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M Zilberstein

Publications and source records attributed to M Zilberstein.

31 records · Page 2Linked to original sources
PubMed1992

Regulatory effect of antiphospholipid antibodies on signal transduction: a possible model for autoantibody-induced reproductive failure.

OBJECTIVES: The broad range of reproductive processes apparently affected by antiphospholipid antibodies suggests that antiphospholipid antibodies have to exert their clinical effects at every basic cellular level. Because phospholipids constitute essential components of every cell membrane, and because signal transduction processes are dependent on second messengers with phospholipid epitopes, we investigated the hypothesis that interference with signal transduction processes by antiphospholipid antibodies could represent such a basic cellular process. STUDY DESIGN: To test this hypothesis we established an in vitro placental culture system in which the stimulation of explants by phospholipase A2 and phospholipase C in the presence of normal human serum resulted in a significant increase in human chorionic gonadotropin secretion. RESULTS: When normal human sera were replaced with antiphospholipid antibody-containing sera from two women with established histories of reproductive failure, this increase in human chorionic gonadotropin production was inhibited 42% to 44% under phospholipase A2 stimulation and by 34% to 39% under phospholipase C stimulation. The hypothesis that antiphospholipid antibodies can interfere with signal transduction processes was further strengthened by the observation that antiphospholipid antibody-positive sera contained significantly higher immunoglobulin A antibody reactivity toward the second messenger inositol 1,4,5-triphosphate than control sera (p less than 0.049). CONCLUSIONS: These observations suggest the possibility that if present at excessive levels antiphospholipid antibodies may exert their adverse effect on reproductive processes through the interception of signal transduction processes.

Antibodies↗
PubMed1991

Expression, action, and steroidal regulation of insulin-like growth factor-I (IGF-I) and IGF-I receptor in the rat corpus luteum: their differential role in the two cell populations forming the corpus luteum.

The overall aim of this investigation was to examine the expression and steroidal regulation of insulin-like growth factor-I (IGF-I) and the IGF-I receptor in the rat corpus luteum and to examine the specificity of IGF-I action in the two luteal cell populations. We first examined whether the corpus luteum expresses the IGF-I and IGF-I receptor genes. Using a solution hybridization/RNase protection assay, IGF-I and IGF-I receptor mRNAs were represented by protected bands 224 and 265 bases in length, respectively. In addition, Northern blot analysis showed that, as in liver, rat IGF-I and IGF-I receptor cDNAs hybridized with 7.5-, 1.8-, and 0.8- to 1.2-kilobase transcripts and an 11-kilobase transcript, respectively. Both IGF-I and IGF-I receptor mRNAs were detected on all days of pregnancy tested (days 5-21). Since the rat corpus luteum increases remarkably in size and steroidogenic capacity at midpregnancy due to estradiol stimulation, we determined whether these developmental changes are accompanied by an increased expression of the IGF-I and/or IGF-I receptor genes. Total RNA was isolated from corpora lutea of day 12 hypophysectomized-hysterectomized rats treated with or without estradiol for 3 days. Estradiol caused a clear and marked reduction in IGF-I and IGF-I receptor mRNA. [125I]IGF-I bound with high specificity and affinity to luteal cell membranes. Large and small cell populations forming corpora lutea of day 3 and 14 pregnant rats were separated by elutriation and used for the determination of binding activity and for cell culture, respectively. IGF-I receptors were found to be localized principally in the large luteal cell population. The small luteal cells had approximately 6.5-fold less IGF-I-binding activity. The difference in binding activity in both cell populations was reflected in the ability of both cell types to respond to IGF-I. IGF-I (25 ng/ml) had a profound effect on the production of progesterone by the large luteal cells. No stimulatory effect of IGF-I on the small luteal cells was observed. Addition of estradiol (10 ng/ml) to the cell culture remarkably enhanced IGF-I stimulation of progesterone biosynthesis by the large luteal cells. In summary, the results of this investigation have revealed that the corpus luteum of the pregnant rat is a major site of expression of both the IGF-I and IGF-I receptor genes.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1990

Expression of the pregnancy-specific beta 1-glycoprotein gene in cultured human trophoblasts.

Human pregnancy-specific beta 1-glycoprotein (PS beta G), the major placental glycoprotein, shares strong sequence similarity with carcinoembryonic antigen and is a member of the immunoglobulin superfamily. To understand the role of PS beta G in placental ontogeny during pregnancy, we examined its synthesis and regulation in primary cultures of trophoblast cells. Freshly plated (12 h) cytotrophoblasts expressed little PS beta G transcripts; however, within 24 h of culture, PS beta G mRNAs became detectable. PS beta G synthesis and mRNA expression increased with time in culture, and maximal synthesis was achieved at 4 days, indicating that primary trophoblasts continue differentiating in vitro. Molecular cloning revealed that PS beta G is an extremely polymorphic protein. Most PS beta G cDNAs identified to date, including the three cDNAs (PSG16, PSG93, and PSG95) isolated in this laboratory, share strong sequence similarity in the 5' (designated PSG-5') and coding regions, but differ in sequences at the 3' region. The PSG-5', PSG93-specific, PSG16/PSG93-3', and PSG95-3' probes, which identify the majority of PS beta G mRNAs, hybridized with three mRNAs of 2.3, 2.2, and 1.7 kilobases in primary trophoblasts and human term placental tissue. Ribonuclease protection analysis demonstrated that primary trophoblasts expressed most of the placental PS beta G transcripts. However, culturing in vitro altered PS beta G gene expression, and the level of PS beta G transcripts containing a PSG95-3' sequence was preferentially increased in primary trophoblasts. Moreover, primary trophoblasts synthesized a 64K PS beta G polypeptide in variable amounts and three PS beta Gs of 72K, 62K, and 54K in roughly equal amounts, whereas purified human term placental PS beta G consists of a major polypeptide of 72K and two minor ones of 64K and 54K. PS beta G gene expression in primary trophoblasts was slightly reduced by 8-bromo-cAMP, but was markedly inhibited by sodium butyrate.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1989

Expression of insulin-like growth factor-I and its receptor by SV40-transformed rat granulosa cells.

Cellular proliferation is a dominant aspect of ovarian follicular development in the rat, and insulin-like growth factor I (IGF-I) has been proposed as a mediator of cellular growth and differentiation in the ovary. An SV40-transformed rat granulosa cell line (RGA-41S) has been established as a model for studies on dividing cells of granulosa origin. Granulosa cells from the ovaries of immature diethylstilbestrol-treated rats were infected with the tsA255 mutant of SV40, followed by cloning in serum-free medium to select transformed cell lines which were serum independent. At the permissive temperature (33 C), RGA-41S cells exhibited a transformed phenotype and rapidly formed high density multilayers of compact cells that readily overgrew nontransformed cells. At the nonpermissive temperature (40 C) cell replication declined and division ceased after 4 days. Furthermore, at 40 C the cells grew as a monolayer and assumed a tetrahedral shape with a high cytoplasm-to-nucleus ratio, and displayed reduced ability to overgrow nontransformed cells. The transformed ovarian cells did not express detectable gonadotropin receptors and steroidogenic activity but retained their epithelial phenotype as demonstrated by cytokeratin staining of the cytoskeleton, the presence of microvilli, and the formation of tight junctions between cells. In support of the proposed autocrine-paracrine actions of IGF-I in the ovary, assay of conditioned serum-free culture medium revealed secretion of IGF-I-immunoreactive material by RGA-41S cells. HPLC-purified IGF-I immunoreactivity from these cells eluted with the same retention time as recombinant human IGF-I. When hybridized with a 32P-labeled rat IGF-I cDNA probe, poly(A)+ mRNA prepared from RGA-41S cells grown at both temperatures showed the typical three size classes of IGF-I mRNA on Northern blots (7.5, 1.7, and 0.8-1.2 kilobase kb), although the levels were somewhat higher at 33 C. The presence of IGF-I receptors in transformed cells was demonstrated by specific 125I-IGF-I binding to intact cells. Scatchard analysis indicated a single class of high affinity receptors at a density of 10(5) binding sites per cell and a dissociation constant (Kd) = 0.52 x 10(-9) M. Furthermore, hybridization of a 32P-labeled IGF-I receptor probe to Northern blots of poly(A+) RNA prepared from cells grown at 33 C and 40 C revealed an 11-kilobase rat IGF-I receptor mRNA. Physiological concentrations of IGF-I increased [3H]aminoisobutyric acid uptake by RGA-41S cells grown at either temperature, attesting to the retention of responsiveness to IGF-I in these transformed granulosa cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Aminobutyrates↗
PubMed1985

Characterization of the maturational changes induced by a GnRH analogue in the rat ovarian follicle.

The GnRH analogue [D-Ser(t-Bu)6]des-Gly10-GnRH-N-ethylamide (GnRHa, 2 micrograms/rat) or hCG (4 i.u./rat) was administered to hypophysectomized, PMSG-primed immature female rats. Oocyte maturation was initially detected by 2 h after GnRHa administration but the response to hCG was observed only after 4 h. Initiation of GnRHa-induced ovulation also preceded the response to hCG by 2 h. Maximal response to both these hormones was obtained at 10 and 14 h after hormone administration for oocyte maturation and ovulation respectively. The number of oocytes ovulated after GnRHa was significantly lower than that with hCG (29 +/- 4 and 50 +/- 7 per rat respectively; P less than 0.05). Expansion of the cumulus mass and secretion of mucoid material, which are characteristic responses to LH, were also observed after GnRHa administration. However, while the action of 5 micrograms ovine LH/ml on the cumulus cells was mediated by cAMP, no accumulation of the nucleotide could be detected in follicles exposed to GnRHa (10(-7) M). We conclude that even though GnRHa and LH/hCG seem to elicit similar responses in the ovarian follicle they differ in their kinetics, their efficiency and the mediator of their action.

Animals↗
PubMed1984

Gonadotropin releasing hormone: regulation of phospholipid turnover and prostaglandin production in ovarian granulosa cells.

The direct effect of gonadotropin releasing hormone (GnRH) upon ovarian function, is initiated by a rapid receptor-mediated increase in phosphatidylinositol (PI) turnover (approximately 5 min) followed by prostaglandin E (PGE, 120 min) and progesterone (120 min) formation, oocyte maturation and induction of ovulation. In contrast, luteinizing hormone (LH) stimulation of oocyte maturation and induction of ovulation is mediated by increased adenosine 3',5'-monophosphate (cAMP, 15 min), progesterone (30 min) and PGE (180 min) production. Both LH and GnRH stimulation of oocyte maturation are inhibited by dibutyryl cAMP and 3-isobutyl-1-methylxanthine, whereas induction of ovulation by the two hormones is blocked by indomethacin. GnRH and LH differ, therefore, in the mechanism leading to PGE formation, but thereafter share a common mechanism responsible for oocyte maturation and independently for induction of ovulation.

Animals↗
PubMed1984

Regulation of prostaglandin E, progesterone, and cyclic adenosine monophosphate production in ovarian granulosa cells by luteinizing hormone and gonadotropin-releasing hormone agonist: comparative studies.

The early direct effects of GnRH on the ovary were investigated using cultured granulosa cells from preovulatory rat follicles, and compared to the known stimulatory effects of LH. Stimulation of ovarian functions by a GnRH agonist include a rapid receptor-mediated phosphatidylinositol turnover (approximately 5 min). On the other hand, LH action on granulosa cells is initiated by increased cAMP production (approximately 10-15 min), consisting of an indomethacin-resistant and indomethacin-sensitive pools (40% and 60%, respectively). The GnRH agonist [D-Ala6] des-Gly10 N-ethylamide (GnRHa) at concentrations of 10(-12)-10(-8) M had no effect on basal or LH-stimulated cAMP production during a 4-h incubation test. Both LH and GnRHa increase progesterone formation (30 and 120 min, respectively) with ED50 values of 2.5 ng/ml and 10(-9) M, respectively and the stimulatory effect is not blocked by indomethacin. LH and GnRHa increase also prostaglandin E (PGE) formation (180 and 120 min, respectively) and the ED50 values were 0.1 microgram/ml and 10(-9) M, respectively. No inhibitory effect of GnRHa on LH actions was observed during 4 h of incubation. It is concluded that: 1) GnRH mimicks LH stimulation of ovarian PGE and progesterone production; 2) cAMP does not play a role in mediating the direct stimulatory effects of GnRH agonists on ovarian PGE and progesterone production; 3) PGE is not involved in mediating GnRH and LH stimulation of progesterone formation. 4) LH-induced cAMP production consists of indomethacin-sensitive and indomethacin-resistant pools.

Animals↗
PubMed1983

Coincidence of down-regulation and desensitization in pituitary gonadotrophs stimulated by gonadotropin releasing hormone.

The dynamics of gonadotropin releasing hormone (GnRH) induced luteinizing hormone (LH) release was studied in vitro by superfusion of cultured pituitary cells. Continuous exposure of the cells to GnRH resulted in desensitization of the gonadotroph responsiveness to further stimulation by the hormone. The refractory state was achieved within 4 hr of hormone introduction (10(-7) M) and was accompanied by down-regulation of GnRH receptors (50%) assayed by equilibration with [125I]iodo-[D-Ala6]des-Gly10-GnRH N-ethylamide. The data indicate that GnRH can regulate the number of its own receptors, and that desensitization is accompanied by down-regulation.

Animals↗
PubMed1983

Dissociation between the direct stimulatory and inhibitory effects of a gonadotropin-releasing hormone analog on ovarian functions.

The paradoxical effects of gonadotropin-releasing hormone (GnRH) on the ovary have hitherto been believed to result from different regimens of administration; an acute treatment was shown to stimulate the ovary while chronic administration of the hormone inhibited LH-induced responses. In the present report we demonstrate that a single injection of a GnRH analog (D-Ala6)des-Gly10-GnRH-N-ethylamide (GnRHa, 2 micrograms/rat) is sufficient to obtain a significant inhibition (75%) of hCG-induced ovulation in PMSG-primed, either intact or hypophysectomized, immature rats. Inhibition of ovarian development, in terms of growth and ovulation, by multiple injections with GnRHa (2 micrograms/rat, twice daily for 3 days) could be obtained only upon administration of the hormone at early stages of follicular development, i.e. concomitantly with the PMSG injection. When administered after PMSG, GnRHa could not inhibit the ovary but rather induced ovulation by itself in the absence of hCG. A 12-24 h delay in initiation of GnRHa treatment triggered 65% of the rats to ovulate while a delay of 48 h resulted in 100% ovulation. Under both regimes of GnRHa administration, either the inhibitory or the stimulatory, the oocytes of the treated rats were induced to resume meiotic maturation. Since under the inhibitory regime ovulation did not occur, maturation was followed by a massive degeneration of the oocytes trapped within their follicles. These findings demonstrate that the follicular stage of development rather than the dose and/or duration of GnRHa administration determines whether GnRHa inhibits ovarian growth and ovulation, while the competence of the oocytes to respond to the GnRHa stimulus and mature is independent of hormonal priming.

Animals↗