MicroRNA4293 基因多態(tài)性與女性肝癌易感性相關

蔡敏　鄭萬威　張駿　程烽濤　李力

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·論著·

MicroRNA4293 基因多態(tài)性與女性肝癌易感性相關

蔡敏鄭萬威張駿程烽濤李力

200090上海同濟大學附屬楊浦醫(yī)院消化科(蔡敏，程烽濤，李力)；復旦大學附屬華山醫(yī)院消化科(鄭萬威，張駿)

【摘要】目的研究microRNA-4293的SNP(rs12220909)與原發(fā)性肝癌易感性的關系。方法收集人體血液樣本1788份，樣本取自健康對照者884名，原發(fā)性肝癌患者904例。抽提血液樣本中基因組DNA，應用MassARRAY法對microRNA-4293的SNP(rs12220909)位點進行基因分型；應用二元邏輯回歸法分析microRNA-4293的SNP(rs12220909)位點與肝癌易感性之間的關系。結果女性肝癌患者microRNA-4293的SNP(rs12220909)中CC基因型較健康對照組顯著增加(P=0.04)，在HBV相關肝癌中顯著性進一步增加(P=0.02)，提示CC基因可能增加女性肝癌尤其是HBV相關肝癌的易感性。而在男性患者中，microRNA-4293的SNP(rs12220909)與肝癌易感性并不相關(P=0.33)。結論microRNA-4293的SNP(rs12220909)與女性肝癌易感性相關。

【關鍵詞】原發(fā)性肝癌；易感性；MicroRNA-4923；SNP(rs12220909)

原發(fā)性肝癌是最常見的肝臟惡性腫瘤，具有發(fā)病率高、病死率高、易復發(fā)、易轉移的特點，已嚴重威脅人類的健康[1,2]。由于大部分肝癌患者確診時病情已進展至晚期，預后較差。盡管肝癌有多種治療方法，如手術切除、介入治療、肝移植等，但因肝功能異常和腫瘤過大等因素，只有5%～15%的患者適合手術切除治療[3]。

microRNA(miRNA)是一類由22 個核苷酸組成的非編碼單鏈RNA 分子，參與轉錄后基因表達調控。miRNA通過對靶標mRNA轉錄后去穩(wěn)定和(或)抑制其翻譯，來調節(jié)細胞增殖、分化、遷移和凋亡等功能[4]。研究發(fā)現(xiàn)，多種與腫瘤相關的miRNA，其靶標基因與腫瘤的發(fā)生、發(fā)展相關[5]。單核苷酸多態(tài)性(single nucleotide polymorphisms，SNP)已成為研究復雜遺傳性疾病、藥物基因組學和人類進化的第三代遺傳標記。因為腫瘤相關基因的SNP常影響該基因的表達及功能[6]，能夠改變腫瘤易感性，故此類基因SNP常作為生物學標志物應用于腫瘤研究[7]。針對肝癌miRNA表達譜的研究發(fā)現(xiàn)，miRNA的微小變化可引起靶mRNA翻譯的顯著變化，從而影響到肝癌的發(fā)生和進展[8]。

mir-4293位于第10號染色體上，目前研究發(fā)現(xiàn)mir-4293的SNP(rs12220909)與食管鱗狀細胞癌易感性相關[9]，但關于該miRNA的研究較少，其與肝癌的相關性仍未知。

本研究對血液樣本進行mir-4293的SNP(rs12220909)檢測，并分析該位點與原發(fā)性肝癌易感性的關系。

資料和方法

一、樣本收集

共收集人體血液樣本1788份。樣本取自健康者884名，原發(fā)性肝癌患者904例，其中HBV相關肝癌701例。健康人群取自江蘇泰州地區(qū)，無腫瘤及肝炎病史；原發(fā)性肝癌病例取自復旦大學附屬華山醫(yī)院、同濟大學附屬楊浦醫(yī)院及東方肝膽醫(yī)院，所有肝癌患者均經病理確診。

二、核酸提取和基因分型

應用血液基因組DNA抽提試劑盒(Axygen Biosciences, Union City, USA)從血液樣本中抽提基因組DNA。在測序分析前，應用NanoDrop 2000c分光光度計測定DNA濃度，并用電泳法測定其純度。應用MassARRAY法對mir-4293的SNP(rs12220909)位點進行基因分型。mir-4293的SNP(rs12220909)的染色體起止位點為chr10：14425199-14425276，SNP位置為14425221(mat)，擴增引物為ACGTTGGATGGCTGGTGACATTGCTAATTCA-CGTTGGATGGGAGAGGGGAAATCCTATTT，延伸引物為：CCCAAGGCTGTTCCTGTCA。

三、數(shù)據(jù)分析

應用二元邏輯回歸法分析mir-4293的SNP(rs12220909)位點與肝癌易感性之間的關系。P<0.05為差異有統(tǒng)計學意義。所有統(tǒng)計學分析采用SPSS 16.0軟件。

結果

健康對照組884名(男性644例，女性240例)，原發(fā)性肝癌患者904例(男性742例，女性162例)。研究對象的基本信息見表1。應用二元邏輯回歸法，校正年齡、吸煙和飲酒等其他因素影響后發(fā)現(xiàn)，mir-4293的SNP(rs12220909)位點的各基因型在肝癌組與對照組差異無統(tǒng)計學意義(P=0.07)，見表2。在女性患者中，肝癌組CC基因型較對照組顯著增加(P=0.04)，在HBV相關肝癌中顯著性進一步增加(P=0.02)，見表3；而在男性患者中各基因型在肝癌組與對照組差異無統(tǒng)計學意義，見表4。

表1　研究對象的基本信息

表2　Mir-4293的SNP(rs12220909)與肝癌易感性分析

表3　女性患者中Mir-4293的SNP(rs12220909)與肝癌易感性分析

表4　男性患者中Mir-4293的SNP(rs12220909)與肝癌易感性分析

討論

早期診斷和有效預防肝癌的已成為世界性的醫(yī)療問題，在目前的臨床應用中迫切需要開發(fā)新型有效的標志物用于肝癌的篩查、早期診斷和判斷預后，并需積極尋找新的治療靶標與方法[10]。研究發(fā)現(xiàn)，miRNA表達紊亂在多種腫瘤，包括原發(fā)性肝癌的發(fā)生、發(fā)展中起到重要作用[11]。而且，一些miRNA在肝癌中特異性表達，并且與多種腫瘤基因如P53、PTENZ和RA等組成了調控網絡，參與肝癌的發(fā)生、侵襲和轉移[12-14]。miRNA的SNP能夠影響miRNA的成熟及miRNA與靶標基因的結合，從而影響靶標基因的功能，進一步影響肝癌、非小細胞肺癌等惡性腫瘤的易感性、治療反應及預后[15,16]。綜上，miRNA及其SNP位點具有極大的潛力成為肝癌篩查、早期診斷及預后判斷的新型標志物，同時為肝癌預防及治療的研究提供新的靶標。故對于miRNA的SNP與肝癌的研究就顯得尤為重要[17-19]。

本研究發(fā)現(xiàn)位于第10號染色體上mir-4293的SNP(rs12220909)與女性肝癌易感性相關，女性肝癌患者中CC基因型較健康對照組顯著增加(P=0.04)，在HBV相關肝癌中顯著性進一步增加(P=0.02)，提示CC基因可能增加女性肝癌尤其是HBV相關肝癌的易感性。而在男性患者中，mir-4293的SNP(rs12220909)與肝癌易感性并不相關。

綜上所述，mir-4293的SNP(rs12220909)中CC基因型能夠顯著增加女性原發(fā)性肝癌及HBV相關肝癌的易感性，而在男性患者中并無顯著作用。本研究可為肝癌發(fā)病性別差異的機制研究提供新方向，同時有助于推動miRNA SNP在肝癌早期診斷上的臨床應用。

參考文獻

[ 1 ]El-Serag HB, Rudolph KL. Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology, 2007, 132:2557-2576.

[ 2 ]Hou W, Bonkovsky HL. Non-coding RNAs in hepatitis C-induced hepatocellular carcinoma: dysregulation and implications for early detection, diagnosis and therapy. World J Gastroenterol 2013, 19:7836-7845.

[ 3 ]de Lope CR, Tremosini S, Forner A, et al. Management of HCC. J Hepatol, 2012, 56 (Suppl 1):S75-S87.

[ 4 ]Bartel DP. MicroRNAs: target recognition and regulatory functions. Cell, 2009, 136:215-233.

[ 5 ]Wang XW, Heegaard NH, Orum H: MicroRNAs in liver disease. Gastroenterology, 2012, 142:1431-1443.

[ 6 ]Chung CC, Chanock SJ. Current status of genome-wide association studies in cancer. Hum Genet, 2011, 130:59-78.

[ 7 ]Pastinen T, Ge B, Hudson TJ. Influence of human genome polymorphism on gene expression. Hum Mol Genet, 2006, 15:R9-R16.

[ 8 ]Selbach M, Schwanhausser B, Thierfelder N, et al. Widespread changes in protein synthesis induced by microRNAs. Nature, 2008, 455:58-63.

[ 9 ]Zhang P, Wang J, Lu T, et al. MiR-449b rs10061133 and miR-4293 rs12220909 polymorphisms are associated with decreased esophageal squamous cell carcinoma in a Chinese population. Tumour Biol, 2015, 36:8789-8795.

[10]Wang X, Zhang A, Sun H. Power of metabolomics in diagnosis and biomarker discovery of hepatocellular carcinoma. Hepatology, 2013, 57:2072-2077.

[11]Wen J, Friedman JR. miR-122 regulates hepatic lipid metabolism and tumor suppression. J Clin Invest, 2012, 122:2773-2776.

[12]Diaz G, Melis M, Tice A, et al. Identification of microRNAs specifically expressed in hepatitis C virus-associated hepatocellular carcinoma. Int J Cancer, 2013, 133:816-824.

[13]Chen JS, Wang Q, Fu XH, et al. Involvement of PI3K/PTEN/AKT/mTOR pathway in invasion and metastasis in hepatocellular carcinoma: Association with MMP-9. Hepatol Res, 2009, 39:177-186.

[14]Hsu HC, Tseng HJ, Lai PL, et al. Expression of p53 gene in 184 unifocal hepatocellular carcinomas: association with tumor growth and invasiveness. Cancer Res, 1993, 53:4691-4694.

[15]Li XD, Li ZG, Song XX, et al. A variant in microRNA-196a2 is associated with susceptibility to hepatocellular carcinoma in Chinese patients with cirrhosis. Pathology, 2010, 42:669-673.

[16]Qi P, Dou TH, Geng L, et al. Association of a variant in MIR 196A2 with susceptibility to hepatocellular carcinoma in male Chinese patients with chronic hepatitis B virus infection. Hum Immunol, 2010, 71:621-626.

[17]Zhang J, Wang R, Ma YY, et al. Association between single nucleotide polymorphisms in miRNA196a-2 and miRNA146a and susceptibility to hepatocellular carcinoma in a Chinese population. Asian Pac J Cancer Prev, 2013, 14:6427-6431.

[18]Zhou J, Yu L, Gao X, et al. Plasma microRNA panel to diagnose hepatitis B virus-related hepatocellular carcinoma. J Clin Oncol, 2011, 29:4781-4788.

[19]Zhang QH, Sun HM, Zheng RZ, et al. Meta-analysis of microRNA-183 family expression in human cancer studies comparing cancer tissues with noncancerous tissues. Gene, 2013, 527:26-32.

(本文編輯：賴榮陶)

通信作者：李力，Email: lli10437@sina.com

Corresponding author:LI Li, Email: lli10437@sina.com

(收稿日期：2016-03-31)

Genetic polymorphisms of microRNA-4293 and hepatocellular carcinoma susceptibility in female

CAIMin,ZHENGWan-wei,ZHANGJun,CHENGFeng-tao,LILi.

TongjiUniversitySchoolofMedicine,YangpuHospital,DepartmentofDigestiveDiseases,Shanghai200090,China

【Abstract】ObjectiveTo investigate the association between single nucleotide polymorphism (SNP) mapping to microRNA-4293 (rs12220909) and hepatocellular carcinoma (HCC) susceptibility. MethodsBlood samples were collected from 1788 cases, including 884 normal control cases and 904 HCC cases. Genomic deoxyribonucleic acid was extracted from blood samples, and SNPs mapping to microRNA-4293 (rs12220909) were detected by MassARRAY. Then association between genotype of rs12220909 and HCC susceptibility was analyzed by binary logistic regression. ResultsIn female HCC patients, CC genotype of rs12220909, especially in hepatitis B virus (HBV)-related HCC patients (P=0.02), was significantly higher than that in healthy control group (P=0.04). It indicated that CC genotype might be a risk factor for female HCC, especially for HBV-related HCC. However, this association did not exist in male patients (P=0.33). ConclusionGenotype of microRNA-4293 (rs12220909) is relevant to HCC in female patients.

【Key words】Hepatocellular carcinoma; Susceptibility; MicroRNA-4923; SNP(rs12220909)