X染色体失活、失活逃逸及相关疾病的研究进展

剂量补偿效应是表观遗传的机制之一,它是指使X连锁基因的表达水平在两性间达到平衡的过程。不同的物种所实现剂量补偿的机制不同,人同其他哺乳动物的剂量补偿机制是雌性个体随即失活一条X染色体。但最近的研究发现,在人和小鼠的失活X染色体上有些基因仍能表达,逃避了失活。目前,许多人类疾病尤其是具有女性发病优势病症的发生与治疗可能与这些逃逸基因密切相关。针对以上内容的最新进展作了相关综述。     

DNA甲基化酶抑制剂对体细胞染色体的影响

在动物的体细胞克隆过程中,供体细胞的细胞核要由高度甲基化状态转变为胚胎细胞的非甲基化状态,从而实现全能性的重新获得。试验利用DNA甲基化酶（5-aza-dc）抑制剂处理供体细胞,尝试建立一种既不影响染色体结构和数目,又能降低供体细胞细胞核的甲基化状态的最适浓度和方法,从而提供一种提高体细胞克隆动物效率的方法。在试验中,用不同浓度的甲基化酶抑制剂（5-aza-dc）处理奶牛成纤维细胞72 h,通过常规染色体核型分析方法制备染色体分裂相,检测其染色体数目、结构等的改变,结果显示,低浓度的甲基化酶抑制剂（0.005和0.01 μmol/L）处理,染色体没有发生明显的畸变（P＞0.05）,染色体能够保持正常的形态,高浓度（0.03与0.05 μmol/L）处理导致染色体畸变率显著增加（P＜0.05）,而0.1 μmol/L的5-aza-dc处理后染色体畸形率增加更为明显（P＜0.01）。总之,此试验的结果说明在对供体细胞做甲基化处理时用0.005和0.01 μmol/L的5-aza-dc处理不会影响细胞染色体结构和数目,但是高浓度处理则会导致染色体畸变率显著增加,不能用于体细胞克隆动物生产。     

Purinergic P2X7 receptor antagonist ameliorates intestinal inflammation in postoperative ileus

Postoperative ileus (POI) is a postsurgical gastrointestinal motility dysfunction caused by mechanical stress to the intestine during abdominal surgery. POI leads to nausea and vomiting reduced patient quality of life, as well as high medical costs and extended hospitalization. Intestinal inflammation caused by macrophages and neutrophils is thought to be important in the mechanism of POI. Surgery-associated tissue injury and inflammation induce the release of adenosine triphosphate (ATP) from injured cells. Released ATP binds the purinergic P2X7 receptor (P2X7R) expressed on inflammatory cells, inducing the secretion of inflammatory mediators. P2X7R antagonists are thought to be important mediators of the first step in the inflammation process, and studies in chemically induced colitis models confirmed that P2X7R antagonists exhibit anti-inflammatory effects. Therefore, we hypothesized that P2X7R plays an important role in POI. POI models were generated from C57BL/6J mice. Mice were treated with P2X7R antagonist A438079 (34 mg/kg) 30 min before and 2 hr after intestinal manipulation (IM). Inflammatory cell infiltration and gastrointestinal transit were measured. A438079 ameliorated macrophage and neutrophil infiltration in the POI model. Impaired intestinal transit improved following A438079 treatment. P2X7R was expressed on both infiltrating and resident macrophages in the inflamed ileal muscle layer. The P2X7R antagonist A438079 exhibits anti-inflammatory effects via P2X7R expressed on macrophages and therefore could be a target in the treatment of POI.     

细菌人工染色体文库基因组DNA制备技术

试验旨在对基因组DNA的制备进行研究,为细菌人工染色体(bacterial artificial chromosome,BAC)文库的构建奠定基础。以豁眼鹅全血为试验材料,提取高质量的基因组DNA,分别采用HindⅢ、EcoRⅠ和BamHⅠ3种限制性内切酶对所提基因组DNA进行部分酶切,并利用控制酶切时间、设置不同的HindⅢ酶切浓度梯度对基因组DNA进行部分酶切。结果表明,HindⅢ为最佳限制性内切酶,并得到了最佳酶切用量(40U/μL)。该方法所制备的基因组DNA质量较好,可用于BAC文库的构建。     

牦牛与普通牛、水牛的X染色体基因编码区比较及密码子偏性分析

【目的】从基因组比对及密码子偏性角度分析牦牛与其他牛亚科动物X染色体,有助于了解牦牛品种差异及系统进化地位,为其适应高原低压低氧环境和密码子优化提供参考。【方法】以牦牛X染色体参考基因编码区序列为参考,与普通牛和江河型水牛X染色体参考基因编码区序列进行基因组比对和基因共线性分析,同时进行基因注释,对过滤后的编码区文件进行相对同义密码子使用频率(RSCU)、ENC-plot、PR2-plot和最优密码子确定等偏性分析。【结果】在牦牛X染色体基因编码区发现了参与气管收缩、肺部呼吸及机体代谢等基因与普通牛和水牛存在差异,如KLHL13、CENPI和PGK1等基因;共线性显示长段由强选择压和突变压下表现出的交换线性区域;密码子分析中3种牛亚科动物密码子使用偏性相似,偏性均较弱,均偏向G/C结尾;强偏性密码子(RSCU≥1.5)均为CUG、GUG、AGA、AGG和UGA;牦牛、普通牛和水牛分别筛选出16、13和9个最优密码子,均以A/U结尾。【结论】牦牛与普通牛、水牛相比面临了更大的选择压力,累计的变异程度更大,三者的密码子偏性受到自然选择作用均大于突变作用。研究结果为牦牛的遗传育种、密码子优化...     

哺乳动物早期胚胎的剂量补偿作用及其对发育的影响

剂量补偿是使X连锁基因的表达水平在两性间达到平衡的过程。哺乳动物是通过随机失活雌性的一条X染色体实现剂量补偿。X染色体失活开始于X染色体失活中心(XIC),然后传播到整条染色体。Xist基因定位于XIC中,参与X染色体失活的启动。本文主要通过分析雄核生殖胚胎、雌核生殖胚胎和正常胚胎发育中X连锁基因表达,概括了X染色体失活(XCI)在哺乳动物早期胚胎发育中作用,综述了哺乳动物早期胚胎的剂量补偿作用及其对发育的影响。     

Effects of Trichostatin A on In Vitro Development and DNA Methylation Level of the Satellite I Region of Swamp Buffalo (Bubalus bubalis) Cloned Embryos

Trichostatin A (TSA), a histone deacetylase inhibitor, has been widely used to improve the cloning efficiency in several species. This brings our attention to investigation of the effects of TSA on developmental potential of swamp buffalo cloned embryos. Swamp buffalo cloned embryos were produced by electrical pulse fusion of male swamp buffalo fibroblasts with swamp buffalo enucleated oocytes. After fusion, reconstructed oocytes were treated with 0, 25 or 50 nM TSA for 10 h. The results showed that there was no significant difference in the rates of fusion (82–85%), cleavage (79–84%) and development to the 8-cell stage (59–65%) among treatment groups. The highest developmental rates to the morula and blastocyst stages of embryos were found in the 25 nM TSA-treated group (42.7 and 30.1%, respectively). We also analyzed the DNA methylation level in the satellite I region of donor cells and in in vitro fertilized (IVF) and cloned embryos using the bisulfite DNA sequencing method. The results indicated that the DNA methylation levels in cloned embryos were significantly higher than those of IVF embryos but approximately similar to those of donor cells. Moreover, there was no significant difference in the methylation level among TSA-treated and untreated cloned embryos. Thus, TSA treatments at 25 nM for 10 h could enhance the in vitro developmental potential of swamp buffalo cloned embryos, but no beneficial effect on the DNA methylation level was observed.     

Experimental approach to prezygotic chromosome screening using only a single pair of gametes in mice

During in vitro embryo production, chromosome screening is essential to prevent pregnancy losses caused by embryonic chromosome aberrations. When the chromosome screening is completed before fertilization, gametes are effectively utilized as genetic resources. The aim of this study was to investigate whether chromosome screening of gametes accompanied by fertilization would be feasible using a single mouse spermatozoon and oocyte. Metaphase II oocytes were divided into a cytoplast and a karyoplast. For genome cloning of the gametes, androgenic and gynogenic embryos were produced by microinjection of sperm into cytoplasts and parthenogenetic activation of karyoplasts, respectively. Pairs of blastomeres from androgenic and gynogenic embryos were fused electrically to produce diploid embryos, which were transferred into pseudopregnant surrogate mothers to examine fetal development. Blastomeres from androgenic and gynogenic embryos were individually treated with calyculin A—a specific inhibitor of type 1 and 2A protein phosphatases—for 2 h to induce premature chromosome condensation. Thereafter, chromosome analysis of blastomeres, reflecting the genetic constitution of individual spermatozoa and oocytes, was performed, and we confirmed that most of the androgenic and gynogenic 2-cell embryos had a haploid set of chromosomes in their sister blastomeres. The reconstructed embryos from blastomeres of androgenic and gynogenic 2-cell embryos could be implanted and develop into live fetuses, albeit at low efficiency. This study indicates that prezygotic chromosome screening and embryo production using a single pair of gametes may be practicable.     

绒山羊X染色体7个微卫星标记的遗传连锁图谱的构建

本研究利用内蒙古白绒山羊的7个父系半同胞家系共794个个体,用X染色体上的7个微卫星标记进行了系谱确认,构建了绒山羊X染色体遗传连锁图。结果表明,7个标记的等位基因数变化范围为9～14,杂合度在0.585～0.918之间,平均杂合度为0.726;多态信息含量在0.759～0.897之间,平均多态信息含量为0.834。构建的内蒙古白绒山羊X染色体遗传连锁图总长度139.4 cM,与英国罗斯林研究所公布的SM4.7绵羊连锁图标记顺序一致,可用于下一步的QTL定位研究。     

Evaluation of the global DNA methylation in canine mast cell tumour samples by immunostaining of 5‐methyl cytosine

Cutaneous mast cell tumours (MCT) are the most common skin tumour in dogs, and to our knowledge, there are no previous studies regarding the global methylation of these tumours. DNA hypomethylation and hypermethylation have been described in several tumours and both mechanisms can lead to carcinogenesis. The purpose of this study was to evaluate the global DNA methylation in canine MCT. A total of 48 MCT samples were classified in grades 1, 2 and 3 or high‐grade or low‐grade. Monoclonal antibodies were used for the immunohistochemical detection of the 5‐methylcytosine. The immunostained nuclei were classified in strong, weak or negative pattern, and these were quantified in five distinct microscopic fields (40× objective) in each slide. The results showed that global DNA hypomethylation was predominant in grade 3, high‐grade, less differentiated MCT. These epigenetic changes in neoplastic mast cells warrant further detailed investigation aiming the establishment of tumour epigenetic therapies.     

营养素对动物表观遗传的影响及其机制

表观遗传是环境因素和细胞内的遗传物质相互作用的结果.表观遗传学是在DNA碱基序列不变的前提下引起的基因表达或细胞表观型变化的一种遗传现象,具体表现在以下3个既相互独立又相互作用的方面:DNA甲基化、组蛋白修饰和microRNA (miRNA)调控.本文就表观遗传的研究进展及营养素对动物表观遗传的影响及其机制进行综述.     

中国鹿科动物染色体研究进展

染色体是生物细胞内遗传物质的主要携带者,其在细胞水平上对生物的性状进行控制,在生物进化过程中对物种的形成起到了重要的作用。近年来,随着细胞遗传学研究方法的不断进步,对鹿科动物染色体的研究也逐渐深入。作者回顾了近年来国内外鹿科动物染色体的研究进展,同时结合国内鹿科动物现状,提出了鹿科动物染色体研究中存在的问题及进一步研究方向。     

五指山猪背最长肌和半腱肌DNA甲基化检测分析

DNA甲基化作为真核生物基因组重要的表观遗传学修饰,对生物体基因的表达有重要的调控作用。为了研究五指山猪肌肉组织基因组DNA的甲基化程度,试验利用MSAP技术,使用筛选的10对引物扩增,检测五指山猪背最长肌和半腱肌的甲基化程度。结果显示:1月龄、2月龄、4月龄五指山猪背最长肌的平均甲基化率分别是44.78%、41.58%、38.92%;1月龄、2月龄、4月龄五指山猪半腱肌的平均甲基化率分别是41.70%、39.39%、38.81%。研究结果为五指山猪生长发育规律、系统选育及矮小机制等方面的研究提供表观遗传学依据。     

DNA甲基化与去甲基化调控肌肉发育研究进展

肌肉发育是一个复杂的生物学过程,其调控机制尚不完善。但近年来表观遗传修饰对肌肉发育的调控作用逐渐成为热点领域,研究发现DNA甲基化与去甲基化修饰对肌肉发生与发育起到重要的调控作用。肌肉干细胞特异位点通过DNA甲基化修饰,影响肌肉发育过程关键基因的表达,进而调控早期发育的生肌过程。本文主要围绕肌肉发育过程中DNA甲基化及去甲基化修饰的变化、重要的甲基转移酶和去甲基化酶以及营养物质通过DNA甲基化修饰影响肌肉发生的作用进行论述。     

羊驼染色体的研究

对于动物染色体的研究，在国内外有许多学者做了大量的工作，但对于羊驼染色体的研究甚少，仅Taylor、Vidal-Rioja及Bianchi报道羊驼正常二倍体染色体数为2n=74。此外，Wilker，Katrin报道美洲驼有性反转、X染色体单体性及染色体间性等畸变现象，Murray报道羊驼在生理结构及遗传特性等方面与美洲驼非常相似，     

Effects of inactive parapoxvirus ovis on cytokine levels in rats

The aim of this study is to determine the effects of iPPOV on pro-inflammatoryand anti-inflammatory cytokine levels in rats. iPPOV (1 ml/rat) wasadministered intraperitoneal route to 49 rats, except for 7 rats (Control, 0 group). Serumsamples were collected from 7 rats at 1st, 2nd, 4th, 8th, 12th, 16th and 24th hr aftertreatments. Levels of TNF-α, IL-6, IL-12 and IL-10 were determined using ELISA.Administration of iPPOV stimulated TNF-α (16th and 24th hr) and IL-6 (12th, 16th and 24thhr) synthesis and caused fluctuations in IL-10 and IL-12 concentrations. In conclusion,increased cytokine levels could be attributed to immunomodulatory activity of iPPOV,however, detailed studies are required to fully understand effects of iPPOV on immunesystem.