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Biomedical subjects

R Webb

Publications and source records attributed to R Webb.

At least 91 records · Page 5Linked to original sources
PubMed1999

Ipratropium bromide nasal spray 0.03% and beclomethasone nasal spray alone and in combination for the treatment of rhinorrhea in perennial rhinitis.

BACKGROUND: Perennial rhinitis is a common condition that affects up to 10% to 20% of the population. Multiple agents are frequently administered since no single agent provides complete relief. Studies assessing the benefit/risk of combined therapy are important especially for newly approved agents such as ipratropium bromide nasal spray 0.03%, a topical anticholinergic agent, approved specifically for the treatment of rhinorrhea in allergic and non-allergic perennial rhinitis. OBJECTIVE: To compare the efficacy and safety of the combined use of ipratropium bromide nasal spray 0.03% (42 microg per nostril tid) and beclomethasone dipropionate nasal spray (84 microg per nostril bid) against that of either active agent alone for the treatment of rhinorrhea. DESIGN: Multicenter, 6-week, double-blind, randomized active- and placebo-controlled, parallel trial. SETTING: Allergist and general practitioner clinical practices. PATIENTS: Five hundred thirty-three patients with perennial rhinitis (279 allergic and 274 non-allergic), 8 to 75 years of age, who had at least a mild degree of severity of rhinorrhea for a minimum of 2 hours per day during the 1 week screening period as well as congestion or sneezing also of at least mild severity. INTERVENTION: Either (1) ipratropium bromide nasal spray 0.03% (42 microg per nostril tid) plus beclomethasone dipropionate nasal spray (84 microg per nostril bid), (2) ipratropium bromide nasal spray 0.03% (42 microg per nostril tid) alone, (3) beclomethasone dipropionate nasal spray (84 microg per nostril bid) alone, or (4) vehicle [matching placebo nasal spray for the ipratropium bromide (2 sprays per nostril tid)] or beclomethasone dipropionate (2 sprays per nostril bid). MAIN OUTCOME MEASURE: Severity and duration of rhinorrhea, and patient and physician global assessment of control of rhinorrhea. RESULTS: Ipratropium bromide nasal spray plus beclomethasone nasal spray was more effective than either active agent alone or vehicle in reducing the average severity and duration of rhinorrhea during 4 weeks of treatment. The advantage of ipratropium bromide plus beclomethasone nasal spray was evident by the first day of combined treatment and continued throughout the 2-week treatment period. Ipratropium bromide nasal spray had a faster onset of action during the first week of treatment and reduced the duration of rhinorrhea more than beclomethasone. Beclomethasone nasal spray was more effective in reducing the severity of congestion and sneezing than ipratropium. In patients who had not responded well to a nasal steroid prior to participation in the study based on a questionnaire administered at screening, ipratropium bromide was as effective in the steroid non-responders as steroid responders, whereas beclomethasone was more effective in steroid responders. Combined active therapy was well tolerated with no increase in adverse events over that seen previously with ipratropium bromide or beclomethasone nasal spray alone. CONCLUSIONS: The combined use of ipratropium bromide nasal spray with beclomethasone dipropionate nasal spray is more effective than either active agent for the treatment of rhinorrhea, and does not result in a potentiation of adverse drug reactions. Ipratropium bromide nasal spray 0.03% alone should be considered in patients for whom rhinorrhea is the primary symptom, and its use in combination with a nasal steroid should be considered in patients where rhinorrhea is one of the predominant symptoms, or in patients with rhinorrhea not fully responsive to other therapy.

Administration, Inhalation↗
PubMed1999

Social class and census-based deprivation scores: which is the best predictor of stillbirth rates?

This study investigates whether social class or a census-based deprivation score is a better predictor of stillbirth rates using data for 1993-5 for residents of South Thames (West) Region. Social class is routinely coded for 10% of live births and 100% of stillbirths. A Townsend deprivation score was assigned to each stillbirth and each live birth with a social class code, according to their electoral ward of residence. In unifactorial analyses of stillbirth rate the relationship was stronger with social class (P = 0.008) than with Townsend score (P = 0.11). Both relationships were strengthened by including those births recorded as social class 'other' ['other' vs. social class I odds ratio (OR) = 2.27, P < 0.001; lower vs. upper septile deprivation score OR = 1.45, P = 0.07)]. When social class and Townsend score were analysed together, the ORs for social class remained similar to before, but the Townsend ORs were reduced and non-significant overall. We conclude that social class, which is based on data on each individual, is a better predictor of stillbirth than Townsend score, which is based on data from the area of residence. We recommend further investigation of the stillbirth risk in the subgroups that make up the 'other' social class.

Analysis of Variance↗
PubMed1999

Long-term ovarian function in sheep after ovariectomy and transplantation of autografts stored at -196 C.

We have previously demonstrated that ovarian function and fertility can be preserved in sheep after castration by autotransplantation of cryopreserved strips of ovarian cortex. In the current experiments we have investigated the long term survival of such grafts by detailed measurements of ovarian function for a period of nearly 2 yr after autotransplantation. After ovariectomy and transplantation of frozen/thawed grafts, the concentrations of FSH and LH rose to castrate levels for about 14 weeks before falling gradually to reach near-normal levels at about 60 weeks. In the breeding season from October 1994 to March 1995, all ewes had 5-10 estrous cycles that were similar in length to those in the 4 control ewes. Luteal function as indicated by the progesterone concentration was identical before and 11 months after transplantation. In contrast, the basal concentrations of FSH and LH were persistently raised throughout the luteal phase, but showed a normal decline during the follicular phase. The concentration of inhibin A in ovarian venous plasma measured at the end of the experiment 22 months after transplantation was significantly lower than that in control ewes (mean +/- SE, 409 +/- 118 vs. 1914 +/- 555 pg/ml; P < 0.004). Transplantation of frozen/thawed ovarian tissue to SCID mice demonstrated that about 28% of primordial follicles survived the procedure. All of the ovaries transplanted into sheep contained large antral follicles and/or cysts, but very few primordial oocytes when recovered at autopsy after 22 months. These results demonstrate that despite a drastic reduction in the total number of primordial follicles, cyclical ovarian function is preserved in sheep after autotransplantation of frozen/thawed ovarian tissue and provide experimental confirmation that such a technique could provide a means of preserving fertility in women undergoing chemo- or radiotherapy for malignant disease.

Animals↗
PubMed1999

Control of early cardiac-specific transcription of Nkx2-5 by a GATA-dependent enhancer.

The homeobox gene Nkx2-5 is the earliest known marker of the cardiac lineage in vertebrate embryos. Nkx2-5 expression is first detected in mesodermal cells specified to form heart at embryonic day 7.5 in the mouse and expression is maintained throughout the developing and adult heart. In addition to the heart, Nkx2-5 is transiently expressed in the developing pharynx, thyroid and stomach. To investigate the mechanisms that initiate cardiac transcription during embryogenesis, we analyzed the Nkx2-5 upstream region for regulatory elements sufficient to direct expression of a lacZ transgene in the developing heart of transgenic mice. We describe a cardiac enhancer, located about 9 kilobases upstream of the Nkx2-5 gene, that fully recapitulates the expression pattern of the endogenous gene in cardiogenic precursor cells from the onset of cardiac lineage specification and throughout the linear and looping heart tube. Thereafter, as the atrial and ventricular chambers become demarcated, enhancer activity becomes restricted to the developing right ventricle. Transcription of Nkx2-5 in pharynx, thyroid and stomach is controlled by regulatory elements separable from the cardiac enhancer. This distal cardiac enhancer contains a high-affinity binding site for the cardiac-restricted zinc finger transcription factor GATA4 that is essential for transcriptional activity. These results reveal a novel GATA-dependent mechanism for activation of Nkx2-5 transcription in the developing heart and indicate that regulation of Nkx2-5 is controlled in a modular manner, with multiple regulatory regions responding to distinct transcriptional networks in different compartments of the developing heart.

Animals↗
PubMed1999

Immune cells and cytokine production in the bovine corpus luteum throughout the oestrous cycle and after induced luteolysis.

Immune cells and their cytokine products have powerful local effects within body tissues. There has been great interest in the potential role of these cells, not only during destruction of the corpus luteum but also during its functional lifespan. In this study, lymphocytes, macrophages and major histocompatibility complex class II molecules were quantified using immunohistochemistry and the reverse transcription-polymerase chain reaction was used to detect mRNA for tumour necrosis factor alpha and interferon gamma within corpora lutea from three groups of cows: (1) corpora lutea collected at an abattoir and assessed visually into four stages (stage I (days 1-5), stage II (days 6-12), stage III (days 13-18) and stage IV (days 19-21) of the oestrous cycle); (2) corpora lutea collected around natural luteolysis (days 14-20); and (3) corpora lutea collected 6, 12 and 24 h after prostaglandin F 2 alpha-induced luteolysis. The numbers of T lymphocytes (CD5+ and CD8+) were significantly higher (P < 0.05) at stage IV and from day 16 onwards, before functional luteolysis. There were significantly higher numbers (P < 0.01) of macrophages at stages I, III and IV compared with stage II in visually staged tissue. Major histocompatibility complex class II molecules were increased (P < 0.05) at stages I and IV compared to stage II and at all times after induced luteolysis. Using reverse transcription-polymerase chain reaction, mRNA encoding tumour necrosis factor alpha and interferon gamma was detected in all luteal tissue collected around natural luteolysis and after induced luteolysis. These findings, particularly the increase in T lymphocytes before functional luteolysis, provide further evidence of a significant role for the immune system in affecting reproductive function in cows.

Animals↗
PubMed1999

Molecular mechanisms regulating follicular recruitment and selection.

Ovarian follicular growth and development is an integrated process encompassing both extraovarian signals, such as gonadotrophins and metabolic hormones, and intraovarian factors. Follicular development has been classified into gonadotrophin-independent and -dependent phases. In the latter, FSH provides the primary drive for follicular recruitment and LH is required for continued development of follicles to the preovulatory stage. A transient increase in circulating FSH precedes the recruitment of a group of follicles, and these recruited follicles are characterized by expression of mRNAs encoding P450scc and P450arom in granulosal cells. As follicles mature, there is a transfer of dependency from FSH to LH, which may be part of the mechanism(s) involved in selection of follicles for continued growth. Indeed, changes in the pattern of expression of mRNA for gonadotrophin receptors and steroid enzymes within follicular cells appear to be closely linked to changes in peripheral concentrations of gonadotrophins. The mechanism of selection of dominant follicles still requires clarification, but seems to be linked to the timing of mRNA expression encoding LHr and 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) in granulosal cells. Additional intraovarian systems, including the ovarian IGF and activin/inhibin systems, also exert a role. For example, it appears that the development of follicular dominance in cows is associated with the FSH-dependent inhibition of the expression of mRNA encoding insulin-like growth factor binding protein 2 (IGFBP-2) in granulosal cells. In conclusion, the integration of these endocrine signals and intraovarian factors within follicles determines whether follicles continue to develop and become dominant or are diverted into apoptotic pathways leading to atresia.

3-Hydroxysteroid Dehydrogenases↗
PubMed1998

Investigation of the relationship between farrowing environment, sex steroid concentrations and maternal aggression in gilts.

Maternal oestrogen and progesterone have been shown to be important in the initiation of maternal behaviour. Thirty-three Large White x Landrace gilts, housed in groups during pregnancy, were observed and aggressive interactions recorded. Individuals had jugular catheters implanted 14.5 (s.e. 0.34) days before their expected parturition date (EPD). Five days before EPD gilts were randomly allocated and moved to either a conventional farrowing crate (C; without straw, 16 gilts) or a pen (P; 2.1 x 3.1 m2; with straw bedding, 17 gilts). Blood samples were taken at frequencies determined by the proximity to farrowing onset. Piglets were removed at birth and returned 2 h after placental expulsion. The reaction of each gilt to her piglets was monitored. Gilts savaging piglets were sedated with azaperone (n = 8). There was no overall effect of farrowing environment on oestradiol and progesterone concentrations. The pre-farrowing ratio of progesterone to oestradiol was higher for (P) gilts (0.45 vs. 0.25, (P) vs. (C); S.E.D. 0.085, P < 0.05) as was their overall maximum oestradiol level (3.39 vs. 2.29 ng/ml, (P) vs. (C); S.E.D. 0.39, P < 0.01). In contrast to progesterone, oestradiol patterns varied considerably between individuals. Dominance rank value during pregnancy, but not levels of aggression, correlated positively to pre-farrowing oestradiol concentrations. Treatment with azaperone was not related to farrowing environment, piglet weight or litter size. Azaperone treated gilts showed a higher pre-farrowing oestradiol to progesterone ratio (0.55 vs. 0.29, +/- azaperone; S.E.D. 0.10, P < 0.05), significantly higher levels of oestradiol post-partum (0.7 vs. 0.19 ng/ml, +/- azaperone; S.E.D. 0.20, P < 0.001) and significantly lower levels of aggression during pregnancy (1.68 vs. 2.23 aggressive interactions/h, +/- azaperone; S.E.D. 0.15, P < 0.001). The results indicate that there are no major effects of farrowing environment on sex steroid concentrations. Maternal aggression under these conditions appears to be negatively related to aggression during pregnancy, but this is not reflected in plasma concentrations of sex steroids around parturition.

Aggression↗
PubMed1998

Specific non-genomic, membrane-localized binding sites for progesterone in the bovine corpus luteum.

Fractionation of bovine corpus luteum (CL) homogenates on continuous sucrose density gradients with and without preincubation with 3H-progesterone demonstrated high levels of tracer binding and high content of endogenous progesterone associated with particulate membrane fractions. Analysis of gradient fractions for a range of luteal plasma membrane and intracellular organelle marker enzyme activities indicated that endogenous progesterone content and 3H-progesterone-binding activity were associated with fractions enriched in luteal plasma membrane markers. This was confirmed by pretreatment of homogenates with the saponin, digitonin, prior to fractionation. Digitonin perturbed the buoyant density of luteal surface membrane markers and 3H-progesterone binding to a similar extent, but did not perturb the buoyant densities of other intracellular markers to the same degree. Interestingly, digitonin pretreatment also increased the proportion of progesterone tracer that entered the gradients. We consistently failed to demonstrate significant binding of 3H-progesterone to membrane fractions incubated with progesterone tracer in vitro. However, when digitonin was included in the in vitro binding assay, we observed a dramatic, dose-dependent stimulation of 3H-progesterone binding by digitonin. Other radiolabeled steroids tested (3H-cortisol, 3H-testosterone) bound poorly in the presence or absence of digitonin. 3H-Progesterone binding in the presence of optimal digitonin concentrations increased linearly with increasing luteal membrane concentration; was dependent on the pH, duration, and temperature of incubation; and low levels of progesterone (68 nM) competed for tracer binding. A range of other steroids tested (androgens, estrogens, corticosteroids, steroid precursors) competed at higher concentrations (10- to 100-fold) or did not compete at all for 3H-progesterone binding. There was no correlation between the hydrophobicity of various steroids and their ability to compete for binding. Moreover, a number of agonists and antagonists specific for the genomic progesterone receptor, agonists of peripheral benzodiazepine receptors, and inhibitors of a range of steroidogenic enzymes did not compete for 3H-progesterone binding. Western blots confirmed that detergent-solubilized progesterone-binding sites could be resolved from cytochrome P450 side-chain cleavage and 3beta-hydroxysteroid dehydrogenase. Moreover, extraction of bound steroid from the binding site and HPLC demonstrated identity to progesterone, suggesting that no metabolism of the progesterone tracer had occurred during incubation. Progesterone binding to membranes of large luteal cells was higher compared with binding to small luteal cells, and levels were similar in membranes prepared from CL at all stages of the luteal phase. We suggest that bovine luteal progesterone-binding sites may play a role either in sequestration of newly synthesized progesterone or in the mediation of autocrine and/or paracrine actions of progesterone in the CL.

Animals↗
PubMed1998

Expression of monocyte chemoattractant protein-1 in the bovine corpus luteum around the time of natural luteolysis.

Monocyte chemoattractant protein (MCP-1) is a specific chemoattractant for monocytes/macrophages that could have a role in the influx of macrophages into the corpus luteum (CL) during structural luteolysis. In this study, reverse transcription-polymerase chain reaction and in situ hybridization were used to investigate MCP-1 mRNA expression in CL collected from 18 heifers between Days 15 and 20 of the estrous cycle. There was expression of mRNA encoding MCP-1 in luteal tissue from all cows; however, expression was greater in animals that had undergone luteolysis at the time of CL collection as compared to animals in which the CL was still functional. Similarly, in situ hybridization showed greater expression of mRNA encoding MCP-1 in CL after functional luteolysis. There was also evidence of increased MCP-1 mRNA expression in an animal with a functional CL where the systemic concentration of prostaglandin F2alpha metabolite was high at the time of tissue collection. T lymphocyte populations, identified by immunohistochemistry, had a distribution similar to that of cells expressing MCP-1 mRNA within the CL, but other cell types were also involved. These results demonstrate an increase in MCP-1 mRNA after functional luteolysis in the cow, which may be related to the influx of macrophages that occurs at this time.

Animals↗
PubMed1998

Insulin-like growth factor binding protein -2 and -4 messenger ribonucleic acid expression in bovine ovarian follicles: effect of gonadotropins and developmental status.

This work is concerned with the role of insulin-like growth factor binding protein (IGFBP)-2 and -4 in the regulation of IGF bioactivity in bovine follicles during the development of dominance. We measured the expression of IGFBP-2 and -4 messenger RNA (mRNA) in small (1-4 mm) gonadotropin-sensitive follicles and medium (4-8 mm) and large (>8 mm) gonadotropin-dependent follicles using in situ hybridization. In healthy nonatretic bovine follicles, IGFBP-2 and -4 mRNA expression was confined to granulosa and theca tissue, respectively. Moreover, during the development of follicular atresia, there were distinct changes in the temporal and spatial expression of these genes. IGFBP-2 immunoactivity was localized in granulosa tissue and the basement membrane of healthy preantral follicles, whereas IGFBP-4 immunoactivity was localized in both theca and granulosa tissue. Of particular interest was the lack of IGFBP-2 mRNA expression in large (>8 mm) gonadotropin-dependent follicles, an observation that was confirmed by the lack of immunoreactive IGFBP-2 in these follicles. The regulation of IGFBP-2 and -4 mRNA expression in granulosa and theca cells was analyzed using a serum-free cell culture system. FSH inhibited the expression of IGFBP-2 mRNA in granulosa cells, whereas LH stimulated IGFBP-4 mRNA expression in theca cells. Our results provide evidence for the existence of different roles for IGFBP-2 and -4 in the developing follicle.

Animals↗
PubMed1998

Requirement of the MADS-box transcription factor MEF2C for vascular development.

The embryonic vasculature develops from endothelial cells that form a primitive vascular plexus which recruits smooth muscle cells to form the arterial and venous systems. The MADS-box transcription factor MEF2C is expressed in developing endothelial cells and smooth muscle cells (SMCs), as well as in surrounding mesenchyme, during embryogenesis. Targeted deletion of the mouse MEF2C gene resulted in severe vascular abnormalities and lethality in homozygous mutants by embryonic day 9.5. Endothelial cells were present and were able to differentiate, but failed to organize normally into a vascular plexus, and smooth muscle cells did not differentiate in MEF2C mutant embryos. These vascular defects resemble those in mice lacking the vascular-specific endothelial cell growth factor VEGF or its receptor Flt-1, both of which are expressed in MEF2C mutant embryos. These results reveal multiple roles for MEF2C in vascular development and suggest that MEF2-dependent target genes mediate endothelial cell organization and SMC differentiation.

Animals↗
PubMed1998

Effects of dose of LH on androgen production and luteinization of ovine theca cells cultured in a serum-free system.

The study reports the development of a serum-free culture system for sheep thecal cells that overcomes the problem of spontaneous luteinization and the use of this system to study the control of proliferation and differentiation. Theca cells were isolated by enzymatic dispersion from small follicles (< 3.5 mm) and the effect of plating densities (25-100 x 10(3) cells per well), LH (0.001-100 micrograms l-1), insulin (1-5000 micrograms l-1), insulin-like growth factor I (IGF-I) analogue (1-100 micrograms LR3-IGF-I l-1) and epidermal growth factor (EGF) (0.005-50 micrograms l-1) on the number of cells and androstenedione and progesterone production were determined. Plating density had a marked effect on the pattern of hormone secretion with densities between 50 and 75 x 10(3) cells per well resulting in a high androstenedione: progesterone ratio at optimum doses of LH (0.1 micrograms l-1: P < 0.001). In the first 48 h, the production of both androstenedione and progesterone was stimulated in a dose-dependent manner by LH (P < 0.001). However, the production of androstenedione was ten times higher than that of progesterone and was more sensitive to LH (ED50 value 0.08 micrograms l-1 for androstenedione and 1 microgram l-1 for progesterone). From 48-144 h of culture higher doses of LH (> 1 ng ml-1) inhibited androstenedione (P < 0.001) and stimulated progesterone (P < 0.001) and resulted in a marked change in cell morphology, thus reflecting both functional and morphological luteinization. At optimum doses of LH, both insulin and IGF stimulated cell proliferation (P < 0.001) and androstenedione production (P < 0.001) in a dose responsive manner and there was a significant (P < 0.001) interaction between them. In contrast, both insulin and IGF-I inhibited (P < 0.001) progesterone production in a dose responsive manner. EGF stimulated cell proliferation (P < 0.001) and progesterone production (P < 0.001), but inhibited androstenedione production (P < 0.001), in a dose responsive manner. In conclusion, this culture system exhibits physiologically relevant responses to known in vivo modulators of follicle development. The biphasic nature of the theca cell response to LH emphasises the exquisite sensitivity of theca cells to LH stimulation and highlights the importance of dose-response relationships in the gonadotrophic control of ovarian function.

Androstenedione↗
PubMed1997

Ultra-structural characteristics of bovine granulosa cells associated with maintenance of oestradiol production in vitro.

We investigated whether the maintenance of oestradiol production by bovine granulosa cells (GC) in vitro was related to GC ultra-structure, and studied the effects of inclusion of serum as a cell attachment factor on oestradiol secretion, cell morphology and ultra-structure. Bovine granulosa cells from medium-sized follicles (4-8 mm diameter), in a serum-free (SF) culture system, maintained oestradiol production for 6 days, whereas oestradiol secretion by cells cultured in serum-coated (SC) wells declined rapidly with time, in culture. SF cells formed clumps consisting of two types of cells. Cells within clumps presented a phenotype similar to GC in vivo, being spherical, tightly joined by extensive gap junctions and interdigitated pseudopodia/microvilli, had abundant rough and smooth endoplasmic reticulum (ER) and mitochondria with trabecular cristae. In contrast, cells cultured in either SC wells or in the flattened base of cell clumps from SF cultures were enlarged, containing less rough ER, had fewer mitochondria (which tended to be round) and contained endosome-like structures, morphological characteristics suggestive of early luteinisation.

Animals↗
PubMed1997

CD44H expression in reactive mesothelium, pleural mesothelioma and pulmonary adenocarcinoma.

Malignant mesotheliomas are known to produce hyaluronic acid, in contrast to most pulmonary adenocarcinomas which produce neutral mucin. CD44H is the major cell surface receptor for hyaluronic acid. The aim of this study was to investigate immunohistochemically the expression of this antigen in reactive mesothelium, pleural mesothelioma and pulmonary adenocarcinoma and to assess its diagnostic utility in distinguishing the two tumours. Diffuse and intense membranous CD44H immunoreactivity was seen in 15 of 20 (75%) mesotheliomas and in all 20 biopsies of reactive mesothelium. In contrast, focal (< 10% tumour) expression of CD44H was seen in only three of 20 (15%) pulmonary adenocarcinomas. We advocate the use of CD44H as a positive mesothelial marker for incorporation alongside other established immunohistochemical markers used to distinguish mesothelioma from adenocarcinoma.

Adenocarcinoma↗
PubMed1997

Development of a long-term bovine granulosa cell culture system: induction and maintenance of estradiol production, response to follicle-stimulating hormone, and morphological characteristics.

The objectives of this study were to develop a serum-free bovine granulosa cell culture system in which FSH-responsive estradiol production could be induced and maintained, and to use this system to evaluate the effects of FSH, insulin, and IGF-I on steroidogenesis and proliferation of bovine granulosa cells from different follicle size categories (< 4-, 4-8, and > 8-mm diameter). In the presence of FSH, granulosa cells from small follicles differentiated in vitro, and estradiol secretion increased with time (p < 0.01) so that by the end of the culture period it was similar to that of cells from large follicles. Granulosa cells from medium and large follicles secreted estradiol throughout the culture period. Cells cultured in plasma-coated culture wells had an increased proliferative response but had lower estradiol production compared to cells cultured under serum-free conditions (p < 0.01). Insulin promoted proliferation and estradiol production by granulosa cells from the three follicle-size categories (p < 0.01). Physiological concentrations of FSH induced proliferation and estradiol secretion (p < 0.01) by granulosa cells in a dose-responsive manner. The inclusion of IGF-I in the culture system enhanced proliferation and estradiol production (p < 0.01), even in the absence of gonadotropic support, demonstrating the gonadotropic characteristics of this growth factor. These results demonstrate the development of a relevant physiological culture system for bovine granulosa cells. This system will permit the detailed study of the key factors controlling the differentiation and proliferation of bovine granulosa cells.

Animals↗
PubMed1997

Extra- and intracellular effects of divergent selection for pituitary responsiveness to gonadotropin-releasing hormone in prepubertal ram lambs.

Divergent selection based on the response of 10-wk-old male lambs to a GnRH challenge has produced two lines of sheep, referred to as high and low lines, that differ in their ability to release LH in response to pharmacological and physiological doses of GnRH. The aim of this study was to determine whether the between-line differences in pituitary sensitivity were related to differences in GnRH receptor number and/or the transduction of the intracellular signal following GnRH receptor activation. Pituitary glands were collected from fourteen 20-wk-old ram lambs from each line, weighed, and sampled for GnRH receptor analysis. The remaining tissue from 9 lambs from each line was dispersed. Of the resultant cell suspension, a sample was stored for measurement of GnRH receptor content and the remainder was plated and cultured for 24 h. The LH responses of cultured cells were measured after exposure to GnRH, A23187, or the phorbol ester phorbol 12,13 dibutyrate (PDB). The results indicated that the pituitary glands of the high line contained significantly higher concentrations of GnRH receptors than did those of the low line and released significantly more LH after stimulation with either GnRH or the Ca2(+)-calmodulin or protein kinase C intracellular second messenger systems. Therefore, the between-line difference in the regulation of pituitary LH secretion occurs at a step distal to the stimulatory sites of action of A23187 and PDB.

Animals↗
PubMed1997

Secretion of inhibin A and follicular dynamics throughout the estrous cycle in the sheep with and without the Booroola gene (FecB).

The identification of a single gene (Booroola FecB) associated with a significant increase in the ovulation rate in sheep provides a powerful tool for the study of factors regulating the selection of preovulatory follicles. The ovarian secretion of dimeric inhibin A was investigated and related to the secretion of ovarian steroids, the concentration of gonadotropins, and the pattern of ovarian follicular development during the follicular and early luteal phases in ewes with an ovarian autotransplant with or without the FecB gene. The secretion of inhibin A was related to the presence of large estrogenic follicles, being high during the follicular phase and falling after the LH surge (P < 0.05) before increasing during the early luteal phase (P < 0.05). There were no differences in the timing of the onset of the LH surge, the concentrations of FSH and progesterone, or the rates of inhibin A, estradiol, and androstenedione secretion during the follicular or luteal phase between ewes with and without the Booroola gene. Although the overall pattern of follicular development was similar between the genotypes, the ovulation rate was higher (P < 0.05) in the gene carrier ewes, and the ovulatory follicles and corpora lutea were significantly smaller (P < 0.05) than those in noncarrier animals. During the luteal phase, the diameter of the large follicles from the first wave was smaller (P < 0.05) in the gene carrier than in noncarrier ewes. Because there are no qualitative or quantitative differences in the pattern of secretion of pituitary gonadotropins or ovarian hormones between the two genotypes, we conclude that is likely that the FecB gene exerts its action at the level of ovary.

Animals↗