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Biomedical subjects

R J Norman

Publications and source records attributed to R J Norman.

At least 19 recordsLinked to original sources
PubMed1992

Tumor necrosis factor alpha in the human ovary: presence in follicular fluid and effects on cell proliferation and prostaglandin production.

OBJECTIVE: To examine the concentration of tumor necrosis factor alpha (TNF alpha) in human follicular fluid (FF) and its effects on cultured human granulosa-lutein cells. DESIGN: The concentration of TNF alpha in FF from hyperstimulated cycles and in conditioned media from cultured granulosa-lutein cells was measured by radioimmunoassay (RIA) and bioassay using L929 cells. The effects of recombinant human TNF alpha (rTNF alpha) on proliferation and production of progesterone (P) and prostaglandin (PG, PGE2, and PGF2 alpha) by cultured human granulosa-lutein cells were assessed. SETTING: In vitro fertilization and embryo transfer (IVF-ET) program at Reproductive Medicine Unit, The Queen Elizabeth Hospital, Woodville, South Australia, Australia. PATIENTS: Twenty-five women undergoing IVF-ET for tubal factor infertility. RESULTS: The concentration of immunoreactive TNF alpha in FF was 0.36 +/- 0.02 microgram/L, and there were no significant correlations between levels of TNF alpha and steroids or FF volume. Bioactivity for TNF alpha was considerably less. Immunoreactive or bioactive TNF alpha was not detected in conditioned media from granulosa-lutein cell culture. Recombinant human TNF alpha dose-dependently stimulated proliferation of cultured granulosa-lutein cells as measured by incorporation of 3H-thymidine, but in contrast to earlier reports, we were not able to demonstrate any effect of rTNF alpha on basal or human chorionic gonadotropin-stimulated P accumulation during culture periods of up to 72 hours. The accumulation of both PGE2 and PGF2 alpha was dose-dependently increased by rTNF alpha during a 48-hour incubation period. Time course studies revealed that maximal levels of both PGE2 and PGF2 alpha were reached within 12 hours of culture. CONCLUSION: Immunoreactive and bioactive TNF alpha is present in FF. Tumor necrosis factor alpha may have a physiological role in stimulating proliferation of follicular cells and PG production at the time of ovulation and formation of the corpus luteum.

Cell Division↗
PubMed1992

Concentrations of immunoreactive interleukin-1 and interleukin-2 in human preovulatory follicular fluid.

The presence of immunoreactive interleukin-1 (IL-1) and interleukin-2 (IL-2) in human follicular fluid obtained at the time of oocyte collection for in-vitro fertilization was ascertained by radioimmunoassay. In group I (20 fluids from 20 patients), the concentrations of IL-1 were 0.9 +/- 0.06 and 1.9 +/- 0.04 (mean +/- SEM) fmol/l in follicular fluid and plasma respectively. A positive correlation existed between IL-1 levels in follicular fluid and plasma (r = 0.56, P less than 0.01). Concentrations of IL-2 were 3.5 +/- 0.2 and 6.1 +/- 0.3 fmol/l in follicular fluid and plasma respectively. A positive correlation of IL-2 levels was also found between follicular fluid and plasma (r = 0.65, P less than 0.01). There was no association between IL-1, IL-2 and steroid levels, regardless of whether they were compared in follicular fluid or plasma. Group II was composed of a series of fluids (two to seven samples for each of seven patients) in which the follicular concentrations of IL-1 and IL-2 did not show a positive correlation with the volume of follicular fluid or the concentrations of follicular fluid steroids. It is concluded that human preovulatory follicular fluid contains immunoreactive IL-1 and IL-2. The role of IL-1 and IL-2 in ovarian physiology remains to be determined.

Estradiol↗
PubMed1992

Distribution of leukocyte subpopulations in the human corpus luteum.

Cytokines, as secreted products of leukocytes, have roles in many organs of the body via paracrine or autocrine mechanisms. In the present study, we demonstrate by immunocytochemistry the leukocytes present in the human corpus luteum in order to investigate further the relationship between leukocytes, cytokines and corpus luteum function. Ten intact corpora lutea were collected from female patients who had no apparent ovarian disease. The mean age of these patients was 37 years (range 23-55 years). Frozen and paraffin sections were subjected to analysis using monoclonal antibodies which were specific to leukocyte marker antigens. The results showed that there are macrophages, cells positive for leukocyte common antigen (LCA), T lymphocytes including T helper/inducer (T4) cells, T cytotoxic/suppressor (T8) cells and activated T (Ta) cells (interleukin-2 receptor-positive cells), monocytes and natural killer (NK) cells but not B lymphocytes present in the human corpus luteum. The distribution of the leukocytes present in the different parts of the corpus luteum was found to be in the order: theca-luteal area greater than loose connective tissue area greater than granulosa-luteal area. Macrophages and T lymphocyte subsets comprised the main components of the total leukocytes in the human corpus luteum. Ta cells were only localized in the loose connective tissue of the corpus luteum. In most cases, macrophages, LCA cells and T4 cells tended to be situated in a single cell layer on the edge of the theca-luteal area and surrounding the granulosa-luteal area. These results suggest that the leukocytes may act to a greater extent in the theca-luteal area than in the granulosa-luteal area.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Comparison of specific immunoassays for detection of the beta-core human chorionic gonadotrophin fragment in body fluids.

We have validated two new methods, one radioimmunoassay (RIA) and one immunoradiometric assay (IRMA), for the detection of beta-core hCG fragment (beta C-hCG) in body fluids. In addition, we have compared their performance with two other assays designed for beta C-hCG quantification. The RIA uses a rabbit polyclonal antibody raised against pure beta C-hCG which has a high affinity constant, is sensitive to 5 pmol/l, and has significant cross-reaction only with the free beta LH subunit. The IRMA, designed in a liquid phase, uses the same polyclonal antibody associated with a 125I-labelled mouse monoclonal antibody (32H2) raised against beta hCG, is sensitive to 1.5 pmol/l, and does not cross-react significantly with any related glycoprotein. Comparison between these two assays and two others previously published was made by measuring beta C-hCG in urine from healthy pregnant women (n = 47) and gave correlation coefficients higher than r = 0.960 with any combination. Analysis of beta C-hCG in urine of non-pregnant subjects (n = 238) showed measurable beta C-hCG in 8.8% (levels ranged from 5 to 34 pmol/l) with the IRMA and 88.3% with the RIA (n = 30; ranging from 28.4 to 228 pmol/l) (P = 0.05). We concluded that, despite different affinities of the antibody involved and different cross-reactivities with related glycoproteins, the four assays we examined may be equally employed to detect beta C-hCG in pregnancy urine. However, the IRMA appears to be more appropriate for beta C-hCG analysis in non-pregnant individuals, specifically in postmenopausal women because of the high cross-reactivity of the RIA with free beta LH or beta fragments of other glycoproteins. These studies have significance for our understanding of the physiology of beta C-hCG in cancer, pregnancy and after the menopause.

Antibodies↗
PubMed1992

Distribution of the beta-core human chorionic gonadotrophin fragment in human body fluids.

The origins of a fragment of the human chorionic gonadotrophin (hCG) molecule, beta-core (beta C-hCG) were studied by analysis of beta C-hCG concentrations in biological fluids. In addition, the ability of the placenta to produce the fragment and the metabolism of hCG to beta C-hCG by human granulosa cells was determined in tissue culture. Finally the conversion of exogenous hCG to beta C-hCG was studied in vivo. The fragment was present in pregnancy urine as well as that from premenopausal and postmenopausal subjects. The highest concentrations were found in pregnant women. Ratios of beta C-hCG to intact hCG were higher in pregnancy urine when radioimmunoassay (RIA) was used compared with immunoradiometric assay (IRMA) (0.67 and 0.37 respectively). Concentrations of beta C-hCG were higher in postmenopausal urine than in premenopausal specimens. A significant amount of a high molecular weight beta C-hCG immunoreactive material was found in serum samples after size separation, and the molar ratio of beta C-hCG/hCG was estimated as 0.019. Amniotic fluid also contained small quantities of two forms of immunoreactive beta C-hCG and the ratio of 0.01 for authentic beta C-hCG/hCG increased to 0.026 when the high molecular weight form was considered. Cultured trophoblastic tissue released material with beta C-hCG immunoreactivity in the medium and chromatographic separation revealed that the majority of this material was of higher molecular weight compared with the authentic beta C-hCG form. beta C-hCG was the principal glycoprotein found in follicular fluid after hyperstimulated folliculogenesis and intramuscular injection of 5000 IU hCG. We also demonstrated that 26% of follicular fluid samples (n = 50) were positive for beta C-hCG; levels ranged from 5.2 to 23.0 pmol/l (13.1 +/- 5.7); S.D.) when a specific IRMA was used. The RIA could detect beta C-hCG in 48 samples (96%), levels ranging from 7.0 to 28.5 pmol/l (19.4 +/- 5.2). Moreover, granulosa cells cultured in the presence of hCG were able to degrade the intact molecule to both high molecular weight and authentic immunoreactive forms of beta C-hCG. After gel filtration, material of molecular weight over a wide range and immunoreactive for beta C-hCG was present in human seminal plasma. Assaying 74 samples of this fluid by IRMA, beta C-hCG was detected in 42 (56.7%), levels ranging between 5.5 and 59.5 pmol/l (24.9 +/- 15.2).(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1992

Urinary concentrations of beta core fragment of hCG throughout pregnancy.

OBJECTIVE: We sought to determine a reference range for urinary immunoreactive beta core fragment of hCG (beta C-hCG) in pregnancy, the ratio between beta C-hCG and intact hCG, and the earliest detectable rise of beta C-hCG in urine. METHODS: Urine was obtained from 741 pregnant women between 6-41 weeks' gestation, as well as from women undergoing donor insemination with timed ovulation peaks. RESULTS: The beta core fragment of hCG reached a maximum between 8-15 weeks, with a decrease between 20-29 weeks. The molar ratio of beta C-hCG to intact hCG was always greater than 1. CONCLUSION: In pregnancy, beta C-hCG concentrations increase in the urine in parallel to intact hCG but at a higher molar ratio, suggesting either placental production of beta C-hCG or enhanced metabolism of hCG to beta C-hCG in peripheral organs.

Chorionic Gonadotropin↗
PubMed1992

Simple quantitative measurement of serum choriogonadotropin compared with immunoradiometric, immunoenzymometric, and chemiluminescent assays.

We evaluated a new simplified quantitative method (Tandem Icon QSR; Hybritech) for choriogonadotropin (hCG), which could theoretically be performed in a ward or with outpatients. The method was compared with immunoradiometric (Bioclone Australia), chemiluminescent (Amerlite; Amersham), and immunoenzymometric assays (Stratus; Dade). We analyzed by each of the methods 104 serum samples from pregnant and nonpregnant patients. For quantitative hCG values indicative of pregnancy (greater than 25 int. units/L), excellent correlation was observed between the Tandem Icon method and the other standard laboratory assays (r = 0.995, 0.990, and 0.992, respectively). Occasional problems arose because of the instability of Tandem Icon QSR reagents at room temperature but this was resolved by storing the reagents at 4 degrees C. We conclude that this simplified quantitative method for hCG is reliable and suitable for use outside of the routine immunoassay laboratory.

Chorionic Gonadotropin↗
PubMed1991

Decrease in luteal gonadotropin concentration in conception cycles after in vitro fertilization/gamete intrafallopian transfer.

The concentrations of the gonadotropins, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) were measured in the luteal phase of the cycle in patients undergoing ovarian hyperstimulation. In nonconception cycles, FSH and LH were increased in the late luteal phase compared with conception cycles in which both gonadotropins were suppressed. Estradiol (E2) and progesterone concentrations increased in pregnancy cycles and may be the sole cause for the decreased gonadotropin concentrations as shown by equivalent concentrations of LH and FSH in both pregnancy and nonpregnancy cycles after matching for E2 concentrations. Subjects who subsequently had twin pregnancy or a spontaneous abortion were compared with those with a successful ongoing singleton conception. There were no significant differences relative to LH and FSH between the three groups, although in twin pregnancy FSH tended to be lower at day 16 from oocyte recovery. It is concluded that suppression of LH and FSH in hyperstimulated pregnancy cycles occurs after the time of the rising human chorionic gonadotropin concentrations in plasma.

Abortion, Spontaneous↗
PubMed1991

Differential effects of gonadotrophin-releasing hormone agonists administered as desensitizing or flare protocols on hormonal function in the luteal phase of hyperstimulated cycles.

The incorporation of gonadotrophin-releasing hormone agonist (GnRHa) in in-vitro fertilization (IVF) stimulation protocols has led to doubt about the quality of the subsequent luteal phase. The effects of two GnRHa stimulation protocols on luteal phase concentrations of oestradiol (E2), progesterone (P), luteinizing hormone (LH) and follicle stimulating hormone (FSH) were compared with the standard clomiphene stimulation regimen. Subjects receiving clomiphene with human menopausal gonadotrophin (HMG, n = 377) showed essentially similar luteal phase P concentrations to those receiving leuprolide acetate/HMG as a desensitization protocol. Subjects receiving concomitant leuprolide and HMG from day 2 to utilize the flare effect of the GnRHa exhibited significantly lower P levels in the luteal phase compared to clomiphene/HMG and leuprolide desensitization protocols despite the addition of HCG support. This occurred despite equivalent E2 concentrations at the time of ovulation and identical numbers of oocytes recovered. LH concentrations in non-conception cycles were suppressed for at least 14 days in the luteal phase in both GnRHa protocols compared to clomiphene stimulation. Differences were less obvious in cycles where conception occurred suggesting that implantation may proceed more favourably when the luteal endocrinology was optimal. It is concluded that flare methods of GnRHa hyperstimulation are associated with significantly different luteal phases compared with clomiphene or desensitization protocols. It is proposed that the use of the flare type of stimulation may significantly influence the response of the granulosa cells to LH or HCG via gonadotrophin receptors or through altered post-receptor function.

Chorionic Gonadotropin↗
PubMed1991

The leuprolide flare regime for in-vitro fertilization/gamete intra-fallopian transfer and embryo cryopreservation.

Gonadotrophin releasing hormone agonists (GnRHa) are now well established as adjuvant agents for in-vitro fertilization (IVF)/gamete intra-Fallopian transfer (GIFT) but several different modes of usage have been proposed. Our experience with 328 cycles of leuprolide used in a flare regime is reviewed. An endocrinologically proven flare effect was associated with a reduction of human menopausal gonadotrophin (HMG) usage (10 versus 16 ampoules) and a lower cycle cancellation/conversion rate (7.4 versus 11.3%). Overall, satisfactory rates of oocyte recovery (93%, mean number of oocytes 7.0), clinical pregnancy (24.4% per oocyte recovery) and pregnancy from frozen/thawed embryo transfers (14%) were achieved. The flare protocol appears to be a satisfactory choice for the majority of subjects but careful monitoring is required to avoid the potential for ovarian hyperstimulation.

Cryopreservation↗
PubMed1991

Evaluation of renal function and fluid homeostasis during recovery from exercise-induced hyponatremia.

Renal function including fluid and electrolyte balance was studied during recovery in eight subjects who developed symptomatic hyponatremia (HN; plasma sodium concentration less than 130 mM) during an 88-km ultramarathon footrace and compared with results for normonatremic runners [NN; n = 18, mean postrace plasma sodium concentration, 138.2 +/- 1.2 (SE) mM]. Estimated fluid intake during the race for HN was 12.5 +/- 1.6 (SE) liters over 9 h 41 min (+/- 28 min). HN excreted a net fluid excess of 2.95 +/- 0.56 (range 1.2-5.9) liters compared with a fluid deficit of 2.7 +/- 0.3% body weight in NN. The sodium deficit was 153 +/- 35 mmol in HN and 187 +/- 37 mmol in NN. Despite the fluid overload, plasma volume was decreased by 24.1 +/- 5.0% in HN compared with 8.2 +/- 2.6% in NN. Serum renin activity (5.1 +/- 2.0 ng.ml-1.h-1), aldosterone concentrations (410 +/- 34 ng/l), creatinine clearances (174.8 +/- 28.2 ml/min), and urine output (6.4 +/- 1.0 ml/min) were markedly elevated in HN during recovery. Thus the hyponatremia of exercise results from fluid retention in subjects who ingest abnormally large fluid volumes during prolonged exercise.

Adult↗
PubMed1991

Lymphokines, including interleukin-2, alter gonadotropin-stimulated progesterone production and proliferation of human granulosa-luteal cells in vitro.

The effects of human interleukin-1 (IL-1) and IL-2 on human granulosa-luteal cell progesterone production were examined with or without hCG stimulation in vitro. Human granulosa-luteal cells were recovered from follicular fluid obtained from women undergoing in vitro fertilization procedures and cultured for up to 7 days before supernatant progesterone level measurement. Lymphokine-rich conditioned medium was prepared from mitogen-stimulated human peripheral blood leukocytes (HPL-CM). The influence of HPL-CM on both granulosa-luteal cell progesterone production and cell growth was inhibitory. In contrast, supernatants of the IL-2-producing cell line MLA-144 (MLA-CM) stimulated both basal progesterone secretion and cell proliferation. Human recombinant IL-2 (from 0.1-100 IU) alone did not change progesterone levels, compared to control values, after 24 h of cell culture. However, 1, 10, and 100 IU IL-2 significantly inhibited progesterone secretion from cells stimulated by 5 IU hCG (P less than 0.01). The enhanced progesterone levels stimulated by forskolin were also significantly inhibited by 10 IU IL-2 (P = 0.01). This effect was not mediated through decreased cAMP, since the forskolin-enhanced cAMP level was not influenced by IL-2, IL-1, with or without hCG, did not show any effect on progesterone production during either 24 or 48 h of cell culture. It is concluded that 1) human recombinant IL-2 significantly inhibits progesterone production stimulated by hCG in human granulosa-luteal cells; 2) IL-2 also had a marked inhibitory effect on forskolin-induced progesterone release, but did not influence the increased cAMP level stimulated by forskolin; 3) the inhibitory influence of IL-2 on progesterone synthesis may be down-stream in the signal transduction pathway from cAMP activation; and 4) HPL-CM and MLA-CM produced inhibitory and stimulatory effects, respectively, on both basal and hCG-stimulated progesterone levels as well as on granulosa-luteal cell proliferation. These activities cannot be completely attributed to IL-2, and other mediators of leukocyte origin may, therefore, exist.

Cell Division↗
PubMed1991

Stability of immunoreactive beta-core fragment of hCG.

The beta core fragment of hCG (beta C-hCG) accounts for a large proportion of hCG immunoreactivity in the urine of pregnant women. It is often increased in the urine of patients with gynecologic tumors and may become an important diagnostic tool in early pregnancy and cancers. Despite the importance of beta C-hCG, little is known about its stability in urine under conditions of differing pH and temperature. This study examined the effect of repeated freeze-thaw cycles; storage at room temperature, 4C, and -20 C over several months; and the effect of alteration of urine pH. The two specific immunoassays for beta C-hCG used do not significantly cross-react with intact hCG or the free alpha subunit. Despite different cross-reactivities of the antibodies to the free beta subunit and higher immunoactivity when the radioimmunoassay was used, there was excellent correlation between the two assays in pregnancy urine. This suggests that there is little free beta subunit in urine from pregnant women. In addition, this study evaluated the stability of intact hCG and beta-hCG under identical conditions. No alterations in their immunoactivities were found under most conditions of storage. It is concluded that, for clinical purposes, beta C-hCG as well as intact hCG and free beta subunit are very stable molecules.

Chorionic Gonadotropin↗
PubMed1991

Choriogonadotropin-mediated thyrotoxicosis in a man.

A 38-year-old man with a metastatic gonadotropin-secreting tumor of unknown primary origin presented with both clinical and biochemical findings of hyperthyroidism in association with markedly increased concentrations of human choriogonadotropin (hCG) in plasma. After chemotherapy, the concentrations of both hCG and free thyroxin decreased and the patient became euthyroid. We discuss the rare occurrence of this presumably hCG-driven hyperthyroidism in men and compare it with the relatively more common eumetabolic hyperthyroidism associated with choriocarcinoma in women.

Adult↗
PubMed1990

The effect of a brain lesion on bone healing.

Using a feline model, the effects of stereotaxic brain lesions producing unilateral spasticity on bilateral fibular osteotomies were observed. Healing in spastic limbs occurred with more new bone formation than on the contralateral (control) side. Union appeared to be slightly faster in spastic limbs. Histomorphometric analysis of resected fibular specimens following union revealed little difference between spastic and non-spastic limbs. When these brain-lesioned animals were compared with a group with identical osteotomies but no brain lesions in respect of various systemic biochemical and endocrine parameters, no differences were found.

Animals↗
PubMed1990

Increased and intermittent prostaglandin release from amnion detected by a new superfusion technique for full thickness fetal membrane.

Prostaglandin E2 (PGE) and F2 alpha (PGF) release by the intact fetal membranes is described using a novel superfusion technique allowing for the independent assessment of prostaglandin release from the amnion and chorio-decidua whilst maintaining the anatomical integrity of the fetal membranes. The effect of labour on prostaglandin release is described. Using this system it was confirmed that the amnion is a major site of prostaglandin release and possibly production. Labour resulted in a significant increase of both PGE and PGF release from the amnion side only (Pre-labour: PGE 918 pg/cm2/3h, PGF 370 pg/cm2/3h; Labour: PGE 2993 pg/cm2/3h, PGF 662 pg/cm2/3h). No change in either PGE or PGF release from the chorio-decidual side was observed in relation to labour. In addition a change in the pattern of prostaglandin release from the amnion was observed in tissues obtained after the onset of labour. In 6 of 8 samples obtained after spontaneous labour an intermittent or pulsatile release of both PGE and PGF was observed from the amnion side as compared to the steady state of prostaglandin release from all 10 samples obtained before labour.

Amnion↗
PubMed1990

Detection of a small molecular species of human chorionic gonadotropin in the urine of patients with carcinoma of the cervix and cervical intraepithelial neoplasia: comparison with other assays for human chorionic gonadotropin and its fragments.

A low-molecular-weight glycoprotein containing sequences of the beta subunit of human chorionic gonadotropin (hCG) has been found in the urine of patients with carcinoma of the cervix using an immunoradiometric assay. This fragment has chromatographic and immunological identity with hCG beta core. This molecule was present in 52 to 77% of all patients with invasive disease, while between 11 and 27% of patients with cervical intraepithelial neoplasia (CIN) also exhibited significant hCG beta-core immunoactivity. Few patients had either a positive assay for intact hCG or a positive assay directed at an epitope on the beta subunit (beta-hCG radioimmunoassay) in serum. However, between 17 and 40% of patients with invasive disease were positive for free beta-subunit immunoactivity in the blood. The origin of the beta-core immunoactivity in the urine is uncertain; while tumor production cannot be excluded, it is possible that the molecule originates from renal metabolism of small quantities of the beta subunit of hCG. Regardless of the source of the molecule, hCG beta core is a far more sensitive marker of hCG production by tumors than is serum hCG.

Carcinoma↗