沉默WNT4基因对稳定转染PAX2基因大鼠肾小管上皮细胞迁移和侵袭能力的影响

doi:10.13481/j.1671-587x.20180618

吉林大学学报(医学版) ›› 2018, Vol. 44 ›› Issue (06): 1212-1217.doi: 10.13481/j.1671-587x.20180618

沉默WNT4基因对稳定转染PAX2基因大鼠肾小管上皮细胞迁移和侵袭能力的影响

李里¹, 宫亮², 吴玉斌³

1. 锦州医科大学附属第一医院风湿免疫科, 辽宁锦州 121000;
2. 锦州医科大学附属第一医院耳鼻喉科, 辽宁锦州 121000;
3. 中国医科大学附属盛京医院小儿肾脏风湿免疫科, 辽宁沈阳 110004

收稿日期:2018-02-05 出版日期:2018-11-28 发布日期:2018-11-28
通讯作者: 吴玉斌,教授,博士研究生导师(Tel:024-23896615,E-mail:772652284@qq.com) E-mail:772652284@qq.com
作者简介:李里(1978-),女,辽宁省锦州市人,副教授,医学博士,主要从事风湿免疫学方面的研究。
基金资助:
辽宁省博士科研启动基金计划项目资助课题（20141136）

Effects of WNT4 gene silencing on migration and invasion of renal tubular epithelial cells of rats stably transfected with PAX2 gene

LI Li¹, GONG Liang², WU Yu bin³

1. Department of Rheumatology and Immunology, First Affiliated Hospital, Jinzhou Medical University, Jinzhou 121000, China;
2. Department of Ear-Nose-Throat, First Affiliated Hospital, Jinzhou Medical University, Jinzhou 121000, China;
3. Department of Pediatric Nephrology and Rheumatology, Shengjing Hospital, China Medical University, Shenyang 110004, China

Received:2018-02-05 Online:2018-11-28 Published:2018-11-28

摘要/Abstract

摘要： 目的：探讨沉默WNT4基因对稳定转染配对盒基因2（PAX2）大鼠肾小管上皮细胞迁移和侵袭能力的影响，阐明PAX2基因是否通过WNT4基因调控细胞生物学活性。方法：选取对数生长期的PAX2基因稳转细胞系，分为稳转组和对照组。稳转组细胞分为稳转对照组（不应用WNTsiRNA沉默）和沉默72 h组（应用WNTsiRNA沉默72h）。Western blotting法检测稳转组和对照组大鼠肾小管上皮细胞中PAX2和WNT4蛋白的相对表达量，Realtime PCR法检测稳转对照组和沉默72 h组大鼠肾小管上皮细胞中WNT4 mRNA相对表达量，细胞划痕实验检测稳转对照组和沉默72 h组大鼠肾小管上皮细胞的细胞迁移率，Transwell实验检测稳转对照组和沉默72 h组大鼠肾小管上皮细胞的穿膜细胞数。结果：Western blotting法检测，稳转组大鼠肾小管上皮细胞中PAX2和WNT4蛋白相对表达量高于对照组（P<0.05）。Real-time PCR法检测，沉默72 h组大鼠肾小管上皮细胞中WNT4 mRNA相对表达量低于稳转对照组（P<0.05）。细胞划痕实验10 h后，沉默72 h组大鼠肾小管上皮细胞的细胞迁移率低于稳转对照组，但组间比较差异无统计学意义（P>0.05）；细胞划痕实验18 h后，沉默72h组大鼠肾小管上皮细胞的细胞迁移率低于稳转对照组（P>0.05）。Transwwell实验，沉默72 h组大鼠肾小管上皮细胞的穿膜细胞数低于稳转对照组（P<0.05）。结论：PAX2基因可能通过WNT4基因调控细胞的生物学活性。

关键词: 配对盒基因2, WNT4基因, 肾小管上皮细胞, 细胞迁移, 细胞侵袭, 转染

Abstract: Objective: To investigate the effects of WNT4 gene silecing on the migration and invasion of renal tubular epithelial cells of the rats stably transfected with paired box gene 2(PAX2) gene,and to clarify whether the PAX2 gene regulates the biological activities of renal tubular epithelial cells through WNT4 gene or not.Methods: The renal tubular epithelial cells transfected with PAX2 gene in the logarithmic phase were divided into stably transfected group and control group. The cells in stably transfected group were divided into stably transfected control group(without WNT4 siRNA silencing) and silencing 72 h group(with WNT4 siRNA silencing for 72 h). The relative expression amounts of PAX2 and WNT4 proteins in renal tubular epithelial cells of the rats in stably transfected group and control group were detected by Western blotting method. The relative expression amounts of WNT4 mRNA in renal tubular epithelial cells of the rats in stably transfected control group and silencing 72 h group were detected by Real time PCR method. The migration rates of renal tubular epithelial cells of the rats in stably transfected control group and silencing 72 h group were detected by cell wound scratch assay. The number of transmembrane renal tubular epithelial cells of the rats in stably transfected control group and silencing 72 h group was detected by Transwell experiment.Results: The Western blotting results showed that the relative expression amounts of PAX2 and WNT4 proteins in renal tubular epithelial cells of the rats in stably transfected group were higher than those in control group(P<0.05). The Real time PCR results showed that the relative expression amount of WNT4 mRNA in renal tubular epithelial cells of the rats in silencing 72 h group was lower than that in stably transfected control group(P<0.05).The cell wound scratch assay results showed that the migration rate of renal tubular epithelial cells in silencing 72 h group after 10 h was lower than that in stably transfected control group,but there was no significant difference (P>0.05); the migration rate of renal tubular epithelial cells in silencing 72 h group after 18 h was lower than that in stably transfected control group(P<0.05).The Transwell experiment results showed that the number of transmembrane renal tubular epithelial cells in silencing 72 h group was lower than that in stably transfected control group(P<0.05).Conclusion: PAX2 gene can regulate the biological activities of renal tubular epithelial cells through WNT4 gene.

Key words: parired box gene 2, WNT4 gene, renal tubular epithelial cells, cell migration, cell invasion, transfection

中图分类号:

李里, 宫亮, 吴玉斌. 沉默WNT4基因对稳定转染PAX2基因大鼠肾小管上皮细胞迁移和侵袭能力的影响[J]. 吉林大学学报(医学版), 2018, 44(06): 1212-1217.

LI Li, GONG Liang, WU Yu bin. Effects of WNT4 gene silencing on migration and invasion of renal tubular epithelial cells of rats stably transfected with PAX2 gene[J]. Journal of Jilin University(Medicine Edition), 2018, 44(06): 1212-1217.

参考文献

[1] Kahraman K, Kiremitci S, Taskin S, et al. Expression pattern of PAX2 in hyperplastic and malignant endometrium[J]. Arch Gynecol Obstet, 2012, 286(1):173-178.
[2] El-Nahas AM. Plasticity of kidney cells:Role in kidney remodeling and scarring[J]. Kindey Int, 2003, 64(5):1553-1563.
[3] Narlis M, Grote D, Gaitan Y, et al.Pax2 and pax8 regulate branching morphogenesis and nephron differentiation in the developing kidney[J]. J Am Soc Nephrol, 2007, 18(4):1121-1129.
[4] Torban E, Goodyer P. What PAX genes do in the kidney[J]. Exp Nephrol, 1998,6(1):7-11.
[5] Torban E, Dziarmaga A, Iglesias D, et al. PAX2 activates WNT4 expression during mammalian kidney development[J]. J Biol Chem, 2006, 281(18):12705-12712.
[6] 李里,南晓娟,吴玉斌. WNT4基因沉默对PAX2诱导肾小管上皮细胞间充质转分化的影响[J]. 吉林大学学报:医学版,2013,39(4):720-725.
[7] 李里,宫亮,吴玉斌. PAX2稳转细胞系构建及对细胞迁移和侵袭能力的作用研究[J]. 中国全科医学, 2015,18(35):4325-4329.
[8] Lyons JP, Mueller UW, Ji H, et al. Wnt-4 activates the canonical beta-catenin-mediated Wnt pathway and binds Frizzled-6 CRD:functional implications of Wnt/beta-catenin activity in kidney epithelial cells[J]. Exp Cell Res, 2004, 298(2):369-387.
[9] Liu Y. New insights into epithelial-mesenchymal transition in kidney fibrosis[J].J Am Soc Nephrol, 2010, 21(2):212-222.
[10] Chilosi M, Poletti V, Zamo A, et al. Aberrant Wnt/beta-catenin pathway activation in idiopathic pulmonary fibrosis[J]. Am J Pathol, 2003, 162(5):1495-1502.
[11] Jiang F, Parsons CJ, Stefanovic B. Gene expression profile of quiescent and activated rat hepatic stellate cells implicates Wnt signaling pathway in activation[J]. J Hepatol, 2006, 45(3):401-409.
[12] Patel D, Shimomura A, Majumdar S, et al. The histone demethylase LSD1 regulates inner ear progenitor differentiation through interactions with Pax2 and the NuRD repressor complex[J]. PLoS One,2018,13(1):e0191689.
[13] Ma X, Ma Z, Jiao X, et al. Functional non-coding polymorphism in an EPHA2 promoter PAX2 binding site modifies expression and alters the MAPK and AKT pathways[J]. Sci Rep,2017,7(1):9992.
[14] Nguyen HT, Thomson AA, Kogan BA, et al. Expression of the Wnt gene family during late nephrogenesis and complete ureteral obstruction[J]. Lab Invest, 1999, 79(6):647-658.
[15] He W, Dai C, Li Y, et al. Wnt/beta-catenin signaling promotes renal interstitial fibrosis[J]. J Am Soc Nephrol, 2009, 20(4):765-776.
[16] Surendran K, McCaul SP, Simon TC. A role for Wnt-4 in renal fibrosis[J].Am J Physiol Renal Physiol, 2002, 282(3):F431-F441.
[17] Surendran K, Schiavi S, Hruska KA. Wnt-dependent beta-catenin signaling is activated after unilateral ureteral obstruction, and recombinant secreted frizzled-related protein 4 alters the progression of renal fibrosis[J]. J Am Soc Nephrol, 2005, 16(8):2373-2384.
[18] Chen Y, Zhang P, Tang P, et al. Wnt4 overexpression promotes thymoma development through a JNK-mediated planar cell polarity-like pathway[J]. Oncol Lett,2018,15(1):83-90.
[19] Wang L, Qing L, Liu H, et al. Mesenchymal stromal cells ameliorate oxidative stress-induced islet endothelium apoptosis and functional impairment via Wnt4-β-catenin signaling[J]. Stem Cell Res Ther,2017,8(1):188-122.
[20] 雷蕾,李良平,张虎.血清胱抑素C联合尿中性粒细胞明胶酶相关脂质运载蛋白对肝硬化腹水继发急性肾损伤的诊断价值[J].临床肝胆病杂志,2018,34(1):101-105.

沉默WNT4基因对稳定转染PAX2基因大鼠肾小管上皮细胞迁移和侵袭能力的影响

Effects of WNT4 gene silencing on migration and invasion of renal tubular epithelial cells of rats stably transfected with PAX2 gene

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