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五羟色胺受体基因和五羟色胺转运体基因交互作用与青年人自杀未遂行为的关系
刘微1, 冉柳毅2, 靳佳佳1, 王苏亚1, 吴同1, 艾明1, 陈建梅1, 况利1     
1. 401331 重庆,重庆医科大学附属大学城医院心理卫生中心;
2. 400016重庆,重庆医科大学附属第一医院心理卫生中心
[摘要] 目的 探讨五羟色胺(5-hydroxy tryptamine,5-HT)系统中五羟色胺1B受体(5-hydroxy tryptamine 1B receptor,5-HTR1B)基因和五羟色胺转运体(5-hydroxy tryptamine transporter,5-HTT)基因的交互作用对青年人自杀未遂行为的影响。方法 选取5-HTR1B基因rs6296和5-HTT基因rs1042173、rs140701、rs2066713多态性位点,采用iMLDRTM多重SNP分型技术对病例组(70例自杀未遂青年人)和对照组(112例健康青年人)进行检验。采用广义多因子降维法(generalized multifactor dimensionality reduction,GMDR)分析基因-基因交互作用。结果 5-HTT基因rs1042173、rs140701、rs2066713的基因型频率和等位基因频率在病例组和对照组中分布差异无统计学意义(P>0.05)。5-HTR1B rs6296等位基因频率在病例组和对照组中分布差异无统计学意义(P>0.05),5-HTR1B rs6296病例组基因型频率(CC 31.4%、CG 31.4%、GG 37.1%)和对照组基因型频率(CC 19.8%、CG 53.2%、GG 27%)分布差异有统计学意义(χ2 =8.327,P=0.016),经Bonferroni校正后差异无统计学意义(P>0.05)。GMDR分析结果显示,5-HTT与5-HTR1B基因交互作用存在于两位点模型(rs1042173、rs6296,测试样本精确度0.6185,交叉验证一致性10/10,P值为0.0210~0.0220)、三位点模型(rs1042173、rs140701、rs6296,测试样本精确度0.644 5,交叉验证一致性10/10,P值为0.003 0~0.004 0)、四位点模型(rs1042173、rs140701、rs2066713、rs6296,测试样本精确度0.6120,交叉验证一致性9/10,P值为0.0320~0.0330)。结论 5-HTR1B和5-HTT基因存在交互作用,并可能与青年人的自杀未遂行为有关。
[关键词] 青年人     自杀未遂     基因-基因交互作用     五羟色胺受体     五羟色胺转运体    
Interaction between 5-hydroxytryptamine receptor 1B gene and serotonin transporter gene is potentially associated with suicide attempts in youths
LIU Wei1 , RAN Liuyi2 , JIN Jiajia1 , WANG Suya1 , WU Tong1 , AI Ming1 , CHEN Jianmei1 , KUANG Li1     
1. Center for Mental Health Services, University-Town Hospital of Chongqing Medical University, Chongqing, 401331;
2. Center for Mental Health Services, the First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016, China
Supported by the General Program of National Natural Science Foundation of China (81671360)
Corresponding author: KUANG Li, E-mail:kuangli0308@163.com
[Abstract] Objective To explore the interaction between 5-hydroxytryptamine (5-HT) receptor 1B (5-HTR1B) gene and 5-HT transporter (5-HTT) gene in the 5-HT system and investigate how this interaction is associated with attempted suicide in youths. Methods iMLDRTM multiplex single nucleotide polymorphism (SNP) typing was used to examine the SNPs at one locus (rs6296) of 5-HTR1B gene and 3 loci (rs1042173, rs140701 and rs2066713) of 5-HTT gene in 70 youths (aged 15-24 years) with attempted suicide (case group) and 112 age-matched healthy subjects (control group). The interaction between 5-HTR1B gene and 5-HTT gene was analyzed using generalized multifactor dimensionality reduction (GMDR). Results No significant differences were found between the case and control groups in the genotype frequencies or allele frequencies at the loci rs1042173, rs140701, or rs2066713 in 5-HTT gene (P>0.05). The allele frequency of rs6296 in 5-HTR1B gene was similar between the 2 groups (P>0.05), but the CC, CG, and GG genotype frequencies of 5-HTR1B rs6296 differed significantly between the case group (31.4%, 31.4%, and 37.1%, respectively) and the control group (19.8%, 53.2%, and 27%, respectively) (Chi-square=8.327, P=0.016); these differences, however, were not statistically significant after Bonferroni correction (P>0.05). GMDR analysis indicated the presence of interactions between 5-HTT gene and 5-HTR1B gene in the two-site model (rs1042173 and rs6296) with a test accuracy of 0.6185 and a cross validation consistency of 10/10 (P=0.021 0-0.022 0), in the three-site model (rs1042173, rs140701, and rs6296) with a test accuracy of 0.644 5 and cross validation consistency of 10/10 (P=0.0030-0.0040) as well as in the four-site model (rs1042173, rs140701, rs2066713, and rs6296) with a test accuracy of 0.6120 and a cross validation consistency of 10/10 (P=0.0320-0.0330). Conclusion There are interactions between 5-HTR1B gene and 5-HTT gene, which may be associated with attempted suicide in youths.
[Key words] youths     attempted suicide     gene-gene interaction     5-hydroxytryptamine receptor 1B     5-hydroxytryptamine transporter    

自杀是指个体蓄意或自愿采取各种手段结束自己生命的行为。根据自杀的结果,分为自杀意念、自杀未遂和自杀完成3种形态。根据世界卫生组织(WHO)2014年调查显示,每年有80万以上的人死于自杀,有自杀企图的人高达10~20倍[1]。在欧洲,自杀是14~24岁年龄组的第2大死因[2]。家系[3]、双生子[4]和寄养子[5]研究均支持遗传是自杀行为的风险因素。一项研究表明,自杀行为的遗传度估计为30%~55%,并在很大程度上独立于其他精神疾病[6]。针对自杀的候选基因的研究大多和5-HT系统相关。研究表明,自杀行为可能与5-羟色胺(5-hydroxy tryptamine,5-HT)的水平降低有关[7]

5-羟色胺1B受体(5-hydroxy tryptamine 1B receptor,5-HTR1B)是突触前的自身受体,调节5-HT的定量释放,位于6q14.1,在基因序列上发现大量的单核苷酸多态性,其中最受关注的是错义突变G861C(rs6296),位于外显子区域。既往的研究已经显示,该变异会增加5-HTR1B的结合性,从而导致其功能改变[8]

5-羟色胺转运体(5-hydroxy tryptamine transporte,5-HTT)在突触中作为信号传导的调节器,在血清素系统中起重要作用。5-HTT位于17q11.1-q12,研究较多的是5-HTT基因启动子多态性(5-HTTLPR,rs4795541),其由短等位基因(S)和长等位基因(L)组成。文献[9-10]报道了S-等位基因与自杀相关联,也有的研究报道差异没有统计学意义。有研究证据表明了rs1042173功能,发现C等位基因破坏了具有miR-590-3p的保守结合位点核苷酸,导致5-HTT表达增加[11]

青年期是个体生理心理发育的特殊时期,大量研究表明青年人自杀与成年人自杀可能具有不同的神经生物学机制[12-14]。课题组前期对重庆市大学生自杀未遂行为的调查分析结果显示,性别、家庭经济情况等对自杀未遂行为有重要影响[15]。本研究以青年人为研究对象,通过基因-基因交互作用分析,探讨5-HTT基因和5-HTR1B基因和自杀未遂行为的关系。

1 对象与方法 1.1 研究对象

病例组70例,选取的是2010年10月起参与网络心理健康调查的重庆市十余所高校大一新生,对调查结果有异常的学生进一步筛查并采集血样,并收集重庆医科大学附属第一医院及附属大学城医院心理卫生中心就诊患者的相关病例。入组标准:①汉族;②年龄15~24岁;③右利手;④小学及以上文化程度;⑤最近1年至少有过1次自杀未遂行为;⑥所有研究对象对本研究知情并同意参加,签署知情同意书。共入组患者70例。排除标准:有重大躯体疾病。

对照组112例,随机抽取同期参与网络心理健康调查的重庆市十余所高校的健康大一新生。入组标准: ①汉族;②年龄15~24岁;③右利手;④小学及以上文化程度;⑤无自杀意念及自杀自伤行为;⑥所有研究对象对本研究知情并同意参加,签署知情同意书。

1.2 方法

1.2.1 基因组DNA提取

采用EDTA抗凝管采集研究对象外周血,每例研究对象采集血样2份,每份2 mL,采用常规盐析法提取DNA。上海天昊生物技术有限公司研发的iMLDRTM多重SNP分型技术进行分型,基因型判读时采用盲法,由2名分别进行基因判读,对判读结果不一样的样本进行重新检测。

1.3 统计学方法

一般资料中年龄采用t检验,性别采用χ2检验。通过Online softwareSHEsis(http://analysis.biox.on)统计软件对病例组和对照组基因型频率分布进行Hardy-Weinberg平衡吻合度检验。采用χ2检验对2组等位基因和基因型频率进行统计分析,进行Bonferroni校正。运用广义多因子降维法软件(generalized multifactor dimensionality reduction,GMDR)分析基因-基因的交互作用,测试样本精确度和交叉验证一致性最高的模型为最佳模型。并采用MDRpt(MDR置换检验)将P值进行1 000次置换检验。检验水准:α=0.05。

2 结果 2.1 一般资料比较

70例病例组中男性38例,女性32例,年龄(18.91±2.68)岁。112例对照组中男性59例,女性53例,年龄(19.09±1.31)岁。病例组与对照组相比,年龄(t=0.510,P=0.611)、性别(χ2=0.450,P=0.200)差异均无统计学意义。

2.2 Hardy-Weinberg平衡检验

对照组5-HTR1B基因1个多态位点rs6296及5-HTT基因3个多态位点rs1042173、rs140701、rs2066713的基因型频率均符合Hardy-Weinberg平衡定律(P> 0.05),样本具有群体代表性。病例组5-HTT rs1042173、rs140701、rs2066713的基因型频率符合Hardy-Weinberg平衡定律(P>0.05),病例组5-HTR1B rs6296基因型频率不符合Hardy-Weinberg平衡定律(χ2=9.550,df=1,P=0.002)。

2.3 基因型和等位基因分布的比较

各基因位点基因型和等位基因在病例组和对照组中频率分布情况见表 1。结果显示5-HTT rs1042173、rs140701、rs2066713基因型频率和等位基因频率在病例组和对照组中分布差异均无统计学意义(P>0.05)。5-HTR1B rs6296等位基因频率在病例组和对照组中分布差异无统计学意义(P>0.05),5-HTR1B rs6296的病例组和对照组基因型频率分布差异有统计学意义(χ2 =8.327,df=2,P=0.016),经Bonferroni校正后差异无统计学意义(P>0.05)。

表 1 病例组与对照组基因型和等位基因分布的比较
SNP 组别 基因型 χ2 P 等位基因 χ2 P
A/A A/C C/C A C
rs1042173 病例组 5(0.071) 23(0.329) 42(0.600) 2.337 0.311 33(0.236) 107(0.764) 0.110 0.740
对照组 3(0.027) 43(0.387) 65(0.586) 49(0.221) 173(0.779)
rs140701 C/C C/T T/T C T
病例组 5(0.071) 22(0.314) 43(0.614) 2.613 0.271 32(0.229) 108(0.771) 0.030 0.861
对照组 3(0.027) 43(0.387) 65(0.586) 49(0.221) 173(0.779)
rs2066713 A/G G/G A G
病例组 14(0.200) 56(0.800) 0.458 0.499 14(0.100) 126(0.900) 0.400 0.527
对照组 27(0.243) 84(0.757) 27(0.122) 195(0.878)
rs6296 C/C C/G G/G C G
病例组 22(0.314) 22(0.314) 26(0.371) 8.327 0.016 66(0.471) 74(0.529) 0.019 0.890
对照组 22(0.198) 59(0.532) 30(0.270) 103(0.464) 119(0.536)
rs1042173位点对照组、rs140701位点对照组、rs2066713位点对照组、rs6296位点对照组分别均有1例未检出

2.4 5-HTT基因和5-HTR1B基因交互作用分析

GMDR分析结果显示,5-HTT rs1042173、rs140701、rs2066713和5-HTR1B rs6296在3种不同位点数的模型中存在基因交互作用,分别是两位点模型(rs1042173、rs6296)、三位点模型(rs1042173、rs140701、rs6296)、四位点模型(rs1042173、rs140701、rs2066713、rs6296)。

5-HTT rs1042173、rs140701和5-HTR1B rs6296组成的三位点模型为最优模型,经1 000次置换检验差异有统计学意义(P<0.01,表 2)。

表 2 5-HTT基因和5-HTR1B基因snp间的GMDR分析
位点数 最佳模型 训练样本精确度 测试样本精确度 交叉验证一致性 1 000次置换检验的P
1 X4 0.610 7 0.611 0 10/10 0.033 0~0.034 0
2 X1,X4 0.630 2 0.618 5 10/10 0.021 0~0.022 0
3 X1,X2,X4 0.666 8 0.644 5 10/10 0.003 0~0.004 0
4 X1a,X2b,X3c,X4d 0.672 7 0.612 0 9/10 0.032 0~0.033 0
a:5-HTT rs1042173;b:5-HTT rs140701;C:5-HTT rs2066713;d:5-HTR1B rs6296

3 讨论

自杀行为的神经生物学研究主要集中在5-HT系统上,包括发现自杀未遂患者脑脊液中检测到5-羟色胺代谢产物5-羟色胺脱乙酸(5-HIAA)水平下降[16],以及在青少年自杀死亡者前额皮层和海马中检测到高表达的五羟色胺2A受体,都将证据指向5-HT系统。鉴于5-HT系统对自杀的重要影响,近年来针对5-HTT和5-HTR1B基因做了大量研究。最近,MURPHY等[17]的研究发现rs6296与自杀企图之间地显着关联。研究指出,自杀意念背后的机制可能与rs6296-C等位基因降低5-HTR1B mRNA水平有关,5-HTR1B mRNA水平较低的人可能会更容易生气,表现出更多的敌意和攻击性,从而产生自杀意念[18-19]。SENEVIRATNE等[20]在HeLa细胞培养物中发现,rs1042173小G等位基因携带者具有较高的5-HTT mRNA和蛋白质水平。因此,我们未来的研究应该不仅仅是启动子区域,而是参与遗传的整个区域。

自杀行为的遗传易感性已逐渐被广泛认识,目前针对自杀相关研究主要关注影响神经递质功能的单个位点。自杀属于一种复杂性行为,全基因组关联研究发现自杀行为与多个基因有关[21],单个基因对该行为的作用是很微弱的,更可能受到多个基因交互作用的影响。本研究发现,5-HTT基因rs1042173、rs140701、rs2066713位点和5-HTR1B基因rs6296位点的基因型和等位基因频率与青年自杀未遂人群无显著性关联。通过GMDR分析结果显示,5-HTT基因和5-HTR1B基因存在多个位点交互模型,由此提示5-HTT和5-HTR1B基因之间的交互作用可能与青年人的自杀未遂风险有关,与复杂性疾病的多基因交互作用理论是一致的。

本研究采用GMDR对青年自杀未遂行为进行基因交互模的建立和分析。2001年RITCHIE等[22]首次提出了MDR,MDR是一种非参数、无须遗传模式的研究方法,适用于病例对照研究设计的交互分析,其优点在于大大降低了建模所需的自由度,弥补了Logistic回归在处理高阶交互作用时的局限性。2007年LOU等[23]提出了一种基于MDR基本原理的扩展方法——GMDR,使其应用范围和预测准确率得到了进一步改善。目前,越来越多的研究运用到MDR和GMDR,包括最近两项关于精神疾病及自杀的研究[24-25]

本研究选用青年人为研究对象,组内同质性高,采用GMDR探究青少年自杀未遂行为的遗传标记,考虑到了基因-基因相互作用对自杀未遂行为的影响,为青年人自杀的遗传学研究提供了一定的参考价值。

本研究不足之处在于样本量较少,选择的基因位点不全面,未考虑到环境的影响。未来的研究应该扩大样本量,结合环境因素进行基因-环境交互分析,更加全面的分析自杀的相关危险因素,增加纵向研究,进一步探讨自杀行为的发生、发展机制,对未来预测和干预自杀高风险人群提供更多的帮助。

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http://dx.doi.org/10.16016/j.1000-5404.201711199
中国人民解放军总政治部、国家科技部及国家新闻出版署批准,
由第三军医大学主管、主办

文章信息

刘微, 冉柳毅, 靳佳佳, 王苏亚, 吴同, 艾明, 陈建梅, 况利.
LIU Wei, RAN Liuyi, JIN Jiajia, WANG Suya, WU Tong, AI Ming, CHEN Jianmei, KUANG Li.
五羟色胺受体基因和五羟色胺转运体基因交互作用与青年人自杀未遂行为的关系
Interaction between 5-hydroxytryptamine receptor 1B gene and serotonin transporter gene is potentially associated with suicide attempts in youths
第三军医大学学报, 2018, 40(12): 1115-1119
Journal of Third Military Medical University, 2018, 40(12): 1115-1119
http://dx.doi.org/10.16016/j.1000-5404.201711199

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收稿: 2017-11-23
修回: 2018-03-15

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