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1.
Fertil Steril ; 54(3): 428-33, 1990 Sep.
Article in English | MEDLINE | ID: mdl-2118859

ABSTRACT

Insulin-like growth factor I (IGF-I) levels were measured in both serum and fluid of preovulatory follicles (n = 156) in 43 women undergoing in vitro fertilization (IVF). The mean IGF-I level in follicular fluid (FF) was significantly lower than in serum (0.52 +/- 0.02 IU/L versus 0.66 +/- 0.23 IU/L), and FF levels were significantly correlated with individual serum IGF-I levels as well as with follicular size and FF volume but not with oocyte maturity, granulosa cell appearance, or IVF. This suggests that FF IGF-I levels cannot serve as a clinical indicator for the degree of oocyte/granulosa cell differentiation or a predictor for IVF. Serum IGF-I levels were inversely correlated with the number of human menopausal gonadotropin ampules administered during treatment, suggesting that IGF-I might enhance ovarian gonadotropic stimulation.


Subject(s)
Follicular Fluid/analysis , Follicular Phase , Insulin-Like Growth Factor I/analysis , Ovarian Follicle/analysis , Somatomedins/analysis , Estradiol/blood , Female , Fertility , Fertilization in Vitro , Follicular Fluid/physiology , Humans , Menotropins/therapeutic use , Ovarian Follicle/cytology
2.
J Clin Endocrinol Metab ; 71(2): 492-6, 1990 Aug.
Article in English | MEDLINE | ID: mdl-2380343

ABSTRACT

The concentrations of melatonin in 112 preovulatory follicular fluid (FF) samples obtained from 60 women undergoing in vitro fertilization and 27 patients at laparotomy during a spontaneous cycle were measured by RIA and compared with those in peripheral serum. The circadian and seasonal variations in FF melatonin were also analyzed. The FF melatonin concentrations in stimulated (mean +/- SEM, 61.9 +/- 6.4 pmol/L) and spontaneous cycles (98.1 +/- 8.9 pmol/L) were significantly higher (P less than 0.005) than those in peripheral serum (25.4 +/- 1.2 and 38.6 +/- 1.8 pmol/L, respectively), and in the stimulated cycles there was a positive correlation between them. The FF melatonin concentration in the morning (58.9 +/- 3.8 pmol/L) was significantly higher (P less than 0.005) than that in the daytime (23.2 +/- 0.8 pmol/L), but the morning concentrations did not differ between the light and the dark seasons of the year, whereas the daytime values were higher (P less than 0.005) during the dark season (27.1 +/- 2.1 pmol/L) than during the light season (21.1 +/- 2.1 pmol/L). The FF melatonin concentration did not correlate with follicular volume, and FF and serum melatonin concentrations showed no significant correlation with the serum concentrations of estradiol, progesterone, testosterone, or PRL. There were also no differences between FF melatonin concentrations in aspirates with or without an ovum. In summary, significant circadian and circannual variations in high FF melatonin concentrations were found, which suggest that melatonin could potentially interfere with the regulation of reproduction in humans at the follicular level.


Subject(s)
Circadian Rhythm , Melatonin/analysis , Ovarian Follicle/analysis , Periodicity , Chromatography, High Pressure Liquid , Embryo Transfer , Female , Fertilization in Vitro , Humans , Melatonin/blood , Seasons
3.
4.
Cell Tissue Res ; 261(2): 367-73, 1990 Aug.
Article in English | MEDLINE | ID: mdl-2205393

ABSTRACT

The present study examined the presence and cellular distribution of angiotensinogen, the precursor to the angiotensin peptides, in the ovary of the normal cycling rat by immunocytochemistry. Angiotensinogen staining was present in the granulosa cells of maturing follicles and to a lesser extent in those undergoing atresia. Staining was not seen in the granulosa cells of primordial or early primary follicles. In maturing follicles intense staining for angiotensinogen was confined to the antral cell layers, cells of the cumulus oophorus and in the follicular fluid. Strong immunostaining was also seen in the germinal epithelium covering the ovary. Lighter angiotensinogen staining was observed in some parts of the cortical and medullary stroma and occasionally in corpora lutea. No variation in the intensity or pattern of angiotensinogen staining was observed throughout the estrous cycle. Comparison of the distribution of angiotensinogen with the previously described localization of renin, AII, angiotensin converting enzyme and AII receptors, suggests that there are a number of intra-ovarian sites at which AII could be produced.


Subject(s)
Angiotensinogen/analysis , Ovary/analysis , Angiotensin II/biosynthesis , Animals , Epithelium/analysis , Estrus , Female , Granulosa Cells/analysis , Immunohistochemistry , Ovarian Follicle/analysis , Ovary/ultrastructure , Rats , Rats, Inbred Strains , Renin/metabolism
5.
Fertil Steril ; 53(4): 627-31, 1990 Apr.
Article in English | MEDLINE | ID: mdl-2108057

ABSTRACT

Levels of immunoreactive luteinizing hormone (LH), bioactive LH, and testosterone (T) were determined in 52 women receiving human menopausal gonadotropins (hMG). In 26 women receiving leuprolide acetate (LA) preceding hMG, there was a significant suppression of immunoreactive LH and bioactive LH. The characteristic increase in serum levels of bioactive LH and T were absent. Follicular fluid estradiol and T concentrations, and serum progesterone were not different. The lower circulating levels of T may reflect reduced LH-stimulated androgen accumulation in smaller nonaspirated follicles and may account for the enhanced follicle recruitment observed during LA. The lack of premature luteinization despite marked rises of bioactive LH in the absence of LA is consistent with normal events during the menstrual cycle and was due to the early termination of hMG stimulation.


Subject(s)
Gonadotropin-Releasing Hormone/analogs & derivatives , Hormones/therapeutic use , Luteinizing Hormone/blood , Reproductive Techniques , Testosterone/blood , Chorionic Gonadotropin/therapeutic use , Estradiol/analysis , Estradiol/blood , Female , Fertilization in Vitro , Gonadotropin-Releasing Hormone/therapeutic use , Humans , Leuprolide , Menotropins/therapeutic use , Oocytes/cytology , Ovarian Follicle/analysis , Progesterone/blood , Testosterone/analysis
6.
Br Poult Sci ; 31(1): 147-54, 1990 Mar.
Article in English | MEDLINE | ID: mdl-2354371

ABSTRACT

1. Eighteen Warren SSL hens of 71 weeks of age were forced-moulted by ad libitum feeding of a high-zinc diet (10,000 ppm zinc for 2 days followed by 5,000 ppm zinc-supplement diet for 4 days). From the start of the treatment, eggs were collected and 3 hens were slaughtered on days 0, 2, 3, 4, 5 and 6 of the study. 2. Zinc analyses were carried out on the different components of the eggs and on liver, pancreas, kidney, different yolky follicles of the ovary and various segments of the oviduct. 3. Seven-, six- and threefold increases in zinc concentration were found in pancreas, liver and kidney, respectively. 4. The shell gland and isthmus, but not the magnum, also showed slight but significant increases in Zn content. 5. Zinc accumulation was also high and almost identical in ovarian follicles F1 to F4 but slightly less in F5 and F6 follicles. 6. In the egg, a significant increase in zinc concentration was only observed in the yolk.


Subject(s)
Chickens/metabolism , Egg Yolk/analysis , Ovarian Follicle/metabolism , Zinc/pharmacokinetics , Animals , Female , Kidney/analysis , Kidney/metabolism , Liver/analysis , Liver/metabolism , Ovarian Follicle/analysis , Oviducts/analysis , Oviducts/metabolism , Pancreas/analysis , Pancreas/metabolism , Tissue Distribution , Zinc/analysis
7.
Domest Anim Endocrinol ; 7(1): 75-82, 1990 Jan.
Article in English | MEDLINE | ID: mdl-2311376

ABSTRACT

Thirty-one gilts were ovariectomized between 21 and 34 hr after the onset of estrus to compare changes in follicular endocrinology with stages of oocyte maturation. Oocytes were recovered from 6 to 8 mm follicles and classified by stage of meiosis. Remaining follicular fluid was assayed for steroids and dermatan sulfate. Amounts of prostaglandin F2 alpha (PGF2 alpha) and E2 (PGE2) were measured in intramural tissues. Coincident with germinal vesicle breakdown, the follicular content of all steroids except testosterone decreased (P less than .05). As oocytes approached metaphase II, the amount of progesterone within follicles increased (P less than .05), and estradiol continued to decrease (P less than .05). The pattern of dermatan sulfate content was biphasic and peaked at germinal vesicle breakdown and anaphase stages. Amounts of PGF2 alpha and PGE2 within intramural tissues increased (P less than .05) throughout oocyte maturation. Follicular atresia was evident during estrus; however, more (P less than .05) atretic follicles were recovered at germinal vesicle than metaphase II stages (20 vs 3%, respectively). Follicular development, within a gilt, was skewed (P less than .05) and classification of follicles by hormone content demonstrated that a majority were more mature than a minority of less mature follicles. These data suggest that follicular maturation and oocyte development are highly correlated in swine. Furthermore, partitioning the follicular variability by hour and stage of oocyte maturation allowed for more precise assessment of follicular endocrinology than previously reported.


Subject(s)
Gonadal Steroid Hormones/analysis , Oocytes/growth & development , Ovarian Follicle/physiology , Swine/physiology , Androstenedione/analysis , Animals , Dermatan Sulfate/analysis , Dinoprost/analysis , Dinoprostone/analysis , Estradiol/analysis , Female , Ovarian Follicle/analysis , Progesterone/analysis , Testosterone/analysis
8.
Biol Cell ; 68(2): 119-27, 1990.
Article in English | MEDLINE | ID: mdl-1694097

ABSTRACT

The changes in distribution and density of mitochondria and the level of mitochondrial RNA during Drosophila oogenesis were studied simultaneously in the 3 cell types ie follicle cells, nurse cells and oocyte, making up the egg chamber. Up to stage 6, mitochondrial density (mitochondrial and cellular areas ratio) was elevated and increased similarly in both follicle and nurse cells. Thereafter the mitochondrial density of follicle cells continued to increase and that of the nurse cells declined markedly while the nurse cell mitochondria assembled in dense groups and decreased in size. This can be related to a transfer of nurse cell cytoplasm, including mitochondria, to the oocyte. In the oocyte from stage 4 to stage 7 we observed a significant decrease of the mitochondrial density due to the absence of mitochondrial biogenesis. Then the cytoplasm transfer caused mitochondrial density to increase up to the level found in the nurse cells at the end of oogenesis. The mature oocyte contains enough mitochondria to supply 15,000 somatic cells. Our results strongly suggest that the variations in size, distribution and density of mitochondria relate to the particular energetic requirements of the different cell types during the first half of oogenesis. Later they relate to the developmental requirements of the nurse cells and the oocyte, in particular the storage of mitochondria in the oocyte. The level of mitochondrial RNA was studied through in situ hybridization. Throughout oogenesis the follicle and nurse cell RNA evolved similarly. Up to stage 9, there was no change in RNA densities in these cells, suggesting a correlation with the cell volume and/or the nuclear DNA content. Thereafter the cellular RNA concentration declined rapidly. In the oocyte the RNA concentration evolved differently especially from stage 10 to the end, the RNA density being stabilized. This can be related to the injection of nurse cell mitochondria, followed by their assignment to reserve status. Our results suggest that the mt RNA density is under extramitochondrial control mechanisms.


Subject(s)
Drosophila/genetics , Mitochondria/physiology , Oogenesis/physiology , Ovary/physiology , Animals , Female , Mitochondria/analysis , Mitochondria/ultrastructure , Nucleic Acid Hybridization , Oocytes/analysis , Oocytes/cytology , Oocytes/ultrastructure , Ovarian Follicle/analysis , Ovarian Follicle/cytology , Ovarian Follicle/ultrastructure , Ovary/cytology , Ovary/ultrastructure , RNA/analysis , RNA/genetics
9.
Acta Endocrinol (Copenh) ; 121(4): 578-80, 1989 Oct.
Article in English | MEDLINE | ID: mdl-2529730

ABSTRACT

The concentrations of immunoreactive C-terminal (ANH-(99-126)) and N-terminal (ANH-(1-98] portions of pro-ANH were measured in follicular fluid and plasma samples from 9 young women undergoing in vitro fertilization. ANH-(99-126) and ANH-(1-98)-like immunoreactivity levels in plasma were 6.0-25.4 (mean 12.2 pmol/l and 184-427 (mean 300) pmol/l, respectively, whereas the corresponding levels in follicular fluid were 3.8-8.0 (mean 4.9) pmol/l and 169-385 (mean 262) pmol/l. The concentrations of both ANH-like peptides were consistently lower (p less than 0.01) in the follicular fluid than in the matched plasma samples, but within the variation found in plasma controls. It is concluded that ANH-like peptides in the follicular fluid, whether secreted locally or derived from circulating ANH, might play a physiological role in the biosynthesis of ovarian steroid hormones or follicular maturation and fluid dynamics.


Subject(s)
Atrial Natriuretic Factor/analysis , Ovarian Follicle/analysis , Peptide Fragments/analysis , Protein Precursors/analysis , Atrial Natriuretic Factor/immunology , Female , Fertilization in Vitro , Humans , Peptide Fragments/immunology , Protein Precursors/immunology
10.
Clin Chim Acta ; 183(2): 115-23, 1989 Aug 15.
Article in English | MEDLINE | ID: mdl-2676243

ABSTRACT

A sandwich-type solid phase time-resolved immunofluorometric assay (IFMA) was developed for endometrial protein PP14 (placental protein 14). The assay utilizes affinity-purified polyclonal antibodies for coating the microtiter wells and for labelling with europium (III) chelate. Maintaining specificity, the 0.6 micrograms/l sensitivity of IFMA is over 25 times higher than that of RIA. The immunofluorometric method enables detection and accurate quantitation of PP14 in all those samples in which PP14 is undetectable by RIA. The method is suitable for quantitative measurement of low PP14 levels in serum of postmenopausal and fertile-aged women and men, as well as in follicular fluid. At 14-16 micrograms/l, which is the sensitivity of radioimmunoassay, the intra-assay variation of IFMA is 6.6% and inter-assay variation 11.4%. In postmenopausal women the PP14 levels are 12.7-56.7 micrograms/l, in fertile-aged women 13.7-113.4 micrograms/l, and in men 3.1-53.1 micrograms/l. The levels in preovulatory follicular fluid are 1.2-20.5 micrograms/l. It is concluded that PP14 IFMA is highly sensitive, accurate and suitable for measurement of protein levels undetectable by other currently available methods.


Subject(s)
Glycoproteins , Pregnancy Proteins/blood , Amniotic Fluid/analysis , Body Fluids/analysis , Cross Reactions , Dose-Response Relationship, Drug , Female , Fluorescent Antibody Technique , Glycodelin , Humans , Menopause/blood , Menstrual Cycle , Ovarian Follicle/analysis , Pregnancy , Radioimmunoassay , Time Factors
11.
Prostaglandins ; 38(2): 197-202, 1989 Aug.
Article in English | MEDLINE | ID: mdl-2549576

ABSTRACT

Prostaglandins are involved in ovulation and in every mammal studied so far, ovulation has been inhibited by prostaglandin inhibition. Information regarding the role of leukotrienes and thromboxanes in ovulation is more limited. In order to study the production of eicosanoids in human pre-ovulatory follicular fluid, follicular aspiration was timed by means of serial ultrasound scans and human chorionic gonadotrophin (hCG) to be immediately pre-ovulatory. 11 women were studied and the eicosanoids measured by radioimmunoassay (RIA). The follicular fluid was found to contain leukotrienes (LT) B4, LTC4 (D4, E4), prostaglandin (PG) E2, PGF2 alpha 6 keto PGF1 alpha k and thromboxane (TX) B2. This is the first published report of leukotrienes in human follicular fluid in spontaneous cycles, and is one of the few reports showing prostaglandins and thromboxanes. The significance of demonstrating leukotrienes in human follicular fluid is discussed as is the correlation between individual eicosanoids in the human ovary.


Subject(s)
Follicular Phase , Leukotrienes/analysis , Ovarian Follicle/analysis , Dinoprostone/analysis , Female , Humans , Leukotriene B4/analysis , Lipoxygenase/metabolism , Ovarian Follicle/enzymology , Prostaglandin-Endoperoxide Synthases/metabolism , Prostaglandins F/analysis , SRS-A/analysis , Thromboxane B2/analysis
12.
Acta Endocrinol (Copenh) ; 121(1): 90-4, 1989 Jul.
Article in English | MEDLINE | ID: mdl-2525858

ABSTRACT

Studies in animals have highlighted a possible role for growth factors, particularly IGF-I on cellular replication and cytodifferentiation in the ovary. At this time, few studies have been performed about IGF-I in the human ovary. From 38 women undergoing in Vitro Fertilization 293 antral antral fluids were collected and assessed for steroids (estradiol and progesterone), FSH and IGF-I. Two induction treatments were compared: clomiphene citrate hMG (group A,N = 15), triptoreline/hMG (group B,N = 23). We also studied relationships between quantitative parameters and oocyte collection or oocyte corona cumulus complex maturity, In group B, the highest antral estradiol levels were found in follicles yielding an oocyte (p less than 0.05). Concerning antral progesterone, higher levels were observed in follicles collected from group A than follicles collected from group B (p less than 0.05): for this parameter, the highest levels were observed when an oocyte was harvested, whatever the treatment (p less than 0.05). Highest antral FSH levels were observed in group B (p less than 0.05). IGF-I levels were higher in follicles collected from group B than in follicles collected from group A (p less than 0.05) and antral IGF-I levels differed between mature and immature oocyte corona cumulus complex in group B (p less than 0.05). These results, which are in keeping with studies about biological action of IGF-I in animal or human follicles or granulosa cells, led us to hypothesize a role for IGF-I in human follicular recruitment and maturation, a role that possible is enhanced during GnRH analogue and gonadotropin therapy.


Subject(s)
Insulin-Like Growth Factor I/analysis , Oocytes/drug effects , Somatomedins/analysis , Cell Division/drug effects , Clomiphene/administration & dosage , Estradiol/analysis , Female , Fertilization in Vitro , Follicle Stimulating Hormone/analysis , Gonadotropin-Releasing Hormone/administration & dosage , Gonadotropin-Releasing Hormone/analogs & derivatives , Humans , Menotropins/administration & dosage , Oocytes/analysis , Ovarian Follicle/analysis , Ovarian Follicle/drug effects , Progesterone/analysis , Triptorelin Pamoate
13.
Hum Reprod ; 4(5): 482-5, 1989 Jul.
Article in English | MEDLINE | ID: mdl-2794009

ABSTRACT

Cigarette smoking is accepted as a risk factor for pregnancy but its effect on fertility is uncertain. In this study we determined the concentration of cotinine, a nicotine metabolite, in follicular fluid and serum from women participating in an in-vitro fertilization and embryo transfer (IVF-ET) programme. Cotinine was undetectable in serum and follicular fluid of non-smokers but ranged from less than 5 to 371 ng/ml in follicular fluid and from 24 to 245 ng/ml in serum of smokers. Granulosa-luteal cells, obtained from IVF patients and cultured for 4 days, secreted progesterone and, when an aromatizable androgen was added, oestradiol-17 beta. The addition of cotinine or nicotine did not alter progesterone or oestradiol-17 beta secretion. However, the presence of cotinine in follicular fluid of women smokers provides evidence for access of at least one component of cigarette smoke to the developing gamete and the cells of the follicle. Further work is required to determine whether fertility is compromised by the presence, in follicular fluid, of contaminants derived from cigarette smoke.


Subject(s)
Cotinine/analysis , Fertilization in Vitro/drug effects , Ovarian Follicle/analysis , Pyrrolidinones/analysis , Cells, Cultured , Cotinine/blood , Cotinine/pharmacology , Estradiol/biosynthesis , Female , Granulosa Cells/metabolism , Humans , Luteal Cells/metabolism , Nicotine/metabolism , Nicotine/pharmacology , Pregnancy , Progesterone/biosynthesis , Smoking/adverse effects
14.
Hum Reprod ; 4(5): 558-67, 1989 Jul.
Article in English | MEDLINE | ID: mdl-2794015

ABSTRACT

The aim of this study was to determine whether certain characteristics of the follicular biochemistry, previously shown by us to be associated with oocyte developmental capacity, also reflected differences in oocyte appearance, and to determine the range of oocyte characteristics induced by ovarian stimulation. A representative sample of 33 human oocytes and associated follicular fluids was obtained after a follicular growth period considered suitable for IVF. Individual follicular fluid protein and proteoglycan levels, and follicular volume were compared with the morphological characteristics of each oocyte. Oocytes which retained the germinal vesicle nuclear status after exposure to human chorionic gonadotrophin tended to occur in small follicles (less than or equal to 2 ml) and to be highly vacuolated and with characteristic predicted a low potential for cleavage. Among those oocytes which had progressed to MII nuclear maturity [in the medium (2.5-6.5 ml) and large (greater than or equal to 7 ml) volume follicles] the degree of oocyte vacuolation was negatively correlated with alpha 1-antitrypsin level, while the degree of organelle clumping was correlated with proteoglycan and immunoglobulin levels. Only five of the oocytes (15%) in this sample had follicular characteristics consistent with a normal potential for pregnancy. These MII oocytes occurred within the medium volume range, had low vacuolation levels, and a low degree of organelle clumping. In contrast, those oocytes with a low potential for cleavage based on their follicular biochemistry, showed high cytoplasmic vacuolation levels.


Subject(s)
Oocytes/cytology , Ovarian Follicle/analysis , Cell Nucleus/ultrastructure , Cytoplasm/metabolism , Cytoplasm/ultrastructure , Female , Humans , Menstrual Cycle , Oocytes/metabolism , Organelles/ultrastructure , Vacuoles/ultrastructure
15.
Gynecol Endocrinol ; 3(2): 125-33, 1989 Jun.
Article in English | MEDLINE | ID: mdl-2510461

ABSTRACT

The efficacy of combined growth hormone (GH)-gonadotropin treatment has been studied in patients previously resistant to sole gonadotropins for induction of superovulation. Eleven patients (aged 26-41) with a mechanical cause of infertility were treated. All were given the same dosage of gonadotropins as in previous cancelled cycles (6-17 ampules/cycle of menofollitropin; 34-80 ampules/cycle of human menopausal gonadotropin) plus a standard dosage of GH (0.1 IU per kg body weight, daily). Younger patients (n = 6, age 26-36) showed a considerable improvement of ovarian response in terms of number of mature follicles aspirated by laparoscopy (performed on day 11-13). Older patients (n = 5, age 39-41) did not show any significant improvement of ovarian response with combined treatment and all had their stimulatory cycle cancelled. Follicular fluid (FF) levels of GH, 17 beta-estradiol (E2) and progesterone (P) were significantly higher in the group of younger GH-treated patients (n = 53 follicles) than in 4 controls treated with gonadotropins only (n = 32 follicles). FF insulin-like growth factor-I (IGF-I) did not significantly differ between the two groups. A significant positive linear correlation has been found between FF GH and IGF-I in the GH-treated group. In conclusion, GH-gonadotropin combined treatment considerably improves ovarian response in protocols for superovulation induction in younger gonadotropin-resistant patients. A local action of GH and IGF-I in the ovaries may be hypothesized.


Subject(s)
Growth Hormone/therapeutic use , Menotropins/therapeutic use , Ovary/drug effects , Ovulation , Superovulation , Adult , Estradiol/blood , Female , Growth Hormone/analysis , Humans , Insulin-Like Growth Factor I/analysis , Luteinizing Hormone/therapeutic use , Ovarian Follicle/analysis , Progesterone/analysis
16.
Fertil Steril ; 51(5): 863-8, 1989 May.
Article in English | MEDLINE | ID: mdl-2707464

ABSTRACT

To test the usefulness of human follicular fluid (FF) in treating male infertility, we incubated washed sperm specimens from 31 couples undergoing intrauterine insemination (IUI), for male and/or unexplained infertility, with either FF or Ham's F-10 medium (Gibco, Grand Island, NY), in alternating cycles in a randomized manner. Semen specimens from 28 men were incubated with either medium or FF. Incubations with FF have increased sperm penetration assay (SPA) scores from 24.8 +/- 17.3 to 34.3 +/- 13.6 (P less than 0.01). Incubation with heat-inactivized FF also has increased SPA scores, although to a lesser extent than noninactivized FF. Seventeen pregnancies occurred in the 31 couples treated by IUI (54.8%), 16 of them in FF-treated cycles (51.6%) and one in "control" IUI cycles (3.2%, P less than 0.01). All pregnancies occurred within four treatment cycles. Thus, IUI after sperm wash and preincubation with FF may be suggested for four to six cycles to couples with male factor and/or unexplained infertility who are reluctant to resort to artificial insemination by donor or adoption, before attempting the more costly and complex in vitro fertilization-embryo transfer procedure.


Subject(s)
Body Fluids/physiology , Infertility, Male/therapy , Ovarian Follicle/analysis , Spermatozoa/physiology , Body Fluids/analysis , Female , Humans , Infertility, Male/physiopathology , Insemination, Artificial , Male , Pregnancy , Pregnancy Outcome , Sperm-Ovum Interactions
17.
J Mol Endocrinol ; 2(3): 189-200, 1989 May.
Article in English | MEDLINE | ID: mdl-2751827

ABSTRACT

Analysis of bovine follicular fluid (FF) using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) coupled with a sensitive immunoblotting procedure resolved several components that were immunoreactive with an antiserum directed against the n-terminus of the alpha subunit of human inhibin (hI alpha(1-32]. Under non-reducing conditions, three intensely stained bands having apparent Mr values of 116,000, 44,000 and 25,000 were present, whilst under reducing conditions only two intensely stained bands (Mr 43,000 and 21,000) were detected. The Mr 44,000 and 25,000 immunoreactive forms (non-reducing conditions) were also demonstrated in bovine utero-ovarian vein and peripheral venous plasma after subjecting samples (40 ml) to immunoaffinity concentration using Sepharose beads coupled to anti-hI alpha(1-32), SDS-PAGE and immunoblotting. The same approach revealed the presence of the smaller (Mr 25,000) form in bovine granulosa cell-conditioned culture medium (GCCM). Gel-permeation chromatography (Sephacryl S-200), immunoaffinity chromatography (Sepharose-anti-hI alpha(1-32] and reversed-phase high-performance liquid chromatography (RP-HPLC; C18 and C8 columns) were employed to isolate from bFF (30 ml, 19.5 g protein) 750 micrograms protein which appeared essentially homogeneous by RP-HPLC and SDS-PAGE and had an Mr of 25,000 (non-reducing conditions)/21,000 (reducing conditions), identical to that of the immunoreactive component of lowest Mr found in bovine FF, utero-ovarian vein plasma, peripheral plasma and GCCM. The isolated material was highly immunoreactive with antisera against both hI alpha(1-32) and purified Mr 32,000 bovine inhibin but was devoid of biological activity when tested in a rat pituitary cell inhibin bioassay. Amino-terminal analysis revealed an amino acid sequence (residues 1-14) identical to that reported elsewhere for the alpha subunit (Mr 20,000/21,000) of bovine inhibin. In conclusion, the present study has revealed that the bovine ovary secretes considerable quantities of monomeric inhibin alpha subunit. The unexpected presence of this material in peripheral blood is likely to hinder attempts to obtain physiologically relevant data on circulating levels of inhibin in cattle using conventional radioimmunoassays.


Subject(s)
Inhibins/metabolism , Ovary/metabolism , Animals , Body Fluids/analysis , Cattle , Cells, Cultured , Electrophoresis, Polyacrylamide Gel , Female , Granulosa Cells/metabolism , Humans , Immunoblotting , Inhibins/blood , Inhibins/isolation & purification , Ovarian Follicle/analysis , Sheep/metabolism , Species Specificity , Swine/metabolism , Veins
18.
Biol Reprod ; 40(5): 1087-93, 1989 May.
Article in English | MEDLINE | ID: mdl-2669983

ABSTRACT

The pituitary gonadotropins and gonadal steroids are required for normal follicular growth and development but neither has been shown to act directly as a granulosa cell mitogen in vitro. A number of polypeptide growth factors, however, are known to have pronounced mitogenic effects on the cells of the follicle. We have localized transforming growth factor-alpha (TGF-alpha), a potent mitogen, in bovine thecal cells via immunoperoxidase staining using a monoclonal antibody for TGF-alpha that does not cross-react with epidermal growth factor. TGF-alpha staining is most intense in the theca of follicles at the discrete physiological stages known to show rapid granulosa cell growth (small follicles of 0.7-2.0 mm diameter). Staining intensity for TGF-alpha declines in large preovulatory follicles, coincident with the known decline in granulosa cell mitosis. These studies provide further evidence for paracrine interactions in the ovary and show that TGF-alpha may play an important role in the regulation of follicular development in the adult bovine ovary.


Subject(s)
Ovarian Follicle/analysis , Ovary/analysis , Transforming Growth Factors/analysis , Animals , Antibodies, Monoclonal , Antibody Specificity , Cattle , Cells, Cultured , Female , Granulosa Cells/analysis , Immunoenzyme Techniques , Immunohistochemistry , Ovarian Follicle/physiology , Theca Cells/analysis , Transforming Growth Factors/physiology
19.
Fertil Steril ; 51(4): 639-43, 1989 Apr.
Article in English | MEDLINE | ID: mdl-2494081

ABSTRACT

Apolipoprotein E (apoE) is an important effector of plasma lipoprotein due to its interaction with cell-surface receptors. In addition, it is secreted by rat and human ovarian granulosa cells in tissue culture, and its production is hormonally modulated. Such observations indicate that apoE should be found in human follicular fluid (FF) and that its abundance therein may be subject to hormonal regulation. The authors have collected FF from seven normally cycling women by needle puncture at the time of laparoscopy and from hyperstimulated ovaries of four women undergoing in vitro fertilization (IVF). Apolipoprotein E is present in human FF. The ratio of apoE concentration in FF to that in serum, as determined by western blot analysis, ranges from 0.43 to 4.2 for the normally cycling patients and from 0.16 to 0.77 for the IVF patients. Serum estrogen levels (range, 42 to 477 pg/ml) for the cycling patients are inversely correlated with the FF-to-serum ratio of apoE (r = 0.91). The content of apoE in FF, relative to serum, thus falls dramatically as the follicle approaches ovulation. These data, plus the known ability of apoE to carry cholesterol and other lipids, suggest an important role for apoE in the ovary.


Subject(s)
Apolipoproteins E/analysis , Estrogens/blood , Ovarian Follicle/analysis , Apolipoproteins E/physiology , Female , Fertilization in Vitro , Follicle Stimulating Hormone/pharmacology , Humans
20.
Hum Reprod ; 4(3): 257-60, 1989 Apr.
Article in English | MEDLINE | ID: mdl-2785532

ABSTRACT

Concentrations of epidermal growth factor (EGF), progesterone (P4) and oestradiol (E2) were determined in follicular fluid (FF) aspirated from 69 preovulatory follicles in 15 women undergoing ovarian hyperstimulation and ovulation induction as part of IVF-ET treatment for infertility. Sixty-nine oocytes were obtained from these FF aspirates and 35 (51%) fertilized and cleaved in vitro. Pregnancy was achieved in five (33%) of the women. Concentrations of EGF in FF ranged between 0.60 and 2.42 ng/ml, corresponding to approximately 50% of the level in serum at the time of follicle aspiration. A significant linear correlation between levels of EGF in FF and in serum was demonstrated. By contrast, no significant relationships were found between intrafollicular levels of EGF and those of P4 and E2, or between EGF in FF and oocyte cleavage rate in vitro and the achievement of pregnancy after ET. It is concluded that EGF in FF of the human preovulatory follicles reflects a passive diffusion into the follicle from the circulation. Moreover, that intrafollicular EGF is not involved in the local regulation of human preovulatory follicle and oocyte maturation. These results, however, do not preclude an auto- and/or paracrine role of intrafollicular EGF in smaller human follicles.


Subject(s)
Epidermal Growth Factor/analysis , Follicular Phase , Ovarian Follicle/analysis , Body Fluids/analysis , Estradiol/analysis , Female , Humans , Progesterone/analysis
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