Please wait a minute...

中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志
中国生物工程杂志  2018, Vol. 38 Issue (3): 9-15    DOI: 10.13523/j.cb.20180302
研究报告     
Herpud1对后肾间充质细胞的作用及其机制的探讨*
李依蔓,周钦()
重庆医科大学检验医学院 重庆 400016
The Effects of Herpud1 on Metanephric Mesenchymal Cells and Its Mechanism
Yi-man LI,Qin ZHOU()
The School of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, China
 全文: PDF(862 KB)   HTML
摘要：

为初步探讨Herpud1在肾脏发育中的生物学作用,过表达Herpud1载体及敲低Herpud1载体被构建。载体被转染进后肾间充质细胞,通过RT-PCR和蛋白质免疫印迹检测了上皮间质的转换过程(EMT)的标志蛋白E-钙黏蛋白(E-cadherin)、波形蛋白(Vimentin)、Snail蛋白及内质网应力的标志蛋白葡萄糖调节蛋白78(GRP78)、真核起始因子2α (eIF2α)的表达变化,通过EDU细胞增殖实验检测了Herpud1对细胞增殖的作用,同时利用细胞划痕实验验证了细胞的迁移能力。结果显示,与空白对照组相比,在Herpud1过表达的实验组中,E-cadherin的表达在mRNA和蛋白质水平均是降低的,而Snail和Vimentin的表达则是上升的,并伴随着细胞增殖活性的降低和细胞迁移能力增加,同时内质网应力相关蛋白GRP78和eIF2α是升高的。在Herpud1敲低的实验组,E-cadherin的表达在mRNA和蛋白质水平均是升高的,而Snail和Vimentin的表达下降,细胞迁移能力是降低并且细胞增殖活性升高,同时内质网应力相关蛋白GRP78和eIF2α也是降低的。这些结果证明Herpud1可以促进MK3细胞EMT过程,提高细胞的迁移能力,抑制细胞的增殖活性,其机制可能与细胞的内质网应力相关。
关键词： Herpud1;  EMT;  细胞增殖;  内质网应力;  细胞迁移    
Abstract:

To explore biological function of Herpud1 in kidney development, overexpression and knockdown vector of Herpud1 gene were transfected into MK3 cells. And then RT-PCR and Western blot were used to detect expression of epithelial mesenchymal transition (EMT) marker genes E-cadherin (epithelial cells marker), Vimentin and Snail (mesenchymal cell marker) and endoplasmic reticulum (ER) stress marker genes (GRP78 and eIF2α). And cell proliferation and migration were detected by EDU cell proliferation experiment and wound healing assay. Our results showed in experimental group with Herpud1 overexpression, E-cadherin was decreased, and expression of Vimentin and Snail was increased at mRNA and protein levels compared with control group. And ER stress markers GRP78 and eIF2α were enhanced at protein levels in MK3 cells with Herpud1 overexpression. In addition, Herpud1 promoted cell migration and inhibited cell proliferation. In Herpud1 knockdown cells, expression of E-cadherin was enhanced at mRNA and protein levels, and expression of Vimentin, Snail, GRP78 and eIF2α is reduced. At the same time, Herpud1 knockdown inhibited cell migration and enhanced ability of cell proliferation. These results demonstrate Herpud1 can promote EMT of MK3 cells, cell migration and inhibit cell proliferation. The mechanism may be associate with ER stress.
Key words: Herpud1    EMT    Cell proliferation    ER stress    Cell migration
收稿日期: 2017-10-30 出版日期: 2018-04-04
ZTFLH:  Q819  
基金资助: * 国家自然科学基金(8157081224)
服务　　
把本文推荐给朋友
加入引用管理器
E-mail Alert
RSS
作者相关文章　　
李依蔓
周钦

引用本文:

李依蔓,周钦. Herpud1对后肾间充质细胞的作用及其机制的探讨*[J]. 中国生物工程杂志, 2018, 38(3): 9-15.

Yi-man LI,Qin ZHOU. The Effects of Herpud1 on Metanephric Mesenchymal Cells and Its Mechanism. China Biotechnology, 2018, 38(3): 9-15.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20180302    或    https://manu60.magtech.com.cn/biotech/CN/Y2018/V38/I3/9

图1  Herpud1过表达或者敲低载体在MK3细胞中的效率
图2  Herpud1能够促进MK3细胞的EMT过程
图3  Herpud1抑制MK3细胞的增殖活性
图4  Herpud1促进MK3细胞的迁移
图5  Herpud1促进内质网应力的相关蛋白表达
[1] Little M H, McMahon A P. Mammalian kidney development: principles, progress, and projections. Cold Spring Harbor Perspectives in Biology, 2012, 4(5):a008300.
doi: 10.1101/cshperspect.a008300 pmid: 22550230
[2] Costantini F, Kopan R.Patterning a complex organ: branching morphogenesis and nephron segmentation in kidney development. Developmental Cell, 2010, 18(5): 698-712.
doi: 10.1016/j.devcel.2010.04.008 pmid: 20493806
[3] Dressler G R.Advances in early kidney specification, development and patterning. Development, 2009, 136(23): 3863-3874.
doi: 10.1242/dev.034876 pmid: 19906853
[4] Faa G, Gerosa C, Fanni D, et al.Morphogenesis and molecular mechanisms involved in human kidney development. Journal of Cellular Physiology, 2012, 227(3): 1257-1268.
doi: 10.1002/jcp.22985 pmid: 21830217
[5] Rumballe B, Georgas K, Wilkinson L, et al.Molecular anatomy of the kidney: what have we learned from gene expression and functional genomics? Pediatric Nephrology, 2010, 25(6): 1005-1016.
doi: 10.1007/s00467-009-1392-6 pmid: 20049614
[6] Vainio S, Lin Y.Coordinating early kidney development: lessons from gene targeting. Nature Reviews Genetics, 2002, 3(7): 533-543.
doi: 10.1038/nrg842 pmid: 12094231
[7] Schroder M, Kaufman R J.The mammalian unfolded protein response. Annual Review of Biochemistry, 2005, 74: 739-789.
doi: 10.1146/annurev.biochem.73.011303.074134
[8] Smith W W, Jiang H, Pei Z, et al.Endoplasmic reticulum stress and mitochondrial cell death pathways mediate A53T mutant alpha-synuclein-induced toxicity. Human Molecular Genetics, 2005, 14(24): 3801-3811.
doi: 10.1093/hmg/ddi396
[9] Yamamoto J, Kamata S, Miura A, et alI. Differential adaptive responses to 1- or 2-day fasting in various mouse tissues revealed by quantitative PCR analysis. FEBS Open Bio, 2015, 5: 357-368.
doi: 10.1016/j.fob.2015.04.012 pmid: 4420774
[10] Lenz B, Bleich S, Beutler S, et al.Homocysteine regulates expression of Herp by DNA methylation involving the AARE and CREB binding sites. Experimental Cell Research, 2006, 312(20): 4049-4055.
doi: 10.1016/j.yexcr.2006.09.004 pmid: 17020760
[11] Valerius M T P L, Witte D P, Potter S S. Microarray analysis of novel cell lines representing two stages of metanephric mesenchyme differentiation. Mech Dev, 2002, 110(1-2): 151-164.
doi: 10.1016/S0925-4773(01)00581-0 pmid: 11744376
[12] Davidson A J. Mouse Kidney Development//StemBook. Cambridge: The President and Fellows of Harvard College, 2009:7-9.
[13] Kokame K, Agarwala K L, Kato H, et al.Herp, a new ubiquitin-like membrane protein induced by endoplasmic reticulum stress. J Biol Chem, 2000, 275(42): 32846-32853.
doi: 10.1074/jbc.M002063200 pmid: 10922362
[14] Hao L, Vassena R, Wu G, et al.The unfolded protein response contributes to preimplantation mouse embryo death in the DDK syndrome. Biol Reprod, 2009, 80(5): 944-953.
doi: 10.1095/biolreprod.108.072546 pmid: 2723760
[15] Kispert A V S, McMahon A P. Wnt-4 is a mesenchymal signal for epithelial transformation of metanephric mesenchyme in the developing kidney. Development, 1998, 125(21): 4225-4234.
doi: 10.1007/s004290050194 pmid: 9753677
[16] Hori O, Ichinoda F, Yamaguchi A, et al.Role of Herp in the endoplasmic reticulum stress response. Genes Cells, 2004, 9(5): 457-469.
doi: 10.1111/gtc.2004.9.issue-5
[17] Zhong Q, Zhou B, Ann D K, et al.Role of endoplasmic reticulum stress in epithelial-mesenchymal transition of alveolar epithelial cells: effects of misfolded surfactant protein. Am J Respir Cell Mol Biol, 2011, 45(3): 498-509.
doi: 10.1165/rcmb.2010-0347OC
[18] Shah P P D T, Siskind L J, Beverly L J. Common cytotoxic chemotherapeutics induce epithelial-mesenchymal transition (EMT) downstream of ER stress. Oncotarget, 2017, 8(14): 22625-39.
doi: 10.18632/oncotarget.15150 pmid: 5410250
[1] 李世荣,陈阳琴,张春盼,齐文杰. RS4651通过上调SMAD7抑制小鼠肝细胞AML12的EMT作用[J]. 中国生物工程杂志, 2021, 41(7): 1-9.
[2] 路玉祥,李元,方丹丹,王学博,杨万鹏,楚元奎,杨华. MiR-5047在乳腺癌细胞增殖迁移中的作用及表达调控*[J]. 中国生物工程杂志, 2021, 41(4): 9-17.
[3] 杨丹,田海山,李校堃. 成纤维细胞生长因子5的研究进展 *[J]. 中国生物工程杂志, 2020, 40(3): 117-124.
[4] 刘善辉,何魏,马文斐,李兰兰,卢建中,陶燕,张静,付生军. KLF8表达修饰及致瘤机制研究进展[J]. 中国生物工程杂志, 2019, 39(8): 80-85.
[5] 苟理尧,刘梦瑶,夏菁,万群,孙恃雷,唐敏,张彦. 骨形成蛋白9对人膀胱癌BIU-87细胞增殖和迁移的影响[J]. 中国生物工程杂志, 2018, 38(5): 10-16.
[6] 冯源, 唐云, 徐蕾, 谭海刚. 海藻多糖通过下调肝癌细胞Hep3B糖酵解途径抑制细胞增殖和迁移[J]. 中国生物工程杂志, 2017, 37(9): 31-40.
[7] 梁姗, 蒋子川, 冯均. 胭脂萝卜花青素提取及对NCI-N87细胞增殖侵袭的影响[J]. 中国生物工程杂志, 2017, 37(11): 101-108.
[8] 汤志雄, 苟德明. miRNA调控成肌分化的研究进展[J]. 中国生物工程杂志, 2017, 37(10): 103-110.
[9] 秦海霞, 崔红凯, 潘莹, 户瑞丽, 朱利红, 王世进. miR-335靶向Rho相关卷曲螺旋形成蛋白激酶1对卵巢癌细胞增殖的影响[J]. 中国生物工程杂志, 2016, 36(6): 24-31.
[10] 刘雪杰, 林巍然, 唐立春, 孙薇, 魏汉东, 姜颖. 慢病毒载体介导RAB27A基因过表达对人HepG2肝癌细胞增殖的影响[J]. 中国生物工程杂志, 2014, 34(9): 16-23.
[11] 权美玉, 郭强, 张坤水, 方瑞, 李翠琳, 杜军. 稳定高表达和干扰Nodal基因黑色素瘤细胞株构建及EMT表型鉴定[J]. 中国生物工程杂志, 2014, 34(3): 1-8.
[12] 陈丽, 曹莹. PKA对斑马鱼前肾发育的影响及其机制研究[J]. 中国生物工程杂志, 2014, 34(10): 41-48.
[13] 季辉, 王琦, 仲蕾, 魏文祥. URI基因稳定表达对肝癌SMMC-7721细胞增殖能力的影响[J]. 中国生物工程杂志, 2013, 33(6): 1-6.
[14] 吴琛, 田焕娜, 王媛媛, 刘芳铭, 张晓康, 李秦剑, 谢园园. GALNT14对人乳腺癌MCF-7细胞迁移的影响[J]. 中国生物工程杂志, 2012, 32(07): 8-15.
[15] 李晓娜, 宋关斌, 罗庆. 机械拉伸对骨髓间充质干细胞迁移行为的影响[J]. 中国生物工程杂志, 2012, 32(05): 1-6.
主管：中国科学院 　主办：中国科学院文献情报中心 　中国生物技术发展中心 　中国生物工程学会