This experiment was undertaken in order to find out if there is any morphological change in oocytes and two-cell embryos whose development have been suppressed by progesterone for six hours in vitro. It can be observed that some part of the outer side of nuclear membrane of the suppressed oocytes was damaged. The number of nuclear pores has decreased in suppressed oocytes and this suggests that progesterone might suppress the transport of intermediary metabolites between cytoplasm and nucleus. Sometimes, closely packed aggregates of parallel or irregular endoplasmic reticula were observed in suppressed oocytes. Microvilli of suppresed oocytes showed signs of degradation and the perivitelline space became apparent. Thus it is presumed that the egg membrane has constricted during cultivation under progesterone in vitro. The other cell organelles such as mitochondria, multivesicular bodies, cortical granules and fibrillar lattices showed no difference in morphology between treated and control (intact) oocytes. In two-cell embryos, there was also no evident morphological change except for the fact that many vacuoles appeared clearly in suppressed embryonal cells. In brief, there was no fundamental morphological change in the oocytes and the embryonal cells exposed to progesterone for six hours even though it inhibits their development. The action of progesterone should be investigated thoroughly.
Animal ; Embryo/cytology* ; Embryo/drug effects ; Female ; In Vitro ; Mice ; Oocytes/drug effects ; Oocytes/ultrastructure* ; Ovum/ultrastructure* ; Progesterone/pharmacology*

OBJECTIVEThe present work aims to investigate the effects of subacute exposure to diesel exhaust particles (DEP) on reproductive function in female mice.

METHODSA total of 168 ICR (Institute of Cancer Research) mice were randomly divided into four groups by numeration table method, including the low (B), middle (C), high (D) dose DEP exposure groups and the control group (A). Each group consisted of 42 mice. Mice were inoculated with 30 µl DEP suspension at 0.8 (B), 3.0 (C), 12.0 (D) µg/µl, respectively, or the same volume of phosphate-buffered saline (A) on pharynx posterior wall per triduum for 4 times. The body weight and ovary weight were tested and ovary weight/body weight ratios were calculated. Rates of survival, germinal vesicle breakdown, extrusion of the first polar body, in-vitro fertilization and quantity of mitochondrial DNA for the oocytes were investigated. Ultrastructural changes of the oocytes were observed.

RESULTSThe ovary weight/body weight ratios were (15.4 ± 7.3) × 10(-5), (14.1 ± 6.8) × 10(-5), (8.2 ± 0.7) × 10(-5) and (7.2 ± 2.5) × 10(-5) in groups A, B, C and D (F = 3.841, P < 0.05). In groups A, B, C and D at 48 h post-insemination, rates of oocyte survival were 64.3%, 56.8%, 39.5% and 32.9% (χ(2) = 21.575, P < 0.05), rates of extrusion of the first polar body were 75.5%, 65.3%, 37.0% and 27.1% (χ(2) = 52.772, P < 0.05), rates of 2-cell embryos were 27.9%, 39.1%, 17.6% and 12.5% (χ(2) = 20.148, P < 0.05), and rates of embryos over 2 cells were 45.3%, 32.2%, 12.5% and 13.9% (χ(2) = 32.135, P < 0.05), respectively, and were significantly lower in groups C and D compared with group A (P < 0.05). Logarithmic values of mitochondrial DNA copy numbers were 6.54 ± 0.13, 6.48 ± 0.09, 5.57 ± 0.15 and 5.41 ± 0.07 in groups A, B, C and D, respectively, and were significantly lower in groups C and D compared with group A or B (F = 89.241, P < 0.05). A number of mitochondria of the oocytes exhibited tremendous tumescence and vacuolization in groups C and D, which was contrast to a roughly normal appearance in groups A and B.

CONCLUSIONSDEP is noxious to murine female reproduction. Subacute exposure to DEP injures the ovary and oocyte resulting in compromised ovarian function and fertilizability of the oocyte.

Animals ; Female ; Mice ; Mice, Inbred ICR ; Oocytes ; drug effects ; Ovary ; cytology ; drug effects ; Vehicle Emissions ; toxicity

OBJECTIVEThis study was aimed to determine the effects of n-hexane on the maturation of mouse oocytes.

METHODSCell culture was used to observe the maturation of mouse oocytes and CLSM was employed to determine their apoptosis.

RESULTSGerminal vesicle breakdown (GVBD) and extrusion of the first polar body in mouse oocytes were significantly inhibited by n-hexane. After fertilization, the number of eggs in the mouse was significantly reduced by n-hexane. Mitochondrial membrane potentials (ΔΨm) were altered in mouse oocytes that were leading to apoptosis of the oocytes.

CONCLUSIONN-hexane might have affected the maturation of oocytes, causing alteration of ΔΨm and leading to apoptosis which maybe one of the most important mechanisms.

Animals ; Apoptosis ; drug effects ; Environmental Pollutants ; toxicity ; Female ; Fertilization ; drug effects ; Hexanes ; toxicity ; Male ; Membrane Potential, Mitochondrial ; drug effects ; Mice ; Mice, Inbred ICR ; Oocytes ; drug effects ; growth & development

Mammalian somatic cloning has got great progress during past few years,however,the efficiency of nuclear trasfer remains low. In order to improve this technique, different perspectives of which are studying. Accordingly, the method of oocyte enucleation also becomes a focus. But the physical enucleation is technically demanding, time-consuming, inherently invasive and clearly damaging to cytoplast spatial organization. As a alternative strategy to traditional method, Demecolcine-induced enucleation(IE) attracted the scientist's eyes. Nuclear transfer procedure assited with IE, factors releated with success rate of IE and effects of IE on the oocytes and cloned embryos are mainly discussed in this review. At the same time, combining with author's research, the existing problems of IE and potential improved measures of some key steps in IE procedure are also elucidated. However, the utility of the demecolcine-induced enucleation protocol will require further deep investigation.
Animals ; Cell Nucleus ; drug effects ; Demecolcine ; pharmacology ; Humans ; Models, Theoretical ; Nuclear Transfer Techniques ; Oocytes ; cytology ; drug effects

AIMTo study whether melatonin has effect on oocyte maturation of mouse in vitro.

METHODSMouse oocytes were cultured in maturation medium, HX-medium, or HX-medium supplemented with FSH, and the effects of MT on meiotic maturation of mouse oocyte were examined.

RESULTS(1) MT at all doses of 0.1 g/L, 0.02 g/L, 0.4 g/L or 0.8 g/L inhibited the formation of PB1 in CEO cultured in maturation medium and had no effect on GVBD. (2) MT could delay GVBD and the extrusion of PB1 in CEOs of mouse oocytes by dynamic curves. In contrast to the control, GVBD and PB1 extrusion of oocytes in the treated groups had been delayed by 8-10 hours and 3-4 hours respectively. (3) MT inhibited the effect of FSH on resumption of meiosis, but no effect on the formation of PB1. (4) MT and HX had cooperation effects on spontaneous oocyte maturation in CEO, but not in DO.

CONCLUSIONMT is able to affect mouse oocyte maturation and the regulation mechanisms may be related to cumulus cells.

Animals ; Female ; In Vitro Oocyte Maturation Techniques ; Melatonin ; pharmacology ; Mice ; Mice, Inbred Strains ; Oocytes ; drug effects ; physiology ; Oogenesis ; drug effects

OBJECTIVETo study the effect of epidermal growth factor(EGF) and different concentrations of gonadotrophin (Gn) on the in vitro maturation of human oocytes.

METHODSEGF was added to the vitro culture medium in order to observe the effect of Gn combined with or without EGF on the result of in vitro maturation. The concentrations of hCG and FSH were changed respectively to observe the difference between the results.

RESULTSAdding EGF to the culture medium improved the maturation rate of oocyte significantly (P < 0.05). There was no difference between the results with different concentrations of hCG and FSH in the culture medium.

CONCLUSIONEGF can improve the results of the in vitro maturation of human oocytes by increasing the maturation rate significantly. Increasing the concentration of Gn does not influence the results of in vitro maturation.

Cells, Cultured ; Dose-Response Relationship, Drug ; Epidermal Growth Factor ; pharmacology ; Female ; Fertilization ; drug effects ; Gonadotropins ; pharmacology ; Humans ; Oocytes ; drug effects ; physiology

AIMTo investigate the effects of c-myb on progesterone-induced mouse germinal vesicle(GV) stage denuded oocyte (DO) maturation in vitro.

METHODSWe used mouse GV stage oocyte cultured with special concentration progesterone, or/and antisense c-myb ODN, or/and db-cAMP, or/and heparin for 24 h, and observed oocyte maturation and analysed the relationship among them.

RESULTSWe cultured DO in the medium 199 for 24 h, and found 10 micromol/L progesterone had more significant effect than 5 micromol/L progesterone (2 h GVBD% P < 0.05, 8 h PB 1% P < 0.05), but had not more significant effect than 20 micromol/L progesterone. We found that 16 micromol/L antisense c-myb ODN significantly inhibited progesterone (10 micromol/L)-induced mouse germinal vesicle stage oocyte maturation in vitro (2 h GVBD% P < 0.05, 8 h PBI% P < 0.01). 1 x 10(-4) micromol/L dbcAMP, 100 microg/ml heparin could single significantly inhibited progesterone-induced mouse GV stage oocyte maturation in vitro (2 h PBI% all P < 0.01, 8 h PBI% all P < 0.01), and could enhanced the inhibition of 16 micromol/L antisense c-myb ODN (2 h GVBD% all P < 0.01, 8h PBI% all P < 0.01).

CONCLUSIONProgesterone, protooncogene c-myb,cAMP and calcium all pay important role in regulating oocyte maturation and the mechanism of progesterone, cAMP and calcium in regulating oocyte maturation may be through the expression of protooncogene c-myb.

Animals ; Cells, Cultured ; Genes, myb ; Meiosis ; Mice ; Mice, Inbred Strains ; Oocytes ; cytology ; drug effects ; Oogenesis ; Progesterone ; pharmacology

OBJECTIVETo observe the effect of Kuntai Capsule (KC) on the number of retrieved oocytes, the quality of high-quality oocytes and embryos in in vitro fertilization of poor ovarian response (POR) patients.

METHODSTotally 70 POR patients preparing for in vitro fertilization-embryo transfer (IVF-ET) were randomly assigned to the observation group and the control group, 35 cases in each group. KC was administered to patients in the observation group in the preparation cycle (i.e., three menstrual cycles before IVF-ET) and during the superovulation process. Those in the control group took placebo during this period. Before and after medication the improvement of Shen yin deficiency syndrome (SYDS) was observed in the two groups. The basal follicle-stimulating hormone (bFSH), luteinizing hormone (LH), estradiol (E2), anti-Mullerian hormone (AMH), the ratio of FSH to LH, and antral follicle count (AFC) were observed. Besides, the E2 level of a single ovum on the day of HCG injection, the number of retrieved oocytes, the high-quality oocyte rate, and the high-quality embryos were observed.

RESULTSCompared with the control group, the SYDS, decreased bFSH and LH levels, increased ACF numbers, the E2 level of a single ovum on the day of HCG injection, the number of retrieved oocytes, high-quality oocytes, and high-quality embryos were superior in the observation group (P < 0.05). There was no statistical difference in the decreased FSH/LH level (P > 0.05). E2 and AMH increased after medication of KC in the observation group, while they decreased after administration of placebos in the control group. There was statistical difference in the post-pre treatment difference of E2 and AMH between the two groups (P < 0.05).

CONCLUSIONKC could increase the number of retrieved oocytes, and elevate the quality of occytes and embryos in the IVF-ET.

Adult ; Drugs, Chinese Herbal ; pharmacology ; Embryo Transfer ; Female ; Fertilization in Vitro ; Humans ; Oocyte Retrieval ; Oocytes ; drug effects ; Pregnancy

OBJECTIVETo investigate the characteristic and effect of cadmium on ATP-activated currents (I(ATP)) mediated by P2X4 purinoceptors.

METHODSTranscribe cDNA coding for the rat P2X4 receptor to cRNA in vitro. Inject the cRNA to oocytes of an xenopus laevis using the microinjection technique. Reveal the effect of cadmium on I(ATP) mediated by P2X4 receptor using the two-electrode whole-cell voltage clamp technique.

RESULTS(1) Within a certain concentration range, cadmium was found to reversibly magnify I(ATP) mediated by P2X4 receptors expressed in oocytes of an xenopus. When the concentration of cadmium reached 30 micromol/L, the increase of I(ATP) was the most significant. I(ATP) turned to decrease when the concentration of cadmium was more than 30 micromol/L; (2) The concentration-response curve was shifted to left by applying cadmium at 10 micromol/L; the EC50 was reduced from (17.1 +/- 1.5) micromol/L to (9.8 +/- 1.8) micromol/L (n = 6, P < 0.01) and the Hill coefficient was increased from 1.14 +/- 0.13 to 1.57 +/- 0.36; (3) The effect of cadmium on I(ATP) showed no dependence on membrane voltage; (4) The magnifying effect on I(ATP) reached maximum when preincubating cadmium for 120 seconds.

CONCLUSIONThe increase I(ATP) by cadmium is reversible, concentration-dependent, time-dependent, and voltage-independent. One reason of this augment effect could be the allosteric modulation on P2X4 receptors.

Adenosine Triphosphate ; metabolism ; Animals ; Cadmium ; toxicity ; Microinjections ; Oocytes ; drug effects ; metabolism ; physiology ; Rats ; Receptors, Purinergic P2X4 ; metabolism ; Xenopus laevis

In order to enhance the efficiency of sheep somatic cell nuclear transfer, we used a chemically assisted enucleation with colchicine to study the effects of the concentration of colchicine, the incubation time of oocytes in colchicine and the maturation time of oocytes on the enucleation rates and the development of reconstructed embryos. The results showed that 1) there were no significant differences in the rates of cytoplast protrusion and enucleation between oocytes that were incubated in colchicine (0.4 microg/mL) for 0.5 h and oocytes that were incubated in colchicine (0.4 microg/mL) for 1 h, and the rate of cytoplast protrusion can be 85.4% while the rate of cytoplast enucleation is 100%. 2) There was no significant difference in oocyte enucleation between oocytes treated with medium containing 0.2 microg/mL colchicine for 0.5 h and oocytes treated with medium containing 0.4 microg/mL colchicine for 0.5 h. 3) A maturation time of 18-23 h did not affect the rates of cytoplast protrusion and enucleation by chemically assisted enucleation, whereas the rate of enucleation of oocytes by blind enucleation was found to decrease with a prolonged incubation time. 4) The development rates of reconstructed embryos could not be influenced by these two enucleation methods, increased from oocytes matured for 21-23 h. These results demonstrate that sheep oocytes can be enucleated fast and effectively by optimized colcholine chemically assisted enucleation, which can enhance the enucleation rate of sheep oocytes and the early development of reconstructed embryos in vitro.
Animals ; Cloning, Organism ; methods ; Colchicine ; pharmacology ; Embryo, Mammalian ; embryology ; Female ; Nuclear Transfer Techniques ; veterinary ; Oocytes ; cytology ; drug effects ; Sheep