酒精對小鼠生殖細胞及植入前胚胎表觀遺傳修飾的影響
發(fā)布時間：2019-03-20 14:49
【摘要】：研究表明酒精可導致后代出現(xiàn)一些典型的生理缺欠,如患者的面部會出現(xiàn)扭曲,中樞神經系統(tǒng)發(fā)育不完善,等他們成年后語言表達障礙,記憶能力明顯的下降及缺乏人際溝通能力,這在醫(yī)學上叫做胎兒酒精綜合征(Fetal Alcohol Spectrum Disorders, FASD)。該病關于表觀遺傳修飾的發(fā)病機制成為研究熱點,而國內外主要集中在對神經系統(tǒng)發(fā)育水平的研究。眾所周知植入前胚胎有序的表觀遺傳模式對后期胚胎的發(fā)育異常的重要,若出現(xiàn)了紊亂,則會導致早期胎兒發(fā)育異常及成年疾病的出現(xiàn)。我們課題組前期工作發(fā)現(xiàn),小鼠植入前胚胎的表觀遺傳修飾易受到環(huán)境因素影響,環(huán)境的變化可使表觀遺傳模式發(fā)生紊亂,于是推測酒精可使早期胚胎表觀遺傳修飾發(fā)生異常。 表觀遺傳修飾主要包括DNA甲基化、組蛋白修飾及非編碼RNA等,一般認為它調控有關細胞命運及胚胎發(fā)育的基因表達。DNA甲基化作為其中一種飾研究較為清楚的表觀遺傳修方式,酒精可干擾蛋氨酸合成與代謝,進而可影響甲基供體的正常供給,并且哺乳動物的組織因酒精攝入而導致DNA甲基化降低,故我們猜測酒精影響植入前胚胎的DNA甲基化的水平。而組蛋白氨基末端的修飾作為另一個重要的表觀遺傳機制,對染色質的結構進行調節(jié)而調控基因的表達,例如,組蛋白的甲基化與乙�；诨虮磉_調控中起著重要作用,并影響生物體的基本發(fā)育進程。在父性、母性孕期酒精暴露研究中,懷孕期間酒精攝入對植入前胚胎DNA甲基化及組蛋白修飾可能造成損害的差異,也為我們預防酒精對胎兒的傷害提供借鑒的作用。 小鼠通常作為研究該病的動物模型,為了研究FASD的表觀遺傳的發(fā)病機制及為治療該病提供一定的實驗依據(jù)。本研究采用活體灌胃法、HE染色法、精子的質量分析技術、體內沖胚技術、胚胎體外培養(yǎng)法及免疫熒光技術,分析酒精對肝臟及精液質量、生殖細胞及體內、外早期胚胎的表觀遺傳修飾的影響。我們分別對雄鼠、雌鼠受孕前后及雙親進行灌酒處理,研究酒精對生殖細胞及植入前胚胎全基因組DNA甲基化水平的影響。后期我們進一步研究雌鼠受孕前后酒精攝入對H3K27三甲基化／乙�；Ｊ降挠绊�。實驗數(shù)據(jù)顯示,灌酒小鼠的生殖細胞及植入前胚胎表觀遺傳修飾模式發(fā)生異常,具體如下：雄鼠灌酒實驗表明酒精分別肝臟和精液的質量的損害,使精子的DNA甲基化水平降低,導致后期植入前胚胎DNA甲基化水平異常。受孕前后雌鼠灌酒實驗結果表明酒精降低卵子及植入前胚胎DNA甲基化水平。雙親灌酒實驗結果表明酒精對植入前胚胎的DNA甲基化損害程度依次為雙親灌酒雌性灌酒雄性灌酒。雌鼠受孕前后酒精攝入對植入前胚胎的H3K27三甲基化影響較不明顯,而酒精對該時期胚胎的H3K27乙酰化影響較明顯。 酒精攝入對小鼠的生殖細胞及植入前胚胎表觀遺傳的的正確建立造成紊亂,為揭示胎兒酒精綜合征發(fā)病機制提供重要的借鑒。為深入理解孕期酒精暴露的危害,揭示受孕酒精暴露對后代長時期的影響提供一定的實驗依據(jù),同時提供預防受孕酒精暴露對子代的傷害提供一種思路。
[Abstract]:The study shows that alcohol can cause some typical physiological defects in the offspring, such as the distortion of the face of the patient, the imperfect central nervous system, the expression of the disorder in the adult post-adult language, the obvious decline of the memory ability and the lack of interpersonal communication ability, This is medically known as the Fetus Alcohol Spectrum Disdders (FASD). The pathogenesis of epigenetic modification has become the focus of the study, and the research on the development level of the nervous system is mainly concentrated at home and abroad. It is well known that the apparent genetic pattern of pre-implantation embryo order is important to the development of later-stage embryos, and if a disorder occurs, the early fetal development and the appearance of adult diseases can be caused. The preliminary work of our research group found that the epigenetic modification of the pre-implantation embryos of the mice is susceptible to environmental factors, and the change of the environment can cause the apparent genetic pattern to be disordered, so that the apparent genetic modification of the early embryo may be abnormal. Epigenetic modification mainly includes DNA methylation, histone modification and non-coding RNA. It is generally considered that it regulates the genes related to cell fate and embryonic development. D. DNA methylation as one of the more clear epigenetic methods in which alcohol can interfere with the synthesis and metabolism of methionine, which in turn affects the normal supply of methyl donor, and the tissue of a mammal results in a DNA methylation down due to alcohol intake. Low, so we suspect that alcohol affects the DNA methylation of the pre-implantation embryos The modification of the amino-terminal of histone is another important epigenetic mechanism, and the structure of chromatin is regulated to regulate the expression of the gene. For example, the methylation of histone and ethylation of histone play an important role in the regulation of gene expression and affect the basic development of the organism. Cheng. During the study of maternal and maternal alcohol exposure, alcohol intake during pregnancy can cause damage to pre-implantation fetal DNA methylation and histone modification, and also provide reference for our prevention of alcohol damage to the fetus. The mouse is usually used as an animal model for the study of the disease. In order to study the pathogenesis of the apparent inheritance of the FASD and to provide some real evidence for the treatment of the disease In this study, the epigenetic modification of the quality of the liver and the semen, the germ cells and the in vivo and in vitro embryos of the liver and the semen, the germ cells and the in vivo and in vitro were analyzed by the method of living in-life, HE staining, the quality analysis of the sperm, the in-vivo embryo-flushing technique, the in vitro culture of the embryo and the immunofluorescence technique. The effects of alcohol on the DNA methylation of the whole genome of the germ cells and the pre-implantation embryos were studied. The effect of alcohol intake on H3K27 trimethylation/ ethylation patterns before and after conception in female rats was further studied in the later stage. The experimental data showed that the germ cells and the epigenetic modification of the pre-implantation embryos of the mice were abnormal, as follows: The experimental data of the male rats showed that the quality of the liver and the semen of the alcohol was damaged, so that the DNA of the sperm was methylated. reduced level, resulting in post-implantation of pre-implantation embryo DNA methylation water The results of the experiment of the female rats before and after conception showed that the alcohol decreased the egg and the pre-implantation embryo DNA methyl group. The results of the experiment of both parents and parents showed that the degree of DNA methylation of alcohol in the pre-implantation embryos was both parent and female. The effect of alcohol intake before and after the conception of female rats on the three-methylation of H3K27 in the pre-implantation embryos was not significant, and the effect of alcohol on the development of H3K27 in the embryo during the period was not significant. Obvious. The correct establishment of the apparent genetic inheritance of the germ cells and pre-implantation embryos of the mice caused by alcohol intake is a disorder, which is provided for the purpose of revealing the pathogenesis of the fetal alcohol syndrome It is an important reference. In order to understand the harm of alcohol exposure during pregnancy, it is revealed that the exposure of the pregnant alcohol to the long period of the offspring provides a certain experimental basis, and also provides the prevention of the exposure of the pregnant alcohol to the offspring.
【學位授予單位】：安徽大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R363
本文編號：2444315
[Abstract]:The study shows that alcohol can cause some typical physiological defects in the offspring, such as the distortion of the face of the patient, the imperfect central nervous system, the expression of the disorder in the adult post-adult language, the obvious decline of the memory ability and the lack of interpersonal communication ability, This is medically known as the Fetus Alcohol Spectrum Disdders (FASD). The pathogenesis of epigenetic modification has become the focus of the study, and the research on the development level of the nervous system is mainly concentrated at home and abroad. It is well known that the apparent genetic pattern of pre-implantation embryo order is important to the development of later-stage embryos, and if a disorder occurs, the early fetal development and the appearance of adult diseases can be caused. The preliminary work of our research group found that the epigenetic modification of the pre-implantation embryos of the mice is susceptible to environmental factors, and the change of the environment can cause the apparent genetic pattern to be disordered, so that the apparent genetic modification of the early embryo may be abnormal. Epigenetic modification mainly includes DNA methylation, histone modification and non-coding RNA. It is generally considered that it regulates the genes related to cell fate and embryonic development. D. DNA methylation as one of the more clear epigenetic methods in which alcohol can interfere with the synthesis and metabolism of methionine, which in turn affects the normal supply of methyl donor, and the tissue of a mammal results in a DNA methylation down due to alcohol intake. Low, so we suspect that alcohol affects the DNA methylation of the pre-implantation embryos The modification of the amino-terminal of histone is another important epigenetic mechanism, and the structure of chromatin is regulated to regulate the expression of the gene. For example, the methylation of histone and ethylation of histone play an important role in the regulation of gene expression and affect the basic development of the organism. Cheng. During the study of maternal and maternal alcohol exposure, alcohol intake during pregnancy can cause damage to pre-implantation fetal DNA methylation and histone modification, and also provide reference for our prevention of alcohol damage to the fetus. The mouse is usually used as an animal model for the study of the disease. In order to study the pathogenesis of the apparent inheritance of the FASD and to provide some real evidence for the treatment of the disease In this study, the epigenetic modification of the quality of the liver and the semen, the germ cells and the in vivo and in vitro embryos of the liver and the semen, the germ cells and the in vivo and in vitro were analyzed by the method of living in-life, HE staining, the quality analysis of the sperm, the in-vivo embryo-flushing technique, the in vitro culture of the embryo and the immunofluorescence technique. The effects of alcohol on the DNA methylation of the whole genome of the germ cells and the pre-implantation embryos were studied. The effect of alcohol intake on H3K27 trimethylation/ ethylation patterns before and after conception in female rats was further studied in the later stage. The experimental data showed that the germ cells and the epigenetic modification of the pre-implantation embryos of the mice were abnormal, as follows: The experimental data of the male rats showed that the quality of the liver and the semen of the alcohol was damaged, so that the DNA of the sperm was methylated. reduced level, resulting in post-implantation of pre-implantation embryo DNA methylation water The results of the experiment of the female rats before and after conception showed that the alcohol decreased the egg and the pre-implantation embryo DNA methyl group. The results of the experiment of both parents and parents showed that the degree of DNA methylation of alcohol in the pre-implantation embryos was both parent and female. The effect of alcohol intake before and after the conception of female rats on the three-methylation of H3K27 in the pre-implantation embryos was not significant, and the effect of alcohol on the development of H3K27 in the embryo during the period was not significant. Obvious. The correct establishment of the apparent genetic inheritance of the germ cells and pre-implantation embryos of the mice caused by alcohol intake is a disorder, which is provided for the purpose of revealing the pathogenesis of the fetal alcohol syndrome It is an important reference. In order to understand the harm of alcohol exposure during pregnancy, it is revealed that the exposure of the pregnant alcohol to the long period of the offspring provides a certain experimental basis, and also provides the prevention of the exposure of the pregnant alcohol to the offspring.
【學位授予單位】：安徽大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R363
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