阿尔茨海默病细胞模型中lncRNA RP11-543N12.1对CDH13表达的调控作用

doi:10.6040/j.issn.1671-7554.0.2016.102

摘要/Abstract

摘要： 目的探讨阿尔茨海默病(AD)细胞模型中长链非编码RNA(lncRNA)RP11-543N12.1对CDH13的表达调控作用。方法采用基因芯片技术筛选AD相关差异表达lncRNA和mRNA,并通过Real-time PCR技术验证AD细胞模型中lncRNA RP11-543N12.1和CDH13 mRNA表达均增加,与芯片结果一致;然后将RP11-543N12.1-siRNA转入SH-SY5Y细胞,并加入β-淀粉样蛋白(Aβ_25-35)作用48 h,接着MTT检测细胞存活率,Real-time PCR技术检测RP11-543N12.1和CDH13在RNA水平的变化,Western blotting检测CDH13蛋白表达,Hoechst33258染色检测细胞核的形态学变化,流式细胞术检测凋亡率。结果在未转染RP11-543N12.1-siRNA的SH-SY5Y细胞中,Aβ_25-35处理48 h后,与正常细胞相比,细胞存活率降低(P<0.05),CDH13 RNA水平增加(P<0.01),CDH13蛋白活性形式表达量增加(P<0.01),细胞核出现凋亡的形态学变化,流式检测细胞凋亡率增加(P<0.01);转入RP11-543N12.1-siRNA后再用 Aβ_25-35处理48 h,细胞存活率、CDH13 RNA水平、蛋白活性形式表达量、细胞凋亡率均趋于正常水平,细胞核凋亡的形态学变化消失。结论 LncRNA RP11-543N12.1能够调节CDH13的表达,降低RP11-543N12.1的表达量能使CDH13表达下调。

关键词: LncRNA RP11-543N12.1, SH-SY5Y细胞, 阿尔茨海默病, CDH13, 表达调控

Abstract: Objective To explore the regulatory function of long noncoding RNA(lncRNA)RP11-543N12.1 on the expression of CDH13 in Alzheimers disease(AD)cell model. Methods Firstly, differently expressed lncRNAs and mRNAs were screened through microarray, and upregulated expressions of both RP11-543N12.1 and CDH13 were verified by real-time PCR. Secondly, SH-SY5Y cells were transfected with RP11-543N12.1-siRNA, and then Aβ_25-35 was added to the transfected cells. After 48 h, the survival rate was detected with MTT; RNA expressions of RP11-543N12.1 and CDH13 were detected with real-time PCR; protein expression of CDH13 was determined with Western blotting; morphological changes were observed with Hoechst33258 staining; cellular apoptosis rate was detected with flow cytometry. Results Compared with control SH-SY5Y cells, cells treated with Aβ_25-35 showed inhibited proliferation(P<0.05), upregulated RNA and protein expressions of CDH13(both P<0.01), and increased cellular apoptosis rate(P<0.01). Apoptotic nuclei were also observed. In cells transfected with RP11-543N12.1-siRNA and treated with Aβ_25-35, cellular survival rate, RNA and protein expressions, and cellular apoptosis rate were all close to normal levels. Nuclei apoptosis disappeared. Conclusion LncRNA RP11-543N12.1 can regulate expression of CDH13, and knock-down of RP11-543N12.1 can downregulate the expression of CDH13.

Key words: CDH13, Expression regulation, SH-SY5Y cell, LncRNA RP11-543N12.1, Alzheimers disease

中图分类号:

R574

李蒙蒙,王苗苗,刁雪琴,田克立,徐霞,任桂杰. 阿尔茨海默病细胞模型中lncRNA RP11-543N12.1对CDH13表达的调控作用[J]. 山东大学学报（医学版）, 2017, 55(3): 12-18.

LI Mengmeng, WANG Miaomiao, DIAO Xueqin, TIAN Keli, XU Xia, REN Guijie. Regulatory effect of lncRNA RP11-543N12.1 on the expression of CDH13 in Alzheimers disease cell model[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2017, 55(3): 12-18.

参考文献

[1] Hebert LE, Weuve J, Scherr PA, et al. Alzheimer disease in the United States(2010—2050)estimated using the 2010 census[J]. Neurology, 2013, 80(19):1778-1783.
[2] Barnes DE, Yaffe K. The projected effect of risk factor reduction on Alzheimers disease prevalence[J]. Lancet Neurol, 2011, 10(9):819-828.
[3] Tan CC, Yu JT, Wang HF, et al. Efficacy and safety of donepezil, galantamine, rivastigmine, and memantine for the treatment of Alzheimers disease: a systematic review and meta-analysis[J]. J Alzheimers Dis, 2014, 41(2):615-631.
[4] Caraci F, Bosco P, Leggio GM, et al. Clinical pharmacology of novel anti-Alzheimer disease modifying medications. Curr. Top[J]. Med Chem, 2013, 13(15):1853-1863.
[5] Esteller M. Non-coding RNAs in human disease[J]. Nat Rev Genet, 2011, 12(4):861-874.
[6] Kaikkonen MU, Lam MT, Glass CK. Non-coding RNAs as regulators of gene expression and epigenetics[J]. Cardiovasc Res, 2011, 90(3):430-440.
[7] Sana J, Faltejskova P, Svoboda M, et al. Novel classes of non-coding RNAs and cancer[J]. J Transl Med, 2012, 10:103. doi:10.1186/1479-5876-10-103.
[8] Zhou H, Hu H, Lai M. Non-coding RNAs and their epigenetic regulatory mechanisms[J]. Biol Cell, 2010, 102(5):645-655.
[9] Wu P, Zuo X, Deng H, et al. Roles of long noncoding RNAs in brain development, functional diversification and neurodegenerative diseases[J]. Brain Res Bull, 2013, 97:69-80. doi:10.1016/j.brainresbull.2013.06.001.
[10] 刁雪芹, 王苗苗, 庄云龙, 等. 嘌呤霉素敏感的氨肽酶对β-淀粉样蛋白诱导的细胞凋亡的影响[J].中华神经医学杂志, 2013, 12(4):347-353. DIAO Xueqin, WANG Miaomiao, ZHUANG Yunlong, et al. Influence of puromycin-sensitive aminopeptidase on apoptosis induced by beta-Amyloid peptides in cells[J]. Chin J Neuromed, 2013, 12(4):347-353.
[11] Sun J, Yan P, Chen Y, et al. MicroRNA-26b inhibits cell proliferation and cytokine secretion in human RASF cells via the Wnt/GSK-3β/β-catenin pathway[J]. Diagn Pathol, 2015, 10:72. doi:10.1186/s13000-015-0309-x.
[12] Huang T, Li J, Zhang C, et al. Distinguishing lung adenocarcinoma from lung squamous cell carcinoma by two hypomethylated and three hypermethylated genes:A meta-analysis[J]. PLoS One, 2016, 11(2):1-13.
[13] Shakeri H, Gharesouran J, Fakhrjou A, et al. DNA methylation assessment as a prognostic factor in invasive breast cancer using methylation-specific multiplex ligation dependent probe amplification[J]. EXCLI J, 2016, 15:11-20. doi:10.17179/excli2015-485. eCollection 2016.
[14] Sheng Y, Hongtao W, Liu D, et al. Methylation of tumor suppressor gene CDH13 and SHP1 promoters and their epigenetic regulation by the UHRF1/PRMT5 complex in endometrial carcinoma[J]. Gynecologic Oncology, 2016, 140(1):145-151.
[15] Angst BD, Marcozzi C, Magee AI. The cadherin superfamily:diversity in form and function[J]. J Cell Sci, 2001, 114(4):629-641.
[16] Toyooka S, Toyooka KO, Maruyama R, et al. DNA methylation profiles of lung Tumors[J]. Mol Cancer Ther, 2001, 1(1):61-67.
[17] Takeichi M. The cadherin superfamily in neuronal connections and interactions[J]. Nat Rev Neurosci, 2007, 8(1):11-20.
[18] Rubina K, Kalinina N, Potekhina A, et al. T-cadherin suppresses angiogenesis in vivo by inhibiting migration of endothelial cells[J]. Angiogenesis, 2007, 10(3):183-195.
[19] Takeuchi T, Misaki A, Liang SB, et al. Expression of T-cadherin(CDH13, H-Cadherin)in human brain and its characteristics as a negative growth regulator of epidermal growth factor in neuroblastoma cells[J]. J Neurochem, 2000, 74(4):1489-1497.
[20] Huang ZY, Wu Y, Hedrick N, et al. T-cadherinmediated cell growth regulation involves G2 phase arrest and requires p21(CIP1/WAF1)expression[J]. Mol Cell Biol, 2003, 23(2):566-578.
[21] Derrien T, Johnson R, Bussotti G, et al. The GENCODE v7 catalog of human long noncoding RNAs: analysis of their gene structure, evolution, and expression[J]. Genome Res, 2012, 22(9):1775-1789.
[22] Faghihi MA, Modarresi F, Khalil, AM, et al. Expression of a noncoding RNA is elevated in Alzheimers disease and drives rapid feed-forward regulation of beta-secretase[J]. Nat Med, 2008, 14(7):723-730.
[23] Wang Y, Pang WJ, Wei N, et al. Identification, stability and expression of Sirt1 antisense long non-coding RNA[J]. Gene, 2014, 539(1):117-124.
[24] Joshi MB, Ivanov D, Philippova M, et al. Integrin-linked kinase is an essential mediator for T-cadherin dependent signaling via Akt and GSK3 beta in endothelial cells[J]. FASEB J, 2007, 21(12):3083-3095.

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