Biomedical subjects

C Féral

Publications and source records attributed to C Féral.

At least 19 recordsLinked to original sources

PubMed2005

[Apoptosis of the follicular cells: its implication in ovarian induction protocols].

Atresia, a degenerative process through which many follicles are removed from the grown pool of follicles involves apoptotic changes in the follicular cells. This review analyses the endocrine regulation of apoptotic cell death in ovarian follicle. FSH is the major survival factor for preovulatory follicle but follicle integrity, in vitro, was necessary to its action on granulosa cell. The role of LH is more ambivalent. FSH and LH exert their activity via activation of the cAMP signal. High levels of intracellular cAMP could enhance steroidogenesis and in the same time induce apoptosis in granulosa cells. Moreover, no correlation between steroidogenesis and apoptosis can be established. During ovarian stimulation in IVF protocol, the use of LH, of coasting and of GnRH agonists and antagonists could be deleterious in follicle survival.

Apoptosis↗

PubMed2003

Involvement of theca cells and steroids in the regulation of granulosa cell apoptosis in rabbit preovulatory follicles.

Follicular atresia is characterized by a rapid loss of granulosa cells and, to a lesser extent, theca cells, via apoptosis. The aim of this study was to investigate the possible involvement of theca cell secretions in the regulation of apoptosis of rabbit granulosa cells. The annexin-V binding method based on externalization of phosphatidylserine to the outer layer of plasma membrane during apoptosis was used to detect apoptotic granulosa cells in flow cytometry. Regulation of apoptosis of granulosa cells was studied in three different culture systems: (i) isolated cultured granulosa cells, (ii) granulosa cells obtained from cultured preovulatory follicles and (iii) granulosa cells co-cultured with theca cells. The results of this study indicate that: (i) the rate of apoptosis of granulosa cells was significantly reduced when granulosa cells were co-cultured with theca cells or obtained from cultured preovulatory follicles in comparison with isolated cultured granulosa cells; (ii) FSH exerts its anti-apoptotic effect only on granulosa cells issued from cultured preovulatory follicles; (iii) ovarian steroids do not affect the percentage of isolated apoptotic granulosa cells; and (iv) the occurrence of an apoptotic process in rabbit theca cells could be upregulated in vitro by hCG and an analogue of the gonadotrophin second messenger cAMP. The results of this study indicate that in rabbits (i) steroids were ineffective in vitro in protecting isolated granulosa cells against apoptosis in comparison with observations in vivo in rats, and (ii) the presence of theca cells was efficient to reduce granulosa cell apoptosis but not sufficient to allow the anti-apoptotic effect of gonadotrophins observed in cultured follicles.

Animals↗

PubMed2002

Quantification of cytochrome P450 aromatase transcripts before and during luteal phase in rabbit.

The aim of the present study was to quantify the promoter II- and I.r-derived transcripts of p450 aromatase gene during follicular stages and during corpus luteum formation in the rabbit. An ovulatory dose of hCG induced, first the disappearance of 90% of aromatase transcripts since 6 h before ovulation, and second a gradual decrease during pseudopregnancy. Individual quantification of both the promoter-derived transcripts showed that promoter II-derived transcript was the main transcript expressed both during follicular phase and pseudopregnancy, but kinetics of disappearance were not similar between both the promoter-derived transcripts. Moreover, hCG up-regulates aromatase expression in vitro in luteal tissue but estradiol, which was without effect on aromatase expression in preovulatory granulosa cells, down-regulates this expression in luteal tissue. In conclusion, the regulation of P450 aromatase in rabbit is mainly under control of promoter II regardless of which cyclic stage is studied. Moreover, we reported an opposite effect of estradiol on aromatase expression in vitro between follicular and luteal cells.

Animals↗

PubMed2002

Hormonal regulation of apoptosis in rabbit granulosa cells in vitro: evaluation by flow cytometric detection of plasma membrane phosphatidylserine externalization.

Annexin V and propidium iodide bivariate analysis and the TUNEL method were used to quantify hormonal regulation of apoptosis in rabbit granulosa cells from preovulatory follicles in vitro. The aim of this study was to analyse comparatively the effects of gonadotrophins and their second messenger in the regulation of granulosa cell apoptosis in (i) cultured isolated granulosa cells and (ii) granulosa cells scraped from cultured follicles. The results showed that increasing doses of FSH had no effect on apoptosis of cultured isolated cells but caused a decrease in the number of apoptotic granulosa cells from preovulatory follicles cultured in serum-free conditions. Unlike FSH, addition of hCG did not modify apoptosis of granulosa cells significantly. In contrast, dibutyryl cAMP had an apoptotic effect in the two cellular models in the presence of serum. Moreover, a biphasic effect of dibutyryl cAMP in isolated granulosa cells was observed with an increase in the incorporation of [(3)H]thymidine into DNA at the lowest dose and an increase in apoptotic cell death at the highest dose. It was concluded that, in rabbits: (i) FSH requires follicle integrity to exert its anti-apoptotic effect in granulosa cells; (ii) dibutyryl cAMP induces a dose-dependent apoptotic effect in granulosa cells cultured alone or obtained from cultured preovulatory follicles; and (iii) cAMP signals induce opposite effects on growth and apoptosis in granulosa cells.

Animals↗

PubMed2001

hH-Rev107, a class II tumor suppressor gene, is expressed by post-meiotic testicular germ cells and CIS cells but not by human testicular germ cell tumors.

By systematic analysis of a human testis library, we have isolated the hH-Rev107-3 cDNA, identical to hH-Rev107-1 cDNA, which was previously described as a class II tumor suppressor gene. In this study, two transcripts (1 and 0.8 kb) were detected by Northern blot in all human tissues, excepted in thymus. The strongest expression was found in testis, skeletal muscle and heart. These two mRNA are probably transcribed from only one gene that we mapped to the q12-q13 region of the chromosome 11. In human testis, hH-Rev107 gene expression was localized, by in situ hybridization, within the round spermatids. To investigate a possible role for hH-Rev107 protein in testicular malignant growth, we examined the expression of this gene in germ cell tumors. A strong hH-Rev107 gene expression was observed in normal testis as well as in samples with preinvasive carcinoma in situ but was completely absent in overt tumors, both seminomas and non-seminomas. By in situ hybridization, CIS was found hH-Rev107 positive and tumor negative. A semi-quantitative assessment of hH-Rev107 mRNA level in testicular germ cell tumors, by RT-PCR, exhibited a ninefold decrease in the gene expression. No gross structural aberrations of hH-Rev107 gene were detected in these human primary tumors. The results suggest that down-regulation of hH-Rev107 may be associated with invasive progression of testicular germ cell tumors.

Amino Acid Sequence↗

PubMed2001

Human testis expresses a specific poly(A)-binding protein.

In testis mRNA stability and translation initiation are extensively under the control of poly(A)-binding proteins (PABP). Here we have cloned a new human testis-specific PABP (PABP3) of 631 amino acids (70.1 kDa) with 92.5% identical residues to the ubiquitous PABP1. A northern blot of multiple human tissues hybridised with PABP3- and PABP1-specific oligonucleotide probes revealed two PABP3 mRNAs (2.1 and 2.5 kb) detected only in testis, whereas PABP1 mRNA (3.2 kb) was present in all tested tissues. In human adult testis, PABP3 mRNA expression was restricted to round spermatids, whereas PABP1 was expressed in these cells as well as in pachytene spermatocytes. PABP3-specific antibodies identified a protein of 70 kDa in human testis extracts. This protein binds poly(A) with a slightly lower affinity as compared to PABP1. The human PABP3 gene is intronless with a transcription start site 61 nt upstream from the initiation codon. A sequence of 256 bp upstream from the transcription start site drives the promoter activity of PABP3 and its tissue-specific expression. The expression of PABP3 might be a way to bypass PABP1 translational repression and to produce the amount of PABP needed for active mRNA translation in spermatids.

Amino Acid Sequence↗

PubMed2001

Expression of the rabbit cytochrome P450 aromatase encoding gene uses alternative tissue-specific promoters.

The aim of the present study was to analyse the tissue-specific expression of various promoter-derived transcripts from the gene encoding rabbit aromatase cytochrome P450. A new promoter, named I.r, was identified, and promoters II and I.r were sequenced. Promoter I.r-derived transcripts were found in preovulatory granulosa cells, corpus luteum, placenta and adipose tissue. An alternative splice variant of this transcript was found with tissue-specific preference. Tissue-specific expression of promoter-derived variants was studied in the ovary before and after ovulation. While the level of promoter II-derived transcript decreased dramatically after ovulation, that of promoter I.r-derived transcript remained unchanged, indicating that promoter II and promoter I.r were not controlled by a single regulation system. The existence of this dual system of regulation suggests that the rabbit ovary could be a useful model to study the promoter-specific regulation of aromatase.

Alternative Splicing↗

PubMed2001

Meiotic human sperm cells express a leucine-rich homologue of Caenorhabditis elegans early embryogenesis gene, Zyg-11.

We cloned a human protein (Hzyg) homologue to Caenorhabditis elegans Zyg-11, an essential protein for cell division at the initial developmental stages of this species, and to a Drosophila melanogaster gene product (Mei-1) which is likely to be involved in meiosis. Hzyg mRNA encodes a protein of 766 amino acids (88 kDa), 14% of which are leucine residues, with some being arranged in four leucine rich repeat motives usually involved in protein-protein interactions. Hzyg is encoded by a single gene, located on chromosome 9q32-q34.1, and transcribed as two mRNA: a 5 kb transcript strongly expressed in testis and skeletal muscle and barely detectable in other human tissues, and an abundant 3.1 kb mRNA detected only in the testis. By using in-situ hybridization and immunohistochemistry, we clearly established the presence of Hzyg expression in pachytene spermatocytes (stage V) and spermatids (stage I and/or II) around the time of meiosis. The cell specific expression of Hzyg transcripts in testis, and the conservation of this gene among distant species, suggest that this protein may have an important role during meiosis.

Adenosine Triphosphatases↗

PubMed1999

Chromosomal localization of three human poly(A)-binding protein genes and four related pseudogenes.

In humans, the poly(A)-binding proteins (PABPs) comprise a small nuclear isoform and a conserved gene family that displays at least three functional proteins: PABP1, inducible PABP (iPABP), and PABP3, plus four pseudogenes (1, 2, 3, and PABP4). In situ hybridization of PABP3 cDNA as the probe on metaphasic chromosomes have revealed five possible loci for this gene family at 2q21-q22, 13q11-q12, 12q13.3-q15, 8q22, and 3q24-q25. Amplifications of specific DNA fragments from a human-rodent somatic cell hybrid panel have allowed us to associate PABP1 and PABP3 with 8q22 and 13q11-q12, respectively. The iPABP gene has been assigned to chromosome 1. This result, compared with radiation hybrid database information, strengthens the location of this gene to 1p32-p36. The pseudogenes PABP4, 1, and 2 have been assigned to chromosomes 15, 4, and 14, respectively. Three loci detected on chromosome spreads are not associated with any amplified fragment. They might represent other related PABP genes not yet identified.

Animals↗

PubMed1998

Inhibition by gonadotropins of interleukin-1 production by rabbit granulosa and theca cells: effects on gonadotropin-induced progesterone production.

Increasing evidence suggests that cytokines may play a role in ovarian processes. The purpose of this study was to determine if interleukin-1 (IL-1) could modulate rabbit pre-ovulatory follicular function and to explore cellular sites of IL-1 biosynthesis in rabbit follicles. Development of rabbit pre-ovulatory follicles was induced by 200 mIU equine chorionic gonadotropin daily for 2 days. Seventy-two hours after the last injection, ovaries were excised and granulosa and theca cells isolated. The two types of cell were pre-incubated for 24 h in Minimum Essential Medium (MEM) with 5% fetal calf serum (FCS), and then incubated for 24 h in MEM with 2.5% FCS with appropriate stimulants. Results showed that rabbit granulosa and theca cell culture supernatants contain IL-1 bioactivity and that this bioactivity was diminished by gonadotropins, FSH and human chorionic gonadotropin, in a dose-dependent manner. Low doses of IL-1 (1 ng/ml) inhibited gonadotropin-induced progesterone production in both cell types and at the same time increased cell numbers. A study of the mechanism of IL-1 action demonstrated that it affects cAMP generation, and also steps distal to cAMP formation. We conclude that in our model gonadotropins, by inhibiting IL-1 production, could control IL-1 modulation of gonadotropin action on steroidogenesis.

1-Methyl-3-isobutylxanthine↗

PubMed1998

Rabbit ovarian production of interleukin-6 and its potential effects on gonadotropin-induced progesterone secretion in granulosa and theca cells.

Recent studies suggest that non-steroid factors, such as cytokines, may play a role in ovarian processes. The purpose of this study was to explore cellular sites of interleukin (IL)-6 biosynthesis in rabbit follicles and to investigate IL-6 modulation in granulosa and theca cell functions. In this report development of rabbit preovulatory follicles was induced by 200 mIU equine chorionic gonadotropin (eCG) daily for 2 days. Seventy-two hours after the last injection ovaries were excised and granulosa and theca cells isolated. The two types of cells were preincubated for 24 h in Minimum Essential Medium (MEM) with 5% fetal calf serum (FCS), and then incubated for 24 h in MEM-2.5% FCS with appropriate stimulants. Results showed that rabbit granulosa and theca cell culture supernatants contained IL-6 bioactivity and that its production was inhibited by FSH and human CG and stimulated by IL-1. IL-6 inhibited gonadotropin-induced progesterone production, but not basal secretion, in both cell types, without a cytotoxic effect. IL-6 affected cAMP generation and steps distal to cAMP formation, but the mechanism of IL-6 action on progesterone differed in granulosa and theca cells. Taken together our results suggest that gonadotropins, by inhibiting IL-6 production, could control, in our model, IL-6 modulation of gonadotropin action on steroidogenesis.

Analysis of Variance↗

PubMed1997

Luteotrophic factors in hyperstimulated pseudopregnant rabbit: I--Evidence for aromatase activity in luteal tissue and luteal cells.

It is well established that the rabbit corpus luteum (CL) function depends upon endogenous oestradiol, the major source of which in the rabbit ovary is considered to be the ovarian follicles. The absence of oestradiol formation by the rabbit CL has been previously reported. In a hyperstimulated pseudopregnant rabbit model used in our laboratory which developed a large number of corpora lutea in response to chorionic gonadotrophin (eCG)/hCG, we observed the survival of corpora lutea in vivo, and normal levels of plasma progesterone throughout pseudopregnancy (PP), despite the scarcity or the absence of follicles as a source of the luteotrophic hormone. Measurement of oestradiol in the plasma indicated that it was at high levels and correlated with the number of corpora lutea. This led us to investigate the luteal origin of oestradiol in this model. PP was induced in rabbits by i.m. injection of 200 IU eCG daily for 2 days followed on day 4 by i.m. injection of 200 IU hCG (day 0 of PP). Luteal tissue obtained at days 5, 9 and 12 of PP and cultured for 24 h synthesized oestradiol and testosterone in addition to progesterone. However, under the same conditions, follicles had limited capacity to secrete oestradiol. The presence of an aromatase activity in luteal tissue was confirmed when exogenous testosterone was added to the culture medium. P450aromatase (P450arom) mRNA was found in luteal tissue at days 5, 9 and 12 of PP. Small or large luteal cells, obtained by enzymatic digestion of the tissue followed by centrifugation in a Percoll density gradient, were cultured during several days with or without gonadotrophin or dibutyryl cAMP (dbcAMP). Both types of cells secreted oestradiol. In small cells and luteal tissue, aromatase activity was stimulated (1.5-2-fold) by hCG and dbcAMP. Large cells exhibited a greater capacity to aromatize testosterone than small cells, but aromatase activity was not modified by hCG or by dbcAMP. FSH had no effect on aromatase activity of either luteal cell type. This intrinsic luteal tissue aromatase capacity and the absence of premature regression of corpora lutea despite the limited support of follicular oestrogen, suggest an autocrine and luteotrophic role for this luteal oestrogen.

Animals↗

PubMed1997

Luteotrophic factors in hyperstimulated pseudopregnant rabbit: II--High sensitivity to hCG of luteal tissue and small luteal cells.

Previous studies on rabbit corpus luteum (CL) led to the conclusion that the luteotrophic complex, in rabbit, may include LH as well as oestradiol for normal luteal function. However, the requirement for LH is controversial. We have recently demonstrated the existence of a human chorionic gonadotrophin (hCG)-stimulated aromatase activity in cultured corpora lutea from a hyperstimulated pseudopregnant rabbit model, which develops a large number of corpora lutea, with only a few or no follicles in the ovaries. The present study was undertaken to investigate the in vitro responsiveness to hCG, dibutyryl cAMP (dbcAMP) and oestradiol of those corpora lutea. Pseudopregnancy (PP) was induced in rabbits by i.m. injection of 200 IU equine chorionic gonadotrophin daily for 2 days followed on day 4 by i.m. injection of 200 IU hCG (day 0 of PP). Luteal tissue and small and large luteal cells obtained at days 5 and 9 of PP were cultured for 24 h or during several days respectively with or without hCG, dbcAMP or oestradiol. Basal progesterone secretion was 3.6- and 22-fold higher in large cells compared with small ones at day 5 and 9 of PP respectively. When stimulated by small doses of hCG, luteal tissue responded by a 5-fold increase in progesterone secretion. Small cells produced four times higher amounts of progesterone than controls in the presence of 1 mIU/ml hCG and more than ten times in the presence of 0.1 IU/ml hCG, whereas large cells were insensitive to hCG stimulation. dbcAMP mimicked the effect of hCG on progesterone secretion by luteal tissue and luteal cells and oestradiol stimulated basal progesterone secretion in both small and large luteal cells. Given the large contribution of non stimulated large cells to luteal progesterone production and the remarkably high sensitivity of luteal tissue to gonadotrophin in vitro it seems that interactions between the two types of cells might occur during LH stimulation. Our results suggest that LH could participate in the luteotrophic complex at least in part through the stimulation of endogenous oestradiol production by luteal cells.

Animals↗

PubMed1996

Angiotensin II receptor type 1 on granulosa and thecal cells of rabbit preovulatory follicles.

Specific, high-affinity angiotensin II (A II) receptors were observed on granulosa and thecal cells of preovulatory ovarian follicles from immature PMSG-treated rabbits. Scatchard analysis of 125I-[Sar1,Ile8]A II binding to freshly prepared cells was indicative of only one class of binding sites. Kd values were 0.26 +/- 0.11 nM and 0.18 +/- 0.02 nM, densities of A II receptors were 0.06 +/- 0.02 fmol/10(5) cells and 0.08 +/- 0.01 fmol/10(5) cells for granulosa and thecal cells, respectively. When cells were incubated for 48 h with hCG, Kd values were of the same order of magnitude, but the amount of A II receptors was increased 2-fold in granulosa and 4-fold in theca. Using subtype specific ligands (Losartan for AT1 and PD 123319 for AT2) in competitive binding experiments, A II receptors were found to be of the AT1 type on both granulosa and thecal cells freshly prepared or incubated 48 h in vitro. These results establishing the existence of high affinity AT1 receptors on the two cell types of the rabbit preovulatory follicles contrast with previous observations showing the presence of AT2 receptors on granulosa or theca from several species.

Angiotensin II↗

PubMed1996

Rapid sequencing of rabbit aromatase cDNA using RACE PCR.

The sequencing of aromatase cDNA from rabbit granulosa cells was obtained by RACE PCR. This cDNA is 2.9 kb long. The first 119 nucleotides correspond to the first untranslated exon. Nucleotides 120 to 1,629 correspond to the coding region (1,509 nucleotides) and the rest of the sequence is non coding and contains a polyadenylation signal. Translation of the cDNA sequence indicates that the protein is composed of 503 amino acids, like in human aromatase. Its molecular weight is 57.4 kDa. The alignment between the rabbit aromatase amino acid sequence and other aromatases already described in the human, mouse, rat, cow, pig, chicken, rainbow trout and teleost fish shows that the rabbit protein exhibits the highest homology with the human one (85%).

Animals↗

PubMed1995

Angiotensin II modulates steroidogenesis in granulosa and theca in the rabbit ovary: its possible involvement in atresia.

Accumulating evidence has shown the ovary of mammals to contain an intrinsic renin-angiotensin system that has been ascribed an autocrine-paracrine role. The present study in the female rabbit ovary investigated the putative in vitro action of angiotensin II (A II) on basal and gonadotropin-induced steroidogenesis. Ovarian follicles from immature female rabbits treated with pregnant mare's serum gonadotropin (PMSG) were dissected out and a complete separation of the theca interna from the granulosa layer was performed, to demonstrate that A II affects separately the two individual cellular components of the follicular wall. We could show that theca is a source of estradiol whose production under human chorionic gonadotropin (hCG) stimulation was reduced by A II. At the same time, A II increased the in vitro hCG-stimulated secretion of testosterone by theca. In granulosa, A II decreased hCG-stimulated aromatization of androstenedione to estradiol but did not alter the release of hCG-stimulated progesterone production. These results suggest that A II could induce locally an increase in follicular fluid androgen/estrogen ratio and possibly participate in causing atresia.

Angiotensin II↗

PubMed1993

[Renin-angiotensin system of the uterus and ovary in mammalian females].

The circulating reninangiotensin system (RAS) participates in the regulation of blood pressure and electrolyte metabolism. Renin, a proteolytic enzyme, synthesized in the kidney from its biological precursor, prorenin, cleaves its substrate angiotensinogen in the blood to form the active octapeptide, angiotensin II (AII). All the RAS components are present in the reproductive system of mammals. During pregnancy, the level of prorenin increases in the plasma. The ovary is the source of this prorenin during early pregnancy and maternal decidua later on. During the menstrual cycle, the thecal of preovulatory follicles synthesize prorenin, renin and AII. Thecal renin synthesis is controlled by LH/hCG as demonstrated in vivo and in vitro in the rabbit. Ovarian renin seems to be identical to kidney renin. Prorenin appears to be the major secretory product rather than renin, which remains intracellular. AT2-type angiotensin II-receptors are expressed in the rat on follicular granulosa cells and could be down-regulated by FSH. The bovine thecal cells also express AT2-receptors, up-regulated by LH. These data are consistent with an autrocrine or paracrine role for ovarian RAS. It has been implicated in neovascularization of the follicle and regulation of steroidogenesis by increasing the androgen/estrogen ratio, an index of follicular atresia.

Animals↗

PubMed1990

Stimulation by hCG of ovarian inactive renin synthesis in rabbit preovulatory theca cells.

The origin of ovarian renin and its regulation by hCG were investigated in rabbit periovulatory follicles and cultured preovulatory follicular cells. Intracellular content of renin in thecal cells was 8-fold greater than of granulosa cells. In vivo, administration of hCG increased intracellular content of renin in thecal but not granulosa cells. Similar results were obtained for cultured follicular cells, from which renin was partly released into the medium. In vitro, hCG increased intracellular renin content of thecal but not granulosa cells, without obvious effect on release. Approximately 95% of ovarian renin was inactive, but could be activated by trypsin. Thecal renin was antagonized in vitro by renin antiserum, indicating a specific renin activity. Our study establishes in the rabbit the thecal cell origin of ovarian inactive renin and demonstrates hCG regulation of its synthesis.

Animals↗