PubMed Health⌕ Search

Biomedical subjects

Y Sadovsky

Publications and source records attributed to Y Sadovsky.

At least 55 records · Page 3Linked to original sources
PubMed1996

Rat uterine myometrium contains the gap junction protein connexin45, which has a differing temporal expression pattern from connexin43.

OBJECTIVE: Our purpose was to determine whether myometrial cell lines and rat myometrial tissue contained additional gap junction proteins besides connexin43. STUDY DESIGNS: Syrian hamster myocytes (SHM-ER) and human SK-UT-1 myometrial cell lines were analyzed for intercellular coupling by microinjection of Lucifer yellow. These cell lines and myometrial tissue isolated from pregnant rats were analyzed for connexin expression by ribonucleic acid blotting and immunofluorescence. RESULTS: SHM-ER and SK-UT-1 cells showed functional gap junctional coupling by intercellular passage of microinjected dye. Both cell lines contained connexin43 and connexin45 messenger ribonucleic acids but did not contain any other detectable connexin messenger ribonucleic acids. Immunofluorescence confirmed the presence of connexin43 and connexin45 proteins in these cells. Connexin 43 and connexin45 messenger ribonucleic acids and immunoreactive proteins were detected in pregnant rat myometrium. Connexin 43 messenger ribonucleic acid levels increased dramatically at term. In contrast, connexin45 messenger ribonucleic acid was present in nonpregnant myometrium, remained relatively constant early in gestation, fell just before term, and more than doubled post partum. CONCLUSIONS: Rat uterine myometrium contains connexin45 and connexin43. Coexpression of connexin45 with connexin43 in uterine myometrium may regulate gap junctional coupling between these cells. The different temporal expression patterns suggest that connexin45 and connexin43 may have different roles or that the ratio of these connexins may be important in the increased cellular coupling coincident with the onset of labor.

Animals↗
PubMed1995

Mice deficient in the orphan receptor steroidogenic factor 1 lack adrenal glands and gonads but express P450 side-chain-cleavage enzyme in the placenta and have normal embryonic serum levels of corticosteroids.

The orphan nuclear receptor steroidogenic factor 1 (SF-1) is expressed in the adrenal cortex and gonads and regulates the expression of several P450 steroid hydroxylases in vitro. We examined the role of SF-1 in the adrenal glands and gonads in vivo by a targeted disruption of the mouse SF-1 gene. All SF-1-deficient mice died shortly after delivery. Their adrenal glands and gonads were absent, and persistent Mullerian structures were found in all genotypic males. While serum levels of corticosterone in SF-1-deficient mice were diminished, levels of adrenocorticotropic hormone (ACTH) were elevated, consistent with intact pituitary corticotrophs. Intrauterine survival of SF-1-deficient mice appeared normal, and they had normal serum level of corticosterone and ACTH, probably reflecting transplacental passage of maternal steroids. We tested whether SF-1 is required for P450 side-chain-cleavage enzyme (P450scc) expression in the placenta, which expresses both SF-1 and P450scc, and found that in contrast to its strong activation of the P450scc gene promoter in vitro, the absence of SF-1 had no effect on P450scc mRNA levels in vivo. Although the region targeted by our disruption is shared by SF-1 and by embryonal long terminal repeat-binding protein (ELP), a hypothesized alternatively spliced product, we believe that the observed phenotype reflects absent SF-1 alone, as PCR analysis failed to detect ELP transcripts in any mouse tissue, and sequences corresponding to ELP are not conserved across species. These results confirm that SF-1 is an important regulator of adrenal and gonadal development, but its regulation of steroid hydroxylase expression in vivo remains to be established.

Adrenal Cortex Hormones↗
PubMed1995

Estrogenic activity of RU 486 (mifepristone) in rat uterus and cultured uterine myocytes.

OBJECTIVE: Our purpose was to determine whether RU 486 (mifepristone) has direct estrogenic activity in uterine myocytes. STUDY DESIGN: Ovariectomized adult rats were treated with RU 486, and its effect on uterine oxytocin receptor concentration, as a marker of estrogenic activity, was measured. Results were compared with the induction by RU 486 of an estrogen-responsive reporter gene in a cultured Syrian hamster uterine myocyte cell line. RESULTS: Baseline oxytocin receptor concentration was 58.8 +/- 7.2 fmol/mg protein (mean +/- SEM) and increased to 227 +/- 49 fmol/mg with 17 beta-estradiol (2.5 micrograms/kg) and to 145 +/- 18 fmol/mg after RU 486 (5 mg/kg) treatment, an effect that was inhibited by the antiestrogen ICI 182,780 (1.5 mg/kg). In the cultured Syrian hamster uterine myocyte cell line cells RU 486 (10(-6) mol/L) caused a 2.17 +/- 0.17-fold increase in the expression of the reporter gene versus 113.0 +/- 7.4-fold with 17 beta-estradiol (10(-8) mol/L). The estrogenic activity of RU 486 was dependent on the presence of both estrogen receptor and the promoter's estrogen response element. CONCLUSION: RU 486 has a weak estrogen-like activity in uterine myocytes. This activity may partly explain the therapeutic effects of RU 486 on this target organ.

Animals↗
PubMed1995

Transcriptional activators differ in their responses to overexpression of TATA-box-binding protein.

We investigated how overexpression of human TATA-box-binding protein (TBP) affects the action of estrogen receptor (ER) and compared the response with that of other activators. When ER activates a simple promoter, consisting of a response element and either the collagenase or tk TATA box, TBP overexpression potentiates transcription. TBP potentiates only estrogen-induced and not basal transcription and does so independent of spacing between response element and TATA box. TBP overexpression also reduces autoinhibition by overexpressed ER, suggesting that one target of the autoinhibition may be TBP itself. Both AF-1 and AF-2 domains of ER are potentiated by TBP, and each domain binds TBP in vitro. Like ER, chimeric GAL4/VP16 and GAL4/Tat activators are also potentiated by TBP, as is the synergistic activation by ER and GAL4/VP16 on a complex promoter. Unlike ER, GAL4/Sp1 and GAL4/NF-I become less potent when TBP is overexpressed. Furthermore, synergy between ER and Sp1 or between ER and NF-I, whether these are supplied by transfected GAL4 fusions or by the endogenous genes, is inhibited by TBP overexpression. Thus, ER resembles VP16 in response to TBP overexpression and is different from Sp1 and NF-I, which predominate over ER in setting the response on complex promoters.

Animals↗
PubMed1995

Adrenocortical function and regulation of the steroid 21-hydroxylase gene in NGFI-B-deficient mice.

The immediate-early gene NGFI-B encodes an orphan nuclear receptor that binds DNA as a monomer and activates transcription through a canonical response element (NBRE). NGFI-B is expressed under basal conditions and in response to external stimuli in many mammalian tissues. In particular, NGFI-B expression is dramatically elevated in the adrenal cortex in response to stress and in Y1 adrenocortical cells in response to adrenocorticotropin. NGFI-B activates transcription through an NBRE of the gene encoding 21-hydroxylase (P450c21) in Y1 cells. Steroidogenic factor 1 (SF-1), a homolog of NGFI-B, also activates the P450c21 promoter. To examine the influence of these factors on P450c21 expression in vivo and the function of the hypothalamic-pituitary-adrenocortical axis as a whole, we generated NGFI-B (-/-) mice. These mice thrive and reproduce normally and maintain normal basal adrenocorticotropin, corticosterone, and P450c21 mRNA levels. In response to increases in adrenocorticotropin, NGFI-B (-/-) and wild-type mice demonstrated equivalent increases in serum corticosterone levels. Furthermore, and in contrast to in vitro results, no increases in P450c21 mRNA levels were observed in response to increases in adrenocorticotropin in NGFI-B (-/-) or wild-type mice. While SF-1 mRNA levels were not increased with increased steroidogenic demand, adrenal expression of Nurr1, a close homolog of NGFI-B, was induced to a greater extent by lipopolysaccharide in NGFI-B (-/-) mice than in wild-type mice. Finally, when the administration of dexamethasone for suppression was stopped, P450c21 mRNA and serum corticosterone levels recovered at the same rate in wild-type and NGFI-B (-/-) mice. Thus, while NGFI-B appears poised to affect the structure and function of the adrenal gland, the gland functions normally in its absence, suggesting that other factors, including Nurr1 and SF-1, are sufficient to drive P450c21 expression in mice and maintain normal steroidogenesis.

Adrenal Cortex↗
PubMed1994

Interleukin-1 beta regulates human cytotrophoblast metalloproteinase activity and invasion in vitro.

During early human pregnancy, fetal cytotrophoblasts rapidly invade the uterus. This process has many similarities to tumor invasion, except that the extent and the timing of cytotrophoblast invasion are carefully regulated. Therefore, this system is particularly useful for studying mechanisms that regulate invasive processes. Previously, we showed that production and activation of the 92-kDa type IV collagenase (matrix metalloproteinase(MMP)-9) is necessary for cytotrophoblast invasion in vitro. In other systems, interleukin (IL)-1 beta is an important regulator of matrix-degrading metalloproteinases. Therefore, we investigated trophoblast production of IL-1 beta and its receptors, as well as the effects of this cytokine on cytotrophoblast metalloproteinase activity and invasion. The results showed that release of IL-1 beta parallels the invasive potential of the cytotrophoblasts; the highest levels are produced by first trimester cells and the lowest levels by term cells. Immunoprecipitation showed that cytotrophoblasts express the 80-kDa type I IL-1 receptor, suggesting that autocrine effects are possible. IL-1 beta stimulated trophoblast MMP-9 secretion (by a mechanism that required nascent mRNA and protein synthesis) as well as metalloproteinase activity and invasion of Matrigel. Increasing (by lipopolysaccharide treatment) or decreasing (by glucocorticoid treatment) IL-1 beta production had parallel effects on MMP-9 secretion, metalloproteinase activity, and invasion. Because IL-1 beta and corticosteroids are present in high concentrations at the maternal-fetal interface, normal trophoblast invasion may be regulated, in part, by their opposing actions. In contrast, stimulation of cytotrophoblast IL-1 beta secretion by lipopolysaccharide may play a role in the sequela of infected fetal membranes.

Collagenases↗
PubMed1993

Restoration of estrogen-dependent progesterone receptor expression in a uterine myocyte cell line.

Although the in vivo effect of estrogen on myometrial differentiation is well documented, estrogen effects on primary myocytes in vitro have been difficult to demonstrate. To construct a stable uterine myocyte system, capable of direct estrogen responsiveness, we used a transformed hamster myometrial cell line. Since these cells expressed a low level of estrogen receptors (ERs), we have stably transfected them with a vector for the human ER. After transfection, ER concentration increased from less than 300 sites per cell to 17,000 +/- 2,000 sites per cell (mean +/- SEM). To test the functional integrity of the transfected receptors, a chloramphenicol acetyltransferase gene linked to an estrogen response element upstream of thymidine kinase promoter was transiently transfected, and the amount of chloramphenicol acetyltransferase activity, an indicator of estrogen responsiveness, was found to increase 20-fold in response to 17 beta-estradiol (1 nM for 48 h). Furthermore, we tested the ability of estrogen to activate endogenous genes by measuring progesterone receptor (PR) induction. PR concentration in the transfected cells was 3,700 +/- 800 and increased 9-fold to 33,000 +/- 6,000 with 17 beta-estradiol (2 nM). This receptor density increase was confirmed by immunoblotting. PR induction was maximal at 16 h, was concentration dependent, and was not elicited by tamoxifen or ICI 164,384. We conclude that transformed hamster myocytes transfected with an ER gene are capable of estrogen-dependent PR expression in vitro and may serve as a useful system to study estrogen effect on myocytes.

Animals↗
PubMed1993

Myometrial characteristics of the Syrian hamster uterine smooth muscle cell line, SHM.

We describe several characteristics of a novel smooth muscle cell line, SHM (Syrian hamster myometrium) derived from a primary uterine leiomyosarcoma which was induced by chronic estrogen plus androgen treatment of a female Syrian (golden) hamster. To determine the usefulness of the SHM cell line as a model for understanding myometrial function and its regulation, we have examined the morphologic and immunocytochemical properties of these cells, and the ability of uterotonic agonists to activate transmembrane signaling via phosphoinositide hydrolysis. The SHM cells exhibited a spindle-shape, smooth musclelike morphology when subconfluent, and a more compact, stellate shape at confluence. Like primary myocytes, SHM cells expressed the intermediate filament desmin and the contractile protein alpha smooth muscle actin, but not the epithelial antigen cytokeratin. Norepinephrine and bradykinin, which stimulate contraction and inositol polyphosphate production in the uterus, also stimulated inositol polyphosphate production in SHM cells. The maximal phosphoinositide signaling responses were lower in SHM cells compared with primary hamster uterine myocytes. We conclude that the SHM cell line exhibits primary uterine myocyte characteristics, and may therefore be a useful system for examining the mechanisms through which myometrial functions are regulated.

Animals↗
PubMed1992

The concentration of estrogen receptors in rabbit uterine myocytes decreases in culture.

OBJECTIVE: The purpose of this study was to determine whether the concentration of estrogen receptors in cultured myocytes is preserved after dispersion. STUDY DESIGN: Primary myocytes were prepared from rabbit myometrium by collagenase dispersion after removing the endometrium and were isolated with Percoll density gradients. The cells were assayed for estrogen receptor concentration at intervals after dispersion by means of a whole-cell binding assay. Unpaired t test was used for comparisons. RESULTS: The concentration of estrogen receptors on the first day after dispersion was 12,058 +/- 1096 sites per cell (mean +/- SEM) and decreased to 4389 +/- 1223 site per cell within 9 to 14 days after dispersion (63% decline, p < 0.001). A similar decrease was observed when 2 nmol/L estradiol was present in the medium. CONCLUSION: The concentration of estrogen receptors in isolated rabbit uterine myocytes decreases after dispersion. This may partly explain the difficulty of demonstrating in vitro estrogen effects on myocytes, which are well established in vivo.

Animals↗
PubMed1992

Cord-containing amniotic fluid pocket: a useful measurement in the management of oligohydramnios.

OBJECTIVE: To test the hypothesis that among patients with a low amniotic fluid (AF) index, those who have a large cord-containing AF pocket will have a lower risk for adverse perinatal outcome than women with a small cord-containing pocket. METHODS: Gravidas with an antepartum AF index no greater than 5 cm were studied prospectively. The vertical diameter of the single largest cord-containing pocket, which was excluded from the calculation of the AF index, was measured. Women with rupture of the membranes, multiple gestation, or fetal anomalies were excluded. RESULTS: Fifty-one women with gestational ages of 35-43 weeks were analyzed. Among 35 who had a cord-containing pocket of no more than 5 cm, eight (23%) had fetal distress necessitating operative delivery and eight neonates had cord arterial pH below 7.20. None of these complications occurred in the 16 women who had a cord-containing pocket above 5 cm (P < .05). The mean AF index (2.9 versus 2.8 cm) was not significantly different between the groups. CONCLUSION: Among women with a low AF index, a cord-containing pocket above 5 cm identifies a subgroup of women at lower perinatal risk compared with those with a smaller cord-containing pocket. The need for intervention may be reduced in women with a cord-containing pocket greater than 5 cm.

Adult↗
PubMed1991

Choriocarcinoma cells increase the number of differentiating human cytotrophoblasts through an in vitro interaction.

The human placenta arises from the zygote through single cell intermediates called cytotrophoblasts that in turn give rise to a syncytium. In culture, mononucleated cytotrophoblasts exhibit little, if any, cell division but are converted to multinucleated cells. Choriocarcinoma, the malignant tumor of placenta trophoblast, comprises a mixed population of dividing cellular intermediates that resemble cytotrophoblasts but are less differentiated. Because the choriocarcinoma intermediates arise from dividing cells, the tumor may contain one or more cell types in abundance not present in the population of isolated placental cells. To study placental differentiation through cell-cell interaction, choriocarcinoma cell lines were co-cultured with placenta-derived cytotrophoblasts, and placental hormone biosynthesis, as a marker of differentiation was examined. We reasoned that intermediates formed by the tumor might interact with and complement those intermediates in the placenta-derived cytotrophoblast population. Co-culturing either the JAr or JEG choriocarcinoma cell lines with cytotrophoblasts elevated the synthesis of the chorionic gonadotropin alpha and beta subunits 10-20 fold, and human placental lactogen 5-fold. The effect was specific for these trophoblast-derived cells, since comparable quantities of Chinese hamster ovary or HeLa cells did not affect the placental cytotrophoblast culture. Further experiments suggested that the source of enhanced synthesis was the cytotrophoblasts. We propose that an interaction between cytotrophoblasts and choriocarcinoma cells occurs, which results in an increased number of differentiating cytotrophoblasts. Such co-cultures may represent a model system for examining choriocarcinoma cell interaction with normal cells, a process known to occur in vivo. The data are also consistent with the hypothesis that the regulated chorionic gonadotropin production in the placenta is determined by interaction among trophoblast cells at different stages of differentiation.

Animals↗
PubMed1991

Tumor necrosis factor alpha is elevated in the peritoneal fluid from women with ruptured ectopic pregnancies.

The purpose of this study was to determine whether exposure of the peritoneum to fetal tissue is associated with elevated tumor necrosis factor alpha (TNF alpha) levels in the peritoneal cavity. We measured TNF alpha levels in the peritoneal fluids and serum from women with ruptured ectopic, unruptured ectopic and intrauterine pregnancies, as well as nonpregnant women undergoing tubal ligation. The results showed that patients with ruptured ectopic pregnancies were more likely to have TNF alpha levels in the peritoneal fluid greater than 40 units/ml (68%), compared with women with unruptured ectopic or intrauterine pregnancies (21%) (p less than 0.05, Fisher exact test). No elevation of peritoneal fluid TNF alpha levels was found in nonpregnant patients. Because TNF alpha is primarily a product of activated macrophages, it is likely that elevated TNF alpha levels in the peritoneal fluid of women with ruptured ectopic pregnancies reflects activation of peritoneal macrophages.

Adolescent↗
PubMed1991

Serum CA-125 levels in women with ectopic and intrauterine pregnancies.

In women with ectopic pregnancies the extrauterine compartments are exposed to fetal tissue. Since such tissue is known to express CA-125, we measured serum levels of CA-125 in patients with ectopic pregnancies and compared them to those in patients with intrauterine pregnancies. A total of 44 pregnant women were included in the study; all were in the first trimester. Twenty-seven of them had ectopic pregnancies, and 17 had intrauterine ones. Our results show that women with ectopic pregnancies, ruptured or unruptured, were more likely to have elevated levels of serum CA-125 than were women who had intrauterine pregnancies.

Adult↗
PubMed1989

Prophylactic amnioinfusion during labor complicated by meconium: a preliminary report.

The purpose of this prospective and randomized study is to evaluate the safety and efficacy of prophylactic transcervical amnioinfusion in the management of labor complicated by meconium. A preexisting intrauterine pressure catheter was used for amnioinfusion. The incidence of thick meconium was significantly lower after amnioinfusion, compared with standard management (5% versus 62% p less than 0.0005). This was also demonstrated with spectrophotometry. Significant differences between the amnioinfusion group and the control group were found for the following: arterial cord pH less than 7.20 (16% versus 38%, p less than 0.05); meconium more than trace below the vocal cords at delivery (0% versus 29%, p less than 0.05) and need for positive pressure ventilation at birth (16% versus 48%, p less than 0.05). No complications related to amnioinfusion were observed. We conclude that (1) transcervical amnioinfusion during labor complicated by meconium is a simple and apparently safe procedure, and that (2) amnioinfusion effectively decreases the frequency of thick meconium, the frequency of neonatal acidemia, the frequency of more than trace meconium below the vocal cords, and the need for positive pressure ventilation.

Adult↗
PubMed1986

Incidence of spontaneous and evoked fetal movements.

Fetal activity is composed of fetal movements (FM), which are either nonreflexive, originating in the fetus itself, or evoked reflexive FM which are stimulated by the fetal environment. The relative ratio of the spontaneous to evoked FM has not yet been documented. Since it is impossible to differentiate in a singleton pregnancy between these two types of FM on the basis of maternal perception alone, the natural model of twin pregnancy was used. It has previously been shown that the number of FM in a singleton pregnancy is similar to that of each individual fetus in a twin pregnancy of the same gestational age. It was assumed that the factors which cause spontaneous and evoked FM are similar in singleton and twin pregnancies. The fetal activity in 44 twin and 76 singleton pregnancies between 28 and 39 weeks of gestation was assessed during 20 min of nonstress test recording. The means of FM in twins were significantly higher than those in singleton pregnancies. The rate of the spontaneous FM was calculated to be 50-59% of all FM in singleton pregnancies and that of the evoked FM 41-50%.

Female↗
PubMedDate not supplied

Imprinting of PEG1/MEST isoform 2 in human placenta.

The PEG1 gene (a.k.a. MEST) is expressed in human placental trophoblast and endothelium, and data from knockout mice show that this gene regulates placental and fetal growth. Isoform 1 of PEG1 mRNA initiates from exon 1c and produces the long form of the MEST protein. This isoform is imprinted, with expression only from the paternal allele in many human and mouse organs, including placenta. In contrast, PEG1 isoform 2, initiating from exon 1a and producing the short form of MEST protein, is biallelically expressed (non-imprinted) in several non-placental organs. Here we show that PEG1 isoform 2 is in fact imprinted in a large subset of human placentae. A CpG island overlapping PEG1 exon 1a is unmethylated in various fetal and adult non-placental tissues, but is often substantially methylated in the placenta, with the extent of methylation in a large series approximating a normal distribution. Bisulfite conversion/sequencing indicates that the inter-individual differences reflect the relative representation of heavily methylated vs. unmethylated alleles, and RT-PCR/RFLP analysis shows strongly biased allelic expression of PEG1 isoform 2 mRNA in a majority of placentae with a high proportion of methylated alleles. These data highlight PEG1 isoform 2 as a marker for future studies of inter-individual epigenetic variation and its relation to placental and fetal growth in humans.

Adult↗