小鼠超数排卵优化试验方案研究

探讨建立一种适合贵州地区、高效、稳定的小鼠超数排卵优化方案。在饲养环境相同的基础上,对激素(PMSG, hCG)不同的剂量组合、注射间隔时间、小鼠周龄等影响因素进行了相关研究。试验结果表明:(1)平均采胚数量组间、平均异常胚组间与平均可用胚组间差异显著(p<0.05),注射10 IU的激素剂量组合获得受精卵最多,且异常胚最少,效果最佳。(2)第1、第2、第3组平均采胚数量组间差异显著(p<0.05),第1组与第2组平均可用胚组间差异不显著(p>0.05),但第1、2组与第3组差异显著(p<0.05),异常胚组间差异不显著(p>0.05),选择4周龄超排效果最佳。(3)第1、第2、第3组平均采胚数量、平均可用胚组间差异显著(p<0.05),平均异常胚组间差异不显著(p>0.05),PMSG,hCG间隔注射时间为48 h为最佳。     

Ultrasonographic fetometry and prenatal fetal sex assessment in camels (Camelus dromedarius)

The aims of this study were to determine the developmental patterns of some fetal parts to achieve a high accuracy level in the assessment of gestational age and to assess the feasibility and accuracy of ultrasonic prenatal fetal sex assessment in camels. Serial ultrasonographic examinations were carried out on seven pregnant dromedary camels. A total of 329 ultrasonographic examinations were conducted between the second and the 54th weeks of pregnancy. Intrauterine fluid accumulation was detected between the second and third weeks of pregnancy. The embryo proper was noticed between the third and fourth weeks. Organization of the embryo was first observed between the sixth and seventh weeks. Ossification was first detected between the seventh and ninth weeks. The accessibility during the total gestational period was 35/329 (10.6%) for crown-rump length, 35/329 (10.6%) for biparietal diameter, 42/329 (12.8%) for abdominal diameter, 42/329 (12.8%) for ruminal length, and 126/329 (38.3%) for eyeball diameter. A high correlation was found between gestational age and each of the studied parameters (P < 0.0001). The highest correlation was found with the crown-rump length and the biparietal diameter during the first trimester and with the eyeball diameter during the third trimester of pregnancy. The overall accuracy of the ultrasonic prenatal fetal sex assessment was 91.7%. The best window was found during the 11th week of pregnancy. It was concluded that sonographic fetometry can be useful for the evaluation of fetal development, the estimation of gestational age, and the prediction of prenatal fetal sex in camels.     

无籽八月桔胚囊及胚胎发育细胞学观察

运用酶解振荡压片技术和常规石蜡切片技术分别研究了无籽八月桔的胚囊育性及无籽八月桔自交和异交(无籽八月桔×台湾椪柑,无籽八月桔×有籽八月桔)的胚胎发育.结果表明:无籽八月桔胚囊可育,成熟胚囊具一个卵细胞、两个助细胞、三个反足细胞以及一个大的含二个极核的中央细胞;其自交和异交的胚胎发育均正常,授粉后2周出现球形胚和少量心形...     

The effect of recipient age on the success of embryo transfer in mice

Eight-hundred-sixteen morulae and blastocysts were transferred to the uteri of 102 recipients of various ages on Day 3 or 4 of pseudopregnancy (Day 1 is the day of the vaginal plug). Day 3 recipients had significantly higher pregnancy rates and embryo survival rates than Day 4 recipients. Recipient age had little effect on pregnancy rates, but had a significant effect on embryo survival in Day 3 recipients. Day 3 recipients of 11-13 weeks of age had the highest pregnancy rate (100%) and embryo survival (75%). The results suggest that recipient age should be considered an important factor in embryo transfer experiments.     

Duration of fertility and hatchability of the common duck (Anas platyrhynchos) in pure- or crossbreeding with Muscovy drakes (Cairina moschata)

A total of 540 common duck dams were used for a comparison of duration of fertility and hatchability between eggs issued from common dams inseminated with sperm (175 x 10(6) dose(-1)) from either common (pure-breeding or PB) or Muscovy (crossbreeding or CB) drakes. Artificial inseminations (AI) were performed at 3 periods of the reproductive season (27-35, 39-43 and 49-56 weeks) with 2 alternate inseminations/period at 3-week intervals (one with semen from common and the other from Muscovy). Fertility was estimated from egg candling while early embryo mortality (EEM), medium embryo mortality (MEM) and late embryo mortality (LEM) was estimated on Days 0-6 (PB+CB), Days 7-25 (PB) or Day 28 (CB) of incubation, and after, respectively. Overall fertility from Days 2-12 after AI was 61.1% in PB and 42.8% in CB. The maximum duration of fertility (time interval between AI and last fertile egg) was 8.1 days in PB versus 6.4 days in CB (p<0.05). The age of the dam influenced this interval, particularly in PB, with a longer duration at 40 weeks compared to 50 (p<0.05). On average, EEM represented 2.5% of fertile eggs while MEM accounted for 5% of surviving embryos on Day 6 and LEM, for 11.5% of hatched eggs. MEM was significantly higher in CB (6.3%) compared to PB (3.9%; p<0.05). Overall, an increase in EEM and MEM was observed in both types of eggs at and after 50 weeks of age. An increase in EEM (regardless of dam's age) and in MEM (only in the oldest females) was observed with sperm storage duration. Sex ratio at hatching (49.2% males in PB vs. 53.0% in CB) was particularly unbalanced on the first fertile day (54.7% and 57.1%, respectively).     

Comparison of fertility and embryo mortality following artificial insemination of common duck females (Anas Platyrhynchos) with semen from common or Muscovy (Cairina Moschata) drakes

The purpose of this study was to compare fertility and early embryo mortality rates (< or = 5 days of incubation) following artificial insemination (AI) of common duck females (Anas Platyrhynchos) with semen from either common or Muscovy (Cairina Moschata) drakes at various periods of the reproductive season (Period I, 27-35 weeks; Period II, 39-43 weeks and Period III, 49-56 weeks). Based on observations performed by stereomicroscopy on eggs laid from Days 2 to 10 after AI, we confirmed that fertility was significantly lower in the interbred compared to the purebred cross at each of the periods tested (purebred 58.1, 61.2 and 54.2 versus crossbred 31.0, 40.4 and 39.5 at Periods I, II and II, respectively; 0.01 < P < 0.001). In a complementary experiment, we demonstrated that the number of perivitelline spermatozoa (NPS) was markedly lower in mule (crossbred) eggs compared to common (purebred) eggs, a strong indication that initial sperm selection occurring in the lower oviduct is probably more intense after crossbred compared to purebred insemination. Comparison of early embryo mortality (EEM) between mule and common duck eggs indicated that increased levels of EEM in mule embryos corresponded to Stages II-IV of the Eyal-Giladi and Kochav classification (EGK). While a similar age-dependent increase in early embryo mortality was observed in eggs from both genetic origins during the latter periods of the reproductive season, it was also established that embryo mortality due to parental age was related rather to Stages X-XIV of the EGK classification in eggs from both genetic origins. It is concluded that the relative subfertility of mule compared to common duck eggs is probably the consequence of a more intense rate of selection of heterologous than homologous spermatozoa occurring in the vaginal portion of the oviduct while the causal origins of EEM in mule duck eggs can at least in part be identified on the basis of precise staging (by stereomicroscopy) of dead embryos.     

ART患者早期流产组织染色体异常及相关因素分析

目的:分析ART患者早期流产组织染色体异常及其相关影响因素。方法:回顾性分析2013-2017年ART患者行早期流产组织染色体检查的409例样本,分析胚胎染色体非整倍性发生及其与女方年龄、不孕年限、不孕因素、促排卵指标之间的关系。结果:ART流产患者中,流产组织染色体非整倍性发生率为57.46%,发生频次以16三体占比最高(23.95%),其次是22三体(13.45%)及Turner(9.24%)。流产组织染色体非整倍性患者平均年龄高于染色体整倍性患者(P0.001)。16三体组患者年龄低于22三体(P0.01)及Turner组(P0.05)。16三体组患者平均Gn使用量低于22三体组(P0.05)。16三体组患者移植15天血HCG值低于22三体(P0.05)及Turner组(P0.01)。结论:ART患者流产组织染色体非整倍性与女方年龄正相关,但16三体及Turner的发生与女方年龄相关性不大,且16三体更容易引发早期流产。     

白及种子萌发与快速繁殖技术的研究

将不同胚龄的白及种子接种到不同培养基中进行培养,对白及种子萌发率、萌发时间、丛生芽增殖、生根等方面进行了研究。结果表明:白及种子萌发率与其胚龄及有胚率成正相关,萌发时间则与胚龄及有胚率成负相关;胚龄16周的种子在1 g/L花宝1号 2 g/L花宝2号的培养基上萌发率可达84%;胚龄等于或大于20周的种子萌发率不受培养基成分的影响,均可达100%,萌发时间只需7 d;丛生芽增殖的最佳培养基为1/2 MS 4.0 mg/L6-BA 0.2 mg/L NAA 100 g/L CM,其增殖倍数达4.41倍;诱导生根较好的培养基为1/2 MS 0.2 mg/L NAA,生根率达90%。     

The repair processes and cell populations relationship in bone marrow of patients with acute radiation injury: morphological study

The results of bone marrow pathomorphologic investigation in patients who died of acute radiation sickness (3.7-13.7 Gy) after the Chernobyl accident are presented. Stratification into three consecutive periods was performed according to the length of postexposure survival (14-96 days) and the morphologic picture dynamics: 1) bone marrow aplasia (2-3 weeks after exposure) with a domination of stromal and plasma cell populations, exhibiting the morphologic characteristics of functional activity; 2) initial repair period (3-4 weeks after exposure) was characterized by the beginning of hematopoietic proliferation with simultaneous occurrence of cell death via apoptosis; 3) active repair period (4 and more weeks after exposure) was defined by increased marrow cellularity (trilineage regeneration), accompanied by decreased number of plasma cells and occurrence of large granular lymphocytes.     

Morphological study on the hypoplastic testis with aplasia of the epididymis, ductus deferens, and gland of the ductus deferens in the TW inbred rat

In the TW inbred rat, about 50% of the males show bilateral or unilateral testicular hypoplasia with aplasia of the ipsilateral epididymis, ductus deferens and gland of the ductus deferens. To investigate the pathogenesis of the testicular abnormality in the TW rats, the weight and morphology of the testes on the aplastic and normal side were studied between one week and one year of age. The weight of the testes on the affected side was greater than those on the normal side at four and five weeks. However, it rose to a plateau at six weeks and then remained at about one half to one third the weight of a normal testis. As for the testicular histology, there were no obvious changes from one day to three weeks of age. The diameter of the seminiferous tubules became larger and the number of germ cells decreased at four and five weeks. At six weeks, degeneration and loss of germ cells were observed and many multinucleated giant cells appeared. Thereafter, the loss of germ cells became more severe, and they eventually disappeared with increasing age, but Sertoli's cells continued to exist. In the interstitial area, edematous changes and proliferation of Leydig's cells were observed. The efferent duct of another strain, with normal testes, was ligated at three weeks of age, and changes of the testis after the operation were examined to investigate whether or not these anomalies of the TW strain were due to the absence of the accessories, which may block the excretion of the testicular fluid.(ABSTRACT TRUNCATED AT 250 WORDS)     

Paternal Diet-Induced Obesity Retards Early Mouse Embryo Development,Mitochondrial Activity and Pregnancy Health

Worldwide, 48% of adult males are overweight or obese. An association between infertility and excessive body weight is now accepted, although focus remains primarily on females. It has been shown that parental obesity results in compromised embryo development, disproportionate changes in embryo metabolism and reduced blastocyst cell number. The aim of this study was to determine whether paternal obesity has negative effects on the resultant embryo. Specifically, using in vitro fertilisation (IVF), we wanted to isolate the functional effects of obesity on sperm by examining the subsequent embryo both pre- and post-implantation. Epididymal sperm was collected from age matched normal and obese C57BL/6 mice and cryopreserved for subsequent IVF with oocytes collected from Swiss females (normal diet/weight). Obesity was induced in male mice by feeding a high fat diet of 22% fat for 10 weeks. Resultant embryos were cultured individually and development monitored using time-lapse microscopy. Paternal obesity resulted in a significant delay in preimplantation embryo development as early as syngamy (P<0.05). Metabolic parameters were measured across key developmental stages, demonstrating significant reduction in mitochondrial membrane potential (P<0.01). Blastocysts were stained to determine trophectoderm (TE) and inner cell mass (ICM) cell numbers, revealing significant differences in the ratio of cell allocation to TE and ICM lineages (P<0.01). Functional studies examining blastocyst attachment, growth and implantation demonstrated that blastocysts derived from sperm of obese males displayed significantly reduced outgrowth on fibronectin in vitro (P<0.05) and retarded fetal development in vivo following embryo transfer (P<0.05). Taken together, these data clearly demonstrate that paternal obesity has significant negative effects on the embryo at a variety of key early developmental stages, resulting in delayed development, reduced placental size and smaller offspring.     

Effects of in ovo injection of bovine lactoferrin before incubation in layer breeder eggs on tibia measurements and performance of laying hens

There is increasing concern about welfare of laying hens in cages, and one aspect of this topic relates to bone fragility. Therefore, bone anabolic components such as bovine lactoferrin (bLF) may be an effective strategy to maintain the integrity and health of bones. A total of 1080 eggs were divided into four groups with three replicates, each comprising 270 eggs; (1) control group was injected with 100 μl of normal saline per egg; (2, 3 and 4) groups including 22.5 (low), 45 (medium) and 67.5 µg (high) of bLF in 100 µl of normal saline per egg. Eggs were incubated and after hatching, chicks were reared to 28 weeks of age. Tibia measurements were obtained at hatch and at 28 weeks of age. Tibia weight at hatch, was not influenced by in ovo injection of bLF in comparison with the control. Eggs injected with the high concentration of bLF (67.5 µg of bLF per egg) showed significant strengthening in laying-hen tibias at 28 weeks of age, as measured by ultimate force and bending stress, compared with the control. Egg weights from hens treated with this concentration of bLF were also significantly greater than the control. Our data suggest that tibia cortical thickness is a suitable variable for evaluating bone status reflecting bone integrity and strength. The present study also shows that bLF (67.5 µg of bLF per egg) injected into layer breeder eggs before incubation can be used to improve bone strength and egg weight of laying hens at 28 weeks of age, while having no detrimental effect on embryo hatchability.     

Embryo-rescue culture of the ‘Galia’ muskmelon (Cucumis melo L. var. reticulatus Ser.) male parental line

In order to obtain a reliable embryo-rescue technique for wild type (WT) and transgenic ‘Galia’ muskmelon male parental line we evaluated three distinct parameters: nutrient media (E20A basic medium and E21, with six new supplements), two culture systems (removing the embryo from the seed or intact seed), and the use of embryos from fruit at increasing days post pollination (DPP). Transgenic muskmelon plants with the ACO gene in antisense orientation were obtained using a protocol previously described. Fruits were harvested at 4, 10, 17, 24 and 30 DPP. The embryos were either removed from the seeds or left in the seeds and placed in E-20A or E-21 medium for 30–35 days. Seedlings (well developed cotyledon) from all treatments were transferred to E-21 elongation medium, incubated 5 weeks, and transferred to soil to evaluate growth. The efficiency of this technique was greater as embryo age (DPP to rescue the embryo) was increased. Embryos 17–30 DPP had the greatest efficiency for embryo rescue, although embryos could be rescued as early as 4 DPP. The number of rescued embryos using an improved medium (E-21) was greater than E-20A basic medium. Survival efficiency rate was the same for WT and transgenic embryos. We have obtained a competent embryo-rescue technique for WT and transgenic ‘Galia’ male parental line with better efficiency rates than others previously reported.     

SPF级小鼠超数排卵效果

目的探求小鼠周龄、品系和季节对SPF级小鼠超数排卵效果的影响。方法在春、夏、冬季对4～10周龄的SPF级BALB／c小鼠和KM小鼠进行超数排卵处理,通过见栓率、产胚率、3．5d胚胎获得量及胚胎类型等方面进行统计学分析。结果小鼠周龄、品系和季节对SPF级BALB／c小鼠和KM小鼠见栓率没有影响;对产胚率、3．5d胚胎获得量和胚胎类型等方面有一定影响。结论SPF级BALB／c小鼠宜用9、10周龄的进行超数排卵处理以供应3．5d胚胎为需要所用,而SPF级KM小鼠使用4～10周龄的都可以;另外,SPF级小鼠超数排卵处理仍宜在春、夏季进行,可以保证获得最佳的效果。     

Development of hamster one-cell embryos recovered under different conditions to the blastocyst stage

One-cell stage embryos, recovered from superovulated golden hamsters (8 to 12 weeks of age) 12 hours after egg activation, were cultured in HECM-1 medium at 37 degrees C and 5% CO(2) in air. The culture conditions investigated were the time and temperature required for embro recovery, the pH shift of the washing medium, and the oxygen concentration of the gas phase during and after embryo recovery. Each condition was assessed by the developmental efficiency of the embryo as determined by morphological criteria. As the time required for embryo recovery was reduced, the developmental rates of the embryos were improved: 2.3% (3 128 ) 26.9% (35 130 ) at 5 and 3 minutes, respectively, as determined by the number of embryos developed to the blastocyst stage. No blastocysts were obtained when more than 10 minutes were required for embryo recovery. As the oxygen concentration was reduced from 40 to 20% or to 5%, rather high developmental rates were obtained even when the time required for embryo recovery was prolonged: 6.9% (9 130 ) and 21.7% (28 129 ) of the embryos developed to the blastocyst stage when they were recovered under 5% oxygen within 10 and 5 minutes, respectively. Neither the temperature during embryo recovery (37 degrees C and 25 degrees C) nor the pH shift (pH 7.22 to 7.52) of the washing medium used in embryo recovery procedures influenced the development of the embryos. These findings suggest that the developmental block in hamster embryos may involve oxidative stress, which may result from exposure to high oxygen concentration and light during the manipulation of oocytes and embryos.     

Rearing of coconut mite Aceria guerreronis and the predatory mite Neoseiulus baraki in the laboratory

A method was developed for the rearing of coconut mite, Aceria guerreronis Keifer (Acari: Eriophyidae), and its predatory mite Neoseiulus baraki (Athias-Henriot) (Acari: Phytoseiidae) on embryo culture seedlings of coconut (Cocos nucifera) in the laboratory. Seedlings in the ages of <2, 2–4 and 4–6 months were infested with 75 field-collected coconut mites and the population growth was determined up to six weeks after introduction. The populations of coconut mites increased exponentially up to five weeks after introduction and declined thereafter on seedlings of all ages with significant differences among the three groups of seedlings occurring over time. At week 5, a significantly higher mean number (±SE) of coconut mites (20,098 ± 3,465) was bred on 4–6-month-old seedlings than on smaller seedlings, and on the largest seedlings the numbers were highest at all time intervals, except at week 2. Neoseiulus baraki was reared on embryo culture seedlings of the three age groups infested with coconut mites, by introduction of five female deutonymphs and one male, three weeks after introducing coconut mites. Predator numbers progressed significantly over time, but the size of seedlings did not significantly influence the numbers. On all groups of seedlings, the mean number of N. baraki increased up to two weeks after introduction on to seedlings and then declined. Many coconut mites were successfully reared in the laboratory for a longer period by this method and it could also be used as an alternative method to rear N. baraki. Development of this method may contribute to the progress of studies on the biology and ecology of coconut mite and its interactions with natural enemies.     

Fertilization and embryo loss in Booroola Merino x South Australian Merino ewes: Effect of the F gene

The effects of Booroola genotype (F+, ++); the number of ovulations per ewe (one, two or three); and the age of a ewe (2.5 yr vs 3.5 to 6.5 yr) on the percentage of ova fertilized, embryo loss and fetal loss were examined in Booroola x South Australian Merino ewes slaughtered on Days 4, 21 and 90 after insemination. Ewes slaughtered on Day 90 were examined by real-time ultrasound imaging (RUI) on Day 45. Fertilization failure was independent of ewe genotype, ovulation rate and age of ewe, and it was not an important source of wastage (F+, 9.4%; ++, 6.7%). Most embryo loss occurred during the first 21 d (F+, 54.7%; ++, 40.3%). Interpretation of the effects of genotype and ovulation rate on embryo wastage measured on Days 21, 45 and 90 was obscured by significant (P < 0.05) genotype and ovulation rate interactions with the day of slaughter/RUI. The effect of age on embryo loss was not significant (P > 0.05). Reasons for the high rate of wastage observed in this experiment require further study.     

NONDEEP COMPLEX MORPHOPHYSIOLOGICAL DORMANCY IN SEEDS OF OSMORHIZA CLAYTONII (APIACEAE)

Freshly matured seeds of Osmorhiza claytonii exhibit a type of morphophysiological dormancy (MPD). Under natural conditions, embryo growth begins in late September and early October and continues until mid***- to late February, with the peak in October and November. Most seeds germinate between mid-February and late March. Embryos did not grow in seeds incubated for 24 weeks at 30/15 (warm stratification) or 5 C (cold stratification). However, in seeds given 12 weeks at 30/15 and then 12 weeks at 5 C, embryo length increased 1,246% while seeds were at 5 C. Zero to 7 days of warm followed by 24 weeks of cold stratification resulted in 2%–27% germination of fresh seeds, whereas 2 to 12 weeks of warm followed by 24 weeks of cold stratification resulted in 80%–98% germination. Warm plus cold stratification was required for embryo growth and germination of seeds that remained undispersed for a year in the field. GA₃ was partially effective in substituting for warm stratification. The name “nondeep complex MPD” is proposed for the type of MPD found in O. claytonii and a few other species, making a total of eight types of MPD presently known.     

Electron microscope studies on the development of the external zone of the mouse median eminence

Summary The development of the external zone of the median eminence of the mouse was studied in the electron microscope. The examination follows the development of the embryo from the 15th day of the gestation period and the juvenile growth until 24 days of age.Single terminals of the tubero-infundibular neurons of the external zone were found to extend to the outer basement membrane of the perivascular space of the portal primary capillary plexus in the 16 day-old embryo. In the 18 day-old embryo a narrow external zone has developed. Organization of the external zone into the adult pattern is accomplished at the age of three to four weeks. Small agranular as well as large granular vesicles are present in the tubero-infundibular nerve terminals even in the 16 day-old embryo.Changes in the organization of the nerve endings along the outer perivascular basement membrane in relation to the ependymal vascular feet were considered.     

Biosynthesis and Degradation of a Wheat Embryo Cytokinin-Binding Protein during Embryogenesis and Germination

The accumulation and degradation of a wheat (Triticum durum) embryo cytokinin-binding protein (CBF-1) was followed during embryo development and germination by its N⁶-benzyladenine (BA) binding activity and immunological reactivity (rocket immunoelectrophoresis and Western blotting). Both BA binding activity and CBF-1 appeared at 2 weeks post-anthesis and rose sharply between 2 to 4 weeks before leveling off to approximately 47 micrograms per embryo (9% of the soluble embryo protein at maturity). In vitro translation of polyadenylated RNA from 20-day-old embryos yielded a polypeptide which was immunoprecipitable with anti-CBF-1 IgG and migrated closely to the 54-kilodalton CBF-1 polypeptide on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Upon germination, both the amount of CBF-1 and BA binding activity dropped to low levels within 3 days. The data are discussed in relation to the possible role of CBF-1 as a regulator of cytokinin availability, and comparisons are drawn between the structural and biosynthetic similarities found between CBF-1 and the vicilin storage proteins of legumes. An improved method for isolating undegraded CBF-1 from whole seeds is also presented.