Biomedical subjects

B N Day

Publications and source records attributed to B N Day.

At least 91 records · Page 5Linked to original sources

PubMed1996

Technical note: porcine non-surgical embryo transfer.

The cervix is a barrier for transferring porcine embryos non-surgically. A non-surgical embryo transfer instrument composed of four parts has been developed that allows the delivery of embryos into the lumen of one uterine horn. Part 1, a modified artificial insemination spirette, is used to produce a cervical lock and to assist in manipulating Part 2 through the cervix. Part 2 is made of stainless steel tubing with a unique curved tip that can be manipulated through the folds of the cervix. Part 3 is a testing bar to ensure that the tip of Part 2 reaches the bifurcation of the uterus. Part 4, a disposable tubing complex for embryo delivery, is composed of a soft plastic leader tubing, a double opened connector for embryo loading and unloading, a semi-hard plastic tubing, and a hard tubing with a water-sealable plunger. Embryos (4-cell to blastocyst stages) were surgically collected and maintained in vitro up to 5 h before the non-surgical embryo transfer. Five of 16 recipients became pregnant (31%) and farrowed an average of 6.2 +/- 3.1 pigs per litter. This easily learned non-surgical embryo transfer technique will simplify embryo transfer in the swine industry and may lead to improvements in swine breeding and scientific research.

Animals↗

PubMed1995

Pronuclear visibility, development and transgene expression in IVM/IVF-derived porcine embryos.

A total of 1550 zygotes was used to assess the timing of pronuclear visibility, embryo development following DNA microinjection, and transgene expression in IVM/IVF-generated porcine embryos. After centrifugation, pronuclei could be seen in 61.6% of zygotes. In 55.3% of these only 1 pronucleus was visible. Pronuclear visibility was highest at 20 h post-insemination. Zygotes were microinjected with 1 of 2 LacZ gene constructs driven by either the SV40 early promoter (pSVON) or the human cytoplasmic beta actin promoter (pbActinLacZ). Development and transgene expression were assessed after either 48 h or 7 d in culture. After 48 h, significantly more zygotes with a single visible pronucleus developed to the 8-cell stage than zygotes in which no pronucleus had been seen (43.0 vs 24.8%), while those with 2 pronuclei were intermediate (31.4%). After 7 d, no difference in development to the morula stage was observed between noninjected control embryos (25.5%) and embryos with 1 (21.0%) or 2 pronuclei (22.5%); however, the proportion of embryos reaching the morula stage in the nonpronuclear group was significantly reduced (9.1%). After 48 h in culture, transgene expression was significantly higher in embryos with 2 pronuclei at the time of injection than in those with 1 (36.4 vs 17.9%). After 7 d in culture, 41.5% of morulae derived from zygotes with 2 pronuclei and 29.97% of thsoe derived from zygotes with 1 pronucleus showed signs of transgene expression. At this stage, significantly more morulae expressed the pbActinLacZ than the pSVON transgene (43.8 vs 25.8%). More than 80% of putative transgenic morulae or blastocysts showed evidence of mosaicism. These results demonstrate that IVM/IVF porcine embryos are able to develop in culture and express a microinjected transgene.

Journal Article↗

PubMed1995

Pronuclear formation and intracellular glutathione content of in vitro-matured porcine oocytes following in vitro fertilisation and/or electrical activation.

Pronuclear formation and intracellular content of glutathione, containing reduced and oxidised forms, in porcine oocytes matured in vitro were determined following insemination and/or electrical stimulation. After insemination, sperm penetration had occurred as early as 3 h and female pronuclei had formed by 6 h with complete development by 12 h. Male pronuclear formation occurred, primarily, between 9 and 12 h after insemination. Glutathione content of the oocytes decreased following sperm penetration and remained at a depressed level until 12 h. After electrical stimulation, oocyte activation had occurred and female pronuclei had formed by 3 and 6 h, respectively. Oocyte glutathione content did not change as a result of oocyte activation. When oocytes were exposed to an electrical pulse and then spermatozoa, female pronuclear formation was observed by 3 h after stimulation/insemination. Sperm penetration was observed between 3 and 9 h. However, the incidence of male pronuclear formation observed at 12 h was extremely low, although sperm decondensation had occurred in some oocytes. Oocyte glutathione content had not decreased by 6 h following electrical activation. These results demonstrate that the changes in glutathione content in porcine oocytes following fertilisation in vitro differ from those due to electrical activation. Further, the decreased intracellular glutathione content in oocytes activated by sperm penetration appears to be due to the presence of a sperm factor.

Animals↗

PubMed1995

Effects of recombinant porcine somatotropin on placental size, fetal growth, and IGF-I and IGF-II concentrations in pigs.

The objective of this study was to determine the effects of recombinant porcine somatotropin (rpST) on placental size, fetal growth, and maternal and fetal IGF-I and IGF-II concentrations. Twenty-four pregnant gilts received daily injections of either 1 mL of saline (control) (n = 12) or 5 mg of rpST (n = 12) from d 30 to 43 of gestation. Gilts were slaughtered on d 44 of gestation, reproductive tracts were removed, and fetal weight and length, placental weight, and implantation length were recorded. There was no effect of rpST on fetal or implantation length. Placental weight increased with rpST administration (71.20 +/- 3.52 vs 58.35 +/- 3.41 g; P < .02), as did fetal weight (18.06 +/- .55 vs 16.44 +/- .53 g; rpST vs control, respectively; P < .05). Implantation lengths were partitioned into quartiles to determine the effect of rpST on fetuses with different implantation lengths. The effect of rpST of fetal weight was greater in the first quartile ( < 137.5 mm) than in the fourth quartile ( > 240 mm) (16.04 vs 13.86 g compared with 19.47 vs 18.21 g, respectively). Analysis using a modified Brody curve suggests that the effect of rpST treatment on fetal weight is equivalent to the effect of increasing implantation length by 58.8 mm. Administration of rpST numerically raised IGF-I (P = .07) and IGF-II (P = .12) concentrations in fetal serum. Although maternal serum IGF-I concentrations were similar at d 30, treatment with rpST increased these concentrations over time (77.76, 247.75, 267.85 vs 82.59, 79.59, 77.97 ng/mL on d 30, 37, 43, respectively; P < .001, SE = 14.09). Maternal IGF-II concentrations were also similar at d 30 but decreased over time with rpST treatment (265.78, 219.61, 191.05 vs 285.44, 284.72, 283.05 ng/mL; P < .03, SE = 14.03). Increased maternal IGF-I concentrations may exhibit negative feedback on maternal IGF-II concentrations. The more pronounced effect of rpST on growth in fetuses with shorter implantation lengths suggests that rpST may increase uptake or utilization of nutrients by fetuses. In addition, nutrient transfer across placental membranes may be enhanced by rpST.

Animals↗

PubMed1994

Developmental ability of porcine oocytes matured and fertilized in vitro.

The developmental abilities of porcine oocytes matured and fertilized in vitro were examined in vivo and in vitro. Cumulus-oocyte complexes were cultured in mM199 supplemented with 10% porcine follicular fluid (PFF) and hormonal supplements (PMSG, hCG and estradiol-17beta) for 20 h and then without hormonal supplements for an additional 20 h. In Experiment 1, oocytes were then co-cultured for 6 h with spermatozoa which had been preincubated with 1% PFF (PFF-treated) or without (control). Oocytes were transferred to oviducts of gilts or cultured in modified Whitten's medium for 5 d. The percentages of oocytes with monospermic penetration (59%, 42 71 ) and with monospermic penetration and male and female pronuclei (32%, 23 71 ) were higher (P < 0.01) in the PFF-treated group than in controls (25%, 18 71 and 8%, 6 71 , respectively). After 5 d, the percentages of oocytes that developed to the morula or blastocyst stages in vitro and in vivo in the PFF-treated group (10%, 28 288 and 13%, 41 318 , respectively) were also higher (P < 0.05) than in controls (2%, 6 284 and 6%, 16 248 , respectively). Whereas some oocytes that were matured and fertilized in vitro developed to the blastocyst stage after 5 d in vivo culture (3%, 9 288 in PFF-treated group and 2%, 6 284 in control), no blastocysts were observed after 5 d when oocytes were cultured in vitro. When the progression of in vitro development of porcine oocytes that were matured and fertilized in vitro was examined in Experiment 2, morulae appeared after 72 h of culture, and 3% (3 100 ) of the oocytes developed to the blastocyst stage after 144 h (6 d) of culture. These results demonstrate that decreasing polyspermic penetration and increasing monospermic male pronuclear formation, as a result of PFF treatment of maturing spermatozoa, improved the developmental ability of porcine oocytes matured and fertilized in vitro. However, development in vitro was delayed by approximately 24 h compared with in vivo development, most of the embryos were blocked at the morula stage.

Journal Article↗

PubMed1994

In vitro development of in vitro-matured porcine oocytes following chemical activation or in vitro fertilization.

Porcine cumulus-oocyte complexes were cultured in BSA-free Whitten's medium or modified Medium 199, each supplemented with porcine follicular fluid (PFF) and hormonal supplements (OMWM and OMM199, respectively) for 20 h; they then were cultured without hormonal supplements for an additional 20 (experiments 1 and 3) or 24 h (experiment 2). At the end of culture (experiment 1), the intracellular glutathione concentration was higher (p < 0.05) in oocytes matured in OMWM vs. OMM199. After activation by Ca2+ ionophore (experiment 2), the incidence of activation in the OMWM group was lower (p < 0.01) than in the OMM199 group. However, the incidence of pronuclear formation was higher (p < 0.01) in the OMWM group than in the OMM199 group at 8 h after activation. The percentage of embryos that developed to the morula stage was higher (p < 0.01) in the group matured in OMWM vs. OMM199 after 5 days of culture. After in vitro fertilization (experiment 3), the incidence of male pronuclear formation and the percentage of monospermic oocytes that formed one male and one female pronuclei were higher (p < 0.05) after maturation in OMWM vs. OMM199. The percentage of cleaved embryos that developed to the 8-cell and morula stages was higher (p < 0.05) in the OMWM group as compared to the OMM199 group. These results indicate that culture in modified Whitten's medium as compared with a standard medium (modified Medium 199) improves cytoplasmic maturation of porcine oocytes as evaluated by intracellular glutathione content, pronuclear formation, and development in vitro after artificial activation or fertilization in vitro.

Animals↗

PubMed1994

Use of low-salt culture medium for in vitro maturation of porcine oocytes is associated with elevated oocyte glutathione levels and enhanced male pronuclear formation after in vitro fertilization.

The effects of sodium chloride (NaCl) in Whitten's medium on intracellular glutathione concentration and on cytoplasmic maturation, as determined by monospermic penetration and male pronuclear formation of porcine oocytes, were examined. Porcine cumulus-oocyte complexes were cultured for 20 h in BSA-free Whitten's medium containing different NaCl concentrations (44.50, 68.49, 92.40, 116.40, or 140.35 mM) and supplemented with 10% porcine follicular fluid and hormonal supplements; the complexes were then cultured without hormonal supplements for an additional 20-h period. The mean width of the perivitelline space of oocytes was increased with decreased concentration of NaCl in the culture medium. Intracellular glutathione concentration was elevated in oocytes cultured in medium with lower NaCl concentrations. After co-culture with spermatozoa for 6 h and culture in modified Whitten's medium for an additional 6 h, there were no differences in maturation and penetration rates among experimental groups. However, the rate of male pronuclear formation was higher in oocytes matured in media with the lower NaCl concentrations. In addition, the rates of monospermic penetration and male pronuclear formation were higher in oocytes matured in medium containing 44.50 mM NaCl (59.3 +/- 8.1 and 70.9 +/- 2.0%, respectively) than in medium containing 68.49 mM NaCl (39.4 +/- 5.5 and 57.1 +/- 4.5%, respectively). These data indicated that decreasing NaCl concentration in maturation medium for porcine oocytes below the customary level improved the quality of the matured oocytes as reflected in higher intracellular glutathione content, wider perivitelline space, higher monospermic penetration rate, and increased frequency of male pronuclear formation.

Animals↗

PubMed1994

Identification of relaxin gene expression and protein localization in the uterine endometrium during early pregnancy in the pig.

The corpus luteum is the primary source of circulating relaxin (RLX) in female pigs. However, a preliminary experiment in our laboratory identified RLX in the uterine endometrium of a day 16 pregnant gilt that had been ovariectomized on day 8 of pregnancy, and the pregnancy maintained via progesterone replacement therapy. Therefore, our objective was to examine the uterus as a potential extraovarian source of RLX during the estrous cycle and early pregnancy in pigs. Reproductive tissues were collected from pregnant (n > or = 3/day) and nonpregnant (n > or = 3/day) gilts on days 10 (n = 6), 12 (n = 6), 14 (n = 6), 16 (n = 8), 18 (n = 6), and 20 (n = 6) of the estrous cycle and pregnancy and on day 42 (n = 2) of gestation. To verify that the RLX identified in uterine tissues was not of ovarian origin, three additional pregnant gilts were ovariectomized on day 8, and the pregnancy was maintained by progesterone replacement therapy until day 16, when the reproductive tissues were collected for immunohistochemistry. The uterine tissues were scored for specific RLX immunostaining and analyzed for the effects day and pregnancy by analysis of variance. Within the uterine endometrium, the luminal epithelium of pregnant pigs contained more RLX than that of nonpregnant pigs (P < or = 0.001). However, there was no difference in either the glandular epithelium or stromal tissues (P > or = 0.10). Increased RLX in the luminal epithelium of pregnant pigs was detected on both days 18 (P < or = 0.05) and 20 (P < or = 0.001) of gestation compared to that in nonpregnant pigs on similar days of the estrous cycle. An effect of day was also observed for RLX immunostaining in the luminal epithelium (P < or = 0.005), but not in glandular or stromal tissues (P > or = 0.10). In both pregnant and nonpregnant pigs, luminal epithelial RLX immunostaining was faint on days 10-12. Thereafter, RLX immunostaining increased on day 14 to reach peak levels on day 16. In nonpregnant pigs, RLX immunostaining dropped to low levels on day 18 and became faint to absent by day 20 of the estrous cycle. In pregnant pigs, RLX immunostaining remained elevated, but appeared to decline slightly by day 20 and became undetectable by day 42 of gestation. RLX immunolocalization in the glandular epithelium was detected on all days in pregnant and nonpregnant pigs, whereas stromal-specific immunostaining was not observed.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1994

Different hormonal requirements of pig oocyte-cumulus complexes during maturation in vitro.

Cytoplasmic maturation as determined by male pronuclear formation following fertilization in vitro was examined in pig oocytes cultured under different hormonal conditions during either the first or second 20 h period of in vitro maturation. Exposure to several combinations of pregnant mares' serum gonadotrophin (PMSG) (10 IU ml-1), hCG (10 IU ml-1) and oestradiol (1 microgram ml-1) for a second 20 h period following culture in a medium supplemented with these hormones for 20 h did not result in differences among treatment groups in maturation rates, penetration rates or polyspermy rates. However, supplementation with PMSG and oestradiol for the last 20 h of culture reduced male pronuclear formation rates significantly. When oocyte-cumulus complexes were cultured in hormone-free media for 20 h after culture in several combinations of supplemental hormones for the first 20 h period, germinal vesicle breakdown rates and maturation rates were lower in oocytes previously exposed to oestradiol alone or no hormonal supplements (68-70% and 45-49%, respectively) than in oocytes previously exposed to PMSG or hCG (89-99% and 71-89%, respectively). Exposure of oocytes to oestradiol alone also reduced the penetration rate (61%) compared with PMSG or hCG (86-99%). Supplementation of media with PMSG alone or together with other hormones increased the male pronuclear formation rate (63-72%) compared with supplementation with oestradiol (33%) or no hormonal supplements (32%). The concentration of oestradiol in maturation medium decreased at filtration (to 216.5 +/- 72.6 ng ml-1) before culture.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Effects of electrical stimulation before or after in vitro fertilization on sperm penetration and pronuclear formation of pig oocytes.

The effects of exposure of pig oocytes to an electrical pulse-on sperm penetration and pronuclear formation were determined before or after in vitro fertilization (IVF). After in vitro maturation (IVM) or after collection from oviducts of unmated gilts, pig oocytes either were not exposed or were exposed to an electrical pulse (a 10 sec pulse at 4.0 Vmm-1 AC followed by a 30 microseconds pulse at 120 V mm-1 DC), followed 30 min later by IVF. The incidence of male pronuclear formation of both IVM and in vivo-matured oocytes at 12 hr after insemination was decreased from 59% and 100%, respectively, to 2% and 36%, respectively, by the electrical pulse, but the penetration rates (88-100%) and polyspermic rates (79-100%) were not affected by exposure to an electrical pulse. Similarly, when pig IVM oocytes were exposed to an electrical pulse at 6 hr after insemination, electrical activation did not decrease penetration rates (93% vs. 90%), polyspermic rates (83% vs. 91%), or number of spermatozoa in penetrated oocytes (4.0 +/- 0.5 vs. 4.6 +/- 0.5) but did decrease the rate of male pronuclear formation from 58% to 18%. When oocytes were examined at 6 hr after insemination, 75% of them had been penetrated and resumed meiotic progression, but all sperm heads in penetrated oocytes were fully condensed or only partially decondensed. The percentage of penetrated eggs with multiple female pronuclei was increased when oocytes were exposed to an electrical pulse in all experimental series.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Effects of media NaCl concentration and osmolarity on the culture of early-stage porcine embryos and the viability of embryos cultured in a selected superior medium.

One- and 2-cell porcine embryos were obtained from oviductal flushes and cultured for 96 hours in media with varied osmolarity that resulted from alterations in NaCl and sorbitol content. The viability of experimental embryos cultured to advanced stages was determined by comparison with that of the controls, noncultured embryos transferred to recipient gilts. The data suggest that variation in embryonic development in the experimental media is related to the NaCl concentration rather than to osmolarity. Increased NaCl concentration impairs development of the embryos to the advanced morula/blastocyst stages (P<0.001). There was no difference in the pregnancy rate between the recipients of cultured (45%) and noncultured (57%) embryos on Day 25. There was, however, a higher embryonic survival rate (P<0.05) within the control gilts.

Journal Article↗

PubMed1993

Effects of different serum supplements in maturation medium on meiotic and cytoplasmic maturation of pig oocytes.

The temporal progression of meiotic and cytoplasmic maturation of pig oocytes cultured in a medium supplemented with 0.4% polyvinylalcohol (PVA), 10% fetal calf serum (FCS), 10% newborn piglet serum (NPS), 10% porcine follicular fluid (PFF) or 10% porcine seminal fluid (PSF) was examined after 20, 30, 40 and 50 hours of culture. There were no differences in germinal vesicle breakdown (GVBD) among FCS and NPS supplements. After 20 hours of culture, the frequency for GVBD was higher (P < 0.05) in FCS and NPS (54% and 52%, respectively) than in PVA (32%) and PFF (33%) culture media but were not different at 40 and 50 hours of culture. Supplementation with PSF resulted in a rapid chromosome condensation of pig oocytes after 20 hours culture, but all GVBD oocytes stopped developing at the condensed germinal vesicle stage. Oocytes were not penetrated by spermatozoa when inseminated following 20 hours of culture, while high penetration (87 to 100%) and polyspermy rates (86 to 100%) were consistently obtained in all the supplement groups when inseminated after 30, 40 or 50 hours of culture. Male pronuclear formation rates at 10 to 12 hours after insemination, following a 50-hour culture period in FCS and NPS, were 28 and 28%, respectively, in comparison with 54% in PVA and 59% in PFF. The results indicate that supplementing maturation media with serum such as FCS and NPS reduced the ability of pig oocytes to form a male pronucleus, and further suggest that the detrimental effects may be due to accelerated progression of maturation events.

Journal Article↗

PubMed1993

Effects of the duration of exposure to hormone supplements on cytoplasmic maturation of pig oocytes in vitro.

The effect of hormone supplements on cytoplasmic maturation in vitro was examined by incubating oocyte-cumulus complexes, in a medium with PMSG (10 iu ml-1), hCG (10 iu ml-1) and oestradiol (1 microgram ml-1) for various periods and then transferring them to medium without added hormones for the remainder of the maturation period. Exposure of oocyte-cumulus complexes to hormone supplements for 2 h improved only germinal vesicle breakdown and maturation rates compared with complexes not exposed to added hormones. The removal of hormone supplements at 20 h after the start of culture enhanced the ability of oocytes to form male pronuclei 10 to 12 h after insemination. Further, the effects of transfer of intact and oocytectomized oocyte-cumulus complexes to hormone-free medium at 20 h on cumulus expansion were examined. The diameter and morphology of the intact oocyte-cumulus complexes were improved after the removal of oocyte-cumulus cell complexes from hormonal exposure. The responses of oocytectomized oocyte-cumulus complexes to hormone were similar to those of intact oocyte-cumulus complexes with the exception of corona radiata expansion. The results suggest that the removal of hormone supplements from maturation media at 20 h after culture enhanced cytoplasmic maturation and cumulus expansion. Further, cumulus expansion does not appear to depend on intercellular communication between cumulus cells and oocytes. Oocytectomy did influence expansion of the corona radiata during culture.

Animals↗

PubMed1993

Reproductive performance in relation to uterine and embryonic traits during early gestation in Meishan, large white and crossbred sows.

Previous studies have shown that females of the Chinese Meishan breed and of their F1 cross with European Large White pigs are very prolific, producing about four more piglets per litter than control Large White females. The main cause of this prolificacy is enhanced prenatal survival for a given ovulation rate in Meishan and F1 females and this is controlled by genes of the mother, not those of the conceptus. The objectives of this study were to determine whether genotypic differences in embryo survival were apparent in the period immediately after attachment and to compare embryonic and uterine development at this time. Sows in their third parity (20 Large White, 14 Meishan, 25 Large White x Meishan F1 and 25 Meishan x Large White F1) were killed 20-22 days after mating and their reproductive tracts recovered for further study. There were significant differences between the purebred sows, and crossbred sows were approximately intermediate for the number of corpora lutea (20.7 +/- 0.9, 27.8 +/- 1.1, 22.4 +/- 0.8 and 23.3 +/- 0.8 for the four genotypes, respectively), the number of embryos (15.2 +/- 0.9, 23.4 +/- 1.1, 17.2 +/- 0.8 and 18.8 +/- 0.8, respectively) and the proportionate embryo survival (0.74 +/- 0.04, 0.84 +/- 0.04, 0.78 +/- 0.03 and 0.82 +/- 0.03, respectively). There was a negative association within genotype between embryo survival and the number of corpora lutea. Adjusting for the genotypic difference in the number of corpora lutea increased the genotypic differences in embryo survival.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Effects of follicular fluid at fertilization in vitro on sperm penetration in pig oocytes.

The effects of porcine follicular fluid (PFF) on sperm penetration of pig oocytes and on prevention of polyspermy were examined and characteristics of spermatozoa exposed to PFF were determined. The addition of PFF at the level of 1 and 10% to the prefertilization and fertilization media decreased penetration rates and the mean number of spermatozoa in penetrated eggs regardless of the origin of PFF. In the presence of BSA, supplementation of 0.1% PFF to prefertilization and fertilization media and 1% PFF to prefertilization media did not decrease the penetration rates but did increase monospermic penetration to 54 and 68%, respectively. When PFF was added to prefertilization media, the number of spermatozoa binding to the zona and the percentage of acrosome-intact spermatozoa decreased with increased PFF concentration (from 43.1 +/- 2.8 and 73.1 +/- 4.9% to 7.2 +/- 1.3 and 15.7 +/- 15.4%, respectively). At the end of prefertilization incubation, sperm agglutination was observed and the degree depended on PFF concentration. Supplementation of fetal calf serum to prefertilization and fertilization media blocked the effects of PFF on sperm penetration and binding of spermatozoa to the zona. These results indicate that the prefertilization incubation of porcine spermatozoa in suitable concentrations of porcine follicular fluid will effectively reduce both the number of spermatozoa that attach to the surface of pig eggs and the incidence of polyspermy.

Acrosome↗

PubMed1992

Immunocytochemical analysis of the association of bovine oviduct-specific glycoproteins with early embryos.

The bovine oviductal epithelium synthesizes and secretes a class of oviduct-specific glycoproteins that is present in the luminal fluid when fertilization and early embryonic development occur. The objective of this study was to determine if these characterized glycoproteins become associated with oviductal embryos. Ovarian ova and oviductal embryos were recovered from super-ovulated cows at 72 h after onset of estrus. Eggs were fixed in 3% paraformaldehyde-1% glutaraldehyde and subsequently embedded in Lowicryl K4M. Sections (1 micron) were processed for peroxidase-antiperoxidase immunocytochemistry. Immunolabeling was not detected in any region of ovarian ova. Oviductal embryos, regardless of cleavage stage, exhibited immunoperoxidase staining localized within their zona pellucidae. Sections (100 nm) obtained from a 4- and an 8-cell embryo were also subjected to colloidal gold immunoelectron microscopy to determine conclusively the subcellular distribution of the oviduct-specific glycoproteins. Gold particles were distributed uniformly throughout the width of the zona pellucida. Also, immunoreactivity was observed associated with flocculent material in the perivitelline space and with the vitelline membrane. These results indicate that the bovine oviduct-specific secretory glycoproteins become associated with oviductal embryos. This association may be biologically important to the developing embryo.

Animals↗

PubMed1992

Influence of ovarian steroids on relaxin-induced uterine growth in ovariectomized gilts.

This study examined the influence of ovarian steroids on the uterotropic actions of relaxin (RLX) in ovariectomized prepubertal gilts. Ovariectomized gilts received (im) corn oil (CO), estradiol benzoate (EB), or EB and progesterone (P) for 0-16 days. Steroid administration was patterned to approximate the plasma concentrations of endogenous ovarian steroids observed during 1) the follicular phase (EB), 2) luteal phase (EB+P), and 3) early pregnancy (EB+P+EB). Half of each group also received PBS or 0.5 mg RLX every 6 h for 54 h, coinciding with the final 2 days of the experimental period. After hysterectomy, uterine tissues were analyzed for water, dry matter, protein, DNA, glycosaminoglycans (GAGs), and collagen contents. Administration of EB or P increased uterine weight 5- to 6-fold, but no differences were observed between EB+P- and EB+P+EB-treated gilts. Cotreatment with RLX enhanced steroid-induced uterine growth 40-70%, and RLX stimulated growth in CO- and EB+CO control gilts 2- to 3-fold. The water content of uterine tissues was greater in EB-, EB+P-, and EB+P+EB-treated gilts than in their respective controls, and this response was augmented by RLX in all treatment groups. Administration of steroids stimulated a 4- to 5-fold increase in uterine dry weight compared to that in controls, with responses not differing between EB+P- and EB+P+EB-treated gilts. In all groups, RLX increased uterine dry weight. Protein and DNA contents of uterine tissue increased with steroid treatment, but neither variable differed between EB+P- and EB+P+EB-treated gilts. Administration of RLX, alone or in combination with steroids, increased protein and DNA contents of uterine tissues. The tissue content of GAGs increased in response to steroids, and coadministration of RLX did not alter this response. Although the uterine tissue concentration of collagen was reduced in steroid- and RLX-treated gilts, the collagen content of the uterus was not affected by the various treatments. The results of this study indicate that RLX is a potent stimulator of uterine growth under a variety of steroidal environments. RLX- or steroid-induced uterine growth was manifest by increased water, dry matter, protein, and DNA and GAG contents, but the uterine content of collagen was not affected. The overall growth-promoting effects of EB and the stimulation of DNA accretion by RLX were not observed when gilts were cotreated with P.(ABSTRACT TRUNCATED AT 400 WORDS)

Analysis of Variance↗

PubMed1991

Effect of cell-free synchronous uterine flushings and microsurgery on the development of porcine embryos in vitro.

Experiment I was designed to determine if cell-free synchronous uterine flushings contain an embryotoxic substance that is normally screened by the intact zona pellucida. Sixty 4-cell embryos were allocated to three treatment groups: 1) control embryos (n = 20) were cultured in Modified Kreb's Ringer Bicarbonate medium + 10% bovine calf serum (mKRB-BCS), 2) UF embryos (n = 20) were cultured in 80% mKRB-BCS + 20% sterile dialyzed uterine flushings (UF), 3) MicroUF embryos (n = 20) received a microsurgical incision in the zona pellucida and were cultured in 80% mKRB-BCS + 20% UF. Following 72 h in culture at 37 degrees C under a 90% N2, 5% CO2, and 5% O2 atmosphere, the number of nuclei/embryo and the incidence of protrusion of the trophoblast through the zona pellucida (PTZ) were recorded. Addition of UF had no effect on embryo development. A greater (P less than .005) proportion of MicroUF embryos exhibited PTZ as compared to UF and control embryos. Experiment II was devised to further characterize the occurrence of PTZ in Micro porcine embryos. Thirty-three 4- to 10-cell embryos and 14 morulae were distributed across two treatments: 1) control embryos (n = 16 and 6, respectively) were cultured as described in Experiment I; and 2) micro embryos were treated similarly to MicroUF embryos in Experiment I but were cultured in mKRB-BCS only. At the onset of PTZ, embryos were immediately fixed and examined. The proportion of embryos exhibiting PTZ was greater (P less than .007) for Micro versus control embryos.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗