成纤维细胞和ES细胞在小鼠和兔去核卵母细胞内发育

以昆明白小鼠成纤维细胞和胚胎干（ES）细胞作为供核细胞,以昆明白小鼠和日本大耳白兔的MⅡ期去核卵母细胞作为受体,采用核移植方法,构楚了克隆胚胎．在同种克隆中,以ES细胞为供核细胞的克隆胚胎卵裂率明显低于以成纤维细胞为供核细胞的克隆胚胎卵裂率（24．4％相对于56．9％,P〈0．05）,1．8％的ES细胞克隆胚胎发育到囊胚阶段,而成纤维细胞克隆胚胎没能发育到囊胚阶段;在异种克隆中,以ES细胞为供核细胞的克隆胚胎卵裂率（89．6％）和囊胚发育率（18．8％）明显高于以成纤维细胞为供核细胞的克隆胚胎卵裂率（54．2％）和囊胚发育率（4．2％）．     

小鼠腔前卵泡无血清体外培养的研究

目的:探索添加物维生素C和促性腺激素对小鼠腔前卵泡卵母细胞体外生长发育的影响及适合小鼠腔前卵泡体外生长发育的培养体系.方法:通过改良的TCM199无血清培养系统,在96孔细胞培养板中,每孔放入1个腔前卵泡等条件下体外培养腔前卵泡卵母细胞.结果:随着培养时间的延长,各组卵泡和卵母细胞的直径都不同程度的有所增长,其中FSH400+LH200组和VC50+FSH400+LH200影响均大于VC50和对照组,存在显著性差异(P0.05),但二者之间无显著性差异.FSH400+LH200质量浓度组卵泡存活率、发生GVBD率和第一极体排出率均最高,分别为68.6%、17.6%、9.8%.但VC50+FSH400+LH 200组卵泡成腔率最高,为48%.结论:在改良的TCM199无血清培养系统中,联合添加促性腺激素和抗氧化剂后,卵泡成腔率大有提高,卵泡存活率、发生GVBD率和第一极体排出率仅次于单独添加促性腺激素组.     

Nitric oxide produced by cumulus cells stimulates maturation of mouse oocytes

In recent years, embryo engineering concerning animal cloning, animal transgene and bioreactor technique has been developed rapidly. Since the operation of each of these techniques needs matured mammalian oocytes, the demand for highly qualified oocytes is expanding. On the other hand, various factors existing in follicle might be involved in oocyte meiotic maturation, and the process involved is extremely complex and inadequately defined. As a result, the mechanism of oocyte maturation regula…     

Nitric oxide produced by cumulus cells stimulates maturation of mouse oocytes

The effect of sodium nitroprusside (SNP), a donor of nitric oxide, on meiotic maturation of mouse oocytes was studied by injecting N^w-nitro-L-arginine methyl ester (L-NAME) intra-peritoneal (ip), a nitric oxide synthase inhibitor, or culturing oocytes in the medium supplemented with L-NAME or hypoxanthine (HX) to arrest the spontaneous oocyte maturation in vitro. The results showed that the inhibitory effect of L-NAME by injecting 10 mg/kg ip on extrusion of the first polar body only could be reversed by injecting 2.5 mg/kg SNP with L-NAME simultaneously (P < 0.05). Half an hour later ten mice died when given 10 mg/kg SNP ip. The treatment of some concentrations of SNP (10^–7, 10^–6, 10^–5mol/L) significantly stimulated meiotic maturation to metaphase Ⅱ stages in cumulus enclosed oocytes in the presence of HX. However, other concentrations of SNP (10^–8, 10^–4, 10^–3 mol/L) had no effect on HX-arrested oocyte meiotic maturation. The optimal concentration of SNP on CEOs had no effect on DOs. The dose of 10^–3 mol/LL-NAME demonstrated a significant suppression in formation of PB₁, but not in GVBD. This inhibition was reversed by the addition of SNP. These results indicated that the physiological levels of NO produced by cumulus cells could stimulate meiotic maturation of mouse oocytes both in vivo and in vitro.     

Cell biology: nondisjunction, aneuploidy and tetraploidy

One simple, widely accepted mechanism for generating an aberrant chromosome number, or aneuploidy, is through nondisjunction--a chromosome distribution error that occurs during mitosis when both copies of a duplicated chromosome are deposited into one daughter cell and none into the other. Shi and King challenge this view, concluding that nondisjunction does not yield aneuploid cells directly, but instead gives rise to tetraploid cells that may subsequently become aneuploid through further division. Here we show that the direct result of chromosome nondisjunction is gain or loss of a single chromosome, which results in near-diploid aneuploidy, not tetraploidy. We suggest that chromatin trapped in the cytokinetic cleavage furrow is the more likely reason for furrow regression and tetraploidization.     

Chromosome nondisjunction yields tetraploid rather than aneuploid cells in human cell lines

Although mutations in cell cycle regulators or spindle proteins can perturb chromosome segregation, the causes and consequences of spontaneous mitotic chromosome nondisjunction in human cells are not well understood. It has been assumed that nondisjunction of a chromosome during mitosis will yield two aneuploid daughter cells. Here we show that chromosome nondisjunction is tightly coupled to regulation of cytokinesis in human cell lines, such that nondisjunction results in the formation of tetraploid rather than aneuploid cells. We observed that spontaneously arising binucleated cells exhibited chromosome mis-segregation rates up to 166-fold higher than the overall mitotic population. Long-term imaging experiments indicated that most binucleated cells arose through a bipolar mitosis followed by regression of the cleavage furrow hours later. Nondisjunction occurred with high frequency in cells that became binucleated by furrow regression, but not in cells that completed cytokinesis to form two mononucleated cells. Our findings indicate that nondisjunction does not directly yield aneuploid cells, but rather tetraploid cells that may subsequently become aneuploid through further division. The coupling of spontaneous segregation errors to furrow regression provides a potential explanation for the prevalence of hyperdiploid chromosome number and centrosome amplification observed in many cancers.     

Effects of maternal aging on localizations and expression levels of DNA methyltransferases and chromosome architecture in in-vivo matured mouse oocytes

This paper investigated the age-related changes in the expression patterns of maintenance methyltransferase (DNMT1) and de novo methyltransferases (DNMT3a, 3b, 3L) and the chromosome architecture in in-vivo matured mouse oocytes using two-photon laser-scanning microscope. Our results showed that (1) DNMT1 and DNMT3a, 3b, 3L in the oocytes of pubertal mice were located in the cortical region of oocyte cytoplasm. In aging groups, DNMT1 was also located in the cortical region. However, DNMT3a, 3b, 3L had a relatively wider distribution in the oocyte cytoplasm and appeared near the chromosomes. These differences between pubertal and aging groups suggested that aging might affect DNA methylation; (2) the expression of DNMT1, and DNMT3a, 3b in aging groups increased significantly compared to pubertal groups, while, the expression of DNMT3L decreased. These results might be explained by the compensation mechanism among DNMTs, which might be impervious to aging; (3) aging caused increased errors in the distribution and three-dimensional morphology of chromosomes, including the increased total volume and surface area, the high ratio of height to diameter of a circular cylinder enclosing the chromosomes (H/D). Our work provided morphological information for the studies of age-related decline in oocyte qualities.     

小鼠白细胞减少症模型的建立

目的为小鼠白细胞减少症模型的建立提供基础资料。方法选用ICR小鼠和C57BL/6J小鼠以5Gy的剂量60Co-γ射线一次性全身照射小鼠,测定小鼠外周血白细胞总数。结果在本实验条件下,ICR小鼠的正常白细胞数量极显著低于C57BL/6J小鼠;雄性C57BL/6J在照射后第3 d白细胞数量显著高于其雌性小鼠。结论小鼠的品系、性别对60Co-γ射线照射建立的白细胞减少症模型具有一定的影响。