chinese Journal of Birth Health & Heredity, 2022, 30(8):1305-1311. doi:10.13404/j.cnki.cjbhh.2022.08.038.
[12]	WANG Q, GALLARDO-MACIAS R, RASHMI, et al. Discovery of novel small-molecule FAK activators promoting mucosal healing[J]. ACS Med Chem Lett, 2021, 12(3):356-364. doi:10.1021/acsmedchemlett.0c00311.
[13]	MATTIOLI F, FERMI M, MOLINARI G, et al. pT3 N0 laryngeal squamous cell carcinoma:oncologic outcomes and prognostic factors of surgically treated patients[J]. Laryngoscope, 2021, 131(10):2262-2268. doi:10.1002/lary.29528.
[14]	AL-HAKAMI H A, AL GARNI M A, ALSUBAYEA H, et al. The incidence of thyroid gland invasion in advanced laryngeal squamous cell carcinoma[J]. Braz J Otorhinolaryngol, 2021, 87(5):533-537. doi:10.1016/j.bjorl.2019.11.003.
[15]	周晓红, 杨雪, 连蕾, 等. 白芍总苷对喉癌Hep-2细胞增殖、侵袭、迁移及PI3K/Akt/GSK3β信号通路的影响[J]. 标记免疫分析与临床, 2022, 29(2):236-241.
	ZHOU X H, YANG X, LIAN L, et al. Effects of total glucosides of paeony on proliferation,invasion,migration and PI3K/Akt/GSK3β signaling pathway of laryngeal cancer Hep-2 cells[J]. Labeled Immunoassays & Clin Med, 2022, 29(2):236-241. doi:10.11748/bjmy.issn.1006-1703.2022.02.012.
[16]	CHEN Y Y, LIANG J J, WANG D L, et al. Nobiletin as a chemopreventive natural product against cancer,a comprehensive review[J]. Crit Rev Food Sci Nutr, 2023, 63(23):6309-6329. doi:10.1080/10408398.2022.2030297.
[17]	JIANG H, CHEN H, JIN C, et al. Nobiletin flavone inhibits the growth and metastasis of human pancreatic cancer cells via induction of autophagy,G0/G1 cell cycle arrest and inhibition of NF-κB signalling pathway[J]. J BUON, 2020, 25(2):1070-1075.
[18]	WU Y, LI Q, LV L L, et al. Nobiletin inhibits breast cancer cell migration and invasion by suppressing the IL-6-induced ERK-STAT and JNK-c-JUN pathways[J]. Phytomedicine, 2023,110:154610. doi:10.1016/j.phymed.2022.154610.
[19]	芮翊馨, 谢红潇, 李丹, 等. 中乌宁通过BAX/BCL-2/Caspase 3信号通路抑制细胞凋亡改善慢性肾衰竭大鼠肾纤维化[J]. 中药药理与临床, 2024, 40(5):42-48.
	RUI Y X, XIE H X, LI D, et al. Mesaconine inhibits apoptosis through the BAX /BCL-2 /Caspase 3 signaling pathway to improve renal fibrosis in rats with chronic renal failure[J]. Pharmacol Clin Chin Mater Med, 2024, 40(5):42-48. doi:10.13412/j.cnki.zyyl.20231117.003.
[20]	BULGURCUOGLU KURAN S, ALTUN A, KARAMUSTAFAOGLU BALCI B, et al. Expression of pro-apoptotic and anti-apoptotic proteins in granulosa cells of women with diminished ovarian reserve[J]. J Assist Reprod Genet, 2022, 39(3):765-775. doi:10.1007/s10815-022-02422-2.
[21]	HU L, DUAN Y T, LI J F, et al. Biglycan enhances gastric cancer invasion by activating FAK signaling pathway[J]. Oncotarget, 2014, 5(7):1885-1896. doi:10.18632/oncotarget.1871.
[22]	DU J, FU L, HAO J, et al. Rab11a is overexpressed in gastric cancer and regulates FAK/AKT signaling[J]. J Oncol, 2020,2020:3494396. doi:10.1155/2020/3494396.
[23]	徐加志, 张芸, 陈大刚, 等. 咖啡因通过FAK/AKT/ROCK通路调控人脑胶质瘤U-373MG细胞的恶性生物学行为[J]. 中国肿瘤生物治疗杂志, 2024, 31(6):573-578.
	XU J Z, ZHNAG Y, CHEN D G, et al. Caffeine modulates the malignant biological behaviors of human brain glioma U-373MG cells by regulating the FAK/AKT/ROCK pathway[J]. Chin J Cancer Biother, 2024, 31(6):573-578. doi:10.3872/j.issn.1007-385x.2024.06.005.
[24]	江小霞, 方燕玲, 李仲均. 褪黑素通过抑制FGA调节FAK/AKT/MMP-2通路对人子宫内膜间质细胞的作用研究[J]. 徐州医科大学学报, 2021, 41(12):865-872.
	JIANG X X, FANG Y L, LI Z J. Effect of melatonin on human endometrial stromal cells by inhibiting FGA to regulate the FAK/AKT/MMP-2 pathway[J]. J Xuzhou Med Univ, 2021, 41(12):865-872. doi:10.3969/j.issn.2096-3882.2021.12.002.
[25]	吴娜萍, 张磊, 方琦, 等. IGSF10通过抑制FAK/PI3K/AKT信号通路降低右美托咪定对乳腺癌促进作用的研究[J]. 中华肿瘤防治杂志, 2022, 29(14):1052-1061.
	WU N P, ZHANG L, FANG Q, et al. IGSF10 reduces dexmedetomidine promotion in breast cancer by inhibiting FAK/PI3K/AKT signaling pathway[J]. Chin J Cancer Prev Treat, 2022, 29(14):1052-1061. doi:10.16073/j.cnki.cjcpt.2022.14.06.

组别	OD值	凋亡率/%
Control组	1.65±0.17	8.67±0.91
L-川陈皮素组	1.27±0.13^a	13.86±1.41^a
M-川陈皮素组	0.86±0.09^ab	18.12±1.86^ab
H-川陈皮素组	0.51±0.06^abc	23.52±2.40^abc
H-川陈皮素+ZINC40099027组	1.31±0.14^d	10.43±1.09^d
F	75.798^＊＊	81.980^＊＊

组别	OD值	凋亡率/%
Control组	1.65±0.17	8.67±0.91
L-川陈皮素组	1.27±0.13^a	13.86±1.41^a
M-川陈皮素组	0.86±0.09^ab	18.12±1.86^ab
H-川陈皮素组	0.51±0.06^abc	23.52±2.40^abc
H-川陈皮素+ZINC40099027组	1.31±0.14^d	10.43±1.09^d
F	75.798^＊＊	81.980^＊＊

组别	划痕愈合率/%	侵袭细胞数/个
Control组	83.62±8.53	186.32±19.12
L-川陈皮素组	68.51±6.92^a	143.68±14.51^a
M-川陈皮素组	51.37±5.15^ab	119.62±12.33^ab
H-川陈皮素组	39.50±4.01^abc	80.55±8.17^abc
H-川陈皮素+ZINC40099027组	70.34±7.11^d	145.27±14.66^d
F	41.968^＊＊	44.681^＊＊

组别	划痕愈合率/%	侵袭细胞数/个
Control组	83.62±8.53	186.32±19.12
L-川陈皮素组	68.51±6.92^a	143.68±14.51^a
M-川陈皮素组	51.37±5.15^ab	119.62±12.33^ab
H-川陈皮素组	39.50±4.01^abc	80.55±8.17^abc
H-川陈皮素+ZINC40099027组	70.34±7.11^d	145.27±14.66^d
F	41.968^＊＊	44.681^＊＊

组别	Bax/GAPDH	Bcl-2/GAPDH	caspase-3/GAPDH
Control组	0.62±0.07	1.42±0.15	0.29±0.03
L-川陈皮素组	0.89±0.09^a	1.13±0.12^a	0.68±0.07^a
M-川陈皮素组	1.08±0.11^ab	0.92±0.10^ab	0.85±0.09^ab
H-川陈皮素组	1.33±0.14^abc	0.61±0.07^abc	0.99±0.11^abc
H-川陈皮素+ ZINC40099027组	0.81±0.09^d	1.21±0.13^d	0.52±0.06^d
F	41.665^＊＊	41.384^＊＊	76.449^＊＊