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Biomedical subjects

J M Sreenan

Publications and source records attributed to J M Sreenan.

At least 37 records · Page 2Linked to original sources
PubMed1999

Immunization of rats and sheep against granulosa cell-inhibitory factor from bovine follicular fluid increases the number of large follicles in rats and the ovulation rate in sheep.

Granulosa cell-inhibitory factor (GCIF), a low molecular weight factor from bovine follicular fluid, inhibits the proliferation of bovine granulosa cells in vitro and the growth of large follicles in rats in vivo. In this study the effects of (1) immunization of rats against GCIF on follicular growth and (2) immunization of sheep against GCIF on ovulation rate were studied. The ability of antiserum from sheep immunized against GCIF to reduce the inhibitory effect of GCIF on bovine granulosa cell proliferation in culture was also examined. Immunization of rats against GCIF increased the number of large follicles (P < 0.001) but decreased the number of small follicles (P < 0.05) per ovary. Ovarian mass (P < 0.05) and uterine wet (P < 0.05) and dry (P < 0.01) masses were increased in immunized rats. Immunization of sheep against GCIF, followed by boosting over two breeding seasons, increased ovulation rate (P < 0.01). Addition of antiserum from sheep immunized against GCIF reduced or abolished the inhibitory effect of GCIF on granulosa cell proliferation (P < 0.01). These data provide further evidence that GCIF has an important role in controlling follicle growth and ovulation in vivo.

Animals↗
PubMed1999

Effect of adenylyl cyclase activation on intracellular and extracellular cAMP and cGMP in preimplantation cattle blastocysts.

The effects of direct and indirect activation of adenylyl cyclase on the production of intracellular and extracellular cAMP and cGMP by 13- to 16-day-old cattle embryos were determined. Embryos were incubated for 2 h in a Krebs Ringer bicarbonate medium containing the phosphodiesterase inhibitor isobutyl-methylxanthine, to which stimulating agents forskolin (100 mumol l-1), cholera toxin (2 micrograms ml-1), or both were added. Total (intra- and extracellular) basal cAMP and cGMP concentrations ranged from 6.65 +/- 0.895 to 3.4 +/- 0.708 fmol microgram-1 protein in 13-day-old embryos and from 4.05 +/- 1.151 to 0.19 +/- 0.041 fmol microgram-1 protein in 16-day-old embryos. Forskolin induced an increase (P < 0.001) in cAMP that ranged from 5.4-fold on day 13 to 2.7-fold on day 16, whereas cholera toxin induced an increase (P < 0.001) that ranged from 30-fold at day 13 to 21-fold at day 16, similar to the effect of forskolin and cholera toxin combined. Individually, forskolin and cholera toxin had no effect on cGMP concentrations, but together they induced an increase (P < 0.05). cAMP (P < 0.01) and cGMP (P < 0.001) concentrations decreased with embryo age from day 13 to day 16 for all treatments; the decrease was greater for cGMP than cAMP (5-24-fold versus 1.6-3.3-fold, respectively). It is concluded that inducible adenylyl cyclase is present in 13- to 16-day-old cattle embryos and that the embryos secrete cAMP and cGMP into the incubation medium. In addition, basal and inducible concentrations of cAMP and cGMP decrease with embryo age from day 13 to day 16. These observations indicate that cAMP and cGMP may have a role in the rapid embryonic cell proliferation that occurs at this time or in signalling to the endometrium.

Adenylyl Cyclases↗
PubMed1998

Effect of calf isolation on follicular wave dynamics, gonadotropin and metabolic hormone changes, and interval to first ovulation in beef cows fed either of two energy levels postpartum.

The effects of postpartum energy intake, restricted suckling, and cow-calf isolation on concentrations of LH, FSH, growth hormone, and insulin-like growth factor-I (IGF-I) and on postpartum anestrous interval were determined by randomly allocating beef cows with a mean body condition score of 2.3 +/- 0.1 to receive either 80 MJ metabolizable energy (low-energy diet [L]; n = 51) or 120 MJ metabolizable energy (high-energy diet [H]; n = 52) per cow per day from calving. At 30 days postpartum, cows within diet were randomized to 1) have continued full access to their calves from birth to weaning (ad libitum suckling: ADLIB), 2) be suckled once-daily with their calves penned adjacent (restricted suckling, adjacent: RESADJ), 3) be isolated from all calves except for a once-daily suckling period (restricted suckling, isolated: RESISO). The mean postpartum interval was similar (p > 0.10) for L and H cows (62 and 63 days, respectively). RESADJ cows had a shorter (p < 0.05) postpartum interval than ADLIB cows, and RESISO cows had a shorter interval (p < 0.05) than RESADJ cows, with all effects independent (p > 0.10) of diet. FSH secretion pattern was not affected by diet, suckling treatment, sequential follicle wave number, or follicle wave retrospectively realigned to emergence of first ovulatory wave. Within 5 days of suckling restriction and calf isolation, the number of LH pulses increased from 0.18 to 0.48 pulses per hour (p < 0.05). Both mean LH and the mean number of LH pulses increased linearly (p < 0.01) during the six follicle waves up to the first ovulatory wave. From 80 days before, until the time of, first ovulation, growth hormone decreased (p < 0.05) while IGF-I increased (p < 0.05), irrespective of treatment. The results indicate that the "suckling effect" in beef cows is the major factor affecting the duration of the postpartum interval and suggests that the maternal bond is more important than suckling in regulating LH pulse frequency, the key endocrine factor determining whether or not a dominant follicles ovulates. Removal of the suckling effect resulted in a rapid increase in LH pulse frequency, which was not dependent on level of postpartum nutrition, at least within the nutritional limits of this study. Mean concentrations of FSH, unlike LH, did not vary with follicle wave number, suggesting that lack of FSH is not a major factor delaying the resumption of ovulation in postpartum beef cows.

Animals↗
PubMed1997

Effect of peptide to carrier ratio on the immune and ovarian response to inhibin immunization in cattle.

We report on the effects of the peptide to carrier ratio on the immune and biological response to inhibin immunization in cattle. A peptide sequence from the alpha C-subunit of bovine inhibin was synthesized and conjugated to human serum albumin (HSA) at ratios of 4.3 moles (L) and 13.1 moles (M) of peptide per mole of HSA. Hereford-cross heifers (n = 6 per group) were injected with 3 mg of one of the peptide conjugates at primary, followed by a booster injection (1.5 mg) 11 weeks later. Control heifers (n = 6) were injected with HSA only. Blood samples were taken at regular intervals to measure antibody titre. Ovulation rate was measured by ultrasonography. Antibodies were generated in both peptide immunized groups. Control heifers and group L heifers had 1 ovulation at all ovulatory cycles monitored. Ovulation rate was increased (P < 0.05) in group M immunized heifers, with four of six heifers having twin ovulations in the first cycle following boost. These data support those of previous studies which indicated that immunization against the alpha C-subunit of bovine inhibin significantly disrupted the mechanism(s) controlling ovulation rate in cattle. It also indicates that both the immune and associated biological response is dependent on the nature of the conjugate used for immunization, specifically the ratio of peptide to carrier.

Animals↗
PubMed1997

Cyclic AMP and cyclic GMP concentrations in, and efflux from, preimplantation cattle embryos.

Basal embryonic cAMP and cGMP concentrations and cAMP and cGMP accumulation in embryos and in the incubation medium were measured in cattle blastocysts recovered at days 14 (n = 23), 15 (n = 29) and 16 (n = 23) of pregnancy. Cyclic AMP and cGMP concentrations were measured by radioimmunoassay and the results expressed per microgram of protein, which was determined by the Pierce Micro BCA protein assay. Cyclic AMP and cGMP were present on each day. Basal embryonic cAMP was similar on days 14, 15, and 16, at 2.3., 2.5 and 2.6 fmol per microgram protein, respectively, while the concentration of cGMP was higher at day 14 (0.14 fmol per microgram protein than at either day 15 or 16 (0.06 and 0.05 fmol per microgram protein, respectively; P < 0.05), which were similar (P > 0.10). Basal embryonic cAMP concentrations were 15 to 60-fold higher than cGMP concentrations. Following a 2 h culture period in the presence or absence of the phosphodiesterase (PDE) inhibitor, isobutylmethylxanthine (IBMX), the accumulated concentrations of cAMP and cGMP were measured in the embryos and in the incubation medium. IBMX did not affect the concentrations of either cAMP or cGMP in the embryos but increased the concentrations of cAMP (P < 0.005) and cGMP (P < 0.01) in the incubation medium. There was no effect of day on either embryonic or medium cAMP, but both embryonic and medium cGMP were higher at day 14 than at day 15 (P < 0.05) or 16 (P < 0.005). Differences between embryonic and medium cAMP and cGMP accumulation were examined. There was no effect of day or treatment. cGMP accumulation in the medium was higher than in the embryos (P < 0.005) whereas cAMP accumulation in the medium was lower than in the embryos (P < 0.05). Whether the efflux of cAMP and cGMP is active or passive is not clear but it may indicate a possible role in embryo-maternal signalling.

1-Methyl-3-isobutylxanthine↗
PubMed1996

Protein content of cattle oocytes and embryos from the two-cell to the elongated blastocyst stage at day 16.

The protein content of cattle oocytes and preimplantation embryos produced in vivo, from the two-cell to the elongated blastocyst at day 16, was determined. From the oocyte to the expanded blastocyst stage (day 8), protein determination was carried out on zona pellucida-enclosed embryos. Protein content was measured by the Pierce Micro BCA protein assay. The mean protein content of oocytes was 0.126 micrograms, with no significant increase at the two-cell stage (0.132 micrograms). Protein content was higher at the morula stage (0.183 micrograms; P < 0.05) with a further increase at the expanded blastocyst stage (0.367 micrograms; P < 0.05). There was a 160-fold increase in protein content from the expanded blastocyst to the hatched day 13 stage. Spherical, ovoid and elongated blastocysts were collected on days 13 and 14. The mean protein content of day 13 (59.8 micrograms) and day 14 (92.4 micrograms) embryos was similar (P > 0.1), but the protein content of the elongated embryos was higher than that of ovoid or spherical embryos collected on the same day. Protein content of day 15 embryos (362.2 micrograms) was higher than that on day 14, with a further increase to 946.6 micrograms by day 16. The correlation between protein content and day of development contained both a linear and a quadratic component. Embryo length and width increased from day 13 (5.24 mm and 0.89 mm, respectively) to day 16 (51.6 mm and 1.82 mm, respectively). From day 13 to day 16, the protein content was correlated with both embryo length and width (r2 = 0.89 and 0.51, respectively; P < 0.001) and was highly correlated (r2 = 0.95) with the product of embryo length by width, indicating that protein content increases as a function of surface area.

Animals↗
PubMed1996

Partial purification from bovine follicular fluid of a factor of low molecular mass with inhibitory effects on the proliferation of bovine granulosa cells in vitro and on rat follicular development in vivo.

Bovine follicular fluid was aspirated from follicles of 2-20 mm in diameter, charcoal-treated to remove steroids and then separated into low and high molecular mass fractions. The low molecular mass (< 10 kDa) fraction was purified on a Sephadex G-25 chromatography column with formic acid as the eluent. Seven peaks were isolated and assayed for biological activity in cultures of bovine granulosa cells at concentrations of 10, 100 and 1000 ng ml-1. One peak (peak 4) inhibited (P < 0.001) the proliferation of granulosa cells when measured by cell counting and by [3H]thymidine incorporation (33-37% inhibition). This peak inhibited proliferation of granulosa cells from both small (< 2 mm) and medium (2-10 mm) follicles, but not large (> 10 mm) follicles. The inhibitory effect of peak 4 was not due to a toxic effect on cells. Administration of peak 4 to rats did not affect liver or kidney masses but did decrease uterine (25%, P < 0.01) and ovarian (35%, P < 0.01) masses. Peak 4 also caused a reduction in the number of large follicles (65%, P < 0.01) but increased the number of small follicles (55%, P < 0.01). We have named the inhibitory factor associated with peak 4, granulosa cell-inhibitory factor (GCIF). The results presented suggest that GCIF may be a factor secreted by dominant follicles that inhibits the development of subordinate follicles.

Animals↗
PubMed1996

Modulation of the effects of FSH, androstenedione, epidermal growth factor (EGF) and insulin-like growth factor I (IGF-I) on bovine granulosa cells by GCIF, a growth-inhibitory factor of low molecular mass from bovine follicular fluid.

A granulosa cell-inhibitory factor (GCIF) of low molecular mass from bovine follicular fluid inhibits the proliferation of bovine granulosa cells in vitro and the growth of large follicles in rats in vivo. The present study examined the ability of GCIF to modulate the effects of FSH, epidermal growth factor (EGF), insulin-like growth factor I (IGF-I) and androstenedione on the proliferation of bovine granulosa cells and on aromatase activity in vitro. Granulosa cell proliferation was assayed by counting haemocytometric cells and by measuring the incorporation of [3H]thymidine into acid-precipitable material. Assay of aromatase activity was based on the conversion of [3H]androstenedione to [3H]H2O. FSH, androstenedione, EGF and IGF-I all stimulated (P < 0.01) granulosa cell proliferation; however, the addition of GCIF reduced (P < 0.01) cell proliferation in their presence. In the case of EGF, the addition of GCIF almost abolished the stimulatory response. FSH and IGF-I, but not EGF, stimulated (P < 0.01) aromatase activity of granulosa cells. The stimulatory effect of IGF-I was decreased by GCIF. The inhibitory effects of GCIF indicate that it may play a significant role in regulating the effects of intraovarian growth factors on granulosa cells and the growth of follicles.

Androstenedione↗
PubMed1995

Effect of immunization against synthetic peptide sequences of the alpha N-subunit of bovine inhibin on ovulation rate, gonadotrophin concentrations and fertility in heifers.

The effects of immunizing cattle against either of two peptides from the amino terminal peptide (alpha N) of the alpha 43-subunit of bovine inhibin on ovulation rate, gonadotrophin concentration and fertility were investigated. Two peptide sequences from the alpha N-subunit of bovine inhibin (P1N, bI alpha-(8-20) and P2N, bI alpha-(153-167)) were synthesized and conjugated to human serum albumin (HSA). Hereford-cross heifers (n = 5 per group) were given an initial injection of 3 mg of one of the peptide conjugates, followed by three booster injections (1.5 mg) at intervals of 11 weeks. Control heifers (n = 5) were injected with HSA only. Blood samples were taken once a week to measure antibody titre and every hour at about the time of the first oestrus and during the mid-luteal phase after the second booster injection, to measure FSH and LH concentrations. Ovulation rate was measured by ultrasonography. Gonadotrophin concentrations were analysed for four periods relative to the peak (time = 0 h) of the preovulatory LH surge as follows: pre-surge: -16 to -5 h; surge: -4 to 4 h; post-surge: 5 to 16 h and a period of 12 h during the mid-luteal (days 10-12) phase. Antibodies that bound to the individual peptides were generated and the ovulation rate increased (P < 0.05) in immunized heifers. Control heifers had one ovulation at all ovulatory cycles monitored. In group P1N, one heifer had two ovulations at each of the six cycles monitored, while another heifer had two ovulations at one cycle.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1993

Effect of immunization against synthetic peptide sequences of bovine inhibin alpha-subunit on ovulation rate and twin-calving rate in heifers.

Three peptide sequences from the bovine inhibin alpha-subunit (P1: 18-30; P2: 63-72 and P3: 107-122) were synthesized and conjugated to human serum albumin (HSA). Hereford cross-heifers (n = 5 per group) were injected with 3 mg of one of the peptide conjugates, followed by three booster injections at intervals of 11 weeks. Control heifers (n = 5) were injected with HSA only. Antibodies recognizing both the individual peptides and 32 kDa bovine inhibin were generated and ovulation rate was increased in peptide immunized heifers. In group P1, 1 of 5 heifers responded with an increased ovulation rate whereas in groups P2 and P3, 5 of 5 and 4 of 5 heifers, respectively, had an increased ovulation rate. In group P2, in the first oestrous cycle following booster injections 2 and 3, 4 of 5 and 3 of 5 heifers, respectively, responded with twin ovulations, whereas a fourth heifer had three ovulations following booster injection 3. After breeding following booster injection 3, 3 of 5 heifers in group P2 and 1 of 5 in group P3 gave birth to twin calves. This study demonstrates the potential of immunizing against synthetic peptide sequences of the alpha-subunit of bovine inhibin to increase ovulation and twinning rates in cattle.

Animals↗
PubMed1992

In vitro preimplantation mouse embryo development with incubation temperatures of 37 and 39 degrees C.

Embryos from two strains of mice were used to assess the effect of incubation temperature on pronuclear and two-cell development to the morula/blastocyst (M/B) stage. Embryos from B6D2F2 and B6SJLF1 strains were cultured in medium M16 at either 37 or 39 degrees C until 120 hr post human chorionic gonadotropin (hCG) or 0, 24, or 48 hr at 37 degrees C and the remaining time at 39 degrees C. Overall M/B development for pronuclear embryos was 0.6, 0, 32.3, and 52.4% for 0-96, 24-72, 48-48, and 96-0 hr at 37 and 39 degrees C, respectively. Only 0-96 and 24-72 hr at 37 and 39 degrees C were not different (P greater than 0.10). Overall M/B development for two-cell embryos was 48.1, 78.1, and 98.0% for 0-72, 24-48, and 72-0 hr at 37 and 39 degrees C, respectively. Percentage development at each time was different (P less than .01) for each category. Additionally, the number of nuclei for morulae and blastocysts tended to be higher for embryos initiating culture at the two-cell stage compared to pronuclear embryos. The first cell cycle was most dramatically affected by a 2 degrees C increase in incubator temperature. More advanced embryos can tolerate slight increases in incubator temperature more readily than pronuclear embryos.

Animals↗
PubMed1991

Effect of immunizing prepuberal lambs of low and high ovulation rate genotypes with inhibin partially purified from bovine follicular fluid.

Active immunization of prepuberal lambs with a partially purified inhibin preparation, isolated from bovine follicular fluid, increased the ovulation rate. In ewe lambs of a low fecundity breed (Suffolk x Galway), the ovulation rate rose from 1.15 to 1.95 (P<0.05) compared with that of the controls. An ovulation rate of 3.38 was recorded for immunized ewe lambs of a high fecundity breed (Finn x Dorset Horn), while the rate for mature ewes from the same flock was 2.29. Immunization did not affect the time of onset of puberty or estrous cycle length. Following immunization, antibodies were produced that bound to a pure preparation of 68kDa bovine inhibin. This report demonstrates the production of antibody to a 68kDa preparation of inhibin following active immunization of sheep using a partially purified preparation. It was concluded that the increased ovulation rate was due to the production of antibodies to inhibin, which may have reduced its negative feedback effect of FSH secretion.

Journal Article↗
PubMed1991

Successful co-culture of 1-4-cell cattle ova to the morula or blastocyst stage.

Bovine ova (n = 326) collected at the 1-4-cell stage were cultured in TCM-199 + 10% foetal calf serum with or without oviducal cells. The bovine oviducal cells were collected and seeded either on the day of ovum recovery (BOC-0) or 3 days earlier (BOC-3). In Exp. 1, the effect of age of oviducal cells in co-culture on ovum development was examined. In the BOC-0 and BOC-3 treatments, respectively, 36/46 (78%) and 30/37 (81%) of ova developed to morulae or blastocysts, while no ova developed past the 8-16-cell stage in the absence of oviducal cells. In Exp. 2, the effect of age of oviducal cells and of physical contact between the oviducal cells and ova on ovum development was examined. In the BOC-0 and BOC-3 treatments, respectively, 29/42 (69%) and 23/43 (53%) of the ova developed to morulae or blastocysts, while 1/42 (2%) developed to the morula stage in the absence of oviducal cells. Physical separation of the ova using a microporous membrane inserted between the oviducal cells and the ova did not affect ovum development, with 26/42 (62%) and 22/42 (52%) of ova developing to morulae or blastocysts in the BOC-0 and BOC-3 treatments, respectively. A high proportion of the morulae and blastocysts in Exp. 1 (57/66, 86%) and Exp. 2 (67/100, 67%) were of quality grades 1 or 2, with mean nuclei counts of 85 for morulae and 111 for blastocysts.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

The efficiency of production, centrifugation, microinjection and transfer of one- and two-cell bovine ova in a gene transfer program.

Forty crossbred beef heifers were superovulated with 2000 IU pregnant mare serum gonadotropin (PMSG) and mated twice by natural service during estrus. Ovulations were counted and ova were recovered during mid-ventral laparotomy between 44 and 54 h after the onset of estrus. The overall donor ovulation rate (M+/-SEM) was 15.2+/-1.3. There was a positive association between ovulation rate and the number of ova recovered (P<0.001), and between ovulation rate and the incidence of ova advanced beyond the two-cell stage of development (P<0.05). When grouped on the basis of superovulation response, the numbers (M+/-SEM) of recovered one-cell, two-cell and more advanced ova were 3.7+/-0.7, 1.0+/-0.3 and 0.5+/-0.3, respectively, for donors with up to 15 ovulations. The corresponding numbers for donors with more than 15 ovulations were 7.2+/-1.8, 6.0+/-1.3 and 2.8+/-1.2, respectively. Following centrifugation, pronuclei were visible in 68% of one-cell ova, and nuclei were visible in 80% of two-cell ova. Approximately 20% of ova were destroyed during DNA microinjection. A total of 66 centrifuged and DNA-injected ova were transferred to the oviducts of 26 crossbred beef heifers, each receiving two, three or four ova. Echography at Day 55 confirmed that 14 (54%) heifers were pregnant with 26 (39%) fetuses. Eleven heifers were held to calve and produced 21 calves.

Journal Article↗
PubMed1989

Effect of a unilateral or bilateral twin embryo distribution on twinning and embryo survival rate in the cow.

Uni- and bilateral twin embryo distributions were effected by the transfer of one embryo on Day 7 to the ipsi- or contralateral uterine horn of previously inseminated heifers (123, Exp. 1) or cows (95, Exp. 2). The embryo transfers were surgical in Exp. 1 and non-surgical in Exp. 2. Transferred and native embryos were distinguished by breed. Embryo survival rate was measured in a proportion (N = 40) of the heifers at Day 53 of gestation and in the remainder of the heifers and all of the cows at term. In the heifers (Exp. 1) overall pregnancy rates of 76% and 75% were recorded after uni- and bilateral twin embryo distributions respectively. Twinning rates of 55% and 60% at Day 53 of gestation and 60% and 60% at term were recorded for uni- and bilateral distributions respectively. In the cows (Exp. 2) calving rates of 61% and 63% and twinning rates of 33% and 38% were recorded following uni- and bilateral twin embryo distributions respectively. When the data from both experiments were combined, overall embryo survival rates were similar for both twin embryo distributions although the ipsilateral transfer of an embryo reduced the survival rate of the native embryo. It is concluded that the confinement of two embryos in one uterine horn on or after Day 7 does not depress pregnancy, twinning or overall survival rate to term.

Animals↗
PubMed1988

Embryo transfer: its uses and recent developments.

The technique of embryo transfer is now routinely employed in several species and with several objectives. In laboratory animals the technique is used mainly in investigations of reproductive biology, whereas in human beings it is used to overcome specific forms of impaired fertility. Because embryo transfer combined with superovulation of the donor can significantly increase the female reproductive rate its greatest application to date has been in farm animals, in which it is widely used in both research and commercial production. Within the past few years there have been many advances in the techniques used in farm animals, particularly in the area of embryo manipulation. The supply of embryos can now be increased by repeated superovulation and by embryo bisection and there has been significant progress in in vitro fertilisation technology. Deep freeze (-196 degrees C) storage of embryos is now routine and may be combined with direct one-step thawing and removal of cryoprotectant. This technique allows the routine non-surgical transfer of embryos in the field. There is also evidence that a routine non-traumatic procedure for sexing embryos may soon be developed. Identical multiplets or clones have been produced by the microdissection of embryos and more recently by the transfer of nuclei from embryos into unfertilized oocytes. Transgenic animals have been produced by the microinjection of recombinant DNA into one of the pronuclei of single-cell unfertilized ova.

Animal Husbandry↗