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1.
J Phys Condens Matter ; 26(7): 075102, 2014 Feb 19.
Article in English | MEDLINE | ID: mdl-24451554

ABSTRACT

We discuss the low energy effective theory of gapless excitations of the mass vortices of systems similar to the Ginzburg-Landau description of superfluid helium-3 in the bulk B-phase. Our approach is to determine the vortex solution by considering a specific ansatz for the order parameter and minimizing the free energy. The conditions on the ßi coefficients required for the stability of the various solutions for the order parameter are calculated. By considering the symmetries that are broken by the vortex solutions we are able to generate the modulus fields associated with the low energy excitations of the vortices. Using these fields we determine the effective free energy describing the dynamics of these excitations.

2.
Reprod Fertil Dev ; 24(7): 962-72, 2012.
Article in English | MEDLINE | ID: mdl-22935157

ABSTRACT

The uterus provides the nurturing environment that supports the growth of the early preimplantation bovine conceptus. To determine critical time points of uterine influence, in vitro-produced Day 7 blastocysts were transferred into synchronous (Day 7) uteri and asynchronous uteri (Days 5 or 9). Embryo growth was evaluated 7 and 15 days after transfer and compared with that of embryos generated by AI. Conceptuses recovered from asynchronous Day 9 transfers were fourfold larger than synchronous transfer or gestational Day 14 AI conceptuses; by 15 days after transfer, differences were less marked. Two-dimensional gel electrophoresis was used to compare the histotroph protein composition of uterine luminal flushings (ULF) on Days 5 and 9 after oestrous to determine any protein differences that would promote embryo growth. The ULF were collected by serially flushing the uteri of the same heifers and mature cows at different times of the cycle. Ten proteins that differed in abundance between Day 5 and 9 were identified by mass spectrometry. Three, namely phosphoserine aminotransferase 1, purine nucleoside phosphorylase and aldose reductase, were verified by western blot analysis as more abundant on Day 9 (P<0.002). Myostatin was present in only in Day 9 ULF, whereas tissue inhibitor of matrix metalloproteinase 2 (TIMP2) and legumain were only detected in Day 14 ULF. Although mature cows had lower progesterone concentrations on Days 5 and 14 (P<0.05) and tended to have less TIMP2 than heifer groups, no other protein differences were detected. Thus, the embryo growth-enhancing environment on Day 9 was associated with temporal changes in the expression of several proteins of the histotroph.


Subject(s)
Blastocyst/metabolism , Embryo Implantation , Embryo Transfer/veterinary , Proteins/metabolism , Proteome , Uterus/metabolism , Aldehyde Reductase/metabolism , Animals , Blotting, Western , Cattle , Cysteine Endopeptidases/metabolism , Electrophoresis, Gel, Two-Dimensional , Embryo Culture Techniques/veterinary , Female , Gestational Age , Insemination, Artificial/veterinary , Myostatin/metabolism , Pregnancy , Progesterone/blood , Proteomics/methods , Purine-Nucleoside Phosphorylase/metabolism , Reproducibility of Results , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Tissue Inhibitor of Metalloproteinase-2/metabolism , Transaminases/metabolism
3.
Reprod Fertil Dev ; 23(6): 737-47, 2011.
Article in English | MEDLINE | ID: mdl-21791175

ABSTRACT

During previous investigations, the capacity of the cow to secrete prostaglandin in response to oxytocin has been linked with pregnancy outcome. The objective of the present study was to evaluate the predictive value of prostaglandin release to identify groups of cows as potentially superior (SR, low prostaglandin release) or inferior (IR, high prostaglandin release) for pregnancy outcome and to utilise these cows to investigate factors that contribute to optimum uterine conditions for early pregnancy. Animals were synchronised and received an in vitro-derived blastocyst on Day 7 post-oestrus. Tissues (trophoblast and endometrial) and uterine luminal fluid (ULF) were recovered 10 days later. Pregnancy rates were 94 and 78% for SR and IR cows, respectively. Of the pregnant SR cows, 69% had larged conceptuses (>24 cm) in contrast to 43% IR of cows. IR cows with small conceptuses (<12 cm) had significantly lower mean Day 3 and 5 post-oestrous progesterone concentrations than cows with large conceptuses. The expression of factors involved in the prostaglandin pathway, pregnancy and conceptus development were analysed via quantitative RT-PCR and Western blot analysis. Investigation of 16 endometrial gene transcripts indicated no differences between IR and SR cows except for osteopontin expression which, in uteri with large conceptuses, was 2-fold greater in SR than IR cows (P=0.02). There was greater expression of CTGF, OXTR, PGES, PGHS2 and UTMP mRNA in uteri of SR and IR cows that had large compared with small conceptuses (P<0.05). More IFNT protein was recovered in SR compared with IR ULF (P<0.03). SR cows with large conceptuses had less TIMP2 and legumain protein in their gravid, compared with their non-gravid horns (P≤0.02) whereas IR cows did not. The predictive value of prostaglandin release in response to oxytocin challenge does not appear to be an effective indicator of subsequent pregnancy rates in cows. Differences between the two groups appear to be associated with subtle differences in progesterone and uterine protein concentrations that may be related to differences in conceptus size.


Subject(s)
Cattle , Embryo, Mammalian/physiology , Pregnancy Outcome/veterinary , Pregnancy, Animal/physiology , Prostaglandins/metabolism , Uterus/physiology , Animals , Blotting, Western/veterinary , Body Size , Breeding/methods , Endometrium/metabolism , Female , Gene Expression Profiling/veterinary , Osteopontin/metabolism , Oxytocin , Pregnancy , Real-Time Polymerase Chain Reaction/veterinary
4.
J Anim Sci ; 89(11): 3551-60, 2011 Nov.
Article in English | MEDLINE | ID: mdl-21666004

ABSTRACT

The focus of this study was to investigate the effect of subclinical endometritis (scEndo) on ovarian follicular steroid concentrations in early postpartum pasture-fed dairy cows. Mixed-age lactating dairy cows (n = 169) were examined to ascertain uterine health status on d 21 postpartum (±3 d). From this herd, a cohort of scEndo and uninfected cows (n = 47) were selected using uterine cytology to determine scEndo. To ensure cows with scEndo were selected for the study, a conservative threshold [>18% polymorphonuclear (PMN) cells among uterine nucleated cells] was chosen as a selection threshold. Ovarian follicular dynamics were assessed by ultrasonography on d 21, 42, and 63 postpartum. On the latter 2 d, all follicles >4 mm in diameter were ablated, and 4 d later, the largest (F1) and second largest (F2) follicles were measured and their follicular fluid aspirated. Hematological variables and plasma metabolites were measured also on these days to further characterize scEndo cows. On d 21, the prevalence of scEndo was approximately 9% in this herd; by d 42 infections had self-resolved in the majority (81%) of those cows classified as having scEndo on d 21. The scEndo cows had a delayed return to cyclicity; however, no effect was evident on ovarian follicle size or growth rate. Weeks after scEndo had self-resolved and cyclicity was restored, decreased (P = 0.07) testosterone and increased (P = 0.07) cortisol concentrations were evident in F1 follicles of scEndo compared with uninfected cows. Progesterone concentrations of F1 increased (P < 0.05) in 11- to 16-mm diameter follicles of scEndo cows, whereas estradiol, androstendione, and dehydroepiandrosterone concentrations were decreased (P < 0.05) in F1 8- to 10-mm diameter follicles of scEndo cows. These 3 steroids also differed (P < 0.05) between F1 follicle size categories of scEndo but not uninfected cows. On d 21, mean plasma albumin concentration was decreased (P = 0.02) in scEndo cows. In summary, early postpartum scEndo had surprisingly long-term influences on the steroid concentrations of ovarian follicles long after infections had self-resolved. This is likely to affect oocyte quality and may partially explain the reduced conception rates and longer interval between calving and conception that are often associated with scEndo, although more detailed investigations are required to substantiate this theory.


Subject(s)
Cattle Diseases/metabolism , Endometritis/veterinary , Follicular Fluid/chemistry , Ovarian Follicle/metabolism , Androstenedione/metabolism , Animals , Cattle , Cattle Diseases/diagnostic imaging , Cohort Studies , Dairying , Dehydroepiandrosterone/metabolism , Endometritis/diagnostic imaging , Endometritis/metabolism , Estradiol/metabolism , Female , Follicular Fluid/metabolism , Hydrocortisone/metabolism , Ovarian Follicle/diagnostic imaging , Postpartum Period , Testosterone/metabolism , Ultrasonography
5.
Anim Reprod Sci ; 122(3-4): 164-73, 2010 Dec.
Article in English | MEDLINE | ID: mdl-20832205

ABSTRACT

Impaired reproduction in farmed animals is a major cost to agriculture, and this is exacerbated by the implementation of intensive production systems. Addressing this has been the focus of a significant body of research. While considerable advances have been made in biological experiments and understanding, a systems insight into the mechanisms that underlie reproductive function in mammals is needed. Mathematical modelling offers a means to develop a systems approach to reproduction by coalescing information and predicting outcomes of interventions. There has been steady progress in the development of mathematical models addressing various issues of reproduction over the last decade, from cell-signalling pathways through to herd management. We review these developments and their insights as well as their limitations. In addition, we identify other areas that need development, and how modelling might usefully contribute to these areas of reproduction science.


Subject(s)
Animals, Domestic/physiology , Models, Biological , Reproduction/physiology , Animals , Breeding/methods , Estrous Cycle/physiology , Estrus Detection , Female , Fertilization , Gonadotropin-Releasing Hormone/physiology , Gonadotropins/physiology , Models, Theoretical , Oocytes/physiology , Ovary/physiology , Oxygen/administration & dosage , Pregnancy
6.
J Theor Biol ; 266(1): 62-9, 2010 Sep 07.
Article in English | MEDLINE | ID: mdl-20553942

ABSTRACT

In this paper we develop a mathematical model of the luteal phase of the reproductive cycle in mammals with the aim to generate a systems understanding of pregnancy recognition. Pregnancy recognition is initiated by the production of interferon tau (IFNtau) by the growing conceptus. This ensures that the maternal corpus luteum (CL) remains viable to secrete progesterone, which is critical for providing a uterine microenvironment suitable for embryonic growth. Our mathematical model describes the interactions among the CL, the reproductive hormones and the hormone receptors in the uterus. It also characterises the complex interactions amongst the uterine oestrogen, progesterone and oxytocin receptors that control the sensitivity of the uterus to oestrogen, progesterone and oxytocin, respectively. The model is represented by a dynamical system and exhibits qualitative features consistent with the known experimental results in sheep. A key factor identified was a time-dependent threshold for the IFNtau signal below which the presence of the embryo might not be recognised and thus pregnancy would likely fail. Furthermore, the model indicated that if the IFNtau signal is later than around day 13 of the cycle, then pregnancy will not be recognised irrespective of the IFNtau concentration. The thresholds in the concentration and time of the IFNtau signal is a screening mechanism whereby only embryos of sufficient quality are able to prevent luteolysis (i.e. regression of the CL). The effect of progesterone secretion rate from the CL on pregnancy recognition was investigated. The model suggests that if the secretion rate is low then the initiation of the IFNtau signal is delayed, which in turn compromises the likelihood of a pregnancy being recognised by the CL. Furthermore, pregnancy recognition does not occur below a critical threshold in the progesterone secretion rate. In summary, the model can be used to identify the most favourable conditions for pregnancy recognition.


Subject(s)
Mammals/metabolism , Models, Biological , Pregnancy/metabolism , Algorithms , Animals , Computer Simulation , Corpus Luteum/growth & development , Corpus Luteum/metabolism , Dinoprost/metabolism , Embryo, Mammalian/metabolism , Estrogens/metabolism , Female , Interferon Type I/metabolism , Luteal Phase/metabolism , Luteolysis/metabolism , Oxytocin/metabolism , Pregnancy Proteins/metabolism , Progesterone/metabolism , Receptors, Estrogen/metabolism , Receptors, Oxytocin/metabolism , Receptors, Progesterone/metabolism , Sheep/metabolism , Time Factors , Uterus/metabolism
7.
J Dairy Sci ; 92(8): 3723-30, 2009 Aug.
Article in English | MEDLINE | ID: mdl-19620654

ABSTRACT

Poor reproductive performance limits cow longevity in seasonal, pasture-based dairy systems. Few differences in ovarian dynamics have been reported in different strains of Holstein-Friesian cows, implying that the uterine environment may be a key component determining reproductive success. To test the hypothesis that the uterine environment differs among genetic strains of the Holstein-Friesian cow, endometrial fatty acids (FA) were analyzed from New Zealand (NZ), and North American (NA) Holstein-Friesian cows. The effect of reproductive status was also investigated, with cows from both Holstein-Friesian strains slaughtered on either d 17 of the estrous cycle (termed cyclic) or d 17 of pregnancy (after embryo transfer; termed pregnant). Endometrial tissues were collected from 22 cows (NZ pregnant, n = 6; NZ cyclic, n = 4; NA pregnant, n = 6; NA cyclic, n = 6), and FA composition was analyzed. Daily plasma progesterone concentrations, milk production, milk FA composition, body weight, and body condition score were determined. Milk yield (4% fat-corrected milk) was similar for the NZ (28.5 kg/d) and NA (29.3 kg/d; SE 2.07 kg/d) cows, but NZ cows had a greater mean milk fat percentage. Mean plasma progesterone concentrations were significantly greater in NZ cows. Plasma progesterone concentrations were similar in the pregnant and cyclic groups. Mean length of the trophoblast recovered from the pregnant cows (NZ: 20.8 +/- 2.84 cm; NA: 27.9 +/- 10.23 cm) was not affected by genetic strain. Endometrial tissues from NZ cows contained greater concentrations of C17:0, C20:3n-3, and total polyunsaturated FA. The endometria from pregnant cows contained greater concentrations of C17:0, C20:2, and C20:3n-6, and less C20:1, C20:2, C20:5n-3. The observed changes in endometrial FA between Holstein-Friesian cows of different genetic origins or reproductive states may reflect differences in endometrial function and may affect reproductive function.


Subject(s)
Cattle/physiology , Endometrium/chemistry , Fatty Acids/analysis , Animals , Body Constitution , Body Weight , Breeding , Cattle/genetics , Fats/analysis , Female , Lactation , Milk/chemistry , Milk/metabolism , Pregnancy , Progesterone/blood , Regression Analysis , Trophoblasts/physiology
8.
Anim Reprod Sci ; 111(2-4): 353-60, 2009 Apr.
Article in English | MEDLINE | ID: mdl-18436399

ABSTRACT

Two studies tested the hypothesis that eicosapentaenoic (20:5omega3; EPA), docosahexaenoic acids (22:6omega3; DHA) or linoleic acid (C18:2omega6; LIN) reduced bovine endometrial and trophoblast prostaglandin F(2alpha) (PGF(2alpha)) and prostaglandin E(2) (PGE(2)) release during short-term culture. In Study 1, endometrial tissues were collected from non-lactating, non-pregnant cows and endometrial plus trophoblast tissues from pregnant cows 16 days post-insemination. In Study 2, endometrial and trophoblast tissues were collected on day 17 of pregnancy, from cows synchronised using a double prostaglandin (PG) or Ovagentrade mark synchronisation. Tissues were incubated in medium only (M) or media supplemented with fatty acids: eicosapentaenoic (20:5omega3; EPA), docosahexaenoic acids (22:6omega3; DHA) or linoleic acid (C18:2omega6; LIN). In Study 1, PGE(2) release from 'pregnant' endometria was higher (P=0.094) than from 'non-pregnant' endometria, while PGF(2alpha) concentrations were similar. Fatty acids treatment had no effect on PGF(2alpha) or PGE(2) release from either pregnant or non-pregnant endometria. Individual fatty acid treatments had no effect on the ratio of PGF(2alpha) to PGE(2) from trophoblast tissues, but when the data from the 3 fatty acid treatments were combined (EPA, DHA and LIN treatment groups) the ratio of PGF(2alpha) to PGE(2) was reduced (P=0.026) when compared to medium only. In Study 2, PGE(2) concentrations were higher (P=0.013) from the trophoblast collected from Ovagentrade mark cows as compared to that of the PG synchrony group. When the data from the 3-omega fatty acids were combined (DHA and EPA treatment groups), the 3-omega treatments decreased (P<0.05) PGE(2) biosynthesis from both endometrial and trophoblast tissues from animals synchronised following PG synchrony but not Ovagentrade mark synchrony. Short-term culture with low concentrations of 3-omega fatty acids tended to reduce prostaglandin release from trophoblast collected 16 days after insemination, with the type of synchrony modifying PGE(2) production from the trophoblast tissues collected 17 days after insemination. The ability of exogenous fatty acids to modify embryonic prostaglandin release needs to be examined in the context of supplementing dairy cows with different sources of fats. Synchronisation method altered trophoblast PGE(2) release, highlighting the importance of the hormonal environment in modifying embryonic prostaglandin synthesis and release.


Subject(s)
Cattle/metabolism , Dinoprost/biosynthesis , Dinoprostone/biosynthesis , Dinoprostone/blood , Endometrium/drug effects , Fatty Acids, Omega-3/pharmacology , Trophoblasts/drug effects , Animals , Docosahexaenoic Acids/pharmacology , Eicosapentaenoic Acid/pharmacology , Endometrium/metabolism , Female , Linoleic Acids/pharmacology , Pilot Projects , Pregnancy , Tissue Culture Techniques , Trophoblasts/metabolism
9.
Mol Reprod Dev ; 76(1): 65-74, 2009 Jan.
Article in English | MEDLINE | ID: mdl-18449874

ABSTRACT

The precise mechanism of placentation in the bovine species where a restricted trophoblast invasion occurs to form the synepitheliochorial placenta is not fully understood. This study initially investigated the conceptus-maternal interactions in the peri-attachment period by comparing the proteins present at Days 16 and 18 in uterine luminal fluid (ULF) of pregnant with nonpregnant cows using 2-D gel electrophoresis. Nine protein spots were identified that were present in greater amounts in pregnant compared to nonpregnant ULF: carbonic anhydrase, ezrin, heat shock protein 70, isocitrate dehydrogenase, nucleoside diphosphate kinase, peroxiredoxin 1, purine nucleoside phosphorylase, thioredoxin and triosephosphate isomerase and four proteins that were less abundant in ULF from the gravid compared to the nongravid horns or nonpregnant uteri: cystatin E/M, legumain, retinol-binding protein (RBP) and tissue inhibitor of matrix metalloproteinase 2 (TIMP-2). Successful placentation requires the remodelling of the endometrial surface therefore uterine mRNA and protein expression of legumain, a protease activator, and TIMP-2, a protease inhibitor, was examined in detail during the oestrous cycle and from Days 13 to 31 of pregnancy. Both mRNAs were up-regulated in the endometrium during the luteal phase of the oestrous cycle and during early pregnancy. Although legumain and TIMP-2 mRNA expression levels were similar between uterine horns at the same day of pregnancy, the amount of protein differed between gravid and nongravid horns possibly modulated by interferon-tau or by other factors produced by the conceptus. These events at the conceptus-maternal interface may provide localised control of protease activity necessary for controlling trophoblast invasion of the endometrium.


Subject(s)
Cysteine Endopeptidases/metabolism , Embryo, Mammalian/enzymology , Endometrium/enzymology , Gene Expression Regulation, Enzymologic , Tissue Inhibitor of Metalloproteinase-2/metabolism , Animals , Cattle , Cysteine Endopeptidases/genetics , Electrophoresis, Gel, Two-Dimensional , Female , Pregnancy , RNA, Messenger/genetics , Tissue Inhibitor of Metalloproteinase-2/genetics
10.
Anim Reprod Sci ; 105(3-4): 332-43, 2008 May.
Article in English | MEDLINE | ID: mdl-17482775

ABSTRACT

The temporal relationships among oocyte maturation, gamete transport and fertilisation following the pre-ovulatory luteinsing hormone surge in red deer were established; and secondly, early preimplantation development to the blastocyst stage in relation to the onset of oestrus was determined for red deer. In the first series of observations, oestrus was synchronised in April (N=22), for the fixed time recovery of gametes from 0 to 36 h after the estimated pre-ovulatory LH peak. Matings were observed and the time of the LH peak was determined from the retrospective analysis of blood plasma collected at 3h intervals. Gametes were recovered surgically and the meiotic status of follicular and ovulated oocytes assessed. Spermatozoa were recovered from the oviduct and their motility analysed by videomicroscopy. Nineteen of 22 hinds exhibited a pre-ovulatory LH surge and were observed to mate. Oocyte metaphase I occurred between 11 and 18 h, and metaphase II was completed within the follicle between 20 and 25 h following the pre-ovulatory LH peak. Fertilised ova were recovered from 30 to 36 h in both the ampulla and isthmic portions of the oviduct. Motile spermatozoa were first recovered from the isthmus and the ampulla at 13 and 21 h, respectively, after the LH peak. Hyperactive spermatozoa were observed in both the isthmus and the ampulla flushings but only from the eight hinds that had ovulated. In the second series of observations, 16 mature hinds were synchronised and allocated to groups for embryo collection on days 3, 5 and 7 after oestrus. Eight embryos were recovered; an 8-cell at 90 h, 3 morulae at 137, 138 and 186 h, and 4 blastocysts at 180, 182 and 190 h post-mating. Blastocysts were only recovered from the uterine horns and the mean+/-S.E.M. number of nuclei per blastocyst was 93.5+/-10.0 with a range of 66-114 cells. The results of this study will improve the application of assisted reproductive technologies to red deer as they indicate that oocyte maturation, fertilisation and early embryonic development of the red deer is similar to other domestic ruminants with the exception that the red deer embryo enters the uterus at the blastocyst stage.


Subject(s)
Deer/physiology , Embryonic Development/physiology , Luteinizing Hormone/physiology , Oocytes/physiology , Animals , Blastocyst/physiology , Blastocyst/ultrastructure , Deer/embryology , Estrus/physiology , Estrus Synchronization/physiology , Female , Luteinizing Hormone/blood , Male , Microscopy, Video/veterinary , Oocytes/growth & development , Oocytes/ultrastructure , Pregnancy , Sperm Motility/physiology , Spermatozoa/physiology , Spermatozoa/ultrastructure
11.
Mol Reprod Dev ; 73(9): 1084-93, 2006 Sep.
Article in English | MEDLINE | ID: mdl-16804885

ABSTRACT

In the development of ruminant embryos, the emergence and growth of the allantois is critical for the establishment of the chorioallantoic placenta. The allantoic membrane contributes to all the vasculature that perfuses the placental tissues and the fetal membranes. Using suppressive subtractive hybridization to compare mRNA from Day 13 ovine preimplantation conceptuses (prior to allantoic emergence) with Day 17 allantoic membrane, we identified nine genes whose expression was associated with the emergence of the allantoic sac. Collagen alpha 1 type XII, collagen alpha 2 type I, collagen alpha 2 type V, epsilon 4 beta-globin, osteonectin, and uroplakin were expressed at significantly greater levels in ovine Day 17 allantois compared to Day 13 conceptuses. These genes are associated with the extracellular matrix and most likely are involved in establishing and strengthening the structural integrity of the allantoic sac and in the development of the blood vessels. RalB expression increased with development although at significantly greater levels in the allantois only at Day 19. Hoxa-10 and RhoA showed no differential expression during this period. All these genes showed a similar temporal pattern of expression in bovine conceptuses at equivalent stages of development with significantly greater expression of all these genes, except for Hoxa-10, found in Day 24 allantois compared to Day 14 conceptuses. This suggests that the role they play in allantoic emergence, growth and function is conserved in both ruminant species and that their expression is regulated in a similar manner. The interactions and regulation of this process remains to be fully explained.


Subject(s)
Allantois/metabolism , Cattle/embryology , Cattle/genetics , Gene Expression Regulation, Developmental/genetics , Sheep/embryology , Sheep/genetics , Animals , Female , In Situ Hybridization , Pregnancy , Time Factors
12.
Placenta ; 27(2-3): 307-16, 2006.
Article in English | MEDLINE | ID: mdl-15955560

ABSTRACT

Bovine nuclear transfer pregnancies are characterized by a high incidence of placental abnormalities, notably, increased placentome size and deficiencies in trophoblast cell function and establishment of placental vasculature. Alterations in gene expression during placental growth and development may contribute to the appearance of large placentomes in pregnancies derived from nuclear transfer. The placenta synthesizes a number of cytokines and growth factors, including the transforming growth factor-betas (TGF-betas) that are involved in the establishment, maintenance and/or regulation of pregnancy. All forms of TGF-beta and their receptors are present at the fetal-maternal interface of the bovine placentome, where they are thought to play an important role in regulating growth, differentiation, and function of the placenta. Using real-time RT-PCR, we have examined the expression of TGF-beta1, TGF-beta2, TGF-beta3 and the receptors TGF-betaRI and TGF-betaRII in placentomes of artificially inseminated (AI) and nuclear transfer (NT)-derived bovine pregnancies at days 50, 100 and 150 of gestation. TGF-beta1, TGF-beta2 and TGF-beta3 mRNA expression increased by 2.0-2.8-fold, while TGF-betaRI and TGF-betaRII mRNA expression decreased by 1.7-2.0-fold in NT placentomes compared to AI controls at all gestational ages examined. These findings indicate that NT placentomes may be resistant to the growth suppressive effects of TGF-betas and could contribute to the placental proliferative abnormalities observed in NT-derived placentas. Alternatively, deficiencies in placentation may provide a mechanism whereby TGF-betas are dysregulated in NT pregnancies.


Subject(s)
Activin Receptors, Type I/genetics , Insemination, Artificial , Placenta/metabolism , Receptors, Transforming Growth Factor beta/genetics , Transforming Growth Factor beta/genetics , Animals , Cattle , Female , Gene Expression , Nuclear Transfer Techniques , Pregnancy , Protein Serine-Threonine Kinases , Receptor, Transforming Growth Factor-beta Type I , Receptor, Transforming Growth Factor-beta Type II , Transforming Growth Factor beta1 , Transforming Growth Factor beta2 , Transforming Growth Factor beta3
13.
J Anim Sci ; 82(8): 2329-32, 2004 Aug.
Article in English | MEDLINE | ID: mdl-15318732

ABSTRACT

A mathematical model that describes the recruitment and growth of ovarian follicles was fitted to data on ovulation rate and the measurements of plasma estradiol collected at times during the estrous cycle for individual gilts. The method of least squares was used to obtain estimates of the parameters of the mathematical model. The estimated model parameters were the maximum estradiol production for a follicle, development of each follicle after commitment, and a function describing the initial estradiol production of committed follicles. The estimated parameters for each pig were classified by estrogen receptor (ER) genotype (AA or BB) and analyzed using a multivariate analysis of variance. There were differences between genotypes (P < 0.05) for the parameter that described the initial distribution of individual follicles at recruitment. Gilts with ER genotype BB recruited follicles that varied more in size but had fewer very small follicles, indicating that the ER gene affects the relative estradiol secretion of the follicles at commitment. This analysis is an example of a general approach to genetic studies that uses a mathematical model of the physiology as a statistical basis for estimating gene action.


Subject(s)
Ovarian Follicle/growth & development , Ovulation/physiology , Receptors, Estrogen/genetics , Swine/physiology , Analysis of Variance , Animals , Estradiol/blood , Female , Genotype , Mathematics , Models, Biological , Receptors, Estrogen/metabolism , Swine/genetics
14.
Theriogenology ; 59(2): 687-97, 2003 Jan 15.
Article in English | MEDLINE | ID: mdl-12499012

ABSTRACT

Over the past 20 years the rate of blastocyst development in vitro has improved through the development of sequential defined media, refining the oxygen concentrations during culture and providing substrates to ameliorate free radical accumulation. Despite these advances there has been little progress in improving calving rates after the transfer of in vitro produced embryos. This suggests that the culture conditions have been very effective in enabling those fertilised oocytes to reach the blastocyst stage that otherwise would not occur in vivo. We suggest that the next advance by which the embryo transfer technology gains more acceptance in cattle production will be identifying those cows which are intrinsically superior recipients. This must be coupled to the development of non-invasive assessments of the developmental competence of both the oocyte and the blastocyst. Until these two goals are achieved the ET industry will remain static and unable to overcome the economic loss caused by embryo mortality occurring 7-10 days after transfer.


Subject(s)
Cattle/embryology , Embryo Transfer/veterinary , Abortion, Veterinary , Animals , Cattle/physiology , Embryo, Mammalian/physiology , Embryonic and Fetal Development , Female , Pregnancy , Time Factors
15.
Mol Reprod Dev ; 61(3): 317-26, 2002 Mar.
Article in English | MEDLINE | ID: mdl-11835577

ABSTRACT

In cattle, oocytes obtained from follicles smaller than 3 mm in diameter can undergo maturation in vitro, progressing to MII and undergoing fertilization, but are developmentally incompetent. Cytoplasts were prepared from in vitro matured oocytes aspirated from small (1-3 mm) or large (6-12 mm) follicles and fused to serum starved mural granulosa cells. Following activation, reconstructed embryos were cultured for 7 days and classified G1 to G4, before being processed for nuclei counting or transferred to synchronized recipients. Oocytes from small follicles had lower rates of polar body extrusion (59.6 vs. 69%; 731/1230 vs. 608/857) and fusion (71.4 vs. 78.8%; 360/497 vs. 364/465; P < 0.06). There were no differences in total rate of blastocysts development (60 vs. 59.8%; small vs. large), or any grade classification. A significant interaction was detected between follicle size and embryo grade with G3 embryos from small follicles having a greater cell number. Developmental competence of G1 and G2 embryos did not differ at day 27 (48 vs. 46%; 16/33 vs. 17/37; small vs. large). Although there were no differences in fetal size between the two groups, differences in allantois length (53 vs. 86 mm; small vs. large; P < 0.002) and allantois width (9.5 vs. 13 mm; small vs. large; P < 0.06) were seen. No differences in survival to term (2/13 in each group) were observed. These results indicate that cytoplasts from follicles of 1-3 and 6-12 mm in diameter are equally developmentally competent when used in a nuclear transfer procedure.


Subject(s)
Cloning, Organism/methods , Embryo, Mammalian/cytology , Embryo, Mammalian/embryology , Oocytes/cytology , Oocytes/transplantation , Ovarian Follicle/anatomy & histology , Ovarian Follicle/cytology , Animals , Blastocyst/cytology , Body Weight , Cattle , Cell Nucleus/physiology , Culture Techniques , Cytoplasm/physiology , Cytoplasm/transplantation , Embryo Transfer , Embryonic and Fetal Development , Female , Fertilization in Vitro , Microsatellite Repeats/genetics , Nuclear Transfer Techniques , Oocytes/growth & development , Ovarian Follicle/transplantation , Pregnancy , Ultrasonography, Prenatal
16.
Reproduction ; 122(4): 561-9, 2001 Oct.
Article in English | MEDLINE | ID: mdl-11570963

ABSTRACT

The aim of this study was to examine the function of granulosa cells and hormone concentrations in follicular fluid in bovine ovarian follicles during selection of the first dominant follicle. Ovaries were obtained from beef heifers on days 1-5 after ovulation: follicles > 4 mm in diameter were dissected and follicular fluid and granulosa cells were collected from individual follicles. Oestradiol production by granulosa cells after culture with testosterone was used to determine aromatase activity and responsiveness to gonadotrophins was determined by cAMP production after culture with FSH or LH. Concentrations of oestradiol, progesterone and insulin-like growth factor binding proteins (IGFBPs)-4 and -5 were measured in follicular fluid. Follicles were classified as largest or smaller (days 1 and 2), or dominant or subordinate (days 3-5). Aromatase activity was greater in granulosa cells from the largest follicle than in granulosa cells from smaller follicles on days 1, 3, 4 and 5 (P < 0.05). Responsiveness to LH was not detected in granulosa cells on day 1, but from day 2 to day 5 cells from the largest follicle were significantly more responsive than cells from smaller follicles (P < 0.05). Responsiveness to FSH was detected in granulosa cells from all follicles from day 1 onwards and did not differ between cells from the largest follicle or smaller follicles on any day. Follicular fluid concentrations of oestradiol and the ratio of oestradiol:progesterone were greater and concentrations of IGFBP-4 and -5 were lower in the largest follicle than in smaller follicles from day 2 to day 5 (P < 0.05). In conclusion, selection of the dominant follicle is associated with increased granulosa cell aromatase activity followed by increased cAMP response to LH and follicular fluid oestradiol concentrations, and decreased follicular fluid concentrations of IGFBP-4 and -5 within 2 days after ovulation.


Subject(s)
Aromatase/metabolism , Follicular Phase/metabolism , Gonadotropins, Pituitary/pharmacology , Insulin-Like Growth Factor Binding Proteins/analysis , Ovarian Follicle/metabolism , Animals , Biological Assay , Cattle , Cells, Cultured , Cyclic AMP/biosynthesis , Estradiol/analysis , Estradiol/biosynthesis , Female , Follicle Stimulating Hormone/pharmacology , Follicular Fluid/chemistry , Granulosa Cells/metabolism , Insulin-Like Growth Factor Binding Protein 4/analysis , Insulin-Like Growth Factor Binding Protein 5/analysis , Luteinizing Hormone/pharmacology , Ovarian Follicle/chemistry , Ovarian Follicle/drug effects , Progesterone/analysis , Stimulation, Chemical , Testosterone/pharmacology
17.
Biomaterials ; 22(13): 1779-86, 2001 Jul.
Article in English | MEDLINE | ID: mdl-11396881

ABSTRACT

Polymerization of a tetrafunctional monomer was investigated under a variety of photoinitiation conditions to assess the ability to form thick materials in situ for orthopaedic applications. The major biological concerns include local cell and tissue necrosis due to the polymerization exotherm and low conversions at greater depths due to light attenuation through thick samples. Experimental results indicate that depth of cure and temperature rises are controllable by altering the photoinitiator concentration, initiating light intensity, and type of photoinitiator. For example, no measurable conversion was detected at a 1.0 cm depth when polymerization was initiated with 1.0 wt% DMPA and 100 mW/cm2 ultraviolet light, whereas approximately 40% conversion was obtained when the initiator concentration was lowered to 0.1 wt%. This conversion was further increased to approximately 55% when a photobleaching initiator system was employed. At the highest rate of initiation studied (i.e., 1.0 wt% DMPA irradiated with 100 mW/cm2 ultraviolet light), a maximum temperature of approximately 49 degrees C was reached at the sample surface; however, this temperature dramatically decreased to approximately 33 degrees C when the light intensity was decreased to 25 mW/cm2. Finally, dual initiating systems that synergistically combine the advantages of photoinitiation and thermal initiation were investigated.


Subject(s)
Biocompatible Materials/chemistry , Polymers/chemistry , Photochemistry , Spectrophotometry, Ultraviolet , Temperature
18.
J Reprod Immunol ; 49(1): 1-19, 2001 Jan.
Article in English | MEDLINE | ID: mdl-11137109

ABSTRACT

Investigations of the human Follitropin receptor (hFSHR) have failed to identify the tertiary structure that forms the active hormone-receptor interaction site which is essential to develop an immunocontraceptive based upon the receptor. To identify such a domain of hFSHR, an immunoneutralizing monoclonal antibody (mAb) 106-105 (IgG2b) was generated. Flow cytometry tested whether mAb 106-105 recognized native hFSHR. The epitope of mAb 106-105 was mapped by Western blot and by peptide ELISA. Inhibition of hFSH binding and bioactivity was determined by radioreceptor assay and by cAMP production, respectively. MAb 106-105 bound native hFSHR through an epitope including residues 300-315. MAb 106-105 completely blocked hormone binding to receptor and cAMP production by Y1-R cells expressing hFSHR. These effects were completely reversible by increasing the concentration of hFSH. Coincubation of this antibody with peptide D300-F315 blocked antibody activity. These data demonstrate that a discrete linear hFSHR epitope is a target for interference with hormone activity. These results further demonstrate that antibody binding to the extracellular domain (ECD) of hFSHR and subsequent bioactivation can be modulated through a domain specific hindrance, offering a reversible immunoneutralizing target.


Subject(s)
Receptors, FSH/immunology , Animals , Antibodies, Monoclonal/biosynthesis , Antibodies, Monoclonal/immunology , CHO Cells , Cricetinae , Cyclic AMP/metabolism , Humans , Iodine Radioisotopes , Isotope Labeling , Mice , Neutralization Tests , Peptides/immunology , Tumor Cells, Cultured
19.
J Appl Physiol (1985) ; 89(3): 1233-8, 2000 Sep.
Article in English | MEDLINE | ID: mdl-10956374

ABSTRACT

To determine how rapidly pulmonary capillaries recruit after sudden changes in blood flow, we used an isolated canine lung lobe perfused by two pumps running in parallel. When one pump was turned off, flow was rapidly halved; when it was turned on again, flow immediately doubled. We recorded pulmonary capillary recruitment in subpleural alveoli using videomicroscopy to measure how rapidly the capillaries reached a new steady state after these step changes in blood flow. When flow was doubled, capillary recruitment reached steady state in <4 s. When flow was halved, steady state was reached in approximately 8 s. We conclude that the pulmonary microcirculation responds rapidly to step changes in flow, even in the capillaries that are most distant from the hilum.


Subject(s)
Blood Flow Velocity/physiology , Pulmonary Circulation/physiology , Weightlessness , Animals , Capillaries/physiology , Dogs , Homeostasis , In Vitro Techniques , Male , Microscopy, Video , Perfusion , Pulmonary Alveoli/blood supply , Time Factors
20.
Mol Cell Endocrinol ; 160(1-2): 203-17, 2000 Feb 25.
Article in English | MEDLINE | ID: mdl-10715554

ABSTRACT

Variant splicing of gonadotropin receptor mRNA commonly occurs, however expression of receptor protein variants and their trafficking has yet to be studied in detail. To determine receptor variant trafficking and intracellular processing in mammalian cells, the intracellular fate of intentionally truncated variants of human follicle stimulating hormone receptor (hFSH-R) expressed in CHO cells was examined. Monoclonal antibodies (mAbs) were made against the hFSH-R's extracellular domain (ECD) expressed in insect cells. Four mAbs 106.156, 106.290, 106.318, and 106.263 were chosen as probes. Epitope mapping using synthetic peptides, and truncated hFSH-R variants revealed that mAb 106.156 bound to ECD residues 183-220, while mAbs 106.318, 106.290, 106.263 bound ECD residues 300-331. Immunofluorescence microscopy showed that mAbs 106.318 and 106.156 stained the surface of fixed, intact CHO cells expressing wild type hFSH-R. However, following cell permeabilization all four antibodies stained hFSH-R in Golgi and endoplasmic reticulum. Permeabilized cells expressing truncated variants ECD213 and ECD254 showed staining accumulated in the endoplasmic reticulum/nuclear envelope continuum. ECD335/His was found to accumulate in extended endoplasmic reticulum (ER). The ER location of ECD335/His was confirmed by double labeling experiments with concanavalin A and ECD mAb. Glycosidase digestion followed by Western blot analysis show ECD213 and ECD335/His to be glycosylated, but not ECD254. Both glycosylated truncated hFSH-R variants were sensitive to peptide-N-glycanase F and endoglycosidase H but insensitive to neuraminidase indicating that these variants possess high mannose type oligosaccharides. Thus truncated hFSH-R variants do not reach the medial or trans Golgi where high mannose oligosaccharides are trimmed and sialic acid is added. These data suggest that the conformation the ECD of the wild type receptor is different from the ECD alone expressed in the endoplasmic reticulum. This information suggests that the ECD serves two distinct roles; the first is to bind FSH and the other is likely to contact the endodomain of the receptor, which presumably leads to activation of the endodomain for signal transduction.


Subject(s)
Receptors, FSH/chemistry , Receptors, FSH/genetics , Amino Acid Sequence , Animals , Antibodies, Monoclonal , CHO Cells , Cricetinae , Endoplasmic Reticulum/metabolism , Epitope Mapping , Fluorescent Antibody Technique, Indirect , Genetic Variation , Humans , Mice , Molecular Sequence Data , Protein Conformation , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism , Protein Processing, Post-Translational , Protein Structure, Tertiary/genetics , Receptors, FSH/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Sequence Deletion
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