共查询到20条相似文献,搜索用时 46 毫秒
1.
Independent studies were undertaken at Alberta Livestock Transplants (ALT) and at Select Embryos, Inc. (SEI) to develop procedures for splitting bovine embryos. At both locations embryos were recovered seven days after the onset of estrus and superovulation. Initially, survival after splitting was evaluated by culture for 18 to 24 hours. Culturing half or demi-embryos without a zona pellucida at ALT resulted in 15% survival compared to 35% survival when both halves were in separate zonae. Culturing demi-embryos on a monolayer of luteal cells at SEI did not improve survival . In fertility trials, best results were obtained at ALT when both demi-embryos within separate zonae were nonsurgically transferred into separate uterine horns of the same recipient (55% pregnancy rate) and at SEI when one demi-embryo was surgically transferred into the uterine horn ipsilateral to the corpus luteum (65% pregnancy rate). Culturing demi-embryos more than 4 hours reduced fertility at both locations. Splitting embryos was a worthwhile addition to the commercial ET programs and further trials are in progress to improve survival and pregnancy rates. 相似文献
2.
Embryos were recovered on Day 4 of pregnancy from superovulated random-bred OF1 Swiss albino mice. They were classified into four categories based on their stage of development: expanding blastocyst, blastocyst, early blastocyst, and compacted morula. They were then cooled at 2 °C/min from ?7 to ?25 °C in a freezing medium containing 1.36 M glycerol and 0.25 M sucrose in phosphate-buffered saline (PBS). At ?25 °C, they were plunged into LN2 and thawed a few hours later in water at 20 °C. After washing in PBS, recovered embryos were cultured for 20 to 24 hr and the number of embryos that had developed normally was recorded. The results showed a clear effect of the stage of development on survival. Survival of expanding blastocysts and blastocysts was very low (1.4 and 21.8%, respectively) compared to that of early blastocysts and compacted morulae (69.4 and 73.5%). The more differentiated stage of the blastocyst (two kinds of cells) and the presence of a blastocoelic cavity may explain the differences observed under our cooling conditions. As a further test of viability, 93 blastocysts that had developed in culture for 20 hr from 153 frozen-thawed early blastocysts and compacted morulae (60.8%) were transferred to 8 recipient mice. Seven became pregnant, yielding normal live young (46.3%). 相似文献
3.
L J Van Winkle 《Biochemical and biophysical research communications》1981,98(2):562-568
Chymotrypsin-like enzyme activity increases transiently in the uterine lumen of ovariectomized mice upon administration of progesterone and estrogen (1). This is one of the few known macromolecular changes associated with conditions which result in activation of delayed implantation blastocysts . , α-chymotrypsin (100 μg/ml) was found to shorten the time required for these embryos to attach to the glass culture dish and then form outgrowths in fetal calf serum-supplemented medium. Higher concentrations of the enzyme (250 μg/ml) prevented embryo attachment probably by digesting the fetuin present in fetal calf serum. Nevertheless, 250 μg/ml α-chymotrypsin could apparently replace fetal calf serum as a stimulator of development during the first 24 hours of culture. In contrast, bovine serum albumin (3.0 mg/ml) seemed to slow development of blastocysts . It is suggested that chymotrypsin-like enzyme activity may stimulate development of delayed implantation blastocysts (a) indirectly by removing inhibitory proteins such as albumin and (b) by directly affecting these embryos in a manner yet to be determined. 相似文献
4.
The effects of endometrium on metabolism of [3H]-arachidonic acid ([3H]-AA) by bovine blastocysts recovered on day 19 postmating were studied . Blastocysts (n = 12) and endometrial slices were assigned to four incubation groups. In group 1, blastocysts were incubated alone; group 2, endometrial slices were incubated alone; group 3, blastocysts were incubated with endometrial slices; group 4, blastocysts were incubated in 7.5 ml fresh incubation medium plus 7.5 ml frozen-thawed medium from endometrial incubations. In all groups, tissues were incubated in 15 ml modified minimum essential medium (MEM) containing 5 μCi of [3H]-AA and 200 μg radioinert arachidonic acid for 24 h at 37°C in an atmosphere of 50% N2:45% O2:5% CO2. For incubation controls, 5 μCi of [3H]-AA were added to 15 ml MEM and incubated at the same time as tissues from each cow. To evaluate metabolism of [3H]-AA, [3H]-AA and its metabolites were extracted from aliquots of MEM and separated on columns of Sephadex LH-20. Most (78.3 ± 3.2%) of the radioactivity (dpm) in the incubation controls was recovered as [3H]-AA, indicating that there was little breakdown of [3H]-AA in the absence of tissue. Blastocysts produced compounds that migrated with [3H]-13,14-dihydro-15-keto-PGF2α ([2H]-PGFM), [3H]-PGE2 and [3H]-PGF2α. Endometrial slices metabolized very little of the [3H]-AA. Data from groups 1 and 4 were combined (group ) for analysis because the distribution of dpm did not differ between the two groups. In group 3, blastocysts and endometrial slices incubated together tended(P<.10) to produced more [3H]-PGE2 than did group , there tended to be less (P<.10)_[3H]-PGF2α, and there was more (P<.05) [3H]-PGFM than in group . Neither endometrial secretions nor endometrial slices altered the proportion of [3H]-AA metabolized by blastocysts. Endometrial slices appear capable of metabolizing [3H]-PGF2α synthesized by blastocysts, and capable of directing blastocyst metabolism of [3H]-AA away from synthesis of [3H]-PGF2α and toward synthesis of [3H]-PGE2. It is postulated that the endometrium has an important role in regulating the amounts and ratios of prostaglandins in th uterine lumen during early prenancy in cows. 相似文献
5.
A total of 126 bovine embryos were surgically collected from 16 superovulated donor heifers 5 days after estrus and randomly selected for either immediate transfer to synchronized recipients or culture at 37°C for 24 hours and subsequent transfer. Twenty-four of 56 (42.8%) embryos maintained for 24 hours in Ham's F10 medium supplemented with 10% heat treated fetal calf serum (HTFCS) and transferred to 32 recipients produced live calves. Survival of 70 noncultured embryos transferred to 35 recipients was 55.7% (39 calves). The percentages of recipients that were diagnosed pregnant at 42 days with cultured and control embryos were 59.4% () and 74.3% (), respectively. No statistical difference was observed between the cultured and control embryos for viability following transfer to recipient females.In a second study, Day 7 embryos maintained in Ham's F10 medium supplemented with 10% HTFC serum for various culture periods were tested for viability following nonsurgical transfer to recipient females. A total of , and embryos cultured for 24, 48 and 72 hours, respectively, resulted in pregnant recipients following transfer. 相似文献
6.
Metabolism of radiolabeled arachidonic acid (1AA) by blastocysts and endometrial slices recovered from five gilts 16 days after detection of estrus was studies . Blastocysts from each gilt were divided into four 216 ± 18 mg, and each portion was placed into a separate petri dish containing 15 ml modified minimum essential medium (MEM)_. The incubates from each gilt received either 25, 50, 100 or 200 μg radioinert arachidonic acid (AA). Endometrium was dissected from each uterin horn, sliced and duplicate 509 ± 3 mg portions from each gilt were placed into petri dishes containing 15 ml MEM and 200 μm AA. All incubates received 5 νCi of 1AA (either [14C]-arichidonic acid or [3H]-arichidonic acid). The incubates were rocked at 37°C for 24 h in an atmosphere of 50% n2:45% O2:5% CO2. After incubation, tissues and MEM were separated by centrifugation. Metabolism of 1AA was assessed in extracts of MEM and tissue homogenates by separating 1AA and its metabolites on columns of Sephades LH-20. Blastocysts produced compounds that migrated with [3H]-13,14-dihydro-15-keto-PGF2α (1PGFM), [3H]-PGE2 (1PGE2) and [3H]-PGF2α (1PGF2α). The greatest (P<.05) proportion (35.7 ± 1.8%) of the radioactivity in blastocyst MEM was recovered as PGE2. In blastocyst homogenates, most (66.2 ± 3.3%; P<0.05) of the radioactivity was in a nonporal peak assumed to be arachidonate esters. The concentration of AA ni MEM did not alter metabolism of 1AA by blastocysts. Endometrial slices produced 1PGFM and 1PGE2 but only in small amounts, and they were capable of producing nonpolar, probably esterified, forms of 1AA. It was concluded that porcine blastocysts produced and metabolized prostaglandins and that they make a contribution to the uterine milieu during early pregnancy. 相似文献
7.
Embryonal carcinoma cells from the in vitro teratocarcinoma cell line PSA-1 were combined with normal, eight-cell stage, embryonic cells of the strain . The aggregates compacted and formed apparently normal blastocysts within 48 hr. Glucose phosphate isomerase (GPI) assays of the blastocysts revealed the presence of both PSA-1 and GPI isozymes. Inner cell masses isolated from the blastocysts by immunosurgery expressed predominantly the PSA-1 GPI type. 相似文献
8.
Mouse blastocysts were exposed to a series of ferritin-conjugated lectins during Day 5 (preadhesive) and Day 6 (adhesive; collected Day 5, 24 hr in vitro) of embryogenesis to determine whether there were any changes in lectin binding characteristics that coincided with the acquisition of adhesiveness. After exposure to lectin, the blastocysts were processed for electron microscopy and lectin binding sites were determined by visualization of ferritin particles with the electron microscope. No binding sites were observed for either Dolichos biflorus agglutinin or soybean agglutinin on blastocysts from either stage examined. Binding sites for Ulex europaeus agglutinin, Con A, and wheat germ agglutinin were seen on blastocysts from both stages without apparent increase or reduction in binding sites from either stage. Ricinus communis agglutinin-I (RCA-I) bound heavily to the surface of Day 5 blastocysts and did not bind at all to Day 6 blastocysts and did bind, though with apparent diminution, to Day 6 blastocysts, as compared with the binding observed on Day 5 blastocysts. Peanut agglutinin (PNA) did not bind at all to Day 5 blastocysts but did bind heavily to the surface of Day 6 blastocysts. Both RCA-I and PNA bound to the surface of embryos during Day 5 of delayed implantation, thus indicating that neither the appearance of PNA binding sites on Day 6 blastocysts nor the apparent reduction of RCA-I binding sites on Day 6 blastocysts could be solely implicated in the acquisition of adhesiveness. PNA binding sites were abolished from the surface of Day 6 blastocysts by treatment with Pronase, indicating that the PNA binding molecule was associated with a glycoprotein rather than a glycolipid. 相似文献
9.
Uterine contractions were observed by laparotomy and exposure of the uterus. Ten hours after the beginning of estrus, an average of 39 contractions per 10 min originated in the posterior ends of the uterine horns and moved toward the oviducts, while an average of, 13 contractions originated in the anterior ends of the horns and moved toward the cervix. Two days later (58 hr after the beginning of estrus), the contractions had changed in origin and direction; only 6 contractions originated in the posterior ends of the horns and moved anteriorly, while 39 originated in the anterior ends and moved posteriorly.Experiments were done to determine whether the change in origin of contractions was reflected in the contractility of strips of myometrium in a tissue bath. The number and amplitude of contractions were recorded from strips of circular and of longitudinal myometrium taken from the posterior and anterior ends of the uterine horns at 10 and 58 hr after the beginning of estrus. The myometrial strips contracted approximately 3 to 4 times per minute regardless of the time after the beginning of estrus, the end of the uterine horn from which the tissue was taken, or whether the contracting muscle was circular or longitudinal. Thus, the physiological mechanisms that controlled the number and origin of uterine contractions did not maintain that control over myometrial tissue . 相似文献
10.
Joseph M. Wright 《Theriogenology》1985,23(1):17-29
Bovine embryos were frozen commercially in clear double length cc French straws with the wick and powder plug in the center of the straw. One-half of the double length straw serves as a handle and contains a color coded cc straw around which an adhesive backed label has been applied. After plunging into liquid nitrogen, straws are transferred into goblets on canes while under liquid nitrogen. The straws are stored in the liquid phase of a nitrogen tank and canes containing straws are not transferred from one container to another unless the goblet containing the straws is full of liquid nitrogen.Embryos held for longer than 4 hours after collection prior to freezing showed a steady decline in pregnancy rate related to the length of time held prior to freezing. The percentage of embryos thawed and then evaluated as being transferrable was related to the quality of the embryos prior to freeze (Grade 1–93.6%, Grade 2–87.0%, Grade 3–63.8%). There was no statistical difference in pregnancy rates obtained from prefreeze Grade 1 embryos when comparing advanced blastocysts (45.2%), blastocysts (38.7%), early blastoclyst (43.1%) and advanced morula (41.6%). 相似文献
11.
Y. Takahashi 《Theriogenology》1981,16(6):631-639
Embryos were non-surgically recovered from superovulated donors. Two trials of ipsilateral single embryo transfer were performed with the Japanese AI instrument.For the first trial, neither the AI instrument nor the cervical expander were ensheathed. The overall pregnancy rate was 33.3 % (). Pregnancy rates obtained from three groups with different media were almost identical.In the second trial, both the AI instrument and the cervical expander were covered with a paper sheath to minimize uterine infection. The overall pregnancy rate after the second trial was 59.1 % (). 相似文献
12.
C. H. Spilman K. K. Bergstrom D. C. Beuving 《Prostaglandins & other lipid mediators》1980,20(6):1061-1074
Serum progesterone and uterine levels of diamine oxidase (DAO) activity were determined during pregnancy in hamsters. Progesterone was elevated on Day 1 of pregnancy, had a transient peak on Day 5, remained relatively constant on Days 6–10, and then increased on Days 13 and 14. Uterine DAO activity could not be detected until Day 7 of pregnancy, approximately 1 days after the initiation of implantation. DAO activity was associated with placental tissue, and more than 90% of the activity was localized in the maternal placenta. The temporal relationship between changes in serum concentrations of progesterone and uterine levels of DAO activity following PG administration also was studied. Serum progesterone was significantly depressed by 6 hr after treatment with PGs on Day 7 of pregnancy. However, uterine levels of DAO activity at 6 hr in the treated animals were not different from those in control animals. In contrast, both the serum progesterone concentrations and uterine levels of DAO activity were significantly lower at 24 hr after PG treatment. The effects of PG treatment on uterine DAO activity were completely blocked by concomitant administration of progesterone. However, concomitant administration of Provera® only blocked the effect of one PG analog that was tested (9-deoxo-9-methylene-16,16-dimethyl0-PGE2). The data indicate that changes in uterine DAO activity following treatment with the PGs used here are primarily a consequence of a decrease in peripheral progesterone (i.e. a luteolytic effect of the PG). 相似文献
13.
C.A. Ruder R.G. Sasser R.J. Williams J.K. Ely R.C. Bull J.E. Butler 《Theriogenology》1981,15(6):561-572
The relationship between adequate (0.96 kg per head daily) and deficient (0.32 kg per head daily) intake of crude protein between 150 days prepartum and 110 days postpartum and the incidence of postpartum infections in the bovine uterus was studied. The incidence of infection at 25 days postpartum was 52.2% and 48.1% (P > .10) for cows in the protein adequate and protein deficient groups, respectively. However, at 40 days postpartum, 21.7% of cows in the protein adequate group had infections 51.9% (P < .05) of cows in the protein deficient group. In addition, the incidence of infection within the protein adequate group between sampling times was different (P < .10); whereas, the incidence within the protein deficient group between sampling times was not different (P > .10). Eighty-three bacterial isolates representing 27 species were recovered from the total of 100 samples. and were the most frequently isolated aerobe and anaerobe, respectively. Although there was no difference between diets (P > .10), these two organisms occurred most frequently in cows on the protein deficient diet and were associated with clinically severe uterine infections. These data suggest that the amount of crude protein in the diet affects both the incidence and duration of postpartum infections in the bovine uterus. 相似文献
14.
Specific versus nonspecific target cell destruction by T lymphocytes sensitized in vitro. II. Responsible factors for nonspecific destruction 总被引:1,自引:0,他引:1
Cell-free supernatants of thoracic duct lymphocyte cultures which were stimulated in vitro by horse serum on syngeneic fibroblast monolayers are demonstrated to be cytotoxic on syngeneic embryonic fibroblasts by means of a direct cell count using microtest plates. Experimental supernatants showed up to 100% suppression of fibroblast growth at dilution and up to 96% suppression at dilution as compared to the control supernatants. Evidence is presented indicating that lymphocytes cultured on mosaic monolayers, which were comprised of syngeneic and xenogeneic fibroblasts, were reacting both to xenogeneic cells and horse serum in the medium at the cellular level. A hapten-to-carrier type relationship is suggested between xenogeneic antigen and horse serum. Absence of horse serum in the test cultures using these lymphocytes resulted in the abrogation of nonspecific toxic activity of lymphocytes while the specific activity, though diminished, remained. This again indicates the difference in the mechanisms underlying the specific and nonspecific target cell destruction by T cells. 相似文献
15.
cultures of sp. were exposed to various concentrations of hydroxyurea (HU) during the logarithmic phase. In the presence of 5 × 10?2M HU, cell division was completely blocked after an initial increase in cell numbers by about 20%. Inhibition of incorporation of 3H-thymidine into acid-insoluble material was effective within 1 hr of exposure to the drug (5 × 10?2M) and it reached a level of 80% after 8 hr. At lower concentrations (5 × 10?4M ? 1 × 10?3M), however, incorporation of 3H-thymidine was remarkably increased while cell division remained unaffected indicating that the increase in incorporation was not due to increased DNA synthesis in preparation for cell division. 相似文献
16.
R Kuttan G J Cardinale S Udenfriend 《Biochemical and biophysical research communications》1975,64(3):947-954
An increase in prolyl hydroxylase activity has been effected in sonicates of early log phase L 929 mouse skin fibroblasts from either monolayer or suspension cultures. The requirements for activation are identical to those needed for the hydroxylation reaction itself, i.e., ferrous ion, ascorbate and α-ketoglutarate. Catalase, which is not an absolute requirement for the hydroxylation, is also necessary for activation. The activation is time dependent and, under the conditions used, is complete in 3 hr at 30°. Since ferrous ion also appears necessary for the activation in intact cells and since the same level of activation is achieved in intact cells as in sonicates, it appears that the activation proceeds in the same manner as that seen in cultured cells. 相似文献
17.
Rat embryos were cultured in serum taken from animals dosed with cadmium, or serum with cadmium added in the presence or absence of additional zinc. Embryos explanted at day ten and grown in serum taken from animals sooner than 4 h after dosing had a reduced DNA content after 24 h culture. In one-hour serum, the yolk sac had become thick and brittle. Zinc ameliorated the effects but had no stimulatory effect on post eight-hour serum when serum zinc levels were at their lowest. The hypothesis that cadmium induces a maternal zinc deficiency sufficient to cause teratogenic changes could not be sustained. Embryos explanted at nine days were much more susceptible to cadmium added than ten-day embryos. The principal anomaly, apart from a reduced DNA content, was a thickening of the yolk sac similar to that seen in embryos grown in serum taken from animals one hour after cadmium dosing. Addition of zinc to the medium prevented both of these effects. The suggestion is made that the cadmium-induced dysgenesis of the yolk sac precludes appropriate embryonic nutrition. 相似文献
18.
Wayne A. Fenton Leon E. Rosenberg 《Biochemical and biophysical research communications》1981,98(1):283-289
We show that the reductants present in the assay used to measure the formation of adenosylcobalamin from cob(III)alamin by cell-free extracts of human fibroblasts result in the non-enzymatic reduction of cob(III)alamin to cob(I)alamin. Hence, the assay uniquely estimates the activity of ATP:cob(I)alamin adenosyltransferase (EC 2.5.1.17). Based on additional studies with extracts of fibroblasts from patients in the class of human methylmalonic acidemia and from their parents, we conclude that this mutant class results from a specific deficiency of adenosyltransferase activity which is inherited as an autosomal recessive trait. 相似文献
19.
Ariane Plet Yoheved Berwald-Netter 《Biochemical and biophysical research communications》1980,94(3):744-754
Culture conditions were defined allowing to demonstrate a stimulatory effect of both serum-contained and purified Somatomedin activity on incorporation of [3H]thymidine and replication of cultured normal human fibroblasts. The use of dialyzed human serum in MEM medium supplemented by 0.2 mM serine offered the necessary and sufficient culture conditions. A significant difference between normal and hypopituitary patients sera was found in their effect on the rate of [3H]thymidine incorporation (p < 0.0001) and on cell replication (p < 0.01). Purified Somatomedin-C, in MEM without serum, is a poor mitogen. Its activity was strongly enhanced by the addition of 0.1 % dialyzed serum and 0.2 mM serine without, however, exceeding the stimulatory level of 1 % whole normal serum. The requirement of concomitant presence, for optimal cell growth, of different low and high MW serum components is discussed. 相似文献
20.
The inhibitory effect of synthetic bovine parathyroid hormone fragment [bPTH-(1-34)] on rat uterine contraction was studied . Oxytocin, prostaglandin F2α and acetylcholine produced log dose-related contraction. The addition of bPTH-(1-34) shifted the dose-response curves of the three agonists to the right. Two doses of bPtH-(1-34) were tested. The higher dose (400 ng/ml) caused a greater inhibition of the agonists than did the lower dose (40 ng/ml). bPTH-(1-34) also inhibited the uterine contraction elicited by electrical stimulation of the tissue. We suggest that bPTH-(1-34) has a non-specific depressing effect on the contractile mechanism of the uterine tissue. 相似文献