叶酸代谢通路关键酶基因非编码区多态的先天性心脏病易感性研究及功能分析
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摘要
先天性心脏病(Congenital heart disease, CHD)是我国排名第一的出生缺陷,是导致婴幼儿死亡和残疾的重要原因,严重影响了我国的人口素质,给社会和家庭带来了沉重的负担。近年来临床实验证明孕前补充叶酸可以显著减低CHD的发生,由此可知叶酸及其代谢通路基因与CHD的发生关系重大。尽管国内外在叶酸代谢基因和CHD易感性方面做了大量的研究工作,并鉴定出一些致病的多态位点,如MTHFR c.677 C>T、c.1298 A>C等,但不同研究中结果不一致,且缺乏相关的分子生物学功能验证,仍有许多问题尚待解决：(1)大多数研究将注意力集中在早期文献报道的、一直以来受到广泛关注的SNP上,而不是搜寻新的功能性致病多态,例如对MTHFR c.677 C>T、c.1298 A>C、MTRR c.66 A>G等多态的研究从十多年前持续至今,但却始终得不到一致结果；(2)研究的基因区域侧重基因编码区,关注因氨基酸变化导致的基因功能异常是否会提高先心病易感性,很少对基因非编码区进行系统研究来探讨基因剂量与CHD的关系；(3)随着dbSNP、HapMap数据库的日益完善,研究者们倾向于根据数据库信息挑选tagSNP开展自己的研究,然而不同种族、不同群体的SNP分布和频率存在差异,完全根据数据库信息挑选tagSNP难免会遗漏很多重要遗传信息；(4)先天性心脏病样本很难收集,因此国内外大多数研究样本量都有一定局限,很少有研究募集到500例以上病例进行遗传学方面研究,因此效力有限,且容易出现假阳性。
为了探讨中国人群叶酸代谢途径核心基因和CHD易感性是否存在联系,并在研究方法上解决上述问题,我们在上海、江苏、山东三个省市收集了总计2,340例CHD病例和2,270例健康对照样本,挑选出32个样本对叶酸代谢途径中甲硫氨酸合成酶(MTR)、甲硫氨酸合成还原酶(MTRR)、胱硫醚p合酶(CB5)、胸苷酸合酶(TYMS)四个核心基因的启动子和3'UTR进行了测序,根据测序结果挑选最小等位基因频率大于10%的tagSNP,在三个人群中进行基因分型和关联分析。在找到与疾病易感性相关的阳性位点后,我们进一步对其进行了功能上的分析。因为叶酸代谢与癌症的发生息息相关,因此我们还分析了阳性位点与癌症患病风险的关联。
通过关联分析,我们共找到4个非编码区多态位点与先天性心脏病易感性相关,并在三个人群中得到了一致的结果,其中MTR-186 T>G (rs28372871)、+905 G>A (rs1131450)、MTRR c.56 A>C (rs326119)是风险型SNP, CBS -551 C>G (rs2850144)是保护型SNP。
MTR基因编码甲硫氨酸合成酶,在叶酸代谢中的主要功能是将5-甲基四氢叶酸(5m-THF)的甲基传递给同型半胱氨酸(HCY),生成四氢叶酸(THF)和甲硫氨酸(MET)。该基因中的两个风险因子分别位于启动子区和3'UTR区,二者均能显著增加先心病的患病风险,其中启动子区多态-186 T>G多态的G等位基因为风险因子,其GG基因型能增高1.56倍的患病风险(adjusted OR=1.56,95％CI=1.33.1.83,P=1.32×10-9)。in vivo的qRT-PCR和in vitro的Luciferase实验证明,-186 G等位基因启动转录的活性低于T等位基因,我们通过软件预测、EMSA、SPR、CHIP等手段证实了导致G等位基因转录活性低的原因是该多态T到G的变化使得启动子和转录激活因子USF的亲和力下降。另一方面从表观遗传学视角出发,我们发现-186 G等位基因的甲基化状况比T等位基因有显著升高,因此会进一步降低其转录活性。位于3'UTR的+905 G>A多态的A等位基因为风险因子,其GA、AA基因型分别能提高1.2倍和2.74倍的患病风险(GA: adjusted OR=1.2,95%CI=1.06-1.36;AA:adjusted OR=2.74,95% CI=2.09-3.60,P= 6.35×10-14)。功能研究发现,该多态由G到A的变化,会加大三种microRNA (miR-485、miR-608.miR-1293)对MTR基因的抑制力度,在翻译水平降低MTR的表达水平。我们还在522例健康志愿者血浆中检测了同型半胱氨酸的水平,结果发现-186 G和+905 A能显著增高体内血浆的HCY水平。关联分析以及HCY水平分析的结果表明这两个SNP可以协同作用,进一步增大CHD的患病风险。
MTRR基因编码酶的主要功能是维持足够的活化形式的维生素B12,后者是MTR的辅酶。实验中发现的阳性位点c.56+781 A>C位于MTRR第一内含子,其C等位基因会显著提高CHD易感性,CA基因型和CC基因型分别会将CHD患病风险提高1.4倍和1.84倍(GA:ajust OR=1.4,95%CI=1.23-1.59;CC:ajust OR=1.84,95%CI=1.54-2.2;P=4.34×10-12)。在功能实验中我们通过qRT-PCR检测心组织中MTRR mRNA的量,通过Luciferase报告基因系统in vitro检测不同等位基因的转录活性,二方面的结论一致,表明c.56+781 C等位基因会显著降低转录活性,SPR实验证实C等位基因和核提取物物的亲和力确实低于野生型的A等位基因。我们也发现在健康人群中CC基因型携带者的血浆HCY浓度要高于AA基因型。
CB5的功能是通过硫化作用直接催化HCY与丝氨酸缩合成胱硫醚,是HCY释放的途径之一。在关联分析中我们发现,位于启动子区的-551 C>G多态能降低CHD的患病风险,这种保护效应随着G等位基因数目的增加而增强,其CG基因型和GG基因型个体分别可降低15%、40%的CHD患病风险(CG:adjust OR=0.85,95%CI=0.75-0.96;GG:adjust OR=0.6,95%CI=0.49-0.73;P=6.62×10-7)。通过检测CBS的mRNA水平和Iuciferase报告基因验证,我们发现G等位基因的保护作用可能来源于提高的转录活性和表达量。在后续功能实验上我们发现-551C>G也受到了遗传学和表观遗传学两方面调控,一方面C到G的变化会降低转录抑制因子SP1与启动子的结合,从而激活转录；另一方面我们检测了-551 C>G附近区域的CpG位点甲基化状况,发现G等位基因对应的甲基化程度仅有C等位基因的1/7,无疑会进一步提高转录能力。
以上4个多态主要围绕叶酸代谢通路的三个重要作用,即同型半胱氨酸的移除、甲基化和DNA合成原料的提供。MTR、MTRR中三个风险多态都能大幅度降低基因的表达量,从而使得HCY在体内堆积、THF和MET的供应不足,这种情况在胎儿在心脏发育期间更显严峻,因为其他代偿途径在心脏发育初期并未形成。而CBS的保护作用则强调了HCY在CHD发生中的风险作用,尽管有研究认为CBS在胚胎发育初期表达量仅有成人的20%,但本文中发现的多态能大幅提高CB5的表达量,对CHD的发生有着显著的保护作用。
在实验中我们也对MTHFR c.677 C>T、c.1298A>C、MTR c.2756 A>G和MTRR c.66 A>G四个编码区多态位点的进行了分析,结果除了MTHFR c.1298 A>C在山东、江苏人群中会提高CHD患病风险之外,其他位点均与CHD易感性无关。将这4个编码区多态分型信息与我们发现的4个非编码区多态结果放在一起,进行多元Logistic回归分析后发现MTR-186 T>G、+905 G>A、MTRR c.56+781 A>C、CBS-551 C>G对表型的贡献是独立的。而在HCY队列中,通过多元回归分析发现,上述8个位点总共可以解释中国汉族人群HCY总变异的24.6%。其中MTR-186 T>G、+905 G>A和MTHFR c.677 C>T分别贡献了10.6%、5.7%、6.7%。
叶酸代谢与癌症的发生关系密切,因此我们最后检测了文中所述各多态与前列腺癌、乳腺癌、肺癌、肝癌的易感性关联。结果在关联分析中发现MTR+905 G>A多态与上述四种肿瘤均存在关联,MTR-186 T>G只与前列腺癌和肺癌的发生相关,而CBS-551 C>G则能降低乳腺癌和肺癌的发生风险。
Congenital heart disease (CHD) is the first common type of birth defects in China and the leading cause of infant morbidity and disability, which seriously affects the population living quality and plays heavy burden to families as well as society. Clinical researches in recent decades have suggested that maternal preconception administration of folic acid can reduce the occurrence of CHD. Soit was indicated that the core genes of folate metabolism pathways might be the great candidate genes associated with increased risk of CHD. Association studies between core genes in folate metabolism pathways and susceptibility of CHD have pointed out that some pathogenic polymorphism, such as like MTHFR c.677 C>T and c.1298 A>C. However, the worldwide association results are not pretty consistent. The suspending problems are waiting for being solved:(1) Researches mostly focus on published variants ortag SNPs instead of searching for the potentially functional pathogenic variants; (2) Coding region variants have been extensively explored, while the noncoding variants, which can regulate the dosage of the CHD associated gene expression, have been largely ingored; (3) With increasingly improved database of dbSNP and HapMap, researchers tend to select tagSNP directly and pay less attention tothe diversity of distribution and frequency of SNPs in different ethnics and groups, which will veil some significant genetic information; (4) Sample size limitation and less studies with a sample of more than 500 CHD cases leads to low validity and possibilities of false positive.
To investigate the association between key genes in the folate metabolism pathway and the susceptibility of CHD, we totally recruited 2,340 CHD cases and 2,270 healthy controls from Shanghai, Jiangsu and Shandong. The researched key genes include Methionine synthase (MTR), methionine synthase reductase (MTRR), cystathionineβ-synthase (CBS), and Thymidylate Synthetase (TYMS). We selected common tagSNPs with minor allele frequency >0.1 according to our own resequencing screening results, and carried out genotyping and association analysis among the three pairs of CHD-control groups. Then we conducted functional analysis of the CHD associated variants. Concerning the close relation between folate metabolism and cancer occurrence, we also analyzed the association between positive variants and several types of cancer incidence. The three independent case-control studies identified four CHD susceptible variants. Among the four polymorphisms, MTR-186T>G (rs28372871),+905 G>A (rs1131450), MTRR c.56+781 A>C (rs326119) are associated with increased risk of congenital heart disease, while CBS-551 C>G (rs2850114) is associated with reduced risk of congenital heart disease.
MTR encodes Methionine synthase, which catalyzes the transfer of a methyl group from 5-methyltetrahydrofolate to homocysteine (HCY). The promoter polymorphism-186 G allele in MTR is the risky factor, and the homozygous GG genotype increases a 1.56-fold CHD risk compared with wildtype (adjusted OR=1.56,95% CI=1.33-1.83, P =1.32×10-9). Both in vivo quantitative real-time PCR analysis of MTR hnRNA and mRNA in cardiac tissue samples of CHD and in vitro luciferase assay in transfected cells showed that the -186 G allele remarkably deceases MTR transcription. We confirmed that the reason why G allele leads to lower promoter activity is that the transversion T to G destroys the binding site of the promoter to the transcriptional activator USF. In the cis-regulation, the MTR -186G allele is always associated with elevated methylation ratio allover the promoter CpG islands, which leads to inhibition of MTR expression. It appears that genetic variation could be the primary determinant for the regulation of MTR expression because the methylation status also changed based on the polymorphism. The A allele of 3'UTR polymorphism +905 G>Ain MTR gene is also a CHD risk factor, and the GA and AA genotype can raise 1.2 and 2.74 times of CHD risk respectively (GA:adjusted OR=1.2,95% CI=1.06-1.36; AA:adjusted OR=2.74,95% CI=2.09-3.60, P=6.35×10-14). Functional study showed that the transversion G to A would corroborate the inhibition of three microRNA (miR-485, miR-608, miR-1293) to MTR, thus reduces the expression of MTR at the translating stage. We also measured the HCY level in 522 healthy volunteered controls. The examination result suggested that both-186 T>G and +905 G>A correlates with the elevated plasma HCY level. The further analysis indicated that these two SNPs increase the CHD risk synergistically.
Enzyme coded by MTRR gene acts mainly on maintaining adequate amount of activated vitamin B12, a coenzyme of MTR. Experiments showed that variant c.56+781 A>C in the first intron of MTRR is associated with CHD. The C allele of variant c.56+781 A>C will increases CHD susceptibility significantly, while the CA and CC genotype increase CHD risk by 1.4 and 1.84-fold respectively (GA:adjust OR=1.4, 95%CI=1.23-1.59; CC:ajust OR=1.84,95%CI=1.54-2.2; P=4.34×10-12). Both in vivo quantitative real-time PCR analysis of MTRR mRNA in cardiac tissue samples of CHD and in vitro luciferase assay in transfected cells showed that the c.56+781 C allele remarkably deceases MTRR transcription. Additionally, healthy individuals with homozygous CC genotype have significantly elevated homocysteine level compared with wild-type AA carriers.
CBS catalyzes the condensation of HCY and serine to cystathionine via the sulfidation, which is one of the HCY-releasing pathways. Correlated analysis showed that-551 C>G polymorphisms in promoter region of CBS can reduce the CHD risk. This protective effect is correlated with the copy number of G alleles, among which the CG and GG genotype can reduce 15% and 40% CHD risk respectively (CG:adjust OR=0.85,95%CI=0.75-0.96; GG:adjust OR=0.6,95%CI=0.49-0.73; P=6.62×10-7). We found that the protective function of G allele might result from increased transcriptional activity and expression of CBS, which was verified through both in vivo mRNA level measurement and luciferase assay. Followed function experiments showed that variant -551 C>G tales play genetically and epigenetically on the CBS expression. The conversion C to G reduces the binding affinity of transcriptional repressor SP1 and promoter, and thus activates the transcription as a result. Moreover, the methylation level of CpG sites in CBS promoter region with G allele was only 1/7 compared with that of C allele, which will improve the transcription of G allelewith no doubt.
The above four variants come from the folate metabolism pathways core genes of MTR, MTRR and CBS, which are coordinately responsible for in the removal of HCY, the resource of methyl group for DNA methylation, as well as the supplement of materials for DNA synthesis. Due tothe three risk variants in MTR and MTRR can reduce gene expression, the substrat HCY would accumulate in the body, while,THF as well as MET would be out of supply. This situation can become severer in the developing period of infant heart, for the reason that other alternative pathways haven't developed well, such as BHMT which is expressed mainly in liver and kidney. And the protection of CBS emphasizes the risky function of HCY in the CHD occurrence. Although some studies consider that the expression of CBS in the early period of infant development is only 20% compared with that of an adult, the polymorphism we identified by uscan improve the express of CBS significantly, thus has a significant protection against the occurrence of CHD.
In the experiments we also analyzed four extensively reported folate SNPs, MTHFR c.677 C>T, c.1298 A>C, MTR c.2756 A>G and MTRR c.66 A>G. Except for MTHFR c.1298 A>C raising the CHD risk, all the rest sites proved irrelative with CHD susceptibility. Using multiple Logistic regression analysis on all of the 4 noncoding variants and 4 extensively explored coding SNPs, considering both gender and age, we discovered that MTR-186 T>G,+905 G>A, MTRR c.56+781 A>C and CBS-551 C>G contributed to CHD in Chinese Han population independently. Additionally, all the 8 tested SNPs could explain 24.6% variation of HCY which was evaluated by partial correlation coefficient. The main contribution to the HCY concentration are from MTR-186 T>G (10.6%),+905 G>A (5.7%), MTHFR c.677 C>T (6.7%) respectively.
Since folate metabolism is closely related to the occurrence of cancer, we finally examined the correlation between the above polymorphisms and susceptibilities of several kinds of cancer, including prostatic cancer, breast cancer, liver cancer and lung cancer. In the correlation analysis, we found that MTR +905 G>A variant was strongly correlated to all types of the cancer occurrences, MTR-186 T>G variant was associated with increased risk only in prostatic cancer and liver cancer, while the CBS-551 C>G showed protective effect on breast cancer and lung cancer. These results would provide a new thought for the prognosis of CHD patients.
为了探讨中国人群叶酸代谢途径核心基因和CHD易感性是否存在联系,并在研究方法上解决上述问题,我们在上海、江苏、山东三个省市收集了总计2,340例CHD病例和2,270例健康对照样本,挑选出32个样本对叶酸代谢途径中甲硫氨酸合成酶(MTR)、甲硫氨酸合成还原酶(MTRR)、胱硫醚p合酶(CB5)、胸苷酸合酶(TYMS)四个核心基因的启动子和3'UTR进行了测序,根据测序结果挑选最小等位基因频率大于10%的tagSNP,在三个人群中进行基因分型和关联分析。在找到与疾病易感性相关的阳性位点后,我们进一步对其进行了功能上的分析。因为叶酸代谢与癌症的发生息息相关,因此我们还分析了阳性位点与癌症患病风险的关联。
通过关联分析,我们共找到4个非编码区多态位点与先天性心脏病易感性相关,并在三个人群中得到了一致的结果,其中MTR-186 T>G (rs28372871)、+905 G>A (rs1131450)、MTRR c.56 A>C (rs326119)是风险型SNP, CBS -551 C>G (rs2850144)是保护型SNP。
MTR基因编码甲硫氨酸合成酶,在叶酸代谢中的主要功能是将5-甲基四氢叶酸(5m-THF)的甲基传递给同型半胱氨酸(HCY),生成四氢叶酸(THF)和甲硫氨酸(MET)。该基因中的两个风险因子分别位于启动子区和3'UTR区,二者均能显著增加先心病的患病风险,其中启动子区多态-186 T>G多态的G等位基因为风险因子,其GG基因型能增高1.56倍的患病风险(adjusted OR=1.56,95％CI=1.33.1.83,P=1.32×10-9)。in vivo的qRT-PCR和in vitro的Luciferase实验证明,-186 G等位基因启动转录的活性低于T等位基因,我们通过软件预测、EMSA、SPR、CHIP等手段证实了导致G等位基因转录活性低的原因是该多态T到G的变化使得启动子和转录激活因子USF的亲和力下降。另一方面从表观遗传学视角出发,我们发现-186 G等位基因的甲基化状况比T等位基因有显著升高,因此会进一步降低其转录活性。位于3'UTR的+905 G>A多态的A等位基因为风险因子,其GA、AA基因型分别能提高1.2倍和2.74倍的患病风险(GA: adjusted OR=1.2,95%CI=1.06-1.36;AA:adjusted OR=2.74,95% CI=2.09-3.60,P= 6.35×10-14)。功能研究发现,该多态由G到A的变化,会加大三种microRNA (miR-485、miR-608.miR-1293)对MTR基因的抑制力度,在翻译水平降低MTR的表达水平。我们还在522例健康志愿者血浆中检测了同型半胱氨酸的水平,结果发现-186 G和+905 A能显著增高体内血浆的HCY水平。关联分析以及HCY水平分析的结果表明这两个SNP可以协同作用,进一步增大CHD的患病风险。
MTRR基因编码酶的主要功能是维持足够的活化形式的维生素B12,后者是MTR的辅酶。实验中发现的阳性位点c.56+781 A>C位于MTRR第一内含子,其C等位基因会显著提高CHD易感性,CA基因型和CC基因型分别会将CHD患病风险提高1.4倍和1.84倍(GA:ajust OR=1.4,95%CI=1.23-1.59;CC:ajust OR=1.84,95%CI=1.54-2.2;P=4.34×10-12)。在功能实验中我们通过qRT-PCR检测心组织中MTRR mRNA的量,通过Luciferase报告基因系统in vitro检测不同等位基因的转录活性,二方面的结论一致,表明c.56+781 C等位基因会显著降低转录活性,SPR实验证实C等位基因和核提取物物的亲和力确实低于野生型的A等位基因。我们也发现在健康人群中CC基因型携带者的血浆HCY浓度要高于AA基因型。
CB5的功能是通过硫化作用直接催化HCY与丝氨酸缩合成胱硫醚,是HCY释放的途径之一。在关联分析中我们发现,位于启动子区的-551 C>G多态能降低CHD的患病风险,这种保护效应随着G等位基因数目的增加而增强,其CG基因型和GG基因型个体分别可降低15%、40%的CHD患病风险(CG:adjust OR=0.85,95%CI=0.75-0.96;GG:adjust OR=0.6,95%CI=0.49-0.73;P=6.62×10-7)。通过检测CBS的mRNA水平和Iuciferase报告基因验证,我们发现G等位基因的保护作用可能来源于提高的转录活性和表达量。在后续功能实验上我们发现-551C>G也受到了遗传学和表观遗传学两方面调控,一方面C到G的变化会降低转录抑制因子SP1与启动子的结合,从而激活转录；另一方面我们检测了-551 C>G附近区域的CpG位点甲基化状况,发现G等位基因对应的甲基化程度仅有C等位基因的1/7,无疑会进一步提高转录能力。
以上4个多态主要围绕叶酸代谢通路的三个重要作用,即同型半胱氨酸的移除、甲基化和DNA合成原料的提供。MTR、MTRR中三个风险多态都能大幅度降低基因的表达量,从而使得HCY在体内堆积、THF和MET的供应不足,这种情况在胎儿在心脏发育期间更显严峻,因为其他代偿途径在心脏发育初期并未形成。而CBS的保护作用则强调了HCY在CHD发生中的风险作用,尽管有研究认为CBS在胚胎发育初期表达量仅有成人的20%,但本文中发现的多态能大幅提高CB5的表达量,对CHD的发生有着显著的保护作用。
在实验中我们也对MTHFR c.677 C>T、c.1298A>C、MTR c.2756 A>G和MTRR c.66 A>G四个编码区多态位点的进行了分析,结果除了MTHFR c.1298 A>C在山东、江苏人群中会提高CHD患病风险之外,其他位点均与CHD易感性无关。将这4个编码区多态分型信息与我们发现的4个非编码区多态结果放在一起,进行多元Logistic回归分析后发现MTR-186 T>G、+905 G>A、MTRR c.56+781 A>C、CBS-551 C>G对表型的贡献是独立的。而在HCY队列中,通过多元回归分析发现,上述8个位点总共可以解释中国汉族人群HCY总变异的24.6%。其中MTR-186 T>G、+905 G>A和MTHFR c.677 C>T分别贡献了10.6%、5.7%、6.7%。
叶酸代谢与癌症的发生关系密切,因此我们最后检测了文中所述各多态与前列腺癌、乳腺癌、肺癌、肝癌的易感性关联。结果在关联分析中发现MTR+905 G>A多态与上述四种肿瘤均存在关联,MTR-186 T>G只与前列腺癌和肺癌的发生相关,而CBS-551 C>G则能降低乳腺癌和肺癌的发生风险。
Congenital heart disease (CHD) is the first common type of birth defects in China and the leading cause of infant morbidity and disability, which seriously affects the population living quality and plays heavy burden to families as well as society. Clinical researches in recent decades have suggested that maternal preconception administration of folic acid can reduce the occurrence of CHD. Soit was indicated that the core genes of folate metabolism pathways might be the great candidate genes associated with increased risk of CHD. Association studies between core genes in folate metabolism pathways and susceptibility of CHD have pointed out that some pathogenic polymorphism, such as like MTHFR c.677 C>T and c.1298 A>C. However, the worldwide association results are not pretty consistent. The suspending problems are waiting for being solved:(1) Researches mostly focus on published variants ortag SNPs instead of searching for the potentially functional pathogenic variants; (2) Coding region variants have been extensively explored, while the noncoding variants, which can regulate the dosage of the CHD associated gene expression, have been largely ingored; (3) With increasingly improved database of dbSNP and HapMap, researchers tend to select tagSNP directly and pay less attention tothe diversity of distribution and frequency of SNPs in different ethnics and groups, which will veil some significant genetic information; (4) Sample size limitation and less studies with a sample of more than 500 CHD cases leads to low validity and possibilities of false positive.
To investigate the association between key genes in the folate metabolism pathway and the susceptibility of CHD, we totally recruited 2,340 CHD cases and 2,270 healthy controls from Shanghai, Jiangsu and Shandong. The researched key genes include Methionine synthase (MTR), methionine synthase reductase (MTRR), cystathionineβ-synthase (CBS), and Thymidylate Synthetase (TYMS). We selected common tagSNPs with minor allele frequency >0.1 according to our own resequencing screening results, and carried out genotyping and association analysis among the three pairs of CHD-control groups. Then we conducted functional analysis of the CHD associated variants. Concerning the close relation between folate metabolism and cancer occurrence, we also analyzed the association between positive variants and several types of cancer incidence. The three independent case-control studies identified four CHD susceptible variants. Among the four polymorphisms, MTR-186T>G (rs28372871),+905 G>A (rs1131450), MTRR c.56+781 A>C (rs326119) are associated with increased risk of congenital heart disease, while CBS-551 C>G (rs2850114) is associated with reduced risk of congenital heart disease.
MTR encodes Methionine synthase, which catalyzes the transfer of a methyl group from 5-methyltetrahydrofolate to homocysteine (HCY). The promoter polymorphism-186 G allele in MTR is the risky factor, and the homozygous GG genotype increases a 1.56-fold CHD risk compared with wildtype (adjusted OR=1.56,95% CI=1.33-1.83, P =1.32×10-9). Both in vivo quantitative real-time PCR analysis of MTR hnRNA and mRNA in cardiac tissue samples of CHD and in vitro luciferase assay in transfected cells showed that the -186 G allele remarkably deceases MTR transcription. We confirmed that the reason why G allele leads to lower promoter activity is that the transversion T to G destroys the binding site of the promoter to the transcriptional activator USF. In the cis-regulation, the MTR -186G allele is always associated with elevated methylation ratio allover the promoter CpG islands, which leads to inhibition of MTR expression. It appears that genetic variation could be the primary determinant for the regulation of MTR expression because the methylation status also changed based on the polymorphism. The A allele of 3'UTR polymorphism +905 G>Ain MTR gene is also a CHD risk factor, and the GA and AA genotype can raise 1.2 and 2.74 times of CHD risk respectively (GA:adjusted OR=1.2,95% CI=1.06-1.36; AA:adjusted OR=2.74,95% CI=2.09-3.60, P=6.35×10-14). Functional study showed that the transversion G to A would corroborate the inhibition of three microRNA (miR-485, miR-608, miR-1293) to MTR, thus reduces the expression of MTR at the translating stage. We also measured the HCY level in 522 healthy volunteered controls. The examination result suggested that both-186 T>G and +905 G>A correlates with the elevated plasma HCY level. The further analysis indicated that these two SNPs increase the CHD risk synergistically.
Enzyme coded by MTRR gene acts mainly on maintaining adequate amount of activated vitamin B12, a coenzyme of MTR. Experiments showed that variant c.56+781 A>C in the first intron of MTRR is associated with CHD. The C allele of variant c.56+781 A>C will increases CHD susceptibility significantly, while the CA and CC genotype increase CHD risk by 1.4 and 1.84-fold respectively (GA:adjust OR=1.4, 95%CI=1.23-1.59; CC:ajust OR=1.84,95%CI=1.54-2.2; P=4.34×10-12). Both in vivo quantitative real-time PCR analysis of MTRR mRNA in cardiac tissue samples of CHD and in vitro luciferase assay in transfected cells showed that the c.56+781 C allele remarkably deceases MTRR transcription. Additionally, healthy individuals with homozygous CC genotype have significantly elevated homocysteine level compared with wild-type AA carriers.
CBS catalyzes the condensation of HCY and serine to cystathionine via the sulfidation, which is one of the HCY-releasing pathways. Correlated analysis showed that-551 C>G polymorphisms in promoter region of CBS can reduce the CHD risk. This protective effect is correlated with the copy number of G alleles, among which the CG and GG genotype can reduce 15% and 40% CHD risk respectively (CG:adjust OR=0.85,95%CI=0.75-0.96; GG:adjust OR=0.6,95%CI=0.49-0.73; P=6.62×10-7). We found that the protective function of G allele might result from increased transcriptional activity and expression of CBS, which was verified through both in vivo mRNA level measurement and luciferase assay. Followed function experiments showed that variant -551 C>G tales play genetically and epigenetically on the CBS expression. The conversion C to G reduces the binding affinity of transcriptional repressor SP1 and promoter, and thus activates the transcription as a result. Moreover, the methylation level of CpG sites in CBS promoter region with G allele was only 1/7 compared with that of C allele, which will improve the transcription of G allelewith no doubt.
The above four variants come from the folate metabolism pathways core genes of MTR, MTRR and CBS, which are coordinately responsible for in the removal of HCY, the resource of methyl group for DNA methylation, as well as the supplement of materials for DNA synthesis. Due tothe three risk variants in MTR and MTRR can reduce gene expression, the substrat HCY would accumulate in the body, while,THF as well as MET would be out of supply. This situation can become severer in the developing period of infant heart, for the reason that other alternative pathways haven't developed well, such as BHMT which is expressed mainly in liver and kidney. And the protection of CBS emphasizes the risky function of HCY in the CHD occurrence. Although some studies consider that the expression of CBS in the early period of infant development is only 20% compared with that of an adult, the polymorphism we identified by uscan improve the express of CBS significantly, thus has a significant protection against the occurrence of CHD.
In the experiments we also analyzed four extensively reported folate SNPs, MTHFR c.677 C>T, c.1298 A>C, MTR c.2756 A>G and MTRR c.66 A>G. Except for MTHFR c.1298 A>C raising the CHD risk, all the rest sites proved irrelative with CHD susceptibility. Using multiple Logistic regression analysis on all of the 4 noncoding variants and 4 extensively explored coding SNPs, considering both gender and age, we discovered that MTR-186 T>G,+905 G>A, MTRR c.56+781 A>C and CBS-551 C>G contributed to CHD in Chinese Han population independently. Additionally, all the 8 tested SNPs could explain 24.6% variation of HCY which was evaluated by partial correlation coefficient. The main contribution to the HCY concentration are from MTR-186 T>G (10.6%),+905 G>A (5.7%), MTHFR c.677 C>T (6.7%) respectively.
Since folate metabolism is closely related to the occurrence of cancer, we finally examined the correlation between the above polymorphisms and susceptibilities of several kinds of cancer, including prostatic cancer, breast cancer, liver cancer and lung cancer. In the correlation analysis, we found that MTR +905 G>A variant was strongly correlated to all types of the cancer occurrences, MTR-186 T>G variant was associated with increased risk only in prostatic cancer and liver cancer, while the CBS-551 C>G showed protective effect on breast cancer and lung cancer. These results would provide a new thought for the prognosis of CHD patients.
引文
1. Hoffman JIE, Kaplan S. The incidence of congenital heart disease. Journal of the American College of Cardiology 2002; 39:1890-1900.
2. Hoffman, J. I. Incidence of congenital heart disease:Ⅱ.Prenatal incidence. Pediatr Cardiol 1995; 16:155-65.
3. Miller, S. P. et al. Abnormal brain development in newborns with congenital heart disease.M Engl J Med 2007; 357:1928-1938.
4. Pierpont, M. E. et al. Genetic basis for congenital heart defects:current knowledge: a scientific statement from the American Heart Association Congenital Cardiac Defects Committee, Council on Cardiovascular Disease in the Young:endorsed by the American Academy of Pediatrics. Circulation 115,3015-3038 (2007).
5. Jenkins, K. J. et al. Noninherited risk factors and congenital cardiovascular defects: current knowledge:a scientific statement from the American Heart Association Council on Cardiovascular Disease in the Young:endorsed by the American Academy of Pediatrics. Circulation 115,2995-3014 (2007).
6. Cooper, W. O. et al., Major congenital malformations after first-trimester exposure to ACE inhibitors. N. Engl. J. Med.354,2443-2451 (2006).
7. Olson, E. N. Gene regulatory networks in the evolution and development of the heart. Science 313,1922-1927 (2006).
8. Srivastava, D. Making or breaking the heart:from lineage determination to morphogenesis. Cell 126,1037-1048 (2006).
9. Buckingham, M., Meilhac, S.& Zaffran, S. Building the mammalian heart from two sources of myocardial cells. Nature Rev. Genet.6,826-835 (2005).
10. Kattman, S. J., Huber, T. L.& Keller, G. M. Multipotent flk-1+cardiovascular progenitor cells give rise to the cardiomyocyte, endothelial, and vascular smooth muscle lineages. Dev. Cell 11,723-732 (2006).
11. Wu, S. M. et al. Developmental origin of a bipotential myocardial and smooth muscle cell precursor in the mammalian heart. Cell 127,1137-1150 (2006).
12. Moretti, A. et al. Multipotent embryonic ISL1+progenitor cells lead to cardiac, smooth muscle, and endothelial cell diversification. Cell 127,1151-1165 (2006).
13. Tzahor, E. Wnt/β-catenin signaling and cardiogenesis:timing does matter. Dev. Cell 13,10-13 (2007).
14. Schoenebeck, J. J., Keegan, B. R.& Yelon, D. Vessel and blood specification override cardiac potential in anterior mesoderm. Dev. Cell 13,254-267 (2007).
15. Keegan, B. R., Feldman, J. L., Begemann, G., Ingham, P. W.& Yelon, D. Retinoic acid signaling restricts the cardiac progenitor pool. Science 307,247-249 (2005).
16. Prall, O. W. et al. An Nkx2-5/Bmp2/Smad1 negative feedback loop controls second heart field progenitor specification and proliferation. Cell 128,947-959 (2007).
17. Lickert, H. et al. Baf60c is essential for function of BAF chromatin remodelling complexes in heart development. Nature 432,107-112 (2004).
18. Gottlieb, P. D. et al. Bop encodes a muscle-restricted protein containing MYND and SET domains and is essential for cardiac differentiation and morphogenesis. Nature Genet.31,25-32 (2002).
19. Montgomery, R. L. et al. Histone deacetylases 1 and 2 redundantly regulate cardiac morphogenesis, growth, and contractility. Genes Dev.21,1790-1802 (2007).
20. Basson, C. T. et al. Mutations in human TBX5 cause limb and cardiac malformation in Holt-Oram syndrome. Nature Genet.15,30-35 (1997).
21. Li, Q. Y. et al. Holt-Oram syndrome is caused by mutations in TBX5, a member of the Brachyury (T) gene family. Nature Genet.15,21-29 (1997).
22. Schott, J.-J. et al. Congenital heart disease caused by mutations in the transcription factor NKX2-5. Science 281,108-111 (1998).
23. Benson, D. W. et al. Mutations in the cardiac transcription factor NKX2.5 affect diverse cardiac developmental pathways. J. Clin. Invest.104,1567-1573 (1999).
24. Biben, C. et al. Cardiac septal and valvular dysmorphogenesis in mice heterozygous for mutations in the homeobox gene Nkx2-5. Circ. Res.87,888-895 (2000).
25. Bruneau, B. G. et al. A murine model of Holt-Oram syndrome defines roles of the T-box transcription factor Tbx5 in cardiogenesis and disease. Cell 106,709-721 (2001).
26. Hiroi, Y. et al. Tbx5 associates with Nkx2-5 and synergistically promotes cardiomyocyte differentiation. Nature Genet.28,276-280 (2001).
27. Garg, V. et al. GATA4 mutations cause human congenital heart defects and reveal an interaction with TBX5. Nature 424,443-447 (2003).
28. Lindsay, E. A. et al. Tbx1 haploinsufficiency in the DiGeorge syndrome region causes aortic arch defects in mice. Nature 410,97-101 (2001).
29. Merscher, S. et al. TBX1 is responsible for cardiovascular defects in velo-cardio-facial/DiGeorge syndrome. Cell 104,619-629 (2001).
30. Yagi, H. et al. Role of TBX1 in human del22q11.2 syndrome. Lancet 362, 1366-1373 (2003).
31. Hu, T. et al. Tbx1 regulates fibroblast growth factors in the anterior heart field through a reinforcing autoregulatory loop involving forkhead transcription factors. Development 131,5491-5502 (2004).
32. Xu, H. et al. Tbx1 has a dual role in the morphogenesis of the cardiac outflow tract. Development 131,3217-3227 (2004).
33. Guris, D. L, Duester, G., Papaioannou, V. E.& Imamoto, A. Dose-dependent interaction of Tbx1 and Crkl and locally aberrant RA signaling in a model of del22q11 syndrome. Dev. Cell 10,81-92 (2006).
34. Moon, A. M. et al. Crkl deficiency disrupts Fgf8 signaling in a mouse model of 22q11 deletion syndromes. Dev. Cell 10,71-80 (2006).
35. Al-Baradie, R. et al. Duane radial ray syndrome (Okihiro syndrome) maps to 20q13 and results from mutations in SALL4, a new member of the SAL family. Am. J. Hum. Genet.71,1195-1199 (2002).
36. Kohlhase, J. et al. Okihiro syndrome is caused by SALL4 mutations. Hum. Mol. Genet.11,2979-2987 (2002).
37. Kirk, E. P. et al. Mutations in cardiac T-box factor gene TBX20 are associated with diverse cardiac pathologies, including defects of septation and valvulogenesis and cardiomyopathy. Am. J. Hum. Genet.81,280-291 (2007).
38. Koshiba-Takeuchi, K. et al. Cooperative and antagonistic interactions between Sall4 and Tbx5 pattern the mouse limb and heart. Nature Genet.38,175-183 (2006).
39. Satoda, M. et al. Mutations in TFAP2B cause Char syndrome, a familial form of patent ductus arteriosus. Nature Genet.25,42-46 (2000).
40. Muncke, N. et al. Missense mutations and gene interruption in PROSIT240, a novel TRAP240-like gene, in patients with congenital heart defect (transposition of the great arteries). Circulation 108,2843-2850 (2003).
41. Hove, J. R. et al. Intracardiac fluid forces are an essential epigenetic factor for embryonic cardiogenesis. Nature 421,172-177 (2003).
42. Auman, H. J. et al. Functional modulation of cardiac form through regionally confined cell shape changes. PLoS Biol.5, e53 (2007).
43. Yashiro, K., Shiratori, H.& Hamada, H. Haemodynamics determined by a genetic programme govern asymmetric development of the aortic arch. Nature 450, 285-288 (2007).
44. Ching, Y. H. et al. Mutation in myosin heavy chain 6 causes atrial septal defect. Nature Genet.37,423-428 (2005).
45. Barnett, J. V.& Desgrosellier, J. S. Early events in valvulogenesis:a signaling perspective. Birth Defects Res. C Embryo Today 69,58-72 (2003).
46. Timmerman, L. A. et al. Notch promotes epithelial-mesenchymal transition during cardiac development and oncogenic transformation. Genes Dev.18,99-115 (2004).
47. Garg, V. et al. Mutations in NOTCH1 cause aortic valve disease. Nature 437, 270-274 (2005).
48. Li, L. et al. Alagille syndrome is caused by mutations in human Jaggedl, which encodes a ligand for Notch1. Nature Genet.16,243-251 (1997).
49. Oda, T. et al. Mutations in the human Jaggedl gene are responsible for Alagille syndrome.Nature Genet.16,235-242 (1997).
50. McDaniell, R. et al. NOTCH2 mutations cause Alagille syndrome, a heterogeneous disorder of the notch signaling pathway. Am. J. Hum. Genet.79,169-173 (2006).
51. Gelb, B. D.& Tartaglia, M. Noonan syndrome and related disorders:dysregulated RASmitogen activated protein kinase signal transduction. Hum. Mol. Genet.15 (Spec. No.2), R220-R226 (2006).
52. Pandit, B. et al. Gain-of-function RAF1 mutations cause Noonan and LEOPARD syndromes with hypertrophic cardiomyopathy. Nature Genet.39,1007-1012 (2007).
53. Razzaque, M. A. et al. Germline gain-of-function mutations in RAF1 cause Noonan syndrome. Nature Genet.39,1013-1017 (2007).
54. Roberts, A. E. et al. Germline gain-of-function mutations in SOS1 cause Noonan syndrome. Nature Genet.39,70-74 (2007).
55. Tartaglia, M. et al. Gain-of-function SOS1 mutations cause a distinctive form of Noonan syndrome. Nature Genet.39,75-79 (2007).
56. Araki, T. et al. Mouse model of Noonan syndrome reveals cell type-and gene dosagedependent effects of Ptpn11 mutation. Nature Med.10,849-857 (2004).
57. Ambros, V. The functions of animal microRNAs. Nature 431,350-355 (2004).
58. van Rooij, E.& Olson, E. N. MicroRNAs:powerful new regulators of heart disease and provocative therapeutic targets. J. Clin. Invest.117,2369-2376 (2007).
59. Zhao, Y., Samal, E.& Srivastava, D. Serum response factor regulates a muscle-specific microRNA that targets Hand2 during cardiogenesis. Nature 436, 214-220 (2005).
60. Zhao, Y. et al. Dysregulation of cardiogenesis, cardiac conduction, and cell cycle in mice lacking microRNA-1-2. Cell 129,303-317 (2007).
61. Warnes, C. A. The adult with congenital heart disease:born to be bad? J. Am. Coll. Cordiol.46,1-8 (2005).
62. Bisping, E. et al. Gata4 is required for maintenance of postnatal cardiac function and protection from pressure overload-induced heart failure. Proc. Natl Acad. Sci. USA 103,14471-14476 (2006).
63. Pashmforoush, M. et al. Nkx2-5 pathways and congenital heart disease; loss of ventricular myocyte lineage specification leads to progressive cardiomyopathy and complete heart block. Cell 2004; 117:373-86.
64. Acuna J, Yoon P, Erickson D. The prevention of neural tube defects with folic acid. Atlanta, GA:Centers for Disease Control and Prevention,1999.
65. Health Canada. Congenital anomalies in Canada:a perinatal health report. Ottawa:Minister of Public Works and Governmental Services,2002.
66. Botto LD, Mulinare J, Erickson JD. Do multivitamin or folic acid supplements reduce the risk for congenital heart defects? Evidence and gaps. Am J Med Genet A 2003; 121:95-101.
67. Botto LD, Olney RS, Erickson JD. Vitamin supplements and the risk for congenital anomalies other than neural tube defects. Am J Med Genet C Semin Med Genet 2004; 125:12-21.
68. Huhta JC, Hernandez-Robles JA. Homocysteine, folate, and congenital heart defects. Fetal Pediatr Pathol 2005; 24:71-9.
69. Bailey LB, Berry RJ. Folic acid supplementation and the occurrence ofcongenital heart defects, orofacial clefts, multiple births, andmiscarriage. Am J Clin Nutr 2005; 81:1213-7S.
70. Huhta JC, Linask K, Bailey L. Recent advances in the prevention ofcongenital heart disease. Curr Opin Pediatr 2006; 18:484-9.
71. Jenkins KJ, Correa A, Feinstein JA, et al. Noninherited risk factors and congenital cardiovascular defects:current knowledge:a scientific statement from the American Heart Association Council on Cardiovascular Disease in the Young:endorsed by the American Academy of Pediatrics. Circulation 2007; 115:2995-3014.
72. Lucock M. Folic acid:Nutritional biochemistry, molecular biology, and role in disease processes. Molecular Genetics and Metabolism 2000; 71:121-38.
73.李竹,刘向东,陈亚华.神经管畸形不同发病地区婚检妇女叶酸缺乏率的研究.中国优生优育1996；1：1-4.
74.郝玲,田熠华,唐仪,李竹.我国部分地区岁人群血浆叶酸水平与年龄性别差异比较.营养学报2002；24(4)：352-4.
75. Gideon FNG, Yechiel F, Arie BY, et al. A common mutation A1298C in human methylenetetrahydrofolate reductase gene:association with plasma total homocysteine and folate concentrations. J Nutr 1999;129:1656-61.
76. Rosenquist TH, Ratashak SA, Selhub J. Homocysteine induces congenital defects of the heart and neural tube:effect of folic acid. Proc Natl Acad Sci USA,1996; 93:15227-15232.
77. Peter K, Chiang RG, Jacov T, et al. S-Adenosylmethionine and methylation. Faseb J 1996;10:471-80.
78. Eloranta TO, Kajander EO. Catabolism and lability of S-adenosyl-L-methionine in rat liver extracts. Biochem J 1994; 224:137-44.
79. WA L. S-adenosylmethionine:jack of all trades and master of everything? Biochem Soc Trans 2006 34 (Pt 2):330-3.
80. Fenech M. The role of folic acid and Vitamin B12 in genomic stability of human cells. Mutation Research 2001;475:57-67.
81. Kristina M von Castel-Dunwoody GPK, Karla P Shelnutt, Jaimie D Vaughn, Elizabeth R Griffin,David R Maneval, Douglas W Theriaque, and Lynn B Bailey.
Transcobalamin 776C-G polymorphism negatively affects vitamin B-12 metabolism. Am J Clin Nutr 2005; 81:1436-9.
82. Brunaud L AJ, Ayav A, Gerard P, Namour F, Antunes L, Braun M, Bronowicki JP, Bresler L, Gueant JL. Effects of vitamin B12 and folate deficiencies on DNA methylation and carcinogenesis in rat liver. Clin Chem Lab Med 2003; 41 (8):1012-9.
83. Jacob RA JD, Allman-Farinelli MA, Swendseid ME. Folate nutriture alters choline status of women and men fed low choline diets. J Nutr 1999; 129:712-7.
84. Stephanie E Chiuve ELG, Susan E Hankinson, Steven H Zeisel, Lauren W Dougherty,Walter C Willett, and Eric B Rimm. The association between betaine and choline intakes and the plasma concentrations of homocysteine in women. Am J Clin Nutr 2007; 86:1073-81.
85. Zeisel SH ZT, daCosta K, Pomfret EA. Effect of choline deficiency on S-adenosylmethionine and methionine concentrations in rat liver. Biochem J 1989; 259:725-9.
86. SA. C. Betaine in human nutrition. Am J Clin Nutr 2004;80:539-49.
87. SEITZ GPaHK. Alcohol and Cancer. Alcohol & Alcoholism 2004;39 (3):155-65.
88. Halsted CH, Villanueva JA, Devlin AM, et al. Metabolic interactions of alcohol and folate.J Nutr 2002; 132:2367S-72S.
89. Halsted CH, Robles EA., Mezey E. Intestinal malabsorption in folate-deficient alcoholics.. Gastroenterology 1973; 64:526-32.
90. Villanueva JA, Devlin, A. M.,& Halsted, C. H. Reduced folate carrier:tissue distribution and effects of chronic ethanol intake in the micropig.. Alcohol Clin Exp Res 2001;25:415-20.
91. Shaw S, Jayatilleke E, Herbert V, et al. Cleavage of folates during ethanol metabolism. Role of acetaldehyde/xanthine oxidase-generated superoxide. Biochem J 1989; 257:277-80.
92. Stickel F, Choi SW, Kim YI, et al. Effect of chronic alcohol consumption on total plasma homocysteine level in rats. Alcoholism Clinical and Experimental Research 2000; 24259-64.
93. Ferre N, Gomez F, Camps J, et al. Plasma homocysteine concentrations in patients with liver cirrhosis. Clin Chem 2002; 48:183-5.
94. Halsted CH, Villanueva JA, Devlin AM, et al. Folate deficiency disturbs hepatic methionine metabolism and promotes liver injury in the ethanol-fed micropig. Proc Natl Acad Sci U S A 2002; 99:10072-7.
95. Fell D,& Selhub J. Disruption of thymidylate synthesis and glycine-serine interconversion by L-methionine and L-homocystine in Raji cells. Biochim Biophys Acta 1990; 1033:80-4.
96. Villanueva JA,& Halsted, CH. Hepatic transmethylation reactions in micropigs with alcoholic liver disease. Hepatology 2004; 39:1303-10.
97. Park El, Renduchintala MS, Garrow TA. Diet-induced changes in hepatic betaine-homocysteine methyltransferase activity are mediated by changes in the steady-state level of its RNA. J Nutr Biochem 1997; 8:541-5.
98. Frasca V RB, Matthews RG. In Vitro inactivation of methionine synthase by nitrous oxide. The Journal of Biological Chemistry 1986; 261:15823-6.
99.章斐然,郝玲,王建英,陆渊源,朱敏敏,蒋新液,俞楠.吸烟对血浆叶酸水平的影响.实用预防医学 2003；10(6)：1017.
100. Luciano CG, Santiago RA, Celia MC, et al. Homocysteine Concentrations and Molecular Analysis in Patients with Congenital Heart Defects. Archives of Medical Research 2007; 38:212-218.
101. van beynum IM, Heijer MD, Blom HJ, et al. The MTHFR 677C/T polymorphism and the risk of congenital heart defects:a literature review and meta-analysis. QJ Med 2007; 100:743-753.
102. Anna VH, Johannes B, Fakhredin ST, et al. Hyperhomocysteinemia and MTHFR Polymorphisms in Association With Orofacial Clefts and Congenital Heart Defects:A Meta-Analysis. American Journal of Medical Genetics Part A 2007; 143A:952-960.
103. Van Beynum IM, Kapusta L, den Heijer M, et al. Maternal MTHFR 677C>T is a risk factor for congenital heart defects:effect modification by periconceptional folate supplementation. European Heart Journal 2006; 27:981-7.
104. Hobbs CA, James SJ, Parsian A, et al. Congenital heart defects and genetic variants in the methylenetetrahydrofolate reductase gene. Journal of Medical Genetics 2006; 43:162-6.
105. Junker R, Kotthoff S, Vielhaber H, et al. Infant methylenetetrahydrofolate reductase 677TT genotype is a risk factor for congenital heart disease. Cardiovascular Research 2001; 51:251-4.
106. Shaw GM, lovannisci DM, Yang W, et al. Risks of human conotruncal heart defects associated with 32 single nucleotide polymorphisms of selected cardiovascular disease-related genes. American Journal of Medical Genetics Part A. 2005; 138:21-6.
107. Storti S, Vittorini S, lascone MR, et al. Association between 5,10-methylenetetrahydrofolate reductase C677T and A1298C polymorphisms and conotruncal heart defects. Clinical Chemistry and Laboratory Medicine 2003; 41:276-80.
108. Wenstrom KD, Johanning GL, Johnston KE, et al. Association of the C677T methylenetetrahydrofolate reductase mutation and elevated homocysteine levels with congenital cardiac malformations. American Journal of Obstetrics & Gynecology 2001; 184:806-17.
109. Van Beynum IM, den Heijer M, Blom HJ, et al. The MTHFR 677C 110. Barbosa PR, Stabler SP, Machado ALK, et al. Association between decreased vitamin levels and MTHFR, MTR and MTRR gene polymorphisms as determinants for elevated total homocysteine concentrations in pregnant women. European Journal of Clinical Nutrition 2008; 62:1010-21.
111. Blom HJ, Shaw GM, den Heijer M, et al. Neural tube defects and folate:case far from closed. Nature Reviews Neuroscience 2006; 7:724-31.
112. Juriloff DM,& Harris MJ. Mouse models for neural tube closure defects. Hum Mol Genet 2000; 9:993-1000.
113. Cabrera RM, Hill DS, Etheredge AJ, et al. Investigations into the etiology of neural tube defects. Birth Defects Res. C Embryo Today 2004; 72:330-344.
114. Copp AJ. Neurulation in the cranial region-normal and abnormal. J Anat 2005; 207:623-635.
115. Greene N & Copp AJ. Mouse models of neural tube defects:investigating preventive mechanisms. Am J Med Genet CSemin Med Genet 2005; 135:31-41.
116. Chen Z, et al. Mice deficient in methylenetetrahydrofolate reductase exhibit hyperhomocysteinemia and decreased methylation capacity, with neuropathology and aortic lipid deposition. Hum Mol Genet 2001; 10:433-443.
117. Watanabe, M. et al. Mice deficient in cystathionine P-synthase:animal models for mild and severe homocyst(e)inemia. Proc Natl Acad Sci USA 1995; 92:1585-1589.
118. Swanson, D.A. et al. Targeted disruption of the methionine synthase gene in mice. Molecular and Cellular Biology 2001; 21:1058-1065.
119. Elmore, C.L. et al. Metabolic derangement of methionine and folate metabolism in mice deficient in methionine synthase reductase. Molecular Genetics and Metabolism 2007; 91:85-97.
120. Deng, L. D. et al. Methionine synthase reductase deficiency results in adverse reproductive outcomes and congenital heart defects in mice. Molecular Genetics and Metabolism 2008; 94:336-342.
121. Lentz SR, Sobey CG, Piegors DJ, et al. Vascular dysfunction in monkeys with diet-induced hyperhomocysteinemia. J Clin Investig 1996; 98:24-29.
122. Bernardo A, Julio CR, Maria TC. Metabolism of homocysteine and its relationship with cardiovascular disease. J Thrombosis Thrombolysis 2004; 18(2):75-87.
123. Jakubowski H. Molecular basis of homocysteine toxicity in human. Cell Mol Life Sci 2004; 61:470-487.
124. Marit JB, Regine PM, Robert EP et al. Cardiac outflow tract malformations in chick embryos exposed to homocysteien. Cardiovascular research 2004; 64:365-373.
125. Fisher MC, Zeisel SH, Mar MH, Sadler TW. Perturbations in choline metabolism cause neural tube defects in mouse embryos in vitro. FASEB Journal 2002; 16(6):619-621
126. Benoit G. Bruneau. The developmental genetics of congenital heart disease. Nature,2008; 451(21):943-48.
127. Sturman JA,Gaull G, Raiha NCR. Absence of cystathionase in human fetal liver:is cystine essential? Science 1970; 169:74-6.
128. Mishra PJ, Humeniuk R, Mishra PJ, et al. A miR-24 microRNA binding-site polymorphism in dihydrofolate reductase gene leads to methotrexate resistance. Proceedings of the National Academy of Sciences 2007;104:13513-8.
129. Leclerc DWA, Dumas R, Gafuik C, et al. Cloning and mapping of a cDNA for methionine synthase reductase, a flavoprotein defective in patients with homocystinuria. Proc Nat AcadS ci 1998; 95:3059-64.
130. Wilson APR, Wu Q, Leclerc D, et al. A common variant in methionine synthase reductase combined with low cobalamin (vitamin B12) increases risk for spina bifida. Mol Genet Metab 1999; 67:317-23.
131.李金平,李丽帆,方显明.胱硫醚β-合酶研究进展.西部医学2006；18(5)：657-9.
132. Sperandeo MP dFR, Andria G, Sebastio G. A 68-bp insertion found in a homocystinuric patient is a common variant and is skipped by alternative splicing of the cystathionine β-synthase mRNA. Am J Hum Genet 1996; 59:1391-3.
133.陈卓,单可人,齐晓岚,渊谢,婷张,骄马,吴昌学,毅李,艳赵,雁肖,任锡麟.贵州西江苗族、三都水族胱硫醚合酶844ins68位点的研究.贵阳医学院学报2005；30(5)：403-5.
134. Kluijtmans LAJ BG, Trijbels FJM, et al. A common 844 INS68 insertion variant in the cystathionine β-synthase gene. Biochem Mol Med 1997; 62:23-5.
135. Kozich VK, de Franchis R, Fowler BB, et al. Hyperhomocysteinemia in premature arterial disease:Examination of cystathionineβ-synthase alleles at the molecular level. Hum Mol Genet 1995;4:623-29.
136. Kraus JP, Oliveriusova J, Sokolova J, et al. The human Cystathionine β-synthase (CBS) gene:complete sequence, alternative splicing, and polymorphisms. Genomics 1998; 52:312-24.
137. Leo AJK, Godfried HJ, Jan PK, et al. The Molecular Basis of Cystathionine b-Synthase Deficiency in Dutch Patients with Homocystinuria:Effect of CBS Genotype on Biochemical and Clinical Phenotype and on Response to Treatment. Am J Hum Genet 1999; 65:56-67.
138. Watanabe M, Osada J, Aratani Y, Kluckman K, Reddick R, Malinow MR, Maeda N. Mice deficient in cystathionine betasynthase:animal models for mild and severe homocyst(e)inemia. Proc Natl Acad Sci USA 1995; 92:1585-1589.
139. Dayal S, Bottiglieri T, Arning E, Maeda N, Malinow MR, Sigmund CD, Heistad DD, et al. Endothelial dysfunction and elevation of S-adenosylhomocysteine in cystathionine beta-synthase-deficient mice. Circ Res 2001; 88:1203-1209.
140. Eberhardt RT, Forgione MA, Cap A, Leopold JA, Rudd MA, Trolliet M, Heydrick S, et al. Endothelial dysfunction in a murine model of mild hyperhomocyst(e)inemia. J Clin Invest 2000; 106:483-491.
141. Lentz SR, Erger RA, Dayal S, Maeda N, Malinow MR, Heistad DD, Faraci FM.Folate dependence of hyperhomocysteinemia and vascular dysfunction in cystathionine beta-synthasedeficient mice. Am J Physiol Heart 2000; 279:H970-975.
142. Weiss N, Heydrick S, Zhang YY, Bierl C, Cap A, Loscalzo J. Cellular redox state and endothelial dysfunction in mildly hyperhomocysteinemic cystathionine beta-synthase-deficient mice. Arterioscler Thromb Vasc Biol 2002; 22:34-41.
143. Quere I, Paul V, Rouillac C, Janbon C, London J, Demaille J, Kamoun P, et al. Spatial and temporal expression of the cystathionine beta-synthase gene during early human development. Biochem Biophys Res Commun 1999; 254:127-137.
144. Robert K, Vialard F, Thiery E, et al. Expression of the Cystathionine B Synthase (CBS) Gene During Mouse Development and Immunolocalization in Adult Brain. The Journal of Histochemistry & Cytochemistry 2003; 51(3):363-71.
145. Ladner RD. The role of dUTPase and uracil-DNA repair in cancer chemotherapy. Curr. Protein PeptSci 2001; 2:361-70.
146. E. Chu CJA. The role of thymidylate synthase as an RNA binding protein. BioEssay 1996; 18191-8.
147. Marsh SCD, Li T., Liu X, et al. Ethnic variation in the thymidylate synthase enhancer region polymorphism among Caucasian and Asian populations. Genomics 1999 58 310-2.
148. Luo HR LX, Yao YG, et al. Length polymorphism of thymidylate synthase regulatory region in Chinese population and evolution of the novel alleles. Biochem Genet 2002; 40(1-2):41-51.
149. Kaneda S, Nalbantoglu J, Takeishi K, Shimizu K, Gotoh O, Seno T, Ayusawa D. Structural and functional analysis of the human thymidylate synthase gene. J Biol Chem 1990; 265 (33):20277-84.
150. Kaneda KT, Ayusawa D, Shimizu K, et al. Role in translation of a triple tandemly repeated sequence in the 5'untranslated region of human thymidylate synthase mRNA. Nucleic Acids Res 1987; 15:1259-70.
151. Kawakami KSD, Park, JM, Danenberg KD, et al. Different lengths of a polymorphic repeat sequence in the thymidylate synthase gene affect translational efficiency but not its gene expression Glin Cancer Res 2001; 7:4096-101.
152. Watanabe KKaG. Identification and Functional Analysis of Single Nucleotide Polymorphism in the Tandem Repeat Sequence of Thymidylate Synthase Gene. Cancer Res 2003; 63:6004-7.
153. Michael VM, Susan MW, Gregory C, et al. A Novel Single Nucleotide Polymorphism within the 5'Tandem Repeat Polymorphism of the Thymidylate Synthase Gene Abolishes USF-1 Binding and Alters Transcriptional Activity. Cancer Res 2003; 63:2898-904.
154. Silanes ILd, Esteller MPQaM. Aberrant regulation of messenger RNA 3'-untranslated region in human cancer. Cellular Oncology 2007; 29:1-17.
155. Rudolf PJ, Abdelmohsen KLA, Martindale JL, et al. Differential Stability of Thymidylate Synthase 3'-Untranslated Region Polymorphic Variants Regulated by AUF1. The Journal of Biological Chemistry 2006; 281 (33):23456-63.
156. Mandola MVSJ, Zhang W, Groshen S, Yu MC, Iqbal S, Lenz HJ, Ladner RD. A 6 bp polymorphism in the thymidylate synthase gene causes message instability and is associated with decreased intratumoral TS mRNA levels. Pharmacogenetics 2004; 14 (5):319-27.
157. Kim YI. Folate and DNA methylation:a mechanistic link between folate deficiency and colorectal cancer? Cancer Epidemiol Biomarkers Prev 2004; 13:511-9.
158. Harnack L, Jacobs DR, Nicodemus K, et al. Relationship of folate, vitamin B-6, vitamin B-12, and methionine intake to incidence of colorectal cancers. Nutr Cancer 2002; 43:152-158.
159. Fang JY, Xiao SD. Alteration ofDNA methylation in gastrointestinal carcinogenesis. J Gastroenterol Hepatol 2001; 16 (9):960-8.
160. Song J, Medline A, Mason JB, Gallinger S, Kim YI. Effects of dietary folate on intestinal tumorigenesis in the apcMin mouse. Cancer Res 2000; 60:5434-40.
161. Song J, Sohn KJ, Medline A, Ash C, Gallinger S, Kim YI. Chemopreventive effects of dietary folate on intestinal polyps in Apc/Msh2/mice. Cancer Res 2000; 60: 3191-9.
162. Kim YI. Folate, colorectal carcinogenesis, and DNA methylation:lessons from animal studies. Environ Mol Mutagen 2004; 44:10-25.
163. Martinez CA, Northrup H, Lin JI, et al. Genetic association study of putative functional single nucleotide polymorphisms of genes in folate metabolism and spina bifida. Am J Obstet Gynecol 2009; 201:394.e1-11.
164. Giovannucci E, Stampfer MJ, Colditz GA, et al. Multivitamin use, folate, and colon cancer in women in the Nurses' Health Study. Ann intern Med 1998; 129:517-24.
165. Yu K, Zhang J, Zhang JY, et al. Methionine synthase A2756G polymorphism and cancer risk:a meta-analysis. European Journal of Human Genetics 2010; 18:370-8.
166. Lu MP, Wang F and Qiu JR. Methionine synthase A2756G polymorphism and breast cancer risk:a meta analysis involving 18,953 subjects. Breast Cancer Research and Treatment 2010; 123:213-7.
167. Hu J, Zhou GW, Wang N, et al. MTRR A66G polymorphism and breast cancer risk: a meta-analysis. Breast Cancer Research and Treatment 2010; 124:779-84.
168. Ohnami S, Sato Y, Yoshimura K, et al. His595Tyr polymorphism in the Methionine Synthase Reductase (MTRR) gene is associated with pancreatic cancer risk. Gastroenterology 2008; 135:477-88.
169. Rubini M, Brusati R, Garattini G, et al. Cystathionine beta-synthase c.844ins68 gene variant and non-syndromic cleft lip and palate. American Journal of Medical Genetics 2005; 136A:368-72.
170. Brattstrom L, Israelsson B, Norrving B, et al. Impaired homocysteine metabolism in early-onset cerebral and peripheral occlusive arterial disease effects of pyridoxine and folic acid treatment. Atherosclerosis 1990; 81(1):51-60.
171. Tang B, Mustafa A, Gupta S, et al. Methionine-deficient diet induces post-transcriptional downregulation of cystathionine beta synthase. Nutrition 2010; 26:1170-5.
[1]Quinlivan EP, Gregory JF Ⅲ. Effect of food fortification on folic acid intake in the United States. Am J Clin Nutr 2003; 77:221-5.
[2]Pfeiffer CM, Caudill SP, Gunter EW, Osterloh J, Sampson EJ. Biochemical indicators of B vitamin status in the US population after folic acid fortification:results from the National Health and Nutrition Examination Survey 1999-2000. Am J Clin Nutr 2005; 82:442-50.
[3]Institute of Medicine. Folate. In:IOM, ed. Dietary reference intakes forthiamin, riboflavin, niacin, vitamin B6, folate, vitamin B12, pantothenic acid, biotin, and choline. Washington, DC:National Academies Press,1998.
[4]Wright AJ, Dainty JR, Finglas PM. Folic acid metabolism in human subjects revisited:potential implications for proposed mandatory folic acid fortification in the UK. BrJ Nutr 2007; 98:667-75.
[5]Bailey SW, Syslo MC, Ayling J. An assay for dihydrofolate reductase in human tissues byHPLC with fluorometric detection. FASEB J 2002; 16:A267(abstr).
[6]Steven WB and June EA. The extremely slow and variable activity of dihydrofolate reductase in human liver and its implications for high folic acid intake. P Natl Acad Sci 2009; 106(36):15424-15429.
[7]Troen AM, Mitchell B, Sorensen B, et al. Unmetabolized folic acid in plasma is associated with reduced natural killer cell cytotoxicity among postmenopausal women. J Nutr 2006; 136:189-94.
[8]Pfeiffer CM, Fazili Z, McCoy L, Zhang M, Gunter EW. Determination of folate vitamers in human serum by stable-isotope-dilution tandem mass spectrometry and comparison with radioassay and microbiologic assay. Clin Chem 2004; 50:423-32.
[9]Sweeney MR, McPartlin J, Weir DG, et al. Evidence of unmetabolised folic acid in cord blood of newborn and serum of 4-day-old infants. Br J Nutr 2005; 94:727-30.
[10]Ashokkumar B, Mohammed ZM, Vaziri ND, Said HM. Effect of folate oversupplementation on folate uptake by human intestinal and renal epithelial cells. Am J Clin Nutr 2007; 86:159-166.
[11]Kim YI. Role of folate in colon cancer development and progression. J Nutr 2003; 133(suppl):3731S-9S.
[12]Kim YI. Folate:a magic bullet or a double edged sword for colorectal cancer prevention? Gut 2006; 55:1387-9.
[13]Kim YI. Does a high folate intake increase the risk of breast cancer? Nutr Rev 2006; 64:468-75.
[14]Song J, Medline A, Mason JB, Gallinger S, Kim YI. Effects of dietary folate on intestinal tumorigenesis in the apcMin mouse. Cancer Res 2000; 60:5434-40.
[15]Song J, Sohn KJ, Medline A, Ash C, Gallinger S, Kim YI. Chemopreventive effects of dietary folate on intestinal polyps in Apc/Msh2/mice. Cancer Res 2000; 60: 3191-9.
[16]Kim YI. Folate, colorectal carcinogenesis, and DNA methylation:lessons from animal studies. Environ Mol Mutagen 2004; 44:10-25.
[17]Bingham SA, Norat T, Moskal A, et al. Is the association with fiber from foods in colorectal cancer confounded by folate intake? Cancer Epidemiol Biomarkers Prev 2005; 14:1552-6.
[18]Dameshek W. Editorial:the use of folic acid antagonists in acute leukemia. Blood 1948; 3:1057-8.
[19]Kato I, Dnistrian AM, Schwartz M, et al. Serum folate, homocysteine and colorectal cancer risk in women:a nested case-control study. Br J Cancer 1999; 79: 1917-22.
[20]Van Guelpen B, Hultdin J, Johansson I, et al. Low folate levels may protect against colorectal cancer. Gut 2006; 55:1461-6.
[21]Stolzenberg-Solomon RZ, Chang S-C, Leitzmann MF, et al. Folate intake, alcohol use, and postmenopausal breast cancer risk in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial. Am J Clin Nutr 2006; 83:895-904.
[22]Ulrich CM. Folate and cancer prevention:a closer look at a complex picture. Am J Clin Nutr 2007; 86:271-3.
[23]Tworoger SS, Hecht JL, Giovannucci E, Hankinson SE. Intake of folate and related nutrients in relation to risk of epithelial ovarian cancer. AmJ Epidemiol 2006; 163: 1101-11.
[24]Reynolds E. Vitamin B12, folic acid, and the nervous system. Lancet Neurol 2006; 5:-949-60.
[25]Ulrich CM, Bigler J, Bostick R, Fosdick L, Potter JD. Thymidylate synthase promoter polymorphism, interaction with folate intake, and risk of colorectal adenomas. Cancer Res 2002; 62:3361-4.
[26]Morris MS, Jacques PF, Rosenberg IH, Selhub J. Folate and vitamin B12 status in relation to anemia, macrocytosis, and cognitive impairment among older Americans in the age of folic acid fortification. AmJ Clin Nutr 2007; 85:193-200.
[27]Komatsu M, Tsukamoto I. Effect of folic acid on thymidylate synthase and thymidine kinase in regenerating rat liver after partial hepatectomy. Biochim Biophys Acta 1998; 1379:289-96.
[28]MorrisMC,Evans DA, Bienias JL, et al. Dietary folate and vitamin B12 intake and cognitive decline among community-dwelling older persons. Arch Neurol 2005; 62: 641-5.
[29]Lange H, Suryapranata H, De Luca G, et al. Folate therapy and in-stent restenosis after coronary stenting. N Engl J Med 2004; 350:2673-81.
[30]HOPE. Homocysteine lowering with folic acid and B vitamins in vascular disease. N Engl J Med 2006; 354:1567-77.
[31]Bonaa KH, Njolstad I, Ueland PM, et al. Homocysteine Lowing and cardiovascular events after acute myocardial infarction. N Engl J Med 2006; 354:1578-88.
[32]Yajnik CS, Deshpande SS, Jackson AA, et al. Vitamin B12 and folate concentrations during pregnancy and insulin resistance in the offspring:the Pune Maternal Nutrition Study. Diabetologia 2008; 51:29-38.
[33]Regine PST, Sylvia AOB, Dennis K, et al. Periconceptional maternal folic acid use of 400u.g per day is related to increased methylation of the IGF2 gene in the very young child. Plos One 2009; 4(11)e7845:1-5.
[34]Reyes-Engel A, Munoz E, Gaitan MJ, et al. Implications on human fertility of the 677C>T and 1298A>C polymorphisms of the MTHFR gene:consequences of a possible genetic selection. Mol Hum Reprod 2002; 8:952-7.
[35]Zetterberg H. Methylenetetrahydrofolate reductase and transcobalamin genetic polymorphisms in human spontaneous abortion:biological and clinical implications. Reprod Biol Endocrinol 2004;2:7.
[36]Whitehead AS. Changes in MTHFR genotype frequencies over time. Lancet 1998; 352:1784-5.
[37]Lucock M, Yates Z. Folic acid-itamin and panacea or genetic time bomb? Nat Rev Genet 2005; 6:235-40.
[38]McGuire JJ. Anticancer antifolates:current status and future directions. Curr Pharm Des 2003; 9:2593-613.
[39]Robien K. Folate during antifolate chemotherapy:what we know and do not know. Nutr Clin Pract 2005; 20:411-22.
[40]Hazra A, Kraft P, Lazarus R, et al. Genome-wide significant predictors of metabolites in the one-carbon metabolism pathway. Hum Mol Genet 2009; 18(23): 4677-4687.
2. Hoffman, J. I. Incidence of congenital heart disease:Ⅱ.Prenatal incidence. Pediatr Cardiol 1995; 16:155-65.
3. Miller, S. P. et al. Abnormal brain development in newborns with congenital heart disease.M Engl J Med 2007; 357:1928-1938.
4. Pierpont, M. E. et al. Genetic basis for congenital heart defects:current knowledge: a scientific statement from the American Heart Association Congenital Cardiac Defects Committee, Council on Cardiovascular Disease in the Young:endorsed by the American Academy of Pediatrics. Circulation 115,3015-3038 (2007).
5. Jenkins, K. J. et al. Noninherited risk factors and congenital cardiovascular defects: current knowledge:a scientific statement from the American Heart Association Council on Cardiovascular Disease in the Young:endorsed by the American Academy of Pediatrics. Circulation 115,2995-3014 (2007).
6. Cooper, W. O. et al., Major congenital malformations after first-trimester exposure to ACE inhibitors. N. Engl. J. Med.354,2443-2451 (2006).
7. Olson, E. N. Gene regulatory networks in the evolution and development of the heart. Science 313,1922-1927 (2006).
8. Srivastava, D. Making or breaking the heart:from lineage determination to morphogenesis. Cell 126,1037-1048 (2006).
9. Buckingham, M., Meilhac, S.& Zaffran, S. Building the mammalian heart from two sources of myocardial cells. Nature Rev. Genet.6,826-835 (2005).
10. Kattman, S. J., Huber, T. L.& Keller, G. M. Multipotent flk-1+cardiovascular progenitor cells give rise to the cardiomyocyte, endothelial, and vascular smooth muscle lineages. Dev. Cell 11,723-732 (2006).
11. Wu, S. M. et al. Developmental origin of a bipotential myocardial and smooth muscle cell precursor in the mammalian heart. Cell 127,1137-1150 (2006).
12. Moretti, A. et al. Multipotent embryonic ISL1+progenitor cells lead to cardiac, smooth muscle, and endothelial cell diversification. Cell 127,1151-1165 (2006).
13. Tzahor, E. Wnt/β-catenin signaling and cardiogenesis:timing does matter. Dev. Cell 13,10-13 (2007).
14. Schoenebeck, J. J., Keegan, B. R.& Yelon, D. Vessel and blood specification override cardiac potential in anterior mesoderm. Dev. Cell 13,254-267 (2007).
15. Keegan, B. R., Feldman, J. L., Begemann, G., Ingham, P. W.& Yelon, D. Retinoic acid signaling restricts the cardiac progenitor pool. Science 307,247-249 (2005).
16. Prall, O. W. et al. An Nkx2-5/Bmp2/Smad1 negative feedback loop controls second heart field progenitor specification and proliferation. Cell 128,947-959 (2007).
17. Lickert, H. et al. Baf60c is essential for function of BAF chromatin remodelling complexes in heart development. Nature 432,107-112 (2004).
18. Gottlieb, P. D. et al. Bop encodes a muscle-restricted protein containing MYND and SET domains and is essential for cardiac differentiation and morphogenesis. Nature Genet.31,25-32 (2002).
19. Montgomery, R. L. et al. Histone deacetylases 1 and 2 redundantly regulate cardiac morphogenesis, growth, and contractility. Genes Dev.21,1790-1802 (2007).
20. Basson, C. T. et al. Mutations in human TBX5 cause limb and cardiac malformation in Holt-Oram syndrome. Nature Genet.15,30-35 (1997).
21. Li, Q. Y. et al. Holt-Oram syndrome is caused by mutations in TBX5, a member of the Brachyury (T) gene family. Nature Genet.15,21-29 (1997).
22. Schott, J.-J. et al. Congenital heart disease caused by mutations in the transcription factor NKX2-5. Science 281,108-111 (1998).
23. Benson, D. W. et al. Mutations in the cardiac transcription factor NKX2.5 affect diverse cardiac developmental pathways. J. Clin. Invest.104,1567-1573 (1999).
24. Biben, C. et al. Cardiac septal and valvular dysmorphogenesis in mice heterozygous for mutations in the homeobox gene Nkx2-5. Circ. Res.87,888-895 (2000).
25. Bruneau, B. G. et al. A murine model of Holt-Oram syndrome defines roles of the T-box transcription factor Tbx5 in cardiogenesis and disease. Cell 106,709-721 (2001).
26. Hiroi, Y. et al. Tbx5 associates with Nkx2-5 and synergistically promotes cardiomyocyte differentiation. Nature Genet.28,276-280 (2001).
27. Garg, V. et al. GATA4 mutations cause human congenital heart defects and reveal an interaction with TBX5. Nature 424,443-447 (2003).
28. Lindsay, E. A. et al. Tbx1 haploinsufficiency in the DiGeorge syndrome region causes aortic arch defects in mice. Nature 410,97-101 (2001).
29. Merscher, S. et al. TBX1 is responsible for cardiovascular defects in velo-cardio-facial/DiGeorge syndrome. Cell 104,619-629 (2001).
30. Yagi, H. et al. Role of TBX1 in human del22q11.2 syndrome. Lancet 362, 1366-1373 (2003).
31. Hu, T. et al. Tbx1 regulates fibroblast growth factors in the anterior heart field through a reinforcing autoregulatory loop involving forkhead transcription factors. Development 131,5491-5502 (2004).
32. Xu, H. et al. Tbx1 has a dual role in the morphogenesis of the cardiac outflow tract. Development 131,3217-3227 (2004).
33. Guris, D. L, Duester, G., Papaioannou, V. E.& Imamoto, A. Dose-dependent interaction of Tbx1 and Crkl and locally aberrant RA signaling in a model of del22q11 syndrome. Dev. Cell 10,81-92 (2006).
34. Moon, A. M. et al. Crkl deficiency disrupts Fgf8 signaling in a mouse model of 22q11 deletion syndromes. Dev. Cell 10,71-80 (2006).
35. Al-Baradie, R. et al. Duane radial ray syndrome (Okihiro syndrome) maps to 20q13 and results from mutations in SALL4, a new member of the SAL family. Am. J. Hum. Genet.71,1195-1199 (2002).
36. Kohlhase, J. et al. Okihiro syndrome is caused by SALL4 mutations. Hum. Mol. Genet.11,2979-2987 (2002).
37. Kirk, E. P. et al. Mutations in cardiac T-box factor gene TBX20 are associated with diverse cardiac pathologies, including defects of septation and valvulogenesis and cardiomyopathy. Am. J. Hum. Genet.81,280-291 (2007).
38. Koshiba-Takeuchi, K. et al. Cooperative and antagonistic interactions between Sall4 and Tbx5 pattern the mouse limb and heart. Nature Genet.38,175-183 (2006).
39. Satoda, M. et al. Mutations in TFAP2B cause Char syndrome, a familial form of patent ductus arteriosus. Nature Genet.25,42-46 (2000).
40. Muncke, N. et al. Missense mutations and gene interruption in PROSIT240, a novel TRAP240-like gene, in patients with congenital heart defect (transposition of the great arteries). Circulation 108,2843-2850 (2003).
41. Hove, J. R. et al. Intracardiac fluid forces are an essential epigenetic factor for embryonic cardiogenesis. Nature 421,172-177 (2003).
42. Auman, H. J. et al. Functional modulation of cardiac form through regionally confined cell shape changes. PLoS Biol.5, e53 (2007).
43. Yashiro, K., Shiratori, H.& Hamada, H. Haemodynamics determined by a genetic programme govern asymmetric development of the aortic arch. Nature 450, 285-288 (2007).
44. Ching, Y. H. et al. Mutation in myosin heavy chain 6 causes atrial septal defect. Nature Genet.37,423-428 (2005).
45. Barnett, J. V.& Desgrosellier, J. S. Early events in valvulogenesis:a signaling perspective. Birth Defects Res. C Embryo Today 69,58-72 (2003).
46. Timmerman, L. A. et al. Notch promotes epithelial-mesenchymal transition during cardiac development and oncogenic transformation. Genes Dev.18,99-115 (2004).
47. Garg, V. et al. Mutations in NOTCH1 cause aortic valve disease. Nature 437, 270-274 (2005).
48. Li, L. et al. Alagille syndrome is caused by mutations in human Jaggedl, which encodes a ligand for Notch1. Nature Genet.16,243-251 (1997).
49. Oda, T. et al. Mutations in the human Jaggedl gene are responsible for Alagille syndrome.Nature Genet.16,235-242 (1997).
50. McDaniell, R. et al. NOTCH2 mutations cause Alagille syndrome, a heterogeneous disorder of the notch signaling pathway. Am. J. Hum. Genet.79,169-173 (2006).
51. Gelb, B. D.& Tartaglia, M. Noonan syndrome and related disorders:dysregulated RASmitogen activated protein kinase signal transduction. Hum. Mol. Genet.15 (Spec. No.2), R220-R226 (2006).
52. Pandit, B. et al. Gain-of-function RAF1 mutations cause Noonan and LEOPARD syndromes with hypertrophic cardiomyopathy. Nature Genet.39,1007-1012 (2007).
53. Razzaque, M. A. et al. Germline gain-of-function mutations in RAF1 cause Noonan syndrome. Nature Genet.39,1013-1017 (2007).
54. Roberts, A. E. et al. Germline gain-of-function mutations in SOS1 cause Noonan syndrome. Nature Genet.39,70-74 (2007).
55. Tartaglia, M. et al. Gain-of-function SOS1 mutations cause a distinctive form of Noonan syndrome. Nature Genet.39,75-79 (2007).
56. Araki, T. et al. Mouse model of Noonan syndrome reveals cell type-and gene dosagedependent effects of Ptpn11 mutation. Nature Med.10,849-857 (2004).
57. Ambros, V. The functions of animal microRNAs. Nature 431,350-355 (2004).
58. van Rooij, E.& Olson, E. N. MicroRNAs:powerful new regulators of heart disease and provocative therapeutic targets. J. Clin. Invest.117,2369-2376 (2007).
59. Zhao, Y., Samal, E.& Srivastava, D. Serum response factor regulates a muscle-specific microRNA that targets Hand2 during cardiogenesis. Nature 436, 214-220 (2005).
60. Zhao, Y. et al. Dysregulation of cardiogenesis, cardiac conduction, and cell cycle in mice lacking microRNA-1-2. Cell 129,303-317 (2007).
61. Warnes, C. A. The adult with congenital heart disease:born to be bad? J. Am. Coll. Cordiol.46,1-8 (2005).
62. Bisping, E. et al. Gata4 is required for maintenance of postnatal cardiac function and protection from pressure overload-induced heart failure. Proc. Natl Acad. Sci. USA 103,14471-14476 (2006).
63. Pashmforoush, M. et al. Nkx2-5 pathways and congenital heart disease; loss of ventricular myocyte lineage specification leads to progressive cardiomyopathy and complete heart block. Cell 2004; 117:373-86.
64. Acuna J, Yoon P, Erickson D. The prevention of neural tube defects with folic acid. Atlanta, GA:Centers for Disease Control and Prevention,1999.
65. Health Canada. Congenital anomalies in Canada:a perinatal health report. Ottawa:Minister of Public Works and Governmental Services,2002.
66. Botto LD, Mulinare J, Erickson JD. Do multivitamin or folic acid supplements reduce the risk for congenital heart defects? Evidence and gaps. Am J Med Genet A 2003; 121:95-101.
67. Botto LD, Olney RS, Erickson JD. Vitamin supplements and the risk for congenital anomalies other than neural tube defects. Am J Med Genet C Semin Med Genet 2004; 125:12-21.
68. Huhta JC, Hernandez-Robles JA. Homocysteine, folate, and congenital heart defects. Fetal Pediatr Pathol 2005; 24:71-9.
69. Bailey LB, Berry RJ. Folic acid supplementation and the occurrence ofcongenital heart defects, orofacial clefts, multiple births, andmiscarriage. Am J Clin Nutr 2005; 81:1213-7S.
70. Huhta JC, Linask K, Bailey L. Recent advances in the prevention ofcongenital heart disease. Curr Opin Pediatr 2006; 18:484-9.
71. Jenkins KJ, Correa A, Feinstein JA, et al. Noninherited risk factors and congenital cardiovascular defects:current knowledge:a scientific statement from the American Heart Association Council on Cardiovascular Disease in the Young:endorsed by the American Academy of Pediatrics. Circulation 2007; 115:2995-3014.
72. Lucock M. Folic acid:Nutritional biochemistry, molecular biology, and role in disease processes. Molecular Genetics and Metabolism 2000; 71:121-38.
73.李竹,刘向东,陈亚华.神经管畸形不同发病地区婚检妇女叶酸缺乏率的研究.中国优生优育1996；1：1-4.
74.郝玲,田熠华,唐仪,李竹.我国部分地区岁人群血浆叶酸水平与年龄性别差异比较.营养学报2002；24(4)：352-4.
75. Gideon FNG, Yechiel F, Arie BY, et al. A common mutation A1298C in human methylenetetrahydrofolate reductase gene:association with plasma total homocysteine and folate concentrations. J Nutr 1999;129:1656-61.
76. Rosenquist TH, Ratashak SA, Selhub J. Homocysteine induces congenital defects of the heart and neural tube:effect of folic acid. Proc Natl Acad Sci USA,1996; 93:15227-15232.
77. Peter K, Chiang RG, Jacov T, et al. S-Adenosylmethionine and methylation. Faseb J 1996;10:471-80.
78. Eloranta TO, Kajander EO. Catabolism and lability of S-adenosyl-L-methionine in rat liver extracts. Biochem J 1994; 224:137-44.
79. WA L. S-adenosylmethionine:jack of all trades and master of everything? Biochem Soc Trans 2006 34 (Pt 2):330-3.
80. Fenech M. The role of folic acid and Vitamin B12 in genomic stability of human cells. Mutation Research 2001;475:57-67.
81. Kristina M von Castel-Dunwoody GPK, Karla P Shelnutt, Jaimie D Vaughn, Elizabeth R Griffin,David R Maneval, Douglas W Theriaque, and Lynn B Bailey.
Transcobalamin 776C-G polymorphism negatively affects vitamin B-12 metabolism. Am J Clin Nutr 2005; 81:1436-9.
82. Brunaud L AJ, Ayav A, Gerard P, Namour F, Antunes L, Braun M, Bronowicki JP, Bresler L, Gueant JL. Effects of vitamin B12 and folate deficiencies on DNA methylation and carcinogenesis in rat liver. Clin Chem Lab Med 2003; 41 (8):1012-9.
83. Jacob RA JD, Allman-Farinelli MA, Swendseid ME. Folate nutriture alters choline status of women and men fed low choline diets. J Nutr 1999; 129:712-7.
84. Stephanie E Chiuve ELG, Susan E Hankinson, Steven H Zeisel, Lauren W Dougherty,Walter C Willett, and Eric B Rimm. The association between betaine and choline intakes and the plasma concentrations of homocysteine in women. Am J Clin Nutr 2007; 86:1073-81.
85. Zeisel SH ZT, daCosta K, Pomfret EA. Effect of choline deficiency on S-adenosylmethionine and methionine concentrations in rat liver. Biochem J 1989; 259:725-9.
86. SA. C. Betaine in human nutrition. Am J Clin Nutr 2004;80:539-49.
87. SEITZ GPaHK. Alcohol and Cancer. Alcohol & Alcoholism 2004;39 (3):155-65.
88. Halsted CH, Villanueva JA, Devlin AM, et al. Metabolic interactions of alcohol and folate.J Nutr 2002; 132:2367S-72S.
89. Halsted CH, Robles EA., Mezey E. Intestinal malabsorption in folate-deficient alcoholics.. Gastroenterology 1973; 64:526-32.
90. Villanueva JA, Devlin, A. M.,& Halsted, C. H. Reduced folate carrier:tissue distribution and effects of chronic ethanol intake in the micropig.. Alcohol Clin Exp Res 2001;25:415-20.
91. Shaw S, Jayatilleke E, Herbert V, et al. Cleavage of folates during ethanol metabolism. Role of acetaldehyde/xanthine oxidase-generated superoxide. Biochem J 1989; 257:277-80.
92. Stickel F, Choi SW, Kim YI, et al. Effect of chronic alcohol consumption on total plasma homocysteine level in rats. Alcoholism Clinical and Experimental Research 2000; 24259-64.
93. Ferre N, Gomez F, Camps J, et al. Plasma homocysteine concentrations in patients with liver cirrhosis. Clin Chem 2002; 48:183-5.
94. Halsted CH, Villanueva JA, Devlin AM, et al. Folate deficiency disturbs hepatic methionine metabolism and promotes liver injury in the ethanol-fed micropig. Proc Natl Acad Sci U S A 2002; 99:10072-7.
95. Fell D,& Selhub J. Disruption of thymidylate synthesis and glycine-serine interconversion by L-methionine and L-homocystine in Raji cells. Biochim Biophys Acta 1990; 1033:80-4.
96. Villanueva JA,& Halsted, CH. Hepatic transmethylation reactions in micropigs with alcoholic liver disease. Hepatology 2004; 39:1303-10.
97. Park El, Renduchintala MS, Garrow TA. Diet-induced changes in hepatic betaine-homocysteine methyltransferase activity are mediated by changes in the steady-state level of its RNA. J Nutr Biochem 1997; 8:541-5.
98. Frasca V RB, Matthews RG. In Vitro inactivation of methionine synthase by nitrous oxide. The Journal of Biological Chemistry 1986; 261:15823-6.
99.章斐然,郝玲,王建英,陆渊源,朱敏敏,蒋新液,俞楠.吸烟对血浆叶酸水平的影响.实用预防医学 2003；10(6)：1017.
100. Luciano CG, Santiago RA, Celia MC, et al. Homocysteine Concentrations and Molecular Analysis in Patients with Congenital Heart Defects. Archives of Medical Research 2007; 38:212-218.
101. van beynum IM, Heijer MD, Blom HJ, et al. The MTHFR 677C/T polymorphism and the risk of congenital heart defects:a literature review and meta-analysis. QJ Med 2007; 100:743-753.
102. Anna VH, Johannes B, Fakhredin ST, et al. Hyperhomocysteinemia and MTHFR Polymorphisms in Association With Orofacial Clefts and Congenital Heart Defects:A Meta-Analysis. American Journal of Medical Genetics Part A 2007; 143A:952-960.
103. Van Beynum IM, Kapusta L, den Heijer M, et al. Maternal MTHFR 677C>T is a risk factor for congenital heart defects:effect modification by periconceptional folate supplementation. European Heart Journal 2006; 27:981-7.
104. Hobbs CA, James SJ, Parsian A, et al. Congenital heart defects and genetic variants in the methylenetetrahydrofolate reductase gene. Journal of Medical Genetics 2006; 43:162-6.
105. Junker R, Kotthoff S, Vielhaber H, et al. Infant methylenetetrahydrofolate reductase 677TT genotype is a risk factor for congenital heart disease. Cardiovascular Research 2001; 51:251-4.
106. Shaw GM, lovannisci DM, Yang W, et al. Risks of human conotruncal heart defects associated with 32 single nucleotide polymorphisms of selected cardiovascular disease-related genes. American Journal of Medical Genetics Part A. 2005; 138:21-6.
107. Storti S, Vittorini S, lascone MR, et al. Association between 5,10-methylenetetrahydrofolate reductase C677T and A1298C polymorphisms and conotruncal heart defects. Clinical Chemistry and Laboratory Medicine 2003; 41:276-80.
108. Wenstrom KD, Johanning GL, Johnston KE, et al. Association of the C677T methylenetetrahydrofolate reductase mutation and elevated homocysteine levels with congenital cardiac malformations. American Journal of Obstetrics & Gynecology 2001; 184:806-17.
109. Van Beynum IM, den Heijer M, Blom HJ, et al. The MTHFR 677C
111. Blom HJ, Shaw GM, den Heijer M, et al. Neural tube defects and folate:case far from closed. Nature Reviews Neuroscience 2006; 7:724-31.
112. Juriloff DM,& Harris MJ. Mouse models for neural tube closure defects. Hum Mol Genet 2000; 9:993-1000.
113. Cabrera RM, Hill DS, Etheredge AJ, et al. Investigations into the etiology of neural tube defects. Birth Defects Res. C Embryo Today 2004; 72:330-344.
114. Copp AJ. Neurulation in the cranial region-normal and abnormal. J Anat 2005; 207:623-635.
115. Greene N & Copp AJ. Mouse models of neural tube defects:investigating preventive mechanisms. Am J Med Genet CSemin Med Genet 2005; 135:31-41.
116. Chen Z, et al. Mice deficient in methylenetetrahydrofolate reductase exhibit hyperhomocysteinemia and decreased methylation capacity, with neuropathology and aortic lipid deposition. Hum Mol Genet 2001; 10:433-443.
117. Watanabe, M. et al. Mice deficient in cystathionine P-synthase:animal models for mild and severe homocyst(e)inemia. Proc Natl Acad Sci USA 1995; 92:1585-1589.
118. Swanson, D.A. et al. Targeted disruption of the methionine synthase gene in mice. Molecular and Cellular Biology 2001; 21:1058-1065.
119. Elmore, C.L. et al. Metabolic derangement of methionine and folate metabolism in mice deficient in methionine synthase reductase. Molecular Genetics and Metabolism 2007; 91:85-97.
120. Deng, L. D. et al. Methionine synthase reductase deficiency results in adverse reproductive outcomes and congenital heart defects in mice. Molecular Genetics and Metabolism 2008; 94:336-342.
121. Lentz SR, Sobey CG, Piegors DJ, et al. Vascular dysfunction in monkeys with diet-induced hyperhomocysteinemia. J Clin Investig 1996; 98:24-29.
122. Bernardo A, Julio CR, Maria TC. Metabolism of homocysteine and its relationship with cardiovascular disease. J Thrombosis Thrombolysis 2004; 18(2):75-87.
123. Jakubowski H. Molecular basis of homocysteine toxicity in human. Cell Mol Life Sci 2004; 61:470-487.
124. Marit JB, Regine PM, Robert EP et al. Cardiac outflow tract malformations in chick embryos exposed to homocysteien. Cardiovascular research 2004; 64:365-373.
125. Fisher MC, Zeisel SH, Mar MH, Sadler TW. Perturbations in choline metabolism cause neural tube defects in mouse embryos in vitro. FASEB Journal 2002; 16(6):619-621
126. Benoit G. Bruneau. The developmental genetics of congenital heart disease. Nature,2008; 451(21):943-48.
127. Sturman JA,Gaull G, Raiha NCR. Absence of cystathionase in human fetal liver:is cystine essential? Science 1970; 169:74-6.
128. Mishra PJ, Humeniuk R, Mishra PJ, et al. A miR-24 microRNA binding-site polymorphism in dihydrofolate reductase gene leads to methotrexate resistance. Proceedings of the National Academy of Sciences 2007;104:13513-8.
129. Leclerc DWA, Dumas R, Gafuik C, et al. Cloning and mapping of a cDNA for methionine synthase reductase, a flavoprotein defective in patients with homocystinuria. Proc Nat AcadS ci 1998; 95:3059-64.
130. Wilson APR, Wu Q, Leclerc D, et al. A common variant in methionine synthase reductase combined with low cobalamin (vitamin B12) increases risk for spina bifida. Mol Genet Metab 1999; 67:317-23.
131.李金平,李丽帆,方显明.胱硫醚β-合酶研究进展.西部医学2006；18(5)：657-9.
132. Sperandeo MP dFR, Andria G, Sebastio G. A 68-bp insertion found in a homocystinuric patient is a common variant and is skipped by alternative splicing of the cystathionine β-synthase mRNA. Am J Hum Genet 1996; 59:1391-3.
133.陈卓,单可人,齐晓岚,渊谢,婷张,骄马,吴昌学,毅李,艳赵,雁肖,任锡麟.贵州西江苗族、三都水族胱硫醚合酶844ins68位点的研究.贵阳医学院学报2005；30(5)：403-5.
134. Kluijtmans LAJ BG, Trijbels FJM, et al. A common 844 INS68 insertion variant in the cystathionine β-synthase gene. Biochem Mol Med 1997; 62:23-5.
135. Kozich VK, de Franchis R, Fowler BB, et al. Hyperhomocysteinemia in premature arterial disease:Examination of cystathionineβ-synthase alleles at the molecular level. Hum Mol Genet 1995;4:623-29.
136. Kraus JP, Oliveriusova J, Sokolova J, et al. The human Cystathionine β-synthase (CBS) gene:complete sequence, alternative splicing, and polymorphisms. Genomics 1998; 52:312-24.
137. Leo AJK, Godfried HJ, Jan PK, et al. The Molecular Basis of Cystathionine b-Synthase Deficiency in Dutch Patients with Homocystinuria:Effect of CBS Genotype on Biochemical and Clinical Phenotype and on Response to Treatment. Am J Hum Genet 1999; 65:56-67.
138. Watanabe M, Osada J, Aratani Y, Kluckman K, Reddick R, Malinow MR, Maeda N. Mice deficient in cystathionine betasynthase:animal models for mild and severe homocyst(e)inemia. Proc Natl Acad Sci USA 1995; 92:1585-1589.
139. Dayal S, Bottiglieri T, Arning E, Maeda N, Malinow MR, Sigmund CD, Heistad DD, et al. Endothelial dysfunction and elevation of S-adenosylhomocysteine in cystathionine beta-synthase-deficient mice. Circ Res 2001; 88:1203-1209.
140. Eberhardt RT, Forgione MA, Cap A, Leopold JA, Rudd MA, Trolliet M, Heydrick S, et al. Endothelial dysfunction in a murine model of mild hyperhomocyst(e)inemia. J Clin Invest 2000; 106:483-491.
141. Lentz SR, Erger RA, Dayal S, Maeda N, Malinow MR, Heistad DD, Faraci FM.Folate dependence of hyperhomocysteinemia and vascular dysfunction in cystathionine beta-synthasedeficient mice. Am J Physiol Heart 2000; 279:H970-975.
142. Weiss N, Heydrick S, Zhang YY, Bierl C, Cap A, Loscalzo J. Cellular redox state and endothelial dysfunction in mildly hyperhomocysteinemic cystathionine beta-synthase-deficient mice. Arterioscler Thromb Vasc Biol 2002; 22:34-41.
143. Quere I, Paul V, Rouillac C, Janbon C, London J, Demaille J, Kamoun P, et al. Spatial and temporal expression of the cystathionine beta-synthase gene during early human development. Biochem Biophys Res Commun 1999; 254:127-137.
144. Robert K, Vialard F, Thiery E, et al. Expression of the Cystathionine B Synthase (CBS) Gene During Mouse Development and Immunolocalization in Adult Brain. The Journal of Histochemistry & Cytochemistry 2003; 51(3):363-71.
145. Ladner RD. The role of dUTPase and uracil-DNA repair in cancer chemotherapy. Curr. Protein PeptSci 2001; 2:361-70.
146. E. Chu CJA. The role of thymidylate synthase as an RNA binding protein. BioEssay 1996; 18191-8.
147. Marsh SCD, Li T., Liu X, et al. Ethnic variation in the thymidylate synthase enhancer region polymorphism among Caucasian and Asian populations. Genomics 1999 58 310-2.
148. Luo HR LX, Yao YG, et al. Length polymorphism of thymidylate synthase regulatory region in Chinese population and evolution of the novel alleles. Biochem Genet 2002; 40(1-2):41-51.
149. Kaneda S, Nalbantoglu J, Takeishi K, Shimizu K, Gotoh O, Seno T, Ayusawa D. Structural and functional analysis of the human thymidylate synthase gene. J Biol Chem 1990; 265 (33):20277-84.
150. Kaneda KT, Ayusawa D, Shimizu K, et al. Role in translation of a triple tandemly repeated sequence in the 5'untranslated region of human thymidylate synthase mRNA. Nucleic Acids Res 1987; 15:1259-70.
151. Kawakami KSD, Park, JM, Danenberg KD, et al. Different lengths of a polymorphic repeat sequence in the thymidylate synthase gene affect translational efficiency but not its gene expression Glin Cancer Res 2001; 7:4096-101.
152. Watanabe KKaG. Identification and Functional Analysis of Single Nucleotide Polymorphism in the Tandem Repeat Sequence of Thymidylate Synthase Gene. Cancer Res 2003; 63:6004-7.
153. Michael VM, Susan MW, Gregory C, et al. A Novel Single Nucleotide Polymorphism within the 5'Tandem Repeat Polymorphism of the Thymidylate Synthase Gene Abolishes USF-1 Binding and Alters Transcriptional Activity. Cancer Res 2003; 63:2898-904.
154. Silanes ILd, Esteller MPQaM. Aberrant regulation of messenger RNA 3'-untranslated region in human cancer. Cellular Oncology 2007; 29:1-17.
155. Rudolf PJ, Abdelmohsen KLA, Martindale JL, et al. Differential Stability of Thymidylate Synthase 3'-Untranslated Region Polymorphic Variants Regulated by AUF1. The Journal of Biological Chemistry 2006; 281 (33):23456-63.
156. Mandola MVSJ, Zhang W, Groshen S, Yu MC, Iqbal S, Lenz HJ, Ladner RD. A 6 bp polymorphism in the thymidylate synthase gene causes message instability and is associated with decreased intratumoral TS mRNA levels. Pharmacogenetics 2004; 14 (5):319-27.
157. Kim YI. Folate and DNA methylation:a mechanistic link between folate deficiency and colorectal cancer? Cancer Epidemiol Biomarkers Prev 2004; 13:511-9.
158. Harnack L, Jacobs DR, Nicodemus K, et al. Relationship of folate, vitamin B-6, vitamin B-12, and methionine intake to incidence of colorectal cancers. Nutr Cancer 2002; 43:152-158.
159. Fang JY, Xiao SD. Alteration ofDNA methylation in gastrointestinal carcinogenesis. J Gastroenterol Hepatol 2001; 16 (9):960-8.
160. Song J, Medline A, Mason JB, Gallinger S, Kim YI. Effects of dietary folate on intestinal tumorigenesis in the apcMin mouse. Cancer Res 2000; 60:5434-40.
161. Song J, Sohn KJ, Medline A, Ash C, Gallinger S, Kim YI. Chemopreventive effects of dietary folate on intestinal polyps in Apc/Msh2/mice. Cancer Res 2000; 60: 3191-9.
162. Kim YI. Folate, colorectal carcinogenesis, and DNA methylation:lessons from animal studies. Environ Mol Mutagen 2004; 44:10-25.
163. Martinez CA, Northrup H, Lin JI, et al. Genetic association study of putative functional single nucleotide polymorphisms of genes in folate metabolism and spina bifida. Am J Obstet Gynecol 2009; 201:394.e1-11.
164. Giovannucci E, Stampfer MJ, Colditz GA, et al. Multivitamin use, folate, and colon cancer in women in the Nurses' Health Study. Ann intern Med 1998; 129:517-24.
165. Yu K, Zhang J, Zhang JY, et al. Methionine synthase A2756G polymorphism and cancer risk:a meta-analysis. European Journal of Human Genetics 2010; 18:370-8.
166. Lu MP, Wang F and Qiu JR. Methionine synthase A2756G polymorphism and breast cancer risk:a meta analysis involving 18,953 subjects. Breast Cancer Research and Treatment 2010; 123:213-7.
167. Hu J, Zhou GW, Wang N, et al. MTRR A66G polymorphism and breast cancer risk: a meta-analysis. Breast Cancer Research and Treatment 2010; 124:779-84.
168. Ohnami S, Sato Y, Yoshimura K, et al. His595Tyr polymorphism in the Methionine Synthase Reductase (MTRR) gene is associated with pancreatic cancer risk. Gastroenterology 2008; 135:477-88.
169. Rubini M, Brusati R, Garattini G, et al. Cystathionine beta-synthase c.844ins68 gene variant and non-syndromic cleft lip and palate. American Journal of Medical Genetics 2005; 136A:368-72.
170. Brattstrom L, Israelsson B, Norrving B, et al. Impaired homocysteine metabolism in early-onset cerebral and peripheral occlusive arterial disease effects of pyridoxine and folic acid treatment. Atherosclerosis 1990; 81(1):51-60.
171. Tang B, Mustafa A, Gupta S, et al. Methionine-deficient diet induces post-transcriptional downregulation of cystathionine beta synthase. Nutrition 2010; 26:1170-5.
[1]Quinlivan EP, Gregory JF Ⅲ. Effect of food fortification on folic acid intake in the United States. Am J Clin Nutr 2003; 77:221-5.
[2]Pfeiffer CM, Caudill SP, Gunter EW, Osterloh J, Sampson EJ. Biochemical indicators of B vitamin status in the US population after folic acid fortification:results from the National Health and Nutrition Examination Survey 1999-2000. Am J Clin Nutr 2005; 82:442-50.
[3]Institute of Medicine. Folate. In:IOM, ed. Dietary reference intakes forthiamin, riboflavin, niacin, vitamin B6, folate, vitamin B12, pantothenic acid, biotin, and choline. Washington, DC:National Academies Press,1998.
[4]Wright AJ, Dainty JR, Finglas PM. Folic acid metabolism in human subjects revisited:potential implications for proposed mandatory folic acid fortification in the UK. BrJ Nutr 2007; 98:667-75.
[5]Bailey SW, Syslo MC, Ayling J. An assay for dihydrofolate reductase in human tissues byHPLC with fluorometric detection. FASEB J 2002; 16:A267(abstr).
[6]Steven WB and June EA. The extremely slow and variable activity of dihydrofolate reductase in human liver and its implications for high folic acid intake. P Natl Acad Sci 2009; 106(36):15424-15429.
[7]Troen AM, Mitchell B, Sorensen B, et al. Unmetabolized folic acid in plasma is associated with reduced natural killer cell cytotoxicity among postmenopausal women. J Nutr 2006; 136:189-94.
[8]Pfeiffer CM, Fazili Z, McCoy L, Zhang M, Gunter EW. Determination of folate vitamers in human serum by stable-isotope-dilution tandem mass spectrometry and comparison with radioassay and microbiologic assay. Clin Chem 2004; 50:423-32.
[9]Sweeney MR, McPartlin J, Weir DG, et al. Evidence of unmetabolised folic acid in cord blood of newborn and serum of 4-day-old infants. Br J Nutr 2005; 94:727-30.
[10]Ashokkumar B, Mohammed ZM, Vaziri ND, Said HM. Effect of folate oversupplementation on folate uptake by human intestinal and renal epithelial cells. Am J Clin Nutr 2007; 86:159-166.
[11]Kim YI. Role of folate in colon cancer development and progression. J Nutr 2003; 133(suppl):3731S-9S.
[12]Kim YI. Folate:a magic bullet or a double edged sword for colorectal cancer prevention? Gut 2006; 55:1387-9.
[13]Kim YI. Does a high folate intake increase the risk of breast cancer? Nutr Rev 2006; 64:468-75.
[14]Song J, Medline A, Mason JB, Gallinger S, Kim YI. Effects of dietary folate on intestinal tumorigenesis in the apcMin mouse. Cancer Res 2000; 60:5434-40.
[15]Song J, Sohn KJ, Medline A, Ash C, Gallinger S, Kim YI. Chemopreventive effects of dietary folate on intestinal polyps in Apc/Msh2/mice. Cancer Res 2000; 60: 3191-9.
[16]Kim YI. Folate, colorectal carcinogenesis, and DNA methylation:lessons from animal studies. Environ Mol Mutagen 2004; 44:10-25.
[17]Bingham SA, Norat T, Moskal A, et al. Is the association with fiber from foods in colorectal cancer confounded by folate intake? Cancer Epidemiol Biomarkers Prev 2005; 14:1552-6.
[18]Dameshek W. Editorial:the use of folic acid antagonists in acute leukemia. Blood 1948; 3:1057-8.
[19]Kato I, Dnistrian AM, Schwartz M, et al. Serum folate, homocysteine and colorectal cancer risk in women:a nested case-control study. Br J Cancer 1999; 79: 1917-22.
[20]Van Guelpen B, Hultdin J, Johansson I, et al. Low folate levels may protect against colorectal cancer. Gut 2006; 55:1461-6.
[21]Stolzenberg-Solomon RZ, Chang S-C, Leitzmann MF, et al. Folate intake, alcohol use, and postmenopausal breast cancer risk in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial. Am J Clin Nutr 2006; 83:895-904.
[22]Ulrich CM. Folate and cancer prevention:a closer look at a complex picture. Am J Clin Nutr 2007; 86:271-3.
[23]Tworoger SS, Hecht JL, Giovannucci E, Hankinson SE. Intake of folate and related nutrients in relation to risk of epithelial ovarian cancer. AmJ Epidemiol 2006; 163: 1101-11.
[24]Reynolds E. Vitamin B12, folic acid, and the nervous system. Lancet Neurol 2006; 5:-949-60.
[25]Ulrich CM, Bigler J, Bostick R, Fosdick L, Potter JD. Thymidylate synthase promoter polymorphism, interaction with folate intake, and risk of colorectal adenomas. Cancer Res 2002; 62:3361-4.
[26]Morris MS, Jacques PF, Rosenberg IH, Selhub J. Folate and vitamin B12 status in relation to anemia, macrocytosis, and cognitive impairment among older Americans in the age of folic acid fortification. AmJ Clin Nutr 2007; 85:193-200.
[27]Komatsu M, Tsukamoto I. Effect of folic acid on thymidylate synthase and thymidine kinase in regenerating rat liver after partial hepatectomy. Biochim Biophys Acta 1998; 1379:289-96.
[28]MorrisMC,Evans DA, Bienias JL, et al. Dietary folate and vitamin B12 intake and cognitive decline among community-dwelling older persons. Arch Neurol 2005; 62: 641-5.
[29]Lange H, Suryapranata H, De Luca G, et al. Folate therapy and in-stent restenosis after coronary stenting. N Engl J Med 2004; 350:2673-81.
[30]HOPE. Homocysteine lowering with folic acid and B vitamins in vascular disease. N Engl J Med 2006; 354:1567-77.
[31]Bonaa KH, Njolstad I, Ueland PM, et al. Homocysteine Lowing and cardiovascular events after acute myocardial infarction. N Engl J Med 2006; 354:1578-88.
[32]Yajnik CS, Deshpande SS, Jackson AA, et al. Vitamin B12 and folate concentrations during pregnancy and insulin resistance in the offspring:the Pune Maternal Nutrition Study. Diabetologia 2008; 51:29-38.
[33]Regine PST, Sylvia AOB, Dennis K, et al. Periconceptional maternal folic acid use of 400u.g per day is related to increased methylation of the IGF2 gene in the very young child. Plos One 2009; 4(11)e7845:1-5.
[34]Reyes-Engel A, Munoz E, Gaitan MJ, et al. Implications on human fertility of the 677C>T and 1298A>C polymorphisms of the MTHFR gene:consequences of a possible genetic selection. Mol Hum Reprod 2002; 8:952-7.
[35]Zetterberg H. Methylenetetrahydrofolate reductase and transcobalamin genetic polymorphisms in human spontaneous abortion:biological and clinical implications. Reprod Biol Endocrinol 2004;2:7.
[36]Whitehead AS. Changes in MTHFR genotype frequencies over time. Lancet 1998; 352:1784-5.
[37]Lucock M, Yates Z. Folic acid-itamin and panacea or genetic time bomb? Nat Rev Genet 2005; 6:235-40.
[38]McGuire JJ. Anticancer antifolates:current status and future directions. Curr Pharm Des 2003; 9:2593-613.
[39]Robien K. Folate during antifolate chemotherapy:what we know and do not know. Nutr Clin Pract 2005; 20:411-22.
[40]Hazra A, Kraft P, Lazarus R, et al. Genome-wide significant predictors of metabolites in the one-carbon metabolism pathway. Hum Mol Genet 2009; 18(23): 4677-4687.
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