沉默XIST通过上调miR-488-3p抑制宫颈鳞状细胞癌细胞的顺铂耐药性-现代医学

沉默XIST通过上调miR-488-3p抑制宫颈鳞状细胞癌细胞的顺铂耐药性

单位：1. 西安交通大学第二附属医院妇产科, 陕西西安 710004;
2. 西安交通大学第二附属医院手术室, 陕西西安 710004;
3. 西安交通大学第二附属医院肿瘤科, 陕西西安 710004;
4. 西安交通大学第二附属医院小儿外科, 陕西西安 710004

关键词：宫颈鳞状细胞癌细胞顺铂耐药性长链非编码RNA XIST 微小RNA-488-3p RNA拉下实验

分类号：R737.33

出版年·卷·期（页码）：2019·47·第四期（415-421）

摘要：

目的：探索长链非编码RNA XIST对宫颈鳞状细胞癌细胞顺铂耐药性的作用及机制。方法：建立宫颈鳞状细胞癌细胞顺铂耐药系CASKI/cDDP；构建沉默XIST的慢病毒载体（si-XIST）及miR-488-3p沉默载体miR-inhibitor；将si-XIST单独稳定转入CASKI/cDDP细胞或将si-XIST与miR-inhibitor共转入CASKI/cDDP细胞，qRT-PCR检测XIST、miR-488-3p水平变化；MTT法检测细胞存活率变化，蛋白质印迹法检测多药耐药基因1（MDR1）表达变化，RNA拉下实验确定miR-488-3p与XIST的结合关系。结果：与CASKI相比，CASKI/cDDP中XIST表达升高（P<0.05）；si-XIST转染CASKI/cDDP细胞48 h后，XIST的水平显著下调（P<0.05）、细胞的存活率下降（P<0.05）、MDR1表达水平下降（P<0.05）。CASKI/cDDP细胞XIST沉默可增加miR-488-3p水平（P<0.05）；生物信息学预测宫颈癌关键标志物microRNA-488-3p与XIST结合；RNA拉下实验确认XIST直接结合miR-488-3p。CASKI/cDDP细胞共转染si-XIST与miR-488-3p-inhibitor；miR-488-3p-inhibitor部分逆转si-XIST的功能，提高CASKI/cDDP在顺铂处理下的细胞活性（P<0.05）。结论：沉默XIST通过上调miR-488-3p抑制宫颈鳞状细胞癌细胞的顺铂耐药性。

Objective:To investigate the effect and mechanism of long non-coding(lnc) RNA XIST on the cisplatin(cDDP) resistance in cervical carcinoma cells. Methods:CASKI/cDDP, the cDDP resistant cell line of cervix squamous cell carcinoma was constructed. Additionally, the lentivirus vector for silencing XIST(si-XIST) and miR-488-3p silencing vector(miR-inhibitor) were constructed. Then, si-XIST was stably transfected into CASKI/cDDP cell alone or si-XIST and miR-inhibitor were co-transfected into CASKI/cDDP cell. The changes of XIST and miR-488-3p levels were tested by qRT-PCR. Cell proliferation activity was detected by MTT. The expression of multidrug resistance gene 1(MDR1) was detected by western blot. The relationship between miR-488-3p and XIST was studied by RNA pull down assay. Results:The expression of XIST in CASKI/cDDP was higher than that in CASKI(P<0.05). The expression of XIST and MDR1 and the cell proliferation activity in si-XIST-transfected CASKI/cDDP cells were downregulated(P<0.05). MiR-488-3p, a key marker of cervical cancer, was predicted to bind with XIST by bioinformatics prediction. RNA pull down assay confirmed that XIST binded directly to miR-488-3p. Co-transfection of si-XIST and miR-inhibitor into CASKI/cDDP cells partially reversed the function of si-XIST, and increased the cell viability of CASKI/cDDP treated with cisplatin(P<0.05). Conclusion:Silencing XIST inhibits the cisplatin resistance in cervical squamous cell carcinoma by up-regulating miR-488-3p.

参考文献：

[1] CHEN W,ZHENG R,BAADE P D,et al.Cancer statistics in China,2015[J].CA-Cancer J Clin,2016,66(2):115-132.
[2] ZHOU M,DIAO Z,YUE X,et al.Construction and analysis of dysregulated lncRNA-associated ceRNA network identified novel lncRNA biomarkers for early diagnosis of human pancreatic cancer[J].Oncotarget,2016,7(35):56383-56394.
[3] 陈新莲,王和,张雪梅,等.人宫颈癌顺铂耐药细胞系SiHa/cDDP的建立[J].四川大学学报(医学版),2012,43(2):151-155.
[4] CHANG S,CHEN B,WANG X,et al.Long non-coding RNA XIST regulates PTEN expression by sponging miR-181a and promotes hepatocellular carcinoma progression[J].Bmc Cancer,2017,17(1):248.
[5] SONG J,KIM D,JIN E J.MicroRNA-488 suppresses cell migration through modulation of the focal adhesion activity during chondrogenic differentiation of chick limb mesenchymal cells[J].Cell Biol Int,2011,35(2):179-185.
[6] WEN Q,YAN L,LYU H,et al.Long noncoding RNA GAS5,which acts as a tumor suppressor via microRNA 21,regulates cisplatin resistance expression in cervical cancer[J].Int J Gynecol Cancer,2017,27(6):1096.
[7] CAO L,CHEN J,OU B,et al.GAS5 knockdown reduces the chemo-sensitivity of non-small cell lung cancer(NSCLC) cell to cisplatin(DDP) through regulating miR-21/PTEN axis[J].Biomed Pharmacother,2017,93:570-579.
[8] 刘丽丹,黄浩梁.宫颈癌组织长链非编码RNAXIST、小分子RNAmiR-101-3p表达变化及其意义[J].山东医药,2017,57(35):44-46.
[9] KOBAYASHI R,MIYAGAWA R,YAMASHITA H,et al.Increased expression of long non-coding RNA XIST predicts favorable prognosis of cervical squamous cell carcinoma subsequent to definitive chemoradiation therapy[J].Oncol Lett,2016,12(5):3066.
[10] SUN W,ZU Y,FU X,et al.Knockdown of lncRNA-XIST enhances the chemosensitivity of NSCLC cells via suppression of autophagy[J].Oncol Rep,2017,38(6):3347-3354.
[11] SUN J,PAN L M,CHEN L B,et al.LncRNA XIST promotes human lung adenocarcinoma cells to cisplatin resistance via let-7i/BAG-1 axis[J].Cell Cycle,2017,16(21):2100-2107.
[12] DU P,ZHAO H,PENG R,et al.LncRNA-XIST interacts withmiR-29cto modulate the chemoresistance of glioma cell to TMZ through DNA mismatch repair pathway[J].Bioscience Rep,2017,37(5):BSR20170696.
[13] LI Y,LI S,LUO Y,et al.LncRNA PVT1 regulates chondrocyte apoptosis in osteoarthritis by acting as a sponge for miR-488-3p[J].DNA Cell Biol,2017,36(7):571-580.
[14] SHEN C J,CHENG Y M,WANG C L.LncRNA PVT1 epigenetically silences miR-195 and modulates EMT and chemoresistance in cervical cancer cells[J].J Drug Target,2017,25(7):637.
[15] NING L,YUE M,LI M,et al.MicroRNA-488-3p sensitizes malignant melanoma cells to cisplatin by targeting PRKDC[J].Cell Biol Int,2017,41(6):622-629.
[16] ARVIDSSON Y,REHAMMAR A,BERGSTROM A,et al.miRNA profiling of small intestinal neuroendocrine tumors defines novel molecular subtypes and identifies miR-375 as a biomarker of patient survival[J].Mod Pathol,2018,31(8):1302-1317.
[17] XUE W,CHEN J,LIU X,et al.PVT1 regulates the malignant behaviors of human glioma cells by targeting miR-190a-5p and miR-488-3p[J].Biochim Biophys Acta,2018,1864(5):1783-1794.

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