【摘要】：目的：檢測(cè)HTR3A及HTR3B的21個(gè)標(biāo)記單核苷酸多態(tài)(tag SNP)在患者及對(duì)照者的等位基因及基因型分布,通過(guò)病例-對(duì)照關(guān)聯(lián)研究探討雙相障礙遺傳易感性,尋找雙相障礙的易感基因,探討HTR3A及HTR3BTag SNP基因多態(tài)性、遺傳不平衡性及單倍型與雙相障礙患者疾病特點(diǎn)及認(rèn)知功能的關(guān)聯(lián)性,進(jìn)一步驗(yàn)證認(rèn)知功能損害是否為雙相障礙的內(nèi)表型特征,為理論研究及臨床實(shí)踐提供重要的參考依據(jù),為雙相障礙早期識(shí)別提供理論依據(jù)。方法：病例組研究對(duì)象來(lái)自2013年6月-2015年5月在山東省精神衛(wèi)生中心住院的209例雙相障礙患者,包括雙相障礙Ⅰ型患者107例,雙相Ⅱ型患者102例。健康對(duì)照組為200例同期招募的健康對(duì)照者。收集5m1EDTA抗凝血,采用QIAamp全血DNA試劑盒提取基因組DNA。經(jīng)飛行質(zhì)譜(MALDI-TOF)Sequenom MassArray平臺(tái)進(jìn)行基因分型,采用PLINK1.07及SPSS20.0統(tǒng)計(jì)軟件進(jìn)行卡方檢驗(yàn)及回歸分析等并對(duì)分型成功且符合H-W平衡的SNP進(jìn)行關(guān)聯(lián)分析研究,計(jì)算OR值及p值。通過(guò)Haploview4.2計(jì)算連鎖不平衡性,構(gòu)建HTR3A及HTR3B單倍型,驗(yàn)證HTR3A、HTR3B的多態(tài)性位點(diǎn)與中國(guó)漢族雙相障礙易感性的關(guān)聯(lián)性。對(duì)符合入組標(biāo)準(zhǔn)的雙相障礙患者,采集病例組疾病資料包括家族史、起病年齡,發(fā)作次數(shù)及有無(wú)精神病性癥狀等。對(duì)病例組及健康對(duì)照組入組1周內(nèi)完成認(rèn)知功能成套測(cè)驗(yàn)(包括連線測(cè)驗(yàn)、符號(hào)編碼、言語(yǔ)流暢性、詞語(yǔ)學(xué)習(xí)、視覺(jué)空間記憶、迷宮測(cè)驗(yàn)、stroop色詞測(cè)驗(yàn)及持續(xù)操作測(cè)驗(yàn)等)評(píng)估,分析HTR3A及HTR3B Tag SNP與雙相障礙認(rèn)知功能損害的關(guān)聯(lián)性。結(jié)果：1.基因頻率及基因型頻率結(jié)果顯示HTR3A的位點(diǎn)rs1062613的T等位基因、rs1176722的A等位基因、rs1176719的A等位基因、rs10160548的G等位基因及rs1176713的C等位基因.HTR3B一個(gè)多態(tài)性位點(diǎn)rs1176746的T等位基因在病例組及對(duì)照組之間的差異性具統(tǒng)計(jì)學(xué)意義(p0.05)。HTR3A及HTR3B標(biāo)記多態(tài)位點(diǎn)在病例組與對(duì)照組的基因型比較結(jié)果顯示,HTR3A的rsl 176722、rs1176719、rs10160548及rs1176713位點(diǎn)及HTR3B的位點(diǎn)rs1176746攜帶突變基因型的患者個(gè)體例數(shù)顯著高于野生純合基因型個(gè)體例數(shù),差異性具統(tǒng)計(jì)學(xué)意義(p005)。2.單倍型分析結(jié)果顯示HTR3A及HTR3B的16個(gè)位點(diǎn)成功構(gòu)建與雙相障礙易感性相關(guān)聯(lián)的3個(gè)單倍體,HTR3B的單倍型rs4938056：T,rs12421126：T,rs1176746：C,rs1176744：G,rs2276305：G,rs3782025：T,rs1672717：T,rs12795805：C與雙相障礙發(fā)病的關(guān)聯(lián)性具有顯著意義(p0.05),分層分析結(jié)果顯示,雙相障礙Ⅰ型也構(gòu)建了3個(gè)單倍體,由HTR3A的5個(gè)多態(tài)性位點(diǎn)rs1062613、rs11604247、rs1176722、rs2276302及rs1176719組成的CCGAG、CCGAA這2種單倍型基因頻率在雙相障礙Ⅰ型與對(duì)照組間比較有統(tǒng)計(jì)學(xué)意義(p005),雙相障礙Ⅱ型構(gòu)建了4個(gè)單倍體,各單倍型與對(duì)照組比較不具有統(tǒng)計(jì)學(xué)意義(p0.05)。3.雙相障礙臨床特點(diǎn)與HTR3A及HTR3B標(biāo)記多態(tài)性位點(diǎn)基因型的關(guān)聯(lián)分析結(jié)果顯示HTR3A的rs11604247、rs2276302、rs10160548、rs1182457攜帶突變基因個(gè)體存在精神病性癥狀的比例較野生純合患者個(gè)體顯著增多(p0.05),攜帶HTR3A的rs1062613突變基因個(gè)體存在精神病性癥狀的比例較野生純合患者個(gè)體顯著減少(p0.05),HTR3B除rsl672717及rs12795805以外的九個(gè)位點(diǎn)均與雙相障礙精神病性癥狀存在顯著關(guān)聯(lián)性,其中rs10789970、rs3831455、rs4938056、rsl2421126、rs1176746及rs3782025這六個(gè)位點(diǎn)的突變基因個(gè)體存在精神病性癥狀的比例較野生純合患者個(gè)體顯著增多(p0.05),位點(diǎn)rs3758987、rs1176744及rs2276305突變基因個(gè)體存在精神病性癥狀的比例較野生純合患者個(gè)體顯著減少(p0.05)。HTR3A的位點(diǎn)rs10160548、rs1176713及HTR3B的位點(diǎn)rs4938056、rs3782025及rs1672717攜帶突變基因的基因型組的平均起病年齡均較野生純合基因型組的平均起病年齡早,具有統(tǒng)計(jì)學(xué)意義(p0.05),HTR3B位點(diǎn)rs1176746攜帶突變基因的基因型組的平均起病年齡均較野生純合基因型組的平均起病年齡晚,差異性顯著(p0.05)。HTR3A多數(shù)多態(tài)性位點(diǎn)攜帶突變基因的基因型組的平均病程均純合野生基因型組的平均病程長(zhǎng),其中rs1062613及rs10789980這兩個(gè)位點(diǎn)在兩組之間的差異性有統(tǒng)計(jì)學(xué)意義(p0.05),HTR3B除三個(gè)位點(diǎn)(rs1176744、rs2276305及rs12795805)外的八個(gè)位點(diǎn)攜帶突變基因的基因型組的平均病程均較純合野生基因型組的平均病程短,其中rs10789970、rs12421126及rs2276305這三個(gè)位點(diǎn)在兩組之間的差異性有統(tǒng)計(jì)學(xué)意義(p0.05)。HTR3A的位點(diǎn)rs10789980與發(fā)作次數(shù)呈正相關(guān)(r=0.283,p0.01)位點(diǎn)rs1062613與家族史呈正相關(guān)(r=0.298,p0.01),HTR3A的其余位點(diǎn)與家族史及發(fā)作次數(shù)無(wú)顯著相關(guān)性。HTR3B的的位點(diǎn)rs4938056及rs1176744與雙相障礙家族史無(wú)顯著相關(guān)性,rs12795805與家族史的相關(guān)系數(shù)(r=0.243,p005)稍低之外,其余八個(gè)多態(tài)性位點(diǎn)均與家族史呈顯著相關(guān)性(r=0.262-0.424,p0.01)。HTR3B的多態(tài)性位點(diǎn)中,僅rs2276305與發(fā)作次數(shù)呈顯著相關(guān)性(r=0.370,p0.01)。4.雙相障礙病例組與對(duì)照組認(rèn)知功能比較結(jié)果顯示雙相障礙Ⅰ型、雙相障礙Ⅱ型及對(duì)照組三組在認(rèn)知成套測(cè)驗(yàn)總分及各項(xiàng)認(rèn)知測(cè)驗(yàn)T分的差異性均有統(tǒng)計(jì)學(xué)意義(F=4.525-8.539,p0.05)雙相障礙Ⅰ型及雙相障礙Ⅱ型各項(xiàng)認(rèn)知測(cè)驗(yàn)得分均較對(duì)照組得分明顯減少,組間比較結(jié)果除視覺(jué)空間記憶、持續(xù)操作測(cè)驗(yàn)及迷宮測(cè)試三項(xiàng)認(rèn)知測(cè)驗(yàn)得分在雙相障礙Ⅰ型與雙相障礙Ⅱ型組間比較有統(tǒng)計(jì)學(xué)意義(p0.01)外,其余各項(xiàng)認(rèn)知測(cè)驗(yàn)得分的組間差異性均無(wú)顯著性統(tǒng)計(jì)學(xué)意義(p0.05)。雙相障礙Ⅰ型與對(duì)照組比較結(jié)果顯示,各項(xiàng)認(rèn)知測(cè)驗(yàn)得分均有統(tǒng)計(jì)學(xué)意義(p0.05),而雙相障礙Ⅱ型患者與對(duì)照組在信息處理速度、詞語(yǔ)學(xué)習(xí)、工作記憶及社會(huì)認(rèn)知這個(gè)四個(gè)認(rèn)知領(lǐng)域的得分有統(tǒng)計(jì)學(xué)意義(p0.05),在視覺(jué)學(xué)習(xí)、注意/警覺(jué)性、推理/問(wèn)題解決及認(rèn)知測(cè)驗(yàn)總分這幾個(gè)領(lǐng)域的差異性均無(wú)統(tǒng)計(jì)學(xué)意義。HTR3A及HTR3B基因多態(tài)性與雙相障礙患者認(rèn)知功能的關(guān)聯(lián)研究結(jié)果顯示,HTR3A大部分多態(tài)性位點(diǎn)攜帶突變基因的個(gè)體其認(rèn)知功能測(cè)驗(yàn)得分較低,且在信息處理速度與注意/警覺(jué)性這兩個(gè)認(rèn)知領(lǐng)域的得分較野生基因型個(gè)體顯著降低(p0..05)。HTR3A的rs1176722、rs2276302及rs1182457這三個(gè)標(biāo)記多態(tài)性位點(diǎn)攜帶突變基因的個(gè)體其認(rèn)知功能各領(lǐng)域均較野生純合基因型個(gè)體顯著減退(p0.05)。而HTR3B大部分多態(tài)性位點(diǎn)攜帶突變基因的個(gè)體其認(rèn)知功能測(cè)驗(yàn)得分相對(duì)稍高,尤其在信息處理速度、工作記憶、注意／警覺(jué)性、社會(huì)認(rèn)知這四個(gè)認(rèn)知領(lǐng)域及總體認(rèn)知功能較純合野生基因型個(gè)體的認(rèn)知功能相對(duì)受損較輕(p0.05),除rs10789970、rs2276305及rsl672717這三個(gè)多態(tài)性位點(diǎn)外,其余HTR3B的標(biāo)記多態(tài)性位點(diǎn)攜帶突變基因的個(gè)體其認(rèn)知功能各領(lǐng)域均較野生純合基因型個(gè)體認(rèn)知功能損害顯著減輕(p0.05)。結(jié)論：1.HTR3A的rs1176722、rs1176719、rs10160548及rs1176713及HTR3B的rs1176746突變基因及突變基因型加大了雙相障礙患者的發(fā)病風(fēng)險(xiǎn)。HTR3A的rs1062613的突變基因可能具有致病作用。2.由HTR3A的五個(gè)位點(diǎn)組成的CCGAG、CCGAA在雙相障礙Ⅰ型的患者可能出現(xiàn)連鎖遺傳。雙相障礙的患者中可能存在HTR3B的八個(gè)位點(diǎn)的連鎖遺傳。3.HTR3A個(gè)別位點(diǎn)及HTR3B多數(shù)位點(diǎn)基因多態(tài)性可能與雙相障礙患者出現(xiàn)精神病性癥狀有關(guān)。HTR3A及HTR3B的個(gè)別位點(diǎn)基因多態(tài)性可能與雙相障礙患者早年發(fā)病有關(guān)。HTR3A的多數(shù)位點(diǎn)突變基因可能與雙相障礙患者病程延長(zhǎng)有關(guān)。HTR3B多數(shù)位點(diǎn)突變基因分布可能增加了家族遺傳的風(fēng)險(xiǎn)。HTR3A或HTR3B基因多態(tài)性與雙相障礙發(fā)作次數(shù)無(wú)密切聯(lián)系。4.雙相障礙患者認(rèn)知功能較健康對(duì)照者認(rèn)知功能普遍下降。雙相障礙Ⅰ型患者在信息處理速度、詞語(yǔ)學(xué)習(xí)、工作記憶及社會(huì)認(rèn)知領(lǐng)域的認(rèn)知功能損害相對(duì)較重。HTR3A與加重雙相障礙患者認(rèn)知損害有關(guān)。HTR3B可能對(duì)減輕雙相障礙患者認(rèn)知功能損害有保護(hù)作用。5.信息處理速度、工作記憶、注意／警覺(jué)性、社會(huì)認(rèn)知等領(lǐng)域的認(rèn)知損害可能為雙相障礙患者的內(nèi)表型特征。
[Abstract]:Objective: to detect the allelic and genotype distribution of 21 labeled single nucleotide polymorphisms (tag SNP) of HTR3A and HTR3B in patients and controls, to explore the genetic susceptibility of bipolar disorder by case control association study and to search for the susceptibility genes of bipolar disorder, and to explore the polymorphism of HTR3A and HTR3BTag SNP genes, genetic imbalances and haplotypes and haplotypes. The relationship between the characteristics of disease and cognitive function in patients with bipolar disorder, further verifying whether cognitive impairment is an internal phenotype of bipolar disorder, provides an important reference for theoretical research and clinical practice, and provides a theoretical basis for the early recognition of bipolar disorder. Methods: the subjects of the case group were from May -2015 June 2013. 209 patients with bipolar disorder hospitalized in Shandong mental health center included 107 patients with bipolar type I and 102 biphasic type II patients. 200 healthy controls were recruited at the same time in the healthy control group. 5m1EDTA anticoagulant was collected, and QIAamp whole blood DNA kit was used to extract genomic DNA. via MALDI-TOF Sequenom MassArray level. The PLINK1.07 and SPSS20.0 statistical software were used to carry out the chi square test and regression analysis, and the SNP and the OR value and the p value were calculated. The OR value and the p value were calculated. The linkage disequilibrium was calculated by Haploview4.2 and the HTR3A and HTR3B single times were constructed to verify the HTR3A, HTR3B polymorphic loci and China. The correlation of the susceptibility to bipolar disorder in the Han nationality. For patients with bipolar disorder, the data included family history, age of onset, frequency of onset and symptoms of psychosis. The case group and healthy control group completed the cognitive function test (including connection test, symbol coding, speech flow) within 1 weeks. Unobstructed, verbal learning, visual spatial memory, maze test, Stroop Color Word Test and continuous operation test, and the correlation between the cognitive impairment of HTR3A and HTR3B Tag SNP and bipolar disorder. Results: the 1. gene frequency and genotype frequency results showed the T allele of the HTR3A site rs1062613, rs1176722 A allele, rs117 6719 A allele, rs10160548 G allele and rs1176713 C allele rs1176746 T allele between the case group and the control group were statistically significant (P0.05).HTR3A and HTR3B marker polymorphic loci in the case group and the control group, the results showed that HTR3A's 176722 The number of individual cases of rs1176719, rs10160548 and rs1176713 loci and HTR3B loci rs1176746 carrying mutant genotype was significantly higher than that of wild homozygous genotype, and the difference was statistically significant (P005).2. haplotype analysis showed that 16 sites of HTR3A and HTR3B were successfully constructed with 3 single links associated with the susceptibility to bipolar disorder. Ploidy, the association of HTR3B's haplotype rs4938056:T, rs12421126:T, rs1176746:C, rs1176744:G, rs2276305:G, rs3782025:T, rs1672717:T, and rs12795805:C was significant (P0.05). The stratified analysis showed that the biphasic disorder type I also constructed 3 haploidy, and the RS10 was 5 polymorphic loci of HTR3A. 62613, rs11604247, rs1176722, rs2276302 and rs1176719 composed of CCGAG, CCGAA, the frequency of the 2 haplotype genes was statistically significant (P005) between the two phase disorder type I and the control group (P005), and the biphasic barrier type II type had 4 haplotypes. The different haplotypes were not statistically significant (P0.05) the clinical characteristics of.3. biphasic disorder and HTR3. The correlation analysis between A and HTR3B markers showed that HTR3A rs11604247, rs2276302, rs10160548, the proportion of psychosis in rs1182457 carrying mutant genes increased significantly (P0.05) than those in the wild homozygous individuals (P0.05), and the proportion of psychotic symptoms in the individuals carrying HTR3A rs1062613 mutant genes was more wild than those in the wild. The patients with homozygous homozygous individuals decreased significantly (P0.05), and the nine loci other than rsl672717 and rs12795805 were significantly associated with bipolar disorder, and the proportion of individuals with rs10789970, rs3831455, rs4938056, rsl2421126, rs1176746 and rs3782025 in the six loci were more than the wild. Patients with homozygous individuals increased significantly (P0.05), the proportion of rs3758987, rs1176744 and rs2276305 mutant genes was significantly lower than those of the wild homozygous individuals (P0.05).HTR3A sites rs10160548, rs1176713 and HTR3B loci rs4938056, rs3782025, and rs1672717 carrying mutant genes. The age of the onset was earlier than the average age of the wild homozygous genotypic group (P0.05). The average onset age of the genotype group carrying the mutant gene in the HTR3B site rs1176746 was later than the average age of the wild homozygous genotype group, and the difference was significant (P0.05).HTR3A most polymorphic loci carrying the mutation gene. The average course of the homozygous wild genotypic group was long in the average course of the type group, and the difference between the two sites of rs1062613 and rs10789980 was statistically significant (P0.05), and the average course of the eight loci, except three loci (rs1176744, rs2276305 and rs12795805), was more than that of the homozygous group. The average course of the wild genotyping group was short, of which the three loci of rs10789970, rs12421126 and rs2276305 were statistically significant between the two groups (P0.05) the.HTR3A site rs10789980 was positively correlated with the number of episodes (r=0.283, P0.01) rs1062613 and family history (r=0.298, P0.01), the rest of the HTR3A sites and family history The site rs4938056 and rs1176744 had no significant correlation with the bipolar disorder family, and the correlation coefficient of rs12795805 and family history (r=0.243, P005) was slightly lower, and the other eight polymorphic loci were significantly correlated with family history (r= 0.262-0.424, P0.01).HTR3B (r= 0.262-0.424, P0.01).HTR3B, only rs227630. 5 a significant correlation with the frequency of seizures (r=0.370, P0.01).4. bipolar disorder case group and the control group compared with the control group, the results of cognitive function showed that the two phase obstacle type I, the two phase obstacle II and the control group three groups in the cognitive test total score and the difference of the cognitive test T scores were statistically significant (F=4.525-8.539, P0.05) biphasic disorder I type and The scores of each cognitive test of bipolar disorder type II were significantly lower than those of the control group. The scores of three cognitive tests, except visual space memory, continuous operation test and labyrinth test, were statistically significant (P0.01), but the difference between the scores of the other cognitive tests was poor. There was no significant statistical significance in the opposite sex (P0.05). The results of the biphasic disorder I and the control group showed that all the scores of the cognitive tests were statistically significant (P0.05), while the scores of the four cognitive domains, such as the speed of information processing, the learning of words, the working memory and the social cognition, were statistically significant in the biphasic disorder type II patients and the control group (the control group). P0.05), in visual learning, attention / alertness, reasoning / problem solving, and cognitive test total score, there is no statistically significant difference in.HTR3A and HTR3B gene polymorphism and cognitive function in patients with bipolar disorder. The results show that most of the individuals with HTR3A polymorphic sites with mutation genes are tested for cognitive function test. The scores of the two cognitive domains of the information processing speed and attention / alertness were significantly lower than those of the wild genotypes (p0..05).HTR3A rs1176722, rs2276302 and rs1182457, which were significantly lower than those of the wild homozygous genotypes (P0.05). While most of the individuals with HTR3B polymorphic loci carrying mutation genes were relatively slightly higher in cognitive function tests, especially in the four cognitive domains, such as information processing speed, working memory, attention / alertness, and social cognition, and the cognitive function of the homozygous wild genotypes was relatively impaired (P0.05), except for rs10789970. Outside the three polymorphic loci of rs2276305 and rsl672717, the other HTR3B marker polymorphic loci that carry the mutant genes are significantly less than those of the wild homozygous genotypes (P0.05). Conclusion: 1.HTR3A's rs1176722, rs1176719, rs10160548, rs1176713 and HTR3B rs1176746 mutants Mutations in the mutant genotype that increase the risk of rs1062613 of.HTR3A in patients with bipolar disorder may have the pathogenicity of CCGAG, which is composed of five sites of HTR3A, and CCGAA in patients with bipolar type I may have a linkage inheritance. There may be a linkage genetic.3.HTR3 of the eight loci of HTR3B in patients with bipolar disorder. A individual loci and HTR3B polymorphisms may be associated with the occurrence of psychosis in patients with bipolar disorder that may be related to the polymorphism of the individual loci of the.HTR3A and HTR3B in the patients with bipolar disorder and the majority of the mutation genes of the.HTR3A in the early onset of the bipolar disorder may be associated with the prolongation of the disease course of the bipolar disorder patient with the multiple point mutation of the.HTR3B. The distribution may increase the risk of family heredity,.HTR3A or HTR3B gene polymorphism is not closely related to the frequency of bipolar disorder. The cognitive function of cognitive function in patients with.4. bipolar disorder is generally decreased. The cognitive function of patients with bipolar disorder I in the field of information processing, word learning, working memory and social cognition .HTR3B may have protective effects on cognitive impairment in patients with bipolar disorder, which may have protective effects on cognitive impairment in bipolar disorder patients. Cognitive impairment in the fields of working memory, attention / alertness and social cognition may be the phenotypic characteristics of the patients with bipolar disorder, which may be associated with cognitive impairment in patients with bipolar disorder.
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類(lèi)號(hào)】：R749.4

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