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At least 145 records · Page 8Linked to original sources
PubMed1986

In vitro development of bovine morulae in bovine serum albumin, normal steer serum and uterine flushings.

One hundred fifty-three excellent and good bovine morulae were cultured in Ham's F-10, supplemented with 10 % steer serum, bovine serum albumin, or uterine flushings (64 mg protein/ml) to compare embryo development. Embryos were observed every 12 h in culture. Treatment differences were evaluated by assigning a numerical value of 0 to 4 to each embryo representing the stage of development reached in vitro. The final morphological developmental score of the embryos was comparable for steer serum (2.66) and bovine serum albumin (2.50), but it differed significantly for uterine flushings obtained from ovariectomized, steroid-supplemented cows (< 0.1) or heat-treated uterine flushings (0.07). Since albumin alone was able to support development, it suggests that the albumin component of steer serum may be responsible for the observed development. Uterine fluids were unable to support growth of embryos, suggesting that incompatibility may be due to asynchrony between the early bovine embryo and uterine constituents, or a concentration of uterine components may exacerbate actions of inhibitory substances.

Journal Article↗
PubMed1988

Production of normal piglets from microsurgically split morulae and blastocysts.

The embryo splitting technique was applied to pig embryos, and the developmental ability of the split embryos was examined by means of in vitro culture and transfer. Morulae, early blastocysts and blastocysts were collected from Landrace x Large White F(1) gilts which had been mated to Duroc boars. The embryos were bisected with a fine glass or alloy (PtIr) needle after the softening of zonae pellucidae. The halved-embryos, which had either been placed in zonae pellucidae or not, were transferred to recipient gilts immediately after the micromanipulation (Experiment 1) or after cultivation for 15 to 20 h (Experiment 2). In Experiment 1, two fetuses were obtained from one of three recipients which had received 12 half-embryos. In Experiment 2, three of five recipients became pregnant, and in one recipient, seven piglets of a litter were obtained from 12 zona-free half-embryos produced from the original seven blastocysts. The results obtained indicate that a simple method not requiring the encasing of split embryos into zonae pellucidae is satisfactory to produce viable half-embryos.

Journal Article↗
PubMed1988

Influence of uterine flushings from superovulated cows on in vitro bovine morulae development.

To evaluate early embryo development, 248 good to excellent bovine morulae were cultured in Ham's F-10 medium, supplemented with 10% steer serum, uterine flushings from Days 6, 10 or 15 following estrus (0.01, 0.1, 1.0 and 10% protein; 64 mg protein/ml), and 1.0% uterine flushings and 10% steer serum. Final development scores for embryos in steer serum were significantly higher (range across experiments was: 4.06 to 4.37) than for embryos cultured in uterine flushings alone (-0.23 to 0.52). Treatment means were not different (P >0.05) when 10% steer serum was added to 1.0% uterine flushings. A higher percentage of embryos in 10% steer serum (92%) than in 10% steer serum plus 1.0% uterine flushing from Day 6 (33%), Day 10 (45%) and Day 15 (50%) developed to hatched blastocysts. Embryos cultured in 1.0% Day 6 uterine flushings plus 10% steer serum required more time to attain the early blastocyst and blastocyst stages, while embryos in 1.0% Day 15 uterine flushings and 10% steer serum developed at the same rate as controls to the expanded blastocyst stage, but hatched sooner (72.8 vs 96.5 h). These results suggest substance(s) in uterine secretions can have inhibitory and stimulatory influences on early bovine embryo development.

Journal Article↗
PubMed1994

Osmotic and cryoprotective effects of a mixture of DMSO and ethylene glycol on rabbit morulae.

Comparisons were made of the osmotic and cryoprotective effects on rabbit embryos preserved by vitrification with 2 solutions and by conventional freezing. Embryos obtained from rabbits killed 70 to 72 h after mating were used in the study (n = 948). Initially, toxicity of the 3 cryoprotectants was studied in fresh (unfrozen) embryos (n = 135). Subsequently, embryos placed in ethylene glycol (EG, 40% v/v; n = 88) and ethylene glycol with dimethyl sulfoxide (EG+DMSO, 20% v/v each, respectively; n = 344) were loaded into straws and plunged directly into liquid nitrogen. Embryos placed in 1.5 M DMSO and 20% heat inactivated rabbit serum were subjected to conventional freezing in a programmable freezer (control group, n = 363). The osmotic effect was estimated by measuring the changes in the embryonic and interzonal volume (crossectional area) and in the thickness of the mucin coat (n = 18). Cryoprotective effectivity was determined by development to the blastocyst stage in vitro, or birth of normal pups after transfer into synchronized recipients. Osmotic effects of cryoprotective solutions on embryonic and interzonal volume and mucin coat thickness were variable and overall not significant. Survival rate of cryopreserved embryos in vitro and development to blastocysts, was worst in the EG-treated embryos. Survival rate at birth was higher in vitrified vs frozen embryos. We conclude that rabbit morulae can be vitrified successfully in EG+DMSO medium.

Journal Article↗
PubMed1978

Albumin protection of mouse morulae and early blastocysts against the toxic effects of cuprous and cupric ions during development in vitro.

Concentrations of cuprous and cupric chloride of 50 micrometer or greater completely inhibited the development of mouse morulae and early blastocysts into blastocysts in vitro. The zona pellucida was completely dissolved at concentrations of 250 micrometer cupric and 500 micrometer cuprous ions. No significant difference in toxicity was found between the cuprous and cupric ion at 0-50 micrometer. The addition of protein, as BSA, to the culture medium partly protected the embryos against the toxic effect of both copper ions. At a concentration of 25 micrometer a significantly higher level of BSA was required to protect against the cupric than the cuprous ion. Increasing the concentration of either ion increased the level of BSA required to exert a protective effect.

Animals↗
PubMed1986

A survey of the effects of proteases and glycosidases on culture of rabbit morulae to blastocysts.

The effects of a range of commercially available proteases and glycosidases on blastocyst development and hatching were examined on rabbit embryos cultured from the morula stage in a defined medium supplemented with charcoal-treated bovine serum albumin. The proteases tested were trypsin, alpha-chymotrypsin, thrombin, elastase, plasmin, papain, clostripain, collagenase, Streptomyces griseus protease and cathepsin C. The glycosidases tested were neuraminidase, alpha-mannosidase, beta-galactosidase and hyaluronidase. None of these enzymes appeared to stimulate blastocyst growth. The only enzymes which digested the embryonic investments, the zona and mucin coat, sufficiently to cause complete blastocyst hatching were trypsin and Streptomyces griseus protease at relatively low concentrations (250 ng/ml) and chymotrypsin and elastase at higher concentrations.

Animals↗
PubMed2001

Morula cells as the major immunomodulatory hemocytes in ascidians: evidences from the colonial species Botryllus schlosseri.

Immunocytochemical methods were used to study the presence and distribution of IL-1-alpha- and TNF-alpha-like molecules in the hemocytes of the colonial ascidian Botryllus schlosseri. Only a few unstimulated hemocytes were positive to both the antibodies used. When the hemocytes were stimulated with either mannan or phorbol 12-mono-myristate, the phagocytes were not significantly changed in their number, staining intensity, or cell morphology. In contrast, stimulated morula cells were intensely labeled, indicating that these cells play an important immunomodulatory role.

Animals↗
PubMedDate not supplied

Survival of pig morula and blastocyst after exposure to vitrification media or vitrification.

The aim of this experiment was to examine the survival of porcine embryos following exposure to vitrification solutions and vitrification. The work was carried out on non-cultured and cultured morulae and blastocysts. The viability of treated embryos was assessed by their ability to develop in in vitro culture. The results showed that the most detrimental step in the vitrification of pig embryos is exposing them specifically to a vitrification medium rather than the vitrification process itself. Moreover, we demonstrated the beneficial effect of culture on the viability of pig embryos after vitrification

Journal Article↗
PubMed1990

Survival of bisected rabbit morulae transferred to synchronous and asynchronous recipients.

The rabbit was used as a model to test the concept that temporal asynchrony is required to establish physiological synchrony when embryos are bisected to produce demiembryos. In preliminary studies with intact embryos it was confirmed that embryos harvested on days 2, 3, 4, or 5 (day 0 = day of breeding) can be transferred with +/- 1 day of asynchrony to the uteri of recipient rabbits. Three experiments were conducted with bisected embryos. In experiment 1, 192 bisected and 194 control day 3 embryos were transferred to uteri of day 2, 2.5, and 3 recipients (ovulated 0, 12, and 24 h after the donors), with 14% of the bisected and 39% of the intact embryos (P less than .05) resulting in young. Only 4% (2/48) of the day 3 bisected embryos vs. 39% (P less than .05) of the intact day 3 embryos survived in the uteri of day 2 recipients. In experiment 2, day 3 bisected and intact embryos were transferred to the oviducts of day 3, 3.5, or 4 recipients, the speculation being that the oviduct might provide a more neutral environment than the uterus. However, embryo survival was very low, except for the intact embryos transferred to synchronized recipients (42% young born). In experiment 3, 150 intact and 162 (81 pairs) bisected day 3 embryos collected from uteri were transferred to uteri of day 2.5, 3.0, and 3.5 recipient does. Significantly more pregnancies (100% vs. 47%, P less than .01) and young born (56% vs. 19%, P less than .01) resulted from intact embryos than from bisected embryos, irrespective of the uterine age.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2000

Effect of Mycoplasma bovis and Mycoplasma bovigenitalium in semen on fertilization and association with in vitro produced morula and blastocyst stage embryos.

Frozen-thawed bovine semen contaminated with Mycoplasma bovis (M. bovis) or Mycoplasma bovigenitalium (M. bovigenitalium) at either a high (10(6) CFU/mL) or low (10(4) CFU/mL) concentration was used for bovine oocyte insemination. The resulting embryos were washed 10 times as recommended by the International Embryo Transfer Society (IETS) prior to isolation of agent. A total of 1494 oocytes was inseminated with contaminated sperm cells and 855 oocytes with uninfected control semen. There was a significantly higher proportion of embryos that developed to the blastocyst stage in control than in the mycoplasma exposed groups (P<0.05). Isolation of motile spermatozoa by swim-up procedure prior to insemination did not render sperm cells free of Mycoplasma spp. Although M. bovis was isolated from all washed embryos after the high exposure level, it was found in only 60% of the samples after the low exposure level. In contrast, M. bovigenitalium was isolated from 70 and 12% of washed embryos exposed to the high and low levels of microorganism, respectively. Using scanning electron microscopy, both microorganisms were detected in association with the surface of zona pellucida-intact embryos and with sperm cells. These results indicate that mycoplasmas present in semen can be transmitted through the IVF system and infect embryos. Furthermore, the experiments showed that supplementation of culture media with standard antibiotics and washing embryos as recommended by IETS were not effective in rendering IVF embryos free from M. bovis and M. bovigenitalium.

Acrosome↗
PubMed2003

Vitrification of in vitro cultured rabbit morulae.

In the present work, we attempt to establish an efficient vitrification procedure for 32-cell rabbit embryos obtained in vitro. In experiment 1, both the effect of the composition of the vitrification solutions and the cryoprotectant addition (either in one or two steps) were studied. For one-step addition, straws with embryos in the final vitrification solution (total time 60s) were plunged into liquid nitrogen. For two-step addition, previously embryos were 2 min pre-equilibrated in 0.5 ml of (1:1) PBS plus 20% FCS: vitrification solution without sucrose. Different solutions of cryoprotectants were compared: 25 vol.% ethylene glycol supplemented with 0.25 M sucrose (25EG+S) and 20% ethylene glycol plus 20% dimethyl sulfoxide, alone (20EG+20DMSO-S) or supplemented with 0.25 M sucrose (20EG+20DMSO+S). Six percent (30/487) of the total of 32-cell embryos obtained by in vitro culture in each experimental session was slow-frozen by a classical method as a technical efficiency control. Only 30% slow-frozen embryos reached blastocyst stage. Significant differences in embryo development were detected between the one-step (25EG+S) and two-step (25EG+S) groups and the one-step (20EG-20DMSO+S) and two-step (20EG-20DMSO-S) groups (0-6% versus 36-50%, respectively). Consequently, in the following experiments only these two vitrification procedures were used. In experiment 2, we attempted to substitute the use of PBS by HEPES-buffered Ham's F-10 (h-CM) in all cryoprotective solutions or media. When h-CM was used, a significant reduction in the in vitro embryo development was observed when the HEPES-buffered groups were compared with one-step (20EG-20DMSO+S) group in s-PBS (35-45% versus 73%). In experiment 3, the one-step (20EG+20DMSO+S) and two-step (20EG+20DMSO-S) procedures were assayed using two FCS levels (20 and 40%) in the PBS-based media. Relative to in vitro development, the highest rates were reached with one step (20EG-20DMSO+S), using PBS plus 20% FCS, which was different from two steps (20EG-20DMSO-S), regardless of percentage of FCS in the PBS-based media (81% versus 41-45%; P<0.05). In conclusion, we propose either the one step (20EG-20DMSO+S) or two steps (20EG-20DMSO-S) prepared in PBS plus 20% serum for use in future works.

Animals↗
PubMed1994

Morulae at compaction and the pattern of protein synthesis in mouse embryos.

Compaction of mouse embryos at the 8-cell stage causes a drastic change in cell form and in cell-to-cell contacts in 3-4 h. We have studied the effect of inhibitors of transcription (alpha-amanitin), DNA replication (aphidicolin) and compaction (cytochalasin D, EGTA, alpha-lactalbumin and Con A) on the pattern of protein synthesis using gel electrophoresis. Our results show that the pattern of protein synthesis is regulated principally by passage through S phase during each early cell cycle rather than by de novo transcription, while changes induced in cell form or contacts do not alter the pattern significantly.

Amanitins↗
PubMed1987

Effects of coculture with uterine epithelial cells on the metabolism of glucose by mouse morulae and early blastocysts.

Coculture of mouse morulae/early blastocysts with isolated endometrial epithelial cells reduced incorporation of glucose carbon into embryonic glycogen but had no significant effect on incorporation into other internal carbon pools during a 5-h culture in serum-supplemented Dulbecco's modification of Eagle's minimum essential medium. Turnover of glycogen pools during 24-h chase culture of pulse-labelled embryos was unaffected by the presence of uterine epithelial cells recovered from day-4 pregnant or non-pregnant mice. However, significantly more label was retained in non-glycogen macromolecules during chase in the presence of endometrium recovered from non-pregnant than from pregnant uteri.

Animals↗