CACNA2D3在顺铂诱导HEI-OC1细胞凋亡中的作用及机制研究

Effect and Mechanism of CACNA2D3 in Cisplatin-induced Apoptosis of HEI-OC1 Cells

田雨鑫;李壮壮;王菁菁;冯艳梅;陈正侬;

1:上海交通大学附属第六人民医院耳鼻咽喉头颈外科

2:上海交通大学耳鼻咽喉科研究所

3:上海市睡眠呼吸障碍疾病重点实验室

摘要
目的 探讨CACNA2D3在顺铂诱导的HEI-OC1细胞凋亡中的作用及机制。方法 利用shRNA干扰技术敲减HEI-OC1细胞中CACNA2D3(sh-Cac3)表达,空白载体(sh-Vector)转染对照细胞,sh-Cac3细胞和sh-Vector细胞分别进行正常培养及顺铂干预。流式细胞术检测线粒体膜电位、凋亡情况、细胞内钙离子浓度;Western blot检测细胞凋亡相关蛋白(Bcl2、Cleaved-PARP1)和自噬标记蛋白LC3Ⅱ;免疫荧光法观察Tunel染色。结果 正常培养下,CACNA2D3敲减会降低细胞内钙离子浓度(P<0.05),但不会影响细胞凋亡和自噬。顺铂干预下,相较于sh-Vector细胞,sh-Cac3细胞的线粒体膜电位降低程度更高(P<0.01);细胞凋亡率更高(9.41±0.66%vs 11.53±0.45%,P<0.01);Bcl2的表达显著降低(P<0.01),而Cleaved-PARP1的表达显著升高(P<0.01);Tunel染色阳性率更高(3.08±0.14%vs 6.40±0.43%,P<0.001)。此外,顺铂干预后sh-Cac3细胞的LC3Ⅱ的表达明显高于sh-Vector细胞(P<0.01)。结论 CACNA2D3敲减通过调节钙相关自噬增强了顺铂诱导的HEI-OC1细胞凋亡;CACNA2D3的表达调控可能是顺铂耳毒性靶点治疗的新策略。
关键词
自噬;凋亡;CACNA2D3;顺铂;耳毒性
基金项目(Foundation):
国家自然基金重点项目(81770998、82071040)
作者
田雨鑫;李壮壮;王菁菁;冯艳梅;陈正侬;
参考文献

1 Karasawa T,Steyger PS.An integrated view of cisplatin-induced nephrotoxicity and ototoxicity[J].Toxicol Lett,2015,237(3):219-227.

2 Gentilin E,Simoni E,Candito M,et al.Cisplatin-induced ototoxicity:updates on molecular targets[J].Trends Mol Med,2019,25(12):1123-1132.

3 Rajput K,Edwards L,Brock P,et al.Ototoxicity-induced hearing loss and quality of life in survivors of paediatric cancer[J].Int J Pediatr Otorhinolaryngol,2020,138:110401.

4 Kamogashira T,Fujimoto C,Yamasoba T.Reactive oxygen species,apoptosis,and mitochondrial dysfunction in hearing loss[J].Biomed Res Int,2015,2015:617207.

5 雷文静,陆菲,米文娟,等.银杏叶提取物保护顺铂致小鼠HEI-OC1细胞损伤的作用及机制研究[J].听力学及言语疾病杂志,2019,27(5):519-523.

6 Nie C,Qin X,Li X,et al.CACNA2D3 enhances the chemosensitivity of esophageal squamous cell carcinoma to cisplatin via inducing Ca2+-mediated apoptosis and suppressing PI3K/Akt pathways[J].Front Oncol,2019,9:185.

7 La Rovere RM,Roest G,Bultynck G,et al.Intracellular Ca2+ signaling and Ca2+ microdomains in the control of cell survival,apoptosis and autophagy[J].Cell Calcium,2016,60(2):74-87.

8 Zhivotovsky B,Orrenius S.Calcium and cell death mechanisms:a perspective from the cell death community[J].Cell Calcium,2011,50(3):211-221.

9 Yao Z,Klionsky DJ.The symphony of autophagy and calcium signaling[J].Autophagy,2015,11(7):973-974.

10 Pirone A,Kurt S,Zuccotti A,et al.α2δ3 is essential for normal structure and function of auditory nerve synapses and is a novel candidate for auditory processing disorders[J].J Neurosci,2014,34(2):434-445.

11 Jin Y,Cui D,Ren J,et al.CACNA2D3 is downregulated in gliomas and functions as a tumor suppressor[J].Mol Carcinog,2017,56(3):945-959.

12 Gogvadze V,Orrenius S,Zhivotovsky B.Multiple pathways of cytochrome c release from mitochondria in apoptosis[J].Biochim Biophys Acta,2006,1757(5-6):639-647.

13 Pinton P,Giorgi C,Siviero R,et al.Calcium and apoptosis:ER-mitochondria Ca2+ transfer in the control of apoptosis[J].Oncogene,2008,27(50):6407-6418.

14 Liu F,Chen J,Wang P,et al.MicroRNA-27a controls the intracellular survival of Mycobacterium tuberculosis by regulating calcium-associated autophagy[J].Nat Commun,2018,9(1):4295.

15 Cao W,Li J,Yang K,et al.An overview of autophagy:Mechanism,regulation and research progress[J].Bull Cancer,2021,108(3):304-322.

16 Yin H,Yang Q,Cao Z,et al.Activation of NLRX1-mediated autophagy accelerates the ototoxic potential of cisplatin in auditory cells[J].Toxicol Appl Pharmacol,2018,343:16-28.

17 Liu T,Zong S,Luo P,et al.Enhancing autophagy by down-regulating GSK-3beta alleviates cisplatin-induced ototoxicity in vivo and in vitro[J].Toxicol Lett,2019,313:11-18.

18 Pang J,Xiong H,Zhan T,et al.Sirtuin 1 and autophagy attenuate cisplatin-induced hair cell Death in them mouse cochlea and zebrafish lateral line[J].Front Cell Neurosci,2019,12:515.

19 He Z,Guo L,Shu Y,et al.Autophagy protects auditory hair cells against neomycin-induced damage[J].Autophagy,2017,13(11):1884-1904.

20 Kong X,Li M,Shao K,et al.Progesterone induces cell apoptosis via the CACNA2D3/Ca2+/p38 MAPK pathway in endometrial cancer[J].Oncol Rep,2020,43(1):121-132.

本文信息

PDF(1384K)

本文关键词相关文章

自噬凋亡CACNA2D3顺铂耳毒性