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PubMed1999

Human granulosa cells express integrin alpha2 and collagen type IV: possible involvement of collagen type IV in granulosa cell luteinization.

Previously, it has been shown that integrin alpha6beta1 expressed on human granulosa cells regulates luteinization in co-operation with its ligand, laminin. In this study, integrin alpha2 was immunohistochemically demonstrated to be expressed on granulosa and large luteal cells. It was also detected on luteinizing theca interna cells after ovulation. Immunoreactive collagen type IV, which is one of the ligands for integrin alpha2beta1, was detected around granulosa cells in the pre-ovulatory follicles and its expression was rapidly increased during ovulation. By flow cytometry, collagen type IV was detected on the cell surface of luteinizing granulosa cells isolated from pre-ovulatory follicles, confirming the physiological interaction between granulosa cells and collagen type IV. Collagen type IV in follicular fluid was positively related with progesterone concentration. In 4-day cultures of granulosa cells, collagen type IV in the media was significantly increased by human chorionic gonadotrophin (HCG). The progesterone production was significantly attenuated when granulosa cells were cultured on collagen type IV-coated dishes, suggesting that collagen type IV suppresses granulosa cell luteinization. These findings show that collagen type IV, a ligand for integrin alpha2beta1, is rapidly produced around luteinizing granulosa cells during ovulation, probably under the control of luteinizing hormone (LH) and suggest that collagen type IV is a new parameter and/or regulator of granulosa cell luteinization in the periovulatory phases.

Antigens, CD↗
PubMed1988

Prediction of ovulation using a dipstick sensitive to urinary luteinising hormone.

Accurate prediction of ovulation is essential for patients receiving donor insemination (AID). In 9 patients, detection of the urinary luteinising hormone (LH) surge by monoclonal antibody immunospecific dipstick tests (Ovustick) for both home and laboratory use was assessed with simultaneous quantitative estimations of plasma and urinary LH. In all patients the Ovustick accurately detected within 24 hours the beginning of the plasma LH surge. This test may prove to be a useful method for ovulation prediction but further evaluation is required.

Adult↗
PubMed1996

Measurement of circulating inhibin forms during the establishment of pregnancy.

This study investigated the forms of inhibin released into the circulation 1) in very early pregnancy, 2) after stimulation of the corpus luteum by exogenous hCG, and 3) in abnormal and failing human pregnancy. Samples were assayed by enzyme-linked immunosorbent assays for inhibin A, inhibin B, and inhibin pro-alphaC-related immunoreactivity (pro-alphaC-RI). The concentration of inhibin A rose steadily during the conception luteal phase to an initial peak 12 days after ovulation (104 +/- 23 pg/mL), then rose rapidly to a further peak 43 days after ovulation 424 +/- 6 pg/mL). The concentration of pro-alphaC-RI exhibited a much larger peak on day 15 after ovulation (1423 +/- 361 pg/mL), but fell thereafter. The concentration of inhibin B was low after ovulation and subsequently barely detectable in pregnancy. hCG treatment resulted in a significant rise in the concentrations of inhibin A and pro-alphaC-RI, but had no effect on the inhibin B concentration. The pro-alphaC-RI concentration was a better indicator of continuing pregnancy viability than either hCG or inhibin A. Early trophoblast secretes proportionately more bioactive inhibin than the corpus luteum. The corpus luteum and trophoblast do not secrete inhibin B into the circulation. These data support the concept of different physiological roles for different inhibin forms.

Abortion, Spontaneous↗
PubMed1993

A differentiation-related molecule on the cell surface of human granulosa cells.

We raised a monoclonal antibody (OG-1) specifically reactive to human ovarian granulosa cells. Indirect immunofluorescence staining of freshly isolated granulosa cells from preovulatory follicles showed the OG-1 antigen on the cell surface of granulosa cells. The antigen was purified from granulosa cells by immunoaffinity chromatography. Under reducing or nonreducing conditions, the sodium dodecyl sulfate-polyacrylamide gel electrophoresis profile of the purified antigen showed two specific protein bands, corresponding to mol wt of 125 and 145 kilodaltons. Immunohistologically, the OG-1 antigen on granulosa cells of primary follicles seemed to increase during follicular development. After ovulation, the antigen was detected on granulosa cells during corpus luteum formation. The antigen expression began to decrease on corpus luteum days 10-12 and disappeared on corpus luteum days 13-14. Luteal cells of pregnancy did not express the OG-1 antigen. Granulosa cells in atretic follicles expressed the OG-1 antigen. On the contrary, thecal or small luteal cells did not express it throughout the menstrual cycle. These results indicate that OG-1 antigen is a differentiation-related surface antigen of human granulosa cells.

Adult↗
PubMed1977

Laparoscopic exposure and sequential observation of the ovary of the cycling bitch.

A method was developed to alter surgically the bursa of the bitch to allow sequential observation of the ovary by means of laparoscopy. Adult anesthetized female dogs of three breeds underwent laparoscopic examinations in which each natural bursa slit was lengthened 1.0-1.5 cm using laparoscopic hook scissors in conjunction with electrocautery. The ovary could be effectively and repeatedly exposed for laparoscopic viewing through the lengthened bursa slit. Neither laparoscopic surgery nor frequent routine laparoscopic examinations of the reproductive organs had an effect on subsequent ovarian activity. All dogs subjected to the laparoscopic ovarian exposure technique exhibited estrus behavior, follicle development, ovulation and corpus luteum formation. Preovulatory follicular development was detected as early as 11 days before ovulation with the most rapid follicular maturation occurring two to three days before ovulation. Follicular collapse and extensive hemorrhage at the rupture site during ovulation did not occur. Release of the cumulus mass from a vesicular follicle was observed in one bitch. The cumulative mass adhered to the follicular wall for an indefinite time following ovulation. The structural size of the bitch corpus luteum was maximal four to eight days following ovulation. Corpora lutea remained visible on the ovarian surface for 16 weeks. No differences in cyclic ovarian morphology were observed among the three breeds of dogs studied.

Animals↗
PubMed2004

Stage-specific formation of the equine blastocyst capsule is instrumental to hatching and to embryonic survival in vivo.

Early embryonic development in the horse is characterized by the formation of an unusual acellular glycoprotein "capsule" between the trophectoderm and the overlying zona pellucida. This structure is first detected between days 6 and 7 after ovulation and completely envelops the spherical conceptus until as late as day 23 of gestation. In the present study, a micromanipulator was used to remove the capsule from 15 embryos on day 6-7 after ovulation. None of these denuded embryos developed into ultrasonographically detectable pregnancies after surgical transfer into recipient mares whereas four of six control embryos handled and transferred similarly but without capsule removal developed normally, thereby demonstrating clearly a role for the capsule in embryonic survival. In addition, observation of the embryonic investments after embryo collection and during micromanipulation led to the hypothesis that hatching of the horse embryo from its zona pellucida is assisted by the force of the expanding capsule, which causes the attenuating zona to literally burst open.

Animals↗
PubMed1987

Evaluation of "Determine; The OvuTest" as a device for identifying optimal time for conception.

The device known as "Determine; The OvuTest" (Ventco Medical Trading Company, Montreal, Quebec, Canada) was appraised for its stated function of determining the optimal time for human conception by detecting the presence of the protein, uteroglobin (UTG), in cervical mucus. Twenty volunteers used the device daily through three cycles in which ovulation was indicated by rise in basal body temperature and/or elevated levels of luteinizing hormone. No correlation was found between the instrument readings and time of ovulation. When tested in vitro, it did not distinguish the presence of UTG. When vaginal flushings, containing cervical mucus, were analyzed for UTG by double immunodiffusion, no lines of precipitation were observed. We conclude that "Determine; The OvuTest" does not identify the approximate time of ovulation and therefore is not a reliable aid to conception. It does not detect UTG, and periovulation cervical mucus does not contain UTG.

Adult↗
PubMed2004

Plasma concentrations of inhibin A in cattle with follicular cysts: relationships with turnover of follicular waves and plasma levels of gonadotropins and steroid hormones.

We investigated the profiles of circulating levels of inhibin A and total inhibin in beef cows with follicular cysts in relation to the patterns of follicular development and circulating gonadotropins and steroid hormones. Turnover of follicular waves was monitored in five cows every 2 days for 70 days from 10 days after detection of estrus without ovulation. The mean interwave intervals were 19.6 +/- 1.0 days (n = 18 waves with cysts from the five cows). Circulating levels of inhibin A were approximately 170 pg/ml before emergence of follicular waves with cysts and increased (P < 0.05) concomitantly with follicle emergence. High concentrations of inhibin A (greater than 300 pg/ml) were noted for 7 days during the growth phase of cystic follicles, but inhibin A levels decreased gradually when development of the cysts reached a plateau. This profile of inhibin A was similar to those of total inhibin and estradiol, but was inversely related to the changes in plasma FSH concentrations. LH pulse frequency and mean concentrations of LH in cows with cysts were higher than those observed in the luteal phase of normal cyclic cows. These results indicate that the capacity to secrete inhibin, as well as estradiol, is maintained in cystic follicles, the growth of which is extended by LH secretion at levels greater than those seen in the normal luteal phase. Inhibin A plays an important role in the extension of interwave intervals by suppressing recruitment of a new cohort of follicles.

Analysis of Variance↗
PubMed1983

Heteroimmunization of squirrel monkeys (Saimiri sciureus) with a purified porcine zona antigen (PPZA): immune response and biologic activity of antiserum.

The potential for utilization as a contraceptive vaccine of a 60,000 Mr glycoprotein component, purified porcine zona antigen (PPZA), isolated from porcine zonae, was investigated in the squirrel monkey. Immunization resulted in production and maintenance of high antibody titers for at least 1 year. Comparable immune profiles were obtained using either monkey or pig zonae in assay systems, but dose-dependent variations in immune response were not observed. In situ antibody binding to monkey zonae was detected, but significantly fewer ovulated eggs were obtained from immunized monkeys than from controls. Exposure of antibody-pretreated pig, monkey, and human zonae to homologous sperm resulted in total inhibition of sperm attachment for the respective species. Thus, the contraceptive potential of PPZA antibodies in these species is demonstrated.

Animals↗
PubMed2003

Effect of treatment with progesterone and oestradiol when starting treatment with an intravaginal progesterone releasing insert on ovarian follicular development and hormonal concentrations in Holstein cows.

Ovarian follicular development and concentrations of gonadotrophin and steroid hormones were studied in non-lactating Holstein cows following administration of progesterone (P(4)) or oestradiol benzoate (ODB) at the start of treatment with an intravaginal progesterone releasing insert (IVP(4)) in a 2 by 2 factorial experiment. Cows were treated at random stages of the oestrous cycle with an IVP(4) device (Day 0) and either no other treatment (n=8), 200 mg of P(4) IM (n=9), 2.0 mg of ODB IM (n=8) or both P(4) and ODB (n=9). Seven days later devices were removed and PGF(2alpha) was administered. Twenty-four hours later 1.0mg of ODB was administered IM. Oestrus was detected in 97.1% and ovulation in 64.7% (effect of treatment, P=0.41) of cows within 96 h of removing inserts. In the cows that ovulated, day of emergence of the ovulatory follicle was delayed (P<0.01) and more precise (P<0.05) in cows treated with ODB compared to the cows treated with P(4). Interval from wave emergence to ovulation and the diameter of the ovulatory follicle was less (P<0.05) in cows treated with ODB compared to cows treated with P(4). Combined treatment with P(4) and ODB at the time of starting treatment with an IVP(4) device did not significantly change the pattern of ovarian follicular development compared to treatment with ODB alone. Concentrations of LH and FSH in plasma were less in cows treated with ODB between Days 0 and 4 (P<0.05) while treatment with P(4) increased concentrations of FSH in plasma between Days 0 and 4 (P<0.05). When anovulatory cows were compared to ovulatory cows, diameters of follicles (P<0.001) and growth rate of follicles (P<0.01) were less in anovulatory cows between Days 7 and 9, while concentrations of FSH in plasma were greater (P<0.01), concentrations of LH similar (P>0.90) and concentrations of oestradiol were less (P=0.01) in the anovulatory cows between Days 4 and 10. Our findings support a hypothesis that ovarian follicular development following administration of P(4) or ODB at the start of treatment with an IVP(4) device differs. Anovulatory oestrus may have been associated with reduced maturity and/or later emergence of ovarian follicles.

Administration, Intravaginal↗
PubMed2003

Effects of suckling frequency and breed on productive performance, follicular dynamics and postpartum interval in beef cows.

One experiment was conducted to determine the influence of suckling frequency (restricted suckling (RS) versus ad libitum suckling (AS)) and breed (Brown Swiss versus Pirenaica) on productive performance, follicular dynamics and postpartum cyclicity resumption in beef cows. Twenty-four spring-calving cows (12 Brown Swiss and 12 Pirenaica) were used. After calving cows were assigned, within breed, to the two suckling frequencies (RS versus AS). During the postpartum period, cows were fed 100% of the daily energy estimated requirements. The study was conducted from calving to either day 90 postpartum or to detection of the second ovulation whenever it came later. The pattern of follicular development was monitored daily by transrectal ultrasonography, and blood samples were collected three times a week to analyse progesterone concentrations by RIA. Body condition score at calving was 2.5 (from 1 to 5). Productive parameters were not significantly affected by suckling frequency. Pirenaica cows gained more weight (P<0.001), while Brown Swiss cows had higher milk yield (P<0.001) and thus their calves grew faster (P<0.05). In the present moderate conditions of nutrition, either suckling frequency or breed did not affect follicular dynamics (on average there were 6.5 follicles in emergence, growth rate of dominant follicle (DF) was 1.3mm per day and maximum diameter of dominant follicle was 13.9mm). There were gradual increases in maximum diameter and growth rate of dominant follicle related to the proximity of the ovulatory waves, and they were defined by third order polynomial equations. Postpartum anoestrous interval (PPI) was markedly affected by the interaction between suckling frequency and breed (P<0.001): ad libitum suckling prolonged significantly the interval from calving to first ovulation in Brown Swiss cows (31.8 day versus 82.7 day, in RS and AS groups, respectively, P<0.05), but not in Pirenaica cows (43.5 day versus 32.6 day, in RS and AS groups, respectively, not statistically significant). It is concluded that in moderate pre- and postpartum nutritional conditions Brown Swiss cows were more susceptible to the suppressor effect of ad libitum suckling on postpartum anoestrous interval than Pirenaica cows, although this breed difference did not seem to be mediated by differences in energy balance or in ovarian follicular dynamics.

Animal Nutritional Physiological Phenomena↗
PubMed2005

Intermittent pollen-tube growth in pistils of alders (Alnus).

In alders, where fertilization occurs approximately 8 weeks after pollination, the pollen tube (male gametophyte) grows intermittently in four steps in close association with the development of the ovary and its ovules. Pollen tubes stop growing in the style, at the ovarian locule, and at the chalaza (ovule), before reaching an embryo sac for fertilization. At the stage when the ovary develops an ovule primordium in each of the two locules, many pollen tubes germinate on the stigma, and a few of them reach the style, where they remain for approximately 7 weeks. Thereafter, a single tube resumes growing; with a short stop in the upper space of the ovarian locule, it reaches the older of the two ovules when it has developed a two-nucleate embryo sac. Except in the last step, where the tube grows from the chalaza to an embryo sac (female gametophyte), an eight-nucleate mature embryo sac is not necessary for pollen-tube guidance in the pistil. Although the intermittent pollen-tube growth appears to play an important role in the selection of a single pollen tube from many and one ovule from two, its detection provides insight into the study of the mechanism of pollen-tube guidance.

Alnus↗
PubMed2003

Mapping quantitative trait loci for bean traits and ovule number in Theobroma cacao L.

Quantitative trait loci (QTL) mapping for bean traits and the number of ovules per ovary was carried out in cocoa (Theobroma cacao L.) using three test-cross progenies derived from crosses between a lower Amazon Forastero male parent (Catongo) and three female parents: one upper Amazon Forastero (IMC78) and two Trinitario (DR1 and S52). RFLP (restriction fragment length polymorphism), microsatellite, and AFLP (amplified fragment lengthpolymorphism) markers were used for mapping. Between one and six QTL for bean traits (length, weight, and shape index) and one and four QTL for the number of ovules per ovary were detected using composite interval mapping (CIM). Individual QTL explained between 5 and 24% of the phenotypic variation. QTL clusters were identified on several chromosomes, but particularly on chromosome 4. QTL related to bean traits were detected in the same region in both Trinitario parents and in a close region in the upper Amazon Forastero parent. In reference to a previous diversity study where alleles specific to Criollo and Forastero genotypes were identified, it was possible to speculate on the putative origin (Criollo or Forastero) of favorable QTL alleles segregating in both Trinitario studied.

Cacao↗
PubMed1991

Effect of mating behavior on luteinizing hormone-releasing hormone release in female rabbits as monitored with push-pull cannulae.

To examine the effect of mating behavior on luteinizing hormone-releasing hormone (LHRH) release, intact New Zealand female rabbits were implanted with push-pull cannulae (PPC) aimed at the tuberal region of the hypothalamus and perfused with modified Krebs-Ringer phosphate medium at 11-13 microliters/min. In the mating experiments, does (n = 10) were initially perfused for a control period of 60-170 min followed by a mating period (100-160 min) which included the introduction of the male rabbit for an average time period of 30 min. Two groups of LHRH release patterns were observed: positive and negative responders. In the positive LHRH responders (n = 5), a clear rapid increase in LHRH release following mounting by the male occurred with a significant increase in the mean LHRH release (1.83 +/- 0.33 to 3.27 +/- 0.80 pg/10 min, p less than 0.040), in the mean LHRH amplitude (1.97 +/- 0.46 to 4.33 +/- 1.29 pg, p less than 0.022) and in the amplitude of the largest LHRH pulse (2.13 +/- 0.43 to 7.58 +/- 3.65 pg, p less than 0.022). In the negative LHRH responders (n = 5), no changes in LHRH release were detected although all rabbits ovulated, with some becoming pregnant. It appears from histological analysis that the difference between these two patterns of responses following mating are due to different cannula placements. In the positive responders, the tip of the PPC was localized in the tuberal region whereas in the negative responders, the placements were more dorsal and, in some cases, anterior.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2006

Gelatinases, endonuclease and Vascular Endothelial Growth Factor during development and regression of swine luteal tissue.

BACKGROUND: The development and regression of corpus luteum (CL) is characterized by an intense angiogenesis and angioregression accompanied by luteal tissue and extracellular matrix (ECM) remodelling. Vascular Endothelial Growth Factor (VEGF) is the main regulator of angiogenesis, promoting endothelial cell mitosis and differentiation. After the formation of neovascular tubes, the remodelling of ECM is essential for the correct development of CL, particularly by the action of specific class of proteolytic enzymes known as matrix metalloproteinases (MMPs). During luteal regression, characterized by an apoptotic process and successively by an intense ECM and luteal degradation, the activation of Ca++/Mg++-dependent endonucleases and MMPs activity are required. The levels of expression and activity of VEGF, MMP-2 and -9, and Ca++/Mg++-dependent endonucleases throughout the oestrous cycle and at pregnancy were analyzed. RESULTS: Different patterns of VEGF, MMPs and Ca++/Mg++-dependent endonuclease were observed in swine CL during different luteal phases and at pregnancy. Immediately after ovulation, the highest levels of VEGF mRNA/protein and MMP-9 activity were detected. On days 5-14 after ovulation, VEGF expression and MMP-2 and -9 activities are at basal levels, while Ca++/Mg++-dependent endonuclease levels increased significantly in relation to day 1. Only at luteolysis (day 17), Ca++/Mg++-dependent endonuclease and MMP-2 spontaneous activity increased significantly. At pregnancy, high levels of MMP-9 and VEGF were observed. CONCLUSION: Our findings, obtained from a precisely controlled in vivo model of CL development and regression, allow us to determine relationships among VEGF, MMPs and endonucleases during angiogenesis and angioregression. Thus, CL provides a very interesting model for studying factors involved in vascular remodelling.

Aging↗
PubMed1989

The nature of embryo reduction in mares with twin conceptuses: deprivation hypothesis.

Ultrasonography was used to determine whether there is embryo reduction in mares with unilaterally fixed twins when a major portion of the vascularized area of the wall of one of the embryonic vesicles is in apposition with the wall of the adjacent vesicle, rather than with the endometrium (deprivation hypothesis). In addition, the effect of ovulatory pattern (synchronous and asynchronous) on the incidence of embryo reduction was studied. Twin vesicles were ultrasonically detected on days 11 to 15 (ovulation = day 0) and were examined daily until there was embryo reduction or until day 40. In 31 mares with twin embryonic vesicles, unilateral fixation (71%) was more frequent (P less than 0.05) than was bilateral fixation (29%). In 28 mares with known ovulatory patterns, synchronous ovulations did not affect the type of fixation (9/17 unilateral and 8/17 bilateral); however, for asynchronous ovulators the frequency of unilateral fixation (10/11) was greater (P less than 0.01) than the frequency of bilateral fixation (1/11). The incidence of embryo reduction was greater (P less than 0.01) for unilateral fixation (14/19) than for bilateral fixation (0/9) and was greater (P less than 0.05) for asynchronous ovulators (9/11) than for synchronous ovulators (5/17). In mares with embryo reduction, the reduction was complete before detection of both embryo propers (early reduction) in 10/14 and after detection of both embryo propers (late reduction) in 4/14.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1998

Progesterone determination in equine plasma using different immunoassays.

Several assay systems (3H radioimmunoassay (RIA) with and without extraction; microplate enzyme-linked immunoassay (ELISA); qualitative ELISA (tube test)] were used to measure plasma progesterone concentration in mare plasma. The direct RIA showed a close correlation (R = 0.94) with the extraction RIA. The direct RIA and the microplate ELISA were compared in two different studies. In the first study 1155 samples of postpartum mares were used for progesterone determination with both assays. The ELISA resulted in more elevated values both in oestrus and dioestrus (0.19+/-0.3 and 2.44+/-3.62 nmol/l for oestrus, n = 436, and 8.94+/-4.29 and 27.88+/-18.34 nmol/l for dioestrus, N = 719, for the RIA and ELISA, respectively, R = 0.71). The evaluation of individual progesterone profiles has revealed that the microplate ELISA detects the time of ovulation at the same time as it is determined by the RIA and clinical examination. The sensitivity and specificity were calculated for different progesterone threshold values. In the second study including 7 non-pregnant, cycling mares the progesterone concentration of 240 samples was determined by both assays. Basal values (Day 0) obtained with the ELISA were higher (1.57 nmol/l) than those of the RIA (0.2 nmol/l). Both curves reached the same maximum concentration (12.11 and 12.45 nmol/l) 5 days after ovulation. The correlation between the RIA and ELISA values was high (R = 0.90). The tube test was compared to the microplate ELISA as reference using 576 plasma samples of 34 non-pregnant, non-cycling mares included in an ovulation induction study. Of these samples 118 had higher and 458 had lower values than 3.18 nmol/l. In most cases the tube test was in complete agreement with the microplate ELISA. The sensitivity, specificity, + predictive and - predictive values for the tube test were 79.7%, 95.4%, 81.7% and 94.8%, respectively.

Animals↗
PubMed1994

[Clinical significance of detection time and site of early gestational sac by ultrasonography].

Ultrasonography was used to determine not only the shape but also the appearance time and site of GS before the formation of the embryo. These factors were analysed for their clinical significance in association with early GS. The following results were obtained. 1. GS with diameters of 2 < or = approximately < or = 4 mm (abr. EGS) were found at 17-25 postovulatory days (gestational age: GA 4w2d-5w3d) in the ovulation presumed group. EGS was detected at GA 4w2d-7w5d in the normal menstrual group and 17.6% of cases were detected at GA 6w or later. 2. 95.3% of EGS were found in the upper 2/3 of the uterine cavity, while only 4.6% were found in the lower 1/3. This difference in incidence was significant. When EGS was found in the lower uterine segment at GA 6w or later, the abortion rate was very high. 3. In cases where gestation continued to term, there was no difference between the ovulation presumed and normal menstrual groups in terms of various fetal conditions. When normal menstrual group cases where EGS was detected at GA 6w or later were compared with the members of the ovulation presumed group, gestation duration was prolonged in the former. From the above mentioned findings, it can be concluded that even in the early stage of gestation, there can be individual growth differences and that in cases with normal menstrual cycles, gestation can take place much later than the expected date of ovulation, and further that the diagnosis of intrauterine gestation should be postponed until the end of GA 6w. Many blighted ova are detected in cases where GS is found at GA 6w or later, but when gestation is carried to term, no fetal growth abnormalities are found. The ideal nidation site is the endometrium near the fundus and when nidation takes place in the lower segment, the incidence of abortion or placenta previa is very high.

Abortion, Spontaneous↗