Strain difference in tolerance to low-temperature treatment of fertilized mouse oocytes

Background and Aims:  The aim of this study was to determine which mouse strains exhibit tolerance to cooling when fertilized oocytes have been stored at 4°C.
Methods:  In-vitro -fertilization-derived oocytes of eight mouse strains were incubated at 4°C in 20 mmol/L Hepes-potassium modified simplex optimized medium (KSOM) medium for 0, 24, 48, 60 or 72 h, and then returned to normal culture conditions at 37°C in KSOM medium. The rates of development of cultured oocytes into blastocysts and cell numbers of blastocysts were examined. In some cases, a Comet assay was carried out to evaluate DNA damage. In addition, the effects of β-mercaptoethanol on the development of the 4°C-treated oocytes were assessed.
Results:  Of the eight strains tested, BDF1, B6C3F1 and FVB/N strains exhibited relatively higher degrees of tolerance to 4°C treatment and approximately 90%, 83% and 78% of oocytes treated at 4°C for 48 h developed to morphologically normal blastocysts, respectively. Comet assay revealed no clear DNA damage in oocytes treated at 4°C. Treatment with β-mercaptoethanol failed to improve the in vitro survival rate of low-temperature-treated oocytes.
Conclusion:  Strain differences were observed in tolerance to cooling treatment when fertilized oocytes were temporarily treated with 4°C, although the reasons for this remain unclear. (Reprod Med Biol 2006; 5 : 43–50)     

Mitotic and meiotic detection of radiation-induced translocations in mouse stem cell spermatogonia using fluorescencein situ hybridization

The efficiency of two methods of detection of translocations induced in mouse stem cell spermatogonia by X-ray doses of 2, 5 and 7 Gy was compared: classical multivalent analysis at diakinesis-metaphase I of meiosis and observation via fluorescencein situ hybridization analysis of mitotic or meiotic stages. Specific DNA libraries for chromosomes 1, 11 and 13 were used. The results obtained indicate that (a) chromosomes 1, 11 and 13 are more involved in multivalent formation than expected on the basis of DNA content and (b) if the mitotic FISH analysis data are corrected for the observed over-representation, the frequencies of induced translocations are similar to those recorded in the classical multivalent studies, suggesting equal scoring efficiencies in both systems.     

烹调油烟致大小鼠肺癌的实验研究

[目的]了解烹调油烟（cooking oil fumes，COF）的动物致癌性。[方法]采用动式染毒法给Balb／c小鼠（雌雄各半）吸入COF浓度为9．09、20．65、38．85mg／m^3，染毒1次／1～2d，30min／次，共150次，计8个月；SD大鼠（雌雄各半）吸入COF浓度为6、88、15．06、35．33mg／m^3，染毒1次／2d，30min／次，共191次，计12．5个月。分别制备COF慢性中毒动物模型；两实验均设空白对照组，吸入与实验组相同温度的清洁空气。[结果]COF诱发Balb／c小鼠实验组肺癌总发生率为18、95％（29／153），低、中、高浓度组肺癌发生率分别为15．09％、20、00％和22．00％，与对照组差异均有显著性。但低、中、高三组间差异无显著性（P〉0．05）；COF诱发SD大鼠肺癌总发生率为9、10％（9／99），低、中、高浓度组肺癌发生率分别为6．45％、8．57％、12．12％，高浓度组肺癌发生率高于对照组（P〈0．05）。各性别组间肺癌发生率的差别无显著性（P〉0．05）。[结论]COF可以诱导Balb／c小鼠和SD大鼠肺癌，诱发的肺癌主要为肺腺癌（小鼠28／29，大鼠7／9），余为小细胞肺癌。     

初诊T2DM短期胰岛素强化治疗40例分析

目的观察初诊T2DM短期胰岛素强化治疗的疗效。方法对40例初诊T2DM患者采用三餐前皮下注射短效胰岛素和睡前注射中效胰岛素2wk，并检测治疗2wk前后FPG、PPG、GHbA1C及血脂等值进行比较。结果两组治疗后FPG、PPG、GHbA1C及血脂各值均显著低于治疗前p〈0．05（x^2=4．940—11．544）。结论胰岛13细胞功能异常和（或）胰岛素抵抗是T2DM发病的基本环节，早期强化胰岛素治疗能获得血糖的有效控制，减少或延缓并发症的发生。     

维生素A酸对小鼠囊胚细胞凋亡的影响

目的:探讨全反式维生素A酸(ATRA)对体外培养小鼠囊胚中内细胞群(ICM)和滋养层(TE)细胞凋亡的影响。方法:获取妊娠3.5d小鼠囊胚,分别培养在含0μmol/l、1μmol/l和10μmol/l的ATRA的M199培养基中24h,用带有荧光(FITC)标记的原位末端标记(TUNEL)检测法,观察ATRA对小鼠囊胚细胞凋亡的效应。结果:ATRA可诱导囊胚细胞凋亡,并增强小鼠囊胚Fas蛋白表达。结论:维生素A酸对小鼠胚胎的发育具有细胞毒性作用。     

Flerobuterol: a potential antidepressant drug related to β-adrenergic agonists. Experimental profile in mice

The potential antidepressant effect of flerobuterol (dl-(fluoro-2 phenyl)-1 t-butylamino-2 ethanol), a new drug related to beta-adrenoceptor agonists, was evaluated and compared with imipramine and salbutamol using classical psychopharmacological tests in mice. Like imipramine and salbutamol, flerobuterol (0.5-32 mg kg-1, ip) fully prevented apomorphine (16 mg kg-1, sc)- and partly reversed reserpine- and oxotremorine-induced hypothermia. At higher doses (16-32 mg kg-1), flerobuterol enhanced the toxic effects of yohimbine. Unlike imipramine, flerobuterol and salbutamol did not reduce immobility duration in the behavioural despair test. Salbutamol and flerobuterol decreased locomotor activity. Flerobuterol did not induce mydriasis, did not prevent oxotremorine-induced tremors or salivary and lacrimal gland secretion and did not reduce reserpine-induced palpebral ptosis. Propranolol (8 mg kg-1, ip) but not alpha-methyl-paratyrosine (75 mg kg-1, ip) prevented the flerobuterol-induced antagonism of apomorphine-induced hypothermia. Our results suggest that flerobuterol demonstrates potential antidepressant activity, which could be related to beta-adrenoceptor activation in mice.     

Golgi study on brain of macular mutant mouse as a model of Menkes kinky hair disease

Summary This study was undertaken to elucidate, using the Golgi method, the neuropathological change in the brain of the macular mutant mouse, whose hemizygote (Ml/y) is considered to be a model of Menkes kinky hair disease (MKHD). The hemizygote mice gradually lost weight after 10 days of age and died with emaciation and seizure around day 15. The normal littermate (+/y) was well developed. In the cerebrum, the arborization of pyramidal neurons in the layer V of the Ml/y was the same as that in the +/y on day 10. However, development of arborization in the Ml/y was delayed in comparison with that in the +/y on days 12 and 14. Purkinje cells with several somal sprouts were observed in the cerebellum in both the Ml/y and +/y on day 7. The somal sprouts in the +/y had regressed gradually by day 12, while they were still in the anterior and middle lobes of the Ml/y on day 14. Additionally, the trunks of Ml/y stem dendrites became thicker and a cactus formation was recognized on the branching portion of the dendrites on day 14. Arborization of these abnormal Purkinje cells was distinctly poor compared with that in the +/y. These results suggest that the growth of the neurons is delayed in the Ml/y and simultaneously their cytoskeletal developments are disturbed, especially in the Purkinje cells. There is a close similarity in many respects to the neuropathological change in MKHD.     

Quantification of Basal and Stimulated ROS Levels as Predictors of Islet Potency and Function

We have developed a luminol-based assay using intact islets, which allows for quantification of reactive oxygen species (ROS). In addition, an index capable of characterizing metabolic and mitochondrial integrity prior to transplantation was created based on the capacity of islets to respond to high glucose and rotenone (mitochondrial respiratory chain complex I inhibitor) by production of ROS. To validate this assay, lipid peroxidation and antioxidative defense capacity were evaluated by detection of malondialdehyde (MDA) levels and glutathione peroxidase activity (GPx), respectively. Also, flow cytometric analyses of ROS (dihydroethidine), apoptosis (Annexin V, active caspases), necrosis (Topro3), and mitochondrial membrane potential (JC-1) were done in parallel to correlate with changes in luminol-measured ROS. ATP/ADP ratios were quantified by HPLC and the predictive value of ROS measurement on islet functional potency was correlated with capacity to reverse diabetes in a streptozotocin-induced diabetic NOD.scid mouse model as well as in human transplant recipients. Our data demonstrate that levels of ROS in islets correlate with the percentage of apoptotic cells and their functional potency in vivo. The ROS indices following glucose and rotenone exposure are indicative of metabolic potency and mitochondrial integrity and can be used as surrogate markers to evaluate the quality of islets prior to transplantation.     

Histopathological characterisation of effects of the mouse Pax6 missense mutation on eye development

Mutations in PAX6/Pax6 lead to a variety of ocular anomalies in humans and mice. The aim of the study was to characterise the ocular abnormalities caused by the missense Pax6^Leca4 mutation and compare them to published observations on Pax6 alleles that are functionally equivalent to Pax6⁻ null alleles (such as Pax6^Sey and Pax6^Sey-Neu) and human inherited eye diseases. Ocular features of homozygous Pax6^Leca4/Leca4 and heterozygous Pax6^Leca4/+ embryos at E12.5-E18.5, heterozygous Pax6^Leca4/+ young mice at P18 and heterozygous Pax6^Leca4/+ adults at 12 weeks were analysed histologically with their wild-type Pax6^+/+ littermates. Homozygous Pax6^Leca4/Leca4 fetuses died perinatally with no eyes although an optic cup rudiment with pigmented cells developed. Pax6^Leca4/+ mice were microphthalmic and a range of other severe ocular phenotypes affected both the anterior and the posterior segments. In contrast to Pax6^+/−, the Pax6^Leca4/+ eyes had no goblet cells in the corneal epithelium, the iris was not hypoplastic and there was no lens-corneal epithelial plug. However, microphthalmia was more severe, corneal vascularisation occurred earlier (during fetal stages), pigmented cells were present in the vitreous and corneal stroma and the ciliary body was malformed or abnormal. These results show that, although Pax6^Leca4/+ lacked some eye abnormalities commonly seen in Pax6^Sey/+ and Pax6^Sey-Neu/+ eyes, in most respects their eyes were more severely affected. These differences probably reflect both differences between the Pax6^Leca4 and the Pax6^Sey-Neu mutations and differences in modifier gene expression in different genetic backgrounds. The presence of pigmented cells in the cornea is a novel observation. Some Pax6^Leca4/+ ocular abnormalities were similar to those present in human Peters' anomaly and persistent hyperplastic primary vitreous (PHPV) so Pax6^Leca4/+ mice provide a useful model for some inherited eye diseases.     

EDTA-induced urothelial cell shedding for the treatment of superficial bladder cancer in the mouse

AIM: The aim of this study was to determine the effect of intravesical EDTA instillation on the development of intravesically implanted tumor cells in normal mice. METHODS: The mouse bladder tumor (MBT-2) model was used in female C3H/eb mice to evaluate the amount of normal urothelial cell shedding, and the degree of tumor growth inhibition following intravesical EDTA instillation in comparison with phosphate-buffered saline (PBS) instillation. RESULTS: At 1 h after instillation, the number of urothelial cells aspirated was 500-1000 per PBS-treated mouse and 10,000-20,000 per EDTA-treated mouse (P < 0.00001). The bladder weight, which reflected the effect of the agent on the tumor, was similar in the untreated and PBS-treated mice (105.46 +/- 46 mg and 106.2 +/- 50 mg, respectively). It was significantly lower in the EDTA-treated mice (80.4 +/- 42 mg) (P = 0.0045). CONCLUSIONS: Intravesical administration of EDTA results in significant normal and neoplastic urothelial cell shedding. Intravesical irrigation with EDTA may prevent adherence of the malignant cells to the bladder wall following tumor resection.