Search PubMed⌕ Search

Biomedical subjects

X C Jia

Publications and source records attributed to X C Jia.

At least 19 recordsLinked to original sources
PubMed1995

Macrophage inflammatory protein 1-alpha mRNA expression in an immortalized microglial cell line and cortical astrocyte cultures.

Macrophage inflammatory protein 1 (MIP-1) is a recently characterized inflammatory and chemokinetic cytokine. Proinflammatory stimuli have been shown to induce expression of MIP-1 by macrophages. We hypothesized that microglia and astrocytes express MIP-1 alpha because of their many immunologic similarities to macrophages. MIP-1 alpha mRNA was examined with quantitative reverse transcription and polymerase chain reaction in an immortalized mouse microglial cell line (BV-2) and in mouse cortical astrocyte cultures. We found that in both the BV-2 microglial cell line and in astrocyte cultures, MIP-1 alpha mRNA was strongly induced by lipopolysaccharide and the phorbol ester PMA. MIP-1 alpha mRNA was reduced by dBcAMP, interferon-gamma, and PGE1. Dexamethasone decreased MIP-1 alpha mRNA levels in astrocyte cultures, but not in BV-2 microglial cells. Interleukin-1 beta, tumor necrosis factor alpha, and MIP-1 alpha had no effect on MIP-1 alpha mRNA expression. These findings demonstrate that MIP-1 alpha mRNA is expressed by cultured glial cells and is regulated by proinflammatory and anti-inflammatory stimuli. MIP-1 alpha may be expressed by microglia and astrocytes in vivo, and may help modulate cerebral inflammation.

Animals↗
PubMed1995

Tumor necrosis factor-alpha and basic fibroblast growth factor decrease glial fibrillary acidic protein and its encoding mRNA in astrocyte cultures and glioblastoma cells.

Tumor necrosis factor-alpha is a pluripotent cytokine that is reportedly mitogenic to astrocytes. We examined expression of the astrocyte intermediate filament component glial fibrillary acidic protein in astrocyte cultures and the U373 glioblastoma cell line after treatment with tumor necrosis factor-alpha. Treatment with tumor necrosis factor-alpha for 72 h resulted in a decrease in content of glial fibrillary acidic protein and its encoding mRNA. At the same time, tumor necrosis factor-alpha treatment increased the expression of the cytokine interleukin-6 by astrocytes. The decrease in glial fibrillary acidic protein expression was greater when cells were subconfluent than when they were confluent. Thymidine uptake studies demonstrated that U373 cells proliferated in response to tumor necrosis factor-alpha, but primary neonatal astrocytes did not. However, in both U373 cells and primary astrocytes tumor necrosis factor-alpha induced an increase in total cellular protein content. Treatment of astrocytes and U373 cells for 72 h with the mitogenic cytokine basic fibroblast growth factor also induced a decrease in glial fibrillary acidic protein content and an increase in total protein level, demonstrating that this effect is not specific for tumor necrosis factor-alpha. The decrease in content of glial fibrillary acidic protein detected after tumor necrosis factor-alpha treatment is most likely due to dilution by other proteins that are synthesized rapidly in response to cytokine stimulation.

Animals↗
PubMed1993

Reverse transcription and polymerase chain reaction technique for quantification of mRNA in primary astrocyte cultures.

The reverse transcription and polymerase chain reaction technique (RT-PCR) was assessed for the quantification of changes in mRNA levels from primary astrocyte cultures. The effects of dibutyryl cyclic AMP (dBcAMP) on glial fibrillary acidic protein (GFAP) mRNA and the effects of tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), and lipopolysaccharide (LPS) on interleukin-6 (IL-6) mRNA were examined. Two quantitative PCR methods were used: one involved carrying out the reaction in the exponential phase and the other involved the coamplification of a competitive target sequence. Increased GFAP mRNA in response to chronic dBcAMP treatment and increased IL-6 mRNA in response to TNF-alpha/IL-1 beta were readily detected. Both RT-PCR techniques were found to be suitable for the detection of large as well as smaller (twofold) changes in mRNA levels. The advantages and limitations of RT-PCR for mRNA quantification are discussed.

Animals↗
PubMed1992

Expression of recombinant human follicle-stimulating hormone receptor: species-specific ligand binding, signal transduction, and identification of multiple ovarian messenger ribonucleic acid transcripts.

The ligand specificity and biochemical properties of the human (h) FSH receptor are poorly characterized due to the low abundance of these receptors and the limited availability of human tissues. Using a fragment of rat FSH receptor cDNA, we screened a human testicular cDNA library and obtained a FSH receptor cDNA covering the entire amino acid-coding region. After transfection of a human fetal kidney cell line (293) with the hFSH receptor cDNA, radioligand receptor analysis revealed the presence of high affinity (Kd, 1.7 x 10(-9) M) FSH-binding sites on the plasma membrane. Both recombinant and wild-type hFSH displaced [125I]hFSH binding, with ED50 values of 25 and 70 ng/ml, respectively, whereas hLH, hCG, and hTSH were ineffective. Although human, rat(r), and ovine FSH as well as equine CG competed for rat testicular FSH receptor binding, only hFSH and rFSH interacted effectively with the recombinant hFSH receptor, suggesting that species-specific ligand recognition exists between human and rodent receptors. After incubation of transfected cells with hFSH, but not recombinant hLH or hCG, a dose-dependent increase (ED50, 10 ng/ml) in extracellular cAMP accumulation was observed, indicating a functional coupling of the expressed human receptor with the endogenous adenyl cyclase. In cells cotransfected with the FSH receptor expression plasmid and a luciferase reporter gene driven by the promoter of a cAMP-responsive gene, treatment with hFSH, but not hCG, resulted in a dose-dependent increase in luciferase activity. Northern blot analysis using a cRNA probe derived from the human receptor cDNA indicated the presence of multiple FSH receptor mRNA transcripts (7.0, 4.2, and 2.5 kilobases) in RNA prepared from human follicular phase ovary, but not from human corpus luteum or placenta. Additionally, two FSH-binding sites of 76 and 112 kilodaltons were detected in transfected 293 cells after ligand/receptor cross-linking and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. These results demonstrate the expression of functional hFSH receptor with unique ligand specificity and provide new data on the biochemical properties of the human receptor at the mRNA and protein levels.

Animals↗
PubMed1991

Ligand-induced down-regulation of testicular and ovarian luteinizing hormone (LH) receptors is preceded by tissue-specific inhibition of alternatively processed LH receptor transcripts.

Down-regulation of plasma membrane receptors by homologous hormones has been found in diverse cell types. In testicular Leydig and ovarian luteal cells, treatment with LH/hCG decreases LH receptor content. Although suppression of LH-binding sites may result from ligand-induced receptor internalization, sequestration, and/or phosphorylation, the gonadotropins may also regulate receptor mRNA levels. We examined the regulation of testis LH receptor mRNAs in adult rats that received 10 or 200 IU hCG, using cRNA probes derived from the 5' extracellular domain (EC) or the 3' transmembrane domain (TM) of the rat receptor cDNA. Probe EC hybridized to predominant signals of 7 and 1.8 kilobases (kb) and weaker signals of 4.2 and 2.5 kb. However, probe TM hybridized to the three larger forms of the LH receptor mRNA, but not to the 1.8-kb species, suggesting that the latter form lacks the transmembrane domain. After 6 and 12 h of treatment with 200 or 10 IU hCG, respectively, hybridization to the larger mRNA species decreased by more than 60%, preceding decreases in testicular [125I]hCG binding. These transcripts were further inhibited (greater than 93%) between 24-72 h after hCG treatment and returned to 40% and 100% of control levels by days 6 and 9, respectively. In contrast, the truncated 1.8-kb LH receptor transcript was not affected by hCG treatment, indicating a differential suppressive effect of the ligand on its receptor mRNA levels. In the ovary, hybridization to probe EC revealed four transcripts with similar sizes as those found in the testes, with a predominant 7-kb species.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Expression of human luteinizing hormone (LH) receptor: interaction with LH and chorionic gonadotropin from human but not equine, rat, and ovine species.

Studies on human LH receptors are difficult due to the limited availability of clinical samples. Recent cloning of rat and porcine LH receptor cDNAs indicated that these binding sites are single polypeptides of the G-protein-coupled receptor family with seven transmembrane domains. Based on the conserved sequences of rat and porcine receptors, we performed reverse transcription polymerase chain reaction, using human ovarian mRNA as template and obtained partial human LH receptor cDNA clones. Further screening of a human ovary cDNA library and subsequent ligation of individual cDNA clones generated a human LH receptor cDNA containing the entire amino acid-coding region. Sequence analysis indicated that the human receptor cDNA displays 89% and 82% homology at the nucleotide level with its porcine and rat counterparts, respectively. A region spanning the second extracellular and third transmembrane domains is highly conserved among the human LH, FSH, and TSH receptors. The ovarian LH receptor clone is, however, significantly different from an incompletely spliced LH receptor cDNA recently obtained from a human thyroid library. Unlike the thyroid clone, the ovarian LH receptor cDNA could be expressed in the human fetal kidney cell line (293), and radioligand receptor assay identified high affinity (Kd, 1.2 x 10(-10) M) LH/hCG-binding sites on the plasma membrane. Binding specificity of the human LH receptor was studied using recombinant human CG, LH, and FSH secreted by CHO cells transfected with the respective genes. Human CG and LH displaced [125I]hCG binding with an ED50 of 4.3 and 4.8 ng/ml, respectively. In contrast, recombinant FSH was not effective. Treatment of transfected cells with recombinant gonadotropins also induced dose-dependent increases in extracellular cAMP production (hCG = LH much greater than FSH; ED50 25, 10, and greater than 3000 ng/ml). Although equine, rat, and ovine LH as well as equine CG competed effectively for rat testicular LH receptor binding, these hormones were unable to displace [125I]hCG binding to the human receptor, suggesting evolutionary changes in receptor binding specificity and the importance of using human receptors for clinical studies. Thus, the cloning and expression of the human LH receptor cDNA allowed analysis of interactions between human LH receptor and gonadotropins from diverse species. The present work should provide the basis for future design of therapeutic agents capable of interacting with the human receptor and for understanding the structural basis for LH receptor binding to different gonadotropins.

Amino Acid Sequence↗
PubMed1990

Synergistic effect of glucocorticoids and androgens on the hormonal induction of tissue plasminogen activator activity and messenger ribonucleic acid levels in granulosa cells.

Tissue-type plasminogen activator (tPA) is secreted by rat granulosa cells in response to treatment with activators of protein kinase A (follitropin, FSH), protein kinase C (gonadotropin-releasing hormone, GnRH) and tyrosine kinase (epidermal growth factor, EGF). Because steroid hormones have been shown to enhance the gonadotropin stimulation of ovarian differentiation, we investigated the effects of steroid hormones, alone or together with various kinase activators, on tPA activities and mRNA levels in cultured rat granulosa cells. Treatment of cells with dexamethasone (DEX; a glucocorticoid agonist) or R1881 (an androgen agonist) caused an increase in tPA secretion and mRNA levels. In addition, the stimulation of tPA activity and mRNA levels by FSH (50 ng/ml) was synergistically enhanced by cotreatment with DEX or R1881 in a time-dependent manner with 2.8- and 1.6-fold increase at 9 h after incubation as compared to cells treated with FSH alone. In contrast, treatment with diethylstilbestrol had no effect on tPA levels. Furthermore, tPA activity and mRNA levels induced by GnRH and EGF were also increased by cotreatment with DEX or R1881 as compared with cells treated with GnRH or EGF alone. Likewise, the stimulation of tPA mRNA levels by dibutyryl cAMP, a protein kinase A activator, and phorbol myristate acetate (PMA), a protein kinase C activator, was enhanced by cotreatment with DEX or R1881. These results demonstrate that glucocorticoid and androgen enhance tPA secretion and mRNA levels stimulated by FSH, GnRH and EGF in granulosa cells. The rat granulosa cells provide a useful model for studying the mechanism of regulation of tPA gene expression by steroid hormones.

Animals↗
PubMed1990

Gonadotropin-induced up- and down-regulation of rat ovarian LH receptor message levels during follicular growth, ovulation and luteinization.

Induction of follicular growth by PMSG is associated with increased ovarian LH receptor content, whereas the preovulatory surge of LH decreases LH binding sites, followed by a secondary increase in receptor numbers coincident with corpora lutea formation. Based on the recently reported LH receptor cDNA sequence, we have performed a reverse transcription-polymerase chain reaction to obtain LH receptor cDNA clones and generated a 32P-labeled cRNA probe to examine the dynamic changes in ovarian LH receptor mRNA levels during gonadotropin induction of follicular growth, ovulation and luteinization. Northern blot analysis of ovary RNA revealed hybridization signals of about 7.0, 4.2, 2.5 and 1.8 kb, with no hybridization to nongonadal tissues. PMSG increased the intensity of all four LH receptor messages at 24 h, preceding an increase in LH receptor number, with peak LH receptor mRNA and receptor content observed at 52 h. Treatment with hCG resulted in decreased LH receptor binding and mRNA levels by 6 h after injection, with maximal inhibition (greater than 85%) of message at 12 to 24 h after hCG treatment. Subsequently, LH receptor message levels increased again at 3 days after hCG, concomitant with increased [125I]hCG binding. A further increase in LH receptor content, but not message levels, was observed 5 days after hCG. These results demonstrate that the induction of LH receptors by PMSG is preceded by increased LH receptor mRNA levels. Furthermore, ligand-induced down-regulation of the LH receptor following an ovulatory dose of hCG is associated with decreased LH receptor message content, followed by increases in LH receptor message levels and binding sites during subsequent luteinization. Thus, the up- and down-regulation of ovarian LH receptors during follicle growth, ovulation and luteinization is probably due, at least in part, to changes in receptor message modulation.

Animals↗
PubMed1988

Bioactive follicle-stimulating hormone levels in serum and urine of male and female rats from birth to prepubertal period.

Using a sensitive in vitro granulosa cell aromatase bioassay (GAB), we determined serum and urinary levels of bioactive follicle-stimulating hormone (bio-FSH) in male and female rats from birth to Day 40 of age. In addition, serum immunoreactive FSH (immuno-FSH) was measured by radioimmunoassay to determine the bio- to immuno-(B/I) ratio of FSH. During the neonatal period (Days 1-7 of age), both sexes had detectable serum bio-FSH levels. In the infantile period (Days 7-21), serum bio-FSH levels initially decreased at Day 10 for both sexes, and then rose steadily, reaching maximum concentrations at Day 14 (males: 68.7 ng/ml; females: 114.6 ng/ml). Subsequently, FSH levels in the females decreased from Day 16 throughout the juvenile (Days 21-35) and prepubertal (Days 35-40) periods. In contrast, FSH levels in the males fluctuated during these periods. In the males, immuno-FSH reflected the bioactive profiles, with a B/I ratio of 2.2 +/- 0.2. In the females, the B/I ratio was approximately 2.5 during the neonatal and infantile periods but declined to approximately 1.0 during the juvenile and prepubertal periods, consistent with earlier observations of heterogeneous forms of pituitary FSH in immature female rats. Morning urine samples were also collected daily and bio-FSH concentrations were determined. In both sexes, urinary bio-FSH profiles were highly correlated (r = 0.93) with serum FSH throughout development. However, the urine concentrations were about 50-fold higher than serum.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1986

Growth hormone enhances follicle-stimulating hormone-induced differentiation of cultured rat granulosa cells.

Suppression of serum GH levels in immature rats is associated with delayed onset of puberty and decreased ovarian steroidogenic responsiveness to FSH. To investigate possible direct effects of GH on the differentiation of ovarian cells, granulosa cells from hypophysectomized estrogen-treated rats were cultured with FSH in the presence or absence of GH for 3 days. FSH stimulated granulosa cell LH receptor formation and steroid production in a dose-dependent manner. Concomitant treatment with GH increased LH receptor content by enhancing the action of low doses of FSH without substantial increases in the maximal response. This increase was due to an elevation in the receptor number rather than changes in their affinity for hCG. At 3 ng/ml FSH, concomitant treatment with ovine or bovine GH increased LH/hCG binding in a dose-dependent manner, with 300 ng/ml GH increasing the FSH action by about 3-fold. LH receptors in the GH-treated cells were functional, as indicated by the enhanced cAMP production of these cells in response to LH treatment. The cellular protein content in the FSH-treated cultures was slightly increased by GH (18%), but cell number and viability were unaffected. The change in cell protein content could not account for the increases in the amount of LH receptors. In addition to its effects on LH/hCG receptor content, GH also augmented FSH-stimulated progesterone and 20 alpha-hydroxy-4-pregnen-3-one production in a dose-dependent manner, with 100 ng/ml GH causing significant increases in FSH-induced progesterone production. In contrast, GH treatment did not significantly affect FSH-stimulated estrogen production. The augmentating effects of GH on LH receptor formation and progestin biosynthesis were associated with an enhancement of FSH-stimulated cAMP production. In addition, GH increased forskolin- and 8-bromo-cAMP-induced LH receptor formation and progestin production. Thus, GH-augmented LH receptor induction and progestin biosynthesis may be due to both increased cAMP production and enhanced action of cAMP. The present data have demonstrated that GH augments gonadotropin-stimulated differentiation of ovarian granulosa cells, suggesting an important regulatory role of GH in follicular growth and pubertal development.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1986

Granulosa cell aromatase bioassay for follicle-stimulating hormone: validation and application of the method.

Stimulation of rat granulosa cell aromatase activity by FSH has recently been used as a sensitive biological end point to develop an in vitro FSH bioassay. The present report provides a detailed validation and application of this assay. In the presence of androstenedione and diethylstilbestrol, FSH stimulated estrogen production in a dose-dependent manner. Although addition of high doses of a phosphodiesterase inhibitor [1-methyl-3-isobutyl xanthine (MIX)] decreased maximal estrogen production, treatment with 0.125 mM MIX increased the sensitivity of granulosa cells to FSH, presumably by minimizing endogenous cAMP breakdown. Addition of insulin and human CG (hCG) further synergistically enhanced granulosa cell sensitivity to FSH. Although inclusion of gonadotropin-free serum obtained from hypophysectomized male rats decreased the assay sensitivity, pretreatment of serum with polyethylene glycol [(PEG) 10-14%] resulted in a dose-dependent decrease in the serum-interfering effect. Studies using exogenous [125I]iodo-rat FSH or RIA measurement indicated recovery of 94-98% FSH after pretreatment of serum with 12% PEG. In the presence of the PEG-pretreated gonadotropin-free serum (4%), ovine, rat, and human FSH preparations induced parallel dose-response curves for estrogen production with minimal detectable doses of 0.12 ng, 0.12 ng, and 0.12 mIU/culture, respectively. In contrast, treatment with GH, PRL, TSH, and ACTH did not affect estrogen production. The apparent stimulatory effect of high doses (greater than 60 ng/culture) of LH and hCG could be attributed to FSH contamination or intrinsic FSH activity in these preparations. Changes in serum bioactive FSH levels were studied in adult male rats after GnRH administration. GnRH (5 micrograms/rat) treatment significantly elevated FSH levels within 30 min after injection. Maximal increases (approximately 2.8-fold) in serum bioactive FSH were observed between 60-120 min. At 8 h after treatment, FSH levels decreased to control levels. Comparison between granulosa cell aromatase bioassay and RIA results indicated no apparent changes in the bio- to immuno- ratio of FSH after GnRH treatment. In conclusion, extreme sensitivity of the bioassay allows the measurement of circulating levels of bioactive FSH. Since rat granulosa cells respond to FSH preparations from different species, the in vitro assay should also provide valuable information on FSH levels in many animal species including those lacking a specific RIA. Measurement of serum levels of bioactive FSH should provide insight regarding the role of FSH in various physiological and pathological conditions.

1-Methyl-3-isobutylxanthine↗
PubMed1986

Serum bioactive follicle-stimulating hormone during the human menstrual cycle and in hyper- and hypogonadotropic states: application of a sensitive granulosa cell aromatase bioassay.

A sensitive in vitro assay based on the stimulation of estrogen production by cultured rat granulosa cells was recently developed for the measurement of biologically active FSH. This bioassay system is specific for FSH, highly sensitive, and capable of measuring basal FSH levels in rat serum. The granulosa cell aromatase bioassay was improved by the use of additives known to enhance FSH activity and by pretreatment of serum with 12% polyethylene glycol to remove inhibitory substances. We applied this method to the measurement of bioactive FSH levels in serum samples from human subjects. As determined in daily blood samples during ovulatory menstrual cycles in seven women, bioactive FSH levels exhibited a pattern closely resembling that of immunoreactive FSH. The mean bioactive serum FSH levels were 29.9, 20.5, 39.2, and 14.8 mIU/ml for the early follicular phase, late follicular phase, preovulatory surge, and luteal phase, respectively. The bio- to immunoratio (B:I) throughout the menstrual cycle ranged from 1.4-3.4, with a mean of 2.5. The ratios for early follicular phase, late follicular phase, preovulatory surge, and luteal phase were 2.7, 2.3, 1.4, and 2.6, respectively. The correlation coefficient (r) of the serum FSH values obtained by bioassay and RIA was 0.91. FSH bioactivity was also measured in patients in each of the following categories with the following mean values: oral contraceptive pill users (undetectable), hypothalamic amenorrhea (18.7 mIU/ml; B:I, 2.6), premature ovarian failure (163 mIU/ml; B:I, 1.7), and postmenopausal women (191 mIU/ml; B:I, 1.6). These findings suggest that measurement of immunoreactive FSH levels correctly reflects the biological activity of FSH in serum of cycling women and patients in certain hyper- and hypogonadotropic states. The granulosa cell aromatase bioassay represents a new tool for future assessments of biologically active FSH in physiological and pathophysiological conditions.

Adult↗
PubMed1985

Autocrine role of estrogens in the augmentation of luteinizing hormone receptor formation in cultured rat granulosa cells.

The effects of estrogens on gonadotropin-stimulated luteinizing hormone (LH) receptor formation were examined in primary cultures of rat granulosa cells. Granulosa cells were cultured for 3 days with increasing concentrations of follicle-stimulating hormone (FSH) in the presence or absence of native and synthetic estrogens. Follicle-stimulating hormone stimulated LH receptor formation in a dose-dependent fashion, and estrogens enhanced the FSH-stimulated LH receptor content by decreasing the apparent ED50 of FSH. At 6.25 ng/ml FSH, the enhancement in LH receptor was estrogen dose dependent, with an ED50 value of about 3 X 10(-9) M for 17 beta-estradiol. The increased LH receptor content seen in cells treated with FSH and estrogen was correlated with increased cAMP production by these cells in response to LH stimulation. Time course studies revealed enhancement of FSH-stimulated LH receptor induction at 48 and 72 h of culture. Granulosa cells were also cultured with FSH for 2 days to induce functional LH receptors, then further cultured for 3 days with LH in the presence or absence of estrogens. At 30 ng/ml LH, increasing concentrations of estrogens maintained LH receptor content in a dose-dependent fashion, with their relative estrogenic potencies in keeping with reported binding affinities to estrogen receptors. An autocrine role of estrogens on LH receptor formation was further tested in granulosa cells treated with FSH and an aromatase substrate (androstenedione) to increase estrogen biosynthesis. Cotreatment with semipurified estrogen antibodies partially blocked the FSH stimulation of LH receptors, whereas nonimmune serum was ineffective. Also, inclusion of diethylstilbestrol prevented the inhibitory effect of the estrogen antibodies. Thus, local estrogens in ovarian follicles may play an autocrine role in granulosa cells to enhance LH receptor formation and to increase granulosa cell responsiveness to the LH surge, with subsequent ovulation and adequate corpus luteum formation.

Animals↗
PubMed1985

Sensitive in vitro bioassay for the measurement of serum follicle-stimulating hormone.

A sensitive in vitro bioassay for measuring serum follicle-stimulating hormone (FSH) levels has been developed based on the stimulation of estrogen production by cultured rat granulosa cells. In the presence of androstenedione, FSH stimulated estrogen production in a dose-dependent manner. Cell sensitivity to FSH was further enhanced by the addition of diethylstilbestrol, a phosphodiesterase inhibitor, insulin, and human chorionic gonadotropin. Although the inclusion of 4% gonadotropin-free serum from hypophysectomized rats inhibited granulosa cell responsiveness, pretreatment of serum with polyethylene glycol (12%) substantially eliminated the serum interfering effect without loss of FSH activity. In normal male and female rats, serum bioactive FSH levels were low, while castration increased these levels 2.6-and 5.7-fold, respectively. These increases were suppressed by treatment with testosterone propionate or estradiol benzoate. This in vitro assay allows the measurement of circulating bioactive FSH and provides a convenient tool to advance studies on the role of FSH in the neuroendocrine control of gonadal maturation and reproductive cycles.

Animals↗
PubMed1985

Androgen inhibition of follicle-stimulating hormone-stimulated luteinizing hormone receptor formation in cultured rat granulosa cells.

Since LH receptors are decreased in atretic follicles known to contain high androgen levels, we have studied the androgen modulation of LH receptor formation in vitro. Granulosa cells from hypophysectomized, diethylstilbestrol-treated rats were cultured for 3 days with FSH in the presence or absence of nonaromatizable androgens, dihydrotestosterone and 5 alpha-androstane-3 alpha, 17 beta-diol, or a synthetic androgen, R1881 (17 beta-hydroxy-17 alpha-methyl-4,9,11-estratrien-3-one). FSH increased LH receptor content in granulosa cells, while concomitant androgen treatment decreased LH receptor content in a dose- and time-dependent manner, without changing the equilibrium dissociation constant (Kd) for human CG. R1881 (10(-7) M), dihydrotestosterone (10(-6) M), and 5 alpha-androstane-3 alpha, 17 beta-diol (10(-6) M) inhibited LH receptor content by 68%, 65%, and 65%, respectively. Similar to earlier findings, these androgens enhanced FSH-stimulated progesterone biosynthesis and aromatase activity in the same cells. To study their LH responsiveness, androgen-treated cells were washed and reincubated for 2 more days with or without LH. Although basal progesterone production was elevated by R1881 pretreatment, the androgen-pretreated cells were less responsive to LH. Treatment with cyanoketone, an inhibitor of 3 beta-hydroxysteroid dehydrogenase, did not alter the inhibitory effects of R1881 on LH receptors, indicating that the androgen action is not mediated by endogenous progestins. Furthermore, R1881 inhibited the stimulation of LH receptor formation by forskolin, cholera toxin, and 8-bromo-cAMP, suggesting that androgens may inhibit LH receptor induction by affecting post-cAMP events. Estrogen treatment enhanced the FSH induction of LH receptor content, while concomitant addition of R1881 also suppressed the estrogen action. Thus, androgens inhibit FSH-induced functional LH receptors in cultured rat granulosa cells. The androgen effect is exerted, at least partially, at post-cAMP sites and is independent of changes in progestin biosynthesis.

8-Bromo Cyclic Adenosine Monophosphate↗