Reproductive Sciences recent issues

Review: Luminescence as a Tool to Assess Pelvic Endometriosis Development in Murine Models

Defrere, S., Colette, S., Lousse, J.-C., Donnez, J., Van Langendonckt, A. — Mon, 16 Nov 2009 19:08:49 PST

Classic murine endometriosis models may be insufficient to evaluate the effect of therapeutic agents on endometriosis development, because the process of identification and measurement of induced lesions is often impeded, as implants are small and embedded in murine tissue. In this context, as summarized in the current review, luminescence techniques have proved useful for identifying and visualizing or quantifying endometriotic transplants. They are also a valuable tool for endometrial cell tracking in live animals, yielding further information by adding spatial and temporal dimensions to biological processes in vivo. Such approaches involve transplanting luminescently labeled murine or human endometrium into animals. Two main strategies are applied to label endometrium before injection: use of genetically modified tissue or tissue labeled with a fluorescent dye. Each model has its advantages and disadvantages, the choice of model depends on the study objectives/design (long- or short-term studies, homologous or heterologous model).

Elafin (SKALP/Trappin-2/proteinase inhibitor-3) Is Produced by the Cervix in Pregnancy and Cervicovaginal Levels Are Diminished in Bacterial Vaginosis

Mon, 16 Nov 2009 19:08:50 PST

Objectives. To examine cervicovaginal elafin production in pregnancy and determine its relationship in bacterial vaginosis. Study Design. Samples of cervicovaginal secretions were collected from women with uncomplicated singleton pregnancies (n = 112) below 20 weeks gestation. Bacterial flora was assessed using Nugent’s criteria, and levels of elafin were measured by enzyme-linked immunosorbent serologic assay (ELISA). Elafin expression in the cervix was also examined by immunohistochemistry. In vitro expression of elafin was examined using cervix and vaginal cell lines. Results. Elafin is expressed in the cervical glandular epithelium. Elafin was found in all 112 samples of cervicovaginal secretions and levels were diminished in women with bacterial vaginosis (P < .05). Interleukin 1β (IL-1β) stimulated elafin expression in cells derived from the endocervix, but not in those derived from the vaginal epithelium. Conclusions. Elafin is a component of cervicovaginal secretions in pregnancy, and levels are diminished in bacterial vaginosis. It may be an important component of innate immunity in the lower genital tract.

Possible Gene-Gene Interaction of KIR2DL4 With its Cognate Ligand HLA-G in Modulating Risk for Preeclampsia

Mon, 16 Nov 2009 19:08:50 PST

Preeclampsia (PE) is a leading cause of maternal and fetal mortality and morbidity that occurs only during pregnancy. Pregnancy is the only physiological situation where killer-cell immunoglobulin-like receptors (KIRs) may meet cognate nonself variants of human leukocyte antigen (HLA) allotypes. We previously reported that presence of fetal HLA-G*0106 was significantly associated with risk for PE in multigravid pregnancies. We have now tested the KIR2DL4 receptor gene for association with PE, as well as for its interaction with HLA-G in modulating disease risk, in a case-control study of 83 PE and 240 normotensive pregnancies. No significant association was observed between alleles of KIR2DL4 and PE in both maternal and fetal groups, either among primigravid or multigravid pregnancies. Alleles of KIR2DL4 and HLA-G were then analyzed together to determine whether particular variant ligand—receptor combinations were associated with an increased risk for PE. Gene-gene interaction analyses suggest that the presence of fetal HLA-G*0106 in combination with maternal KIR2DL4*006 is significantly associated with PE risk in multigravid pregnancies (P < .001). These data provide the first preliminary evidence suggesting that although KIR2DL4 itself is not associated with PE, it may modulate the effect of HLA-G*0106 on risk for PE.

Aberrant Processing of Plasma Vitronectin and High-Molecular-Weight Kininogen Precedes the Onset of Preeclampsia

Blumenstein, M., Prakash, R., Cooper, G. J. S., North, R. A., SCOPE Consortium — Mon, 16 Nov 2009 19:08:50 PST

To date, there is no reliable test to identify women in early pregnancy at risk of developing preeclampsia. Difference gel electrophoresis (DIGE) identified the plasma proteins vitronectin (VN) and high-molecular-weight kininogen (HK) in association with preeclampsia. In a longitudinal proteomics study, the plasma of preeclamptic patients (n = 6) was compared to healthy control participants (n = 6) before the onset of preeclampsia (week 20) and at the time of presentation with clinical disease (weeks 33-36). The 75-kd single-chain VN molecule increased 1.6- to 1.9-fold in preeclampsia, whereas the 65-kd moiety of the 2-chain VN molecule decreased 1.5- to 1.7-fold compared to healthy controls (P < .05). Immunoblots revealed differences in proteolytic processing of VN and/or HK in women who develop preeclampsia or preeclampsia further complicated by small-for-gestational-age. Vitronectin and HK may prove to be useful as early markers of fibrinolytic activity and neutrophil activation, which are known to be associated with preeclampsia.

Transforming Growth Factor {beta}3 Regulates the Versican Variants in the Extracellular Matrix-Rich Uterine Leiomyomas

Mon, 16 Nov 2009 19:08:50 PST

Uterine leiomyoma are common, benign tumors that are enriched in extracellular matrix. The tumors are characterized by a disoriented and loosely packed collagen fibril structure similar to other diseases with disrupted Transforming growth factor β (TGF-β) signaling. Here we characterized TGF-β3 signaling and the expression patterns of the critical extracellular matrix component versican in leiomyoma and myometrial tissue and cell culture. We also demonstrate the regulation of the versican variants by TGF-β3. Using leiomyoma and matched myometrium from 15 patients, messenger RNA (mRNA) from leiomyoma and myometrium was analyzed by semiquantitative real time reverse transcription—polymerase chain reaction (RT-PCR), while protein analysis was done by western blot. Transforming growth factor β3 transcripts were increased 4-fold in leiomyoma versus matched myometrium. Phosphorylated-TGF-β RII and phosphorylated-Smad 2/3 complex were greater in leiomyoma as documented by Western blot. The inhibitor Smad7 transcripts were decreased 0.44-fold. The glycosaminoglycan (GAG)-rich versican variants were elevated in leiomyoma versus myometrial tissue: specifically V0 (4.27 ± 1.12) and V1 (2.01 ± 0.27). Treatment of leiomyoma and myometrial cells with TGF-β3 increased GAG-rich versican variant expression 7 to 12 fold. Neutralizing TGF-β3 antibody decreased the expression of the GAG-rich versican variants 2 to 8 fold in leiomyoma cells. Taken together, the aberrant production of excessive and disorganized extracellular matrix that defines the leiomyoma phenotype involves the activation of the TGF-β signaling pathway and excessive production of GAG-rich versican variants.

Gonadotropin Stimulation Increases the Expression of Angiotensin-(1--7) and Mas Receptor in the Rat Ovary

Mon, 16 Nov 2009 19:08:50 PST

We have previously shown the presence of immunoreactive angiotensin-(1—7) [Ang-(1—7)] in rat ovary homogenate and its stimulatory effect on estradiol and progesterone production in vitro. In the current study, we investigated the presence and cellular distribution of Ang-(1—7) and the Mas receptor, the expression of Mas and angiotensin-converting enzyme 2 (ACE2) messenger RNA (mRNA), and the enzymatic activity in the rat ovary following gonadotropin stimulation in vivo. Immature female Wistar rats (25 days old) were injected subcutaneously (SC) with equine chorionic gonadotropin (eCG, 20 IU in 0.2 mL) or vehicle 48 hours before euthanasia. Tissue distributions of Ang-(1—7), Mas receptor, and ACE2 were evaluated by immunohistochemistry, along with angiotensin II (Ang II) localization, while the mRNA expression levels of Mas receptor and ACE2 were evaluated by real-time polymerase chain reaction (PCR). In addition, we determined the activity of neutral endopeptidase (NEP), prolyl endopeptidase (PEP), and ACE by fluorometric assays. After eCG treatment, we found strong immunoreactivity for Ang-(1—7) and Mas primarily in the theca-interstitial cells, while Ang II appeared in the granulosa but not in the thecal layer. Equine chorionic gonadotropin treatment increased Mas and ACE2 mRNA expression compared with control animals (3.3- and 2.1-fold increase, respectively; P < .05). Angiotensin-converting enzyme and NEP activities were lower, while PEP activity was higher in the eCG-treated rats (P < .05). These data show gonadotropin-induced changes in the ovarian expression of Ang-(1—7), Mas receptor, and ACE2. These findings suggest that the renin-angiotensin system (RAS) branch formed by ACE2/Ang-(1—7)/Mas, fully expressed in the rat ovary and regulated by gonadotropic hormones, could play a role in the ovarian physiology.

Regulation of Human Umbilical Artery Contractility By Different Serotonin and Histamine Receptors

Santos-Silva, A. J., Cairrao, E., Marques, B., Verde, I. — Mon, 16 Nov 2009 19:08:50 PST

We studied the role of several serotonin (5-HT) and histamine receptors in the regulation of human umbilical artery (HUA) contractility. Among the 5-HT agonists used, only the 5-HT_2A and 5HT_1B/D agonists contracts HUA. The 5-HT-induced contractions were fully inhibited by ketanserin (5-HT_2A antagonist). The 5-HT₇-activation also relaxes and increases intracellular cyclic adenosine monophosphate (cAMP). Among the histamine receptor agonists, only betahistine (H₁ agonist) induced significant contractile effect. Histamine-induced contraction was partially relaxed by pyrilamine (H₁ antagonist). Betahistine-induced contraction was partially blocked by dimaprit (H₂ agonist) and by the H₃ agonist when a low concentration of forskolin is present. Both, H₂ and H₃ agonists increased the cAMP intracellular levels in HUA smooth muscle. These findings show that in HUA, 5-HT_2A- and 5-HT_1B/1D-activation lead to vasoconstriction and 5-HT₇-activation induces vasorelaxation. Concerning histamine receptors, H₁-activation induces contraction and H₂- and H₃-activation lead to vasorelaxation.

Enhanced Expression of P2X4 and P2X7 Purinergic Receptors in the Myometrium of Pregnant Rats in Preterm Delivery Models

Urabe, S., Miyoshi, H., Fujiwara, H., Yamaoka, K., Kudo, Y. — Mon, 16 Nov 2009 19:08:50 PST

The expression levels of P2X purinergic receptors were determined in the myometrium of pregnant rats using the quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR). The messenger RNAs (mRNAs) of P2X4 and P2X7 were expressed most strongly. The expression levels of these receptors increased during the late stages of pregnancy; at the time of delivery, the mRNA levels of P2X4 and P2X7 had increased to 1.9 and 3.2 times the day 19 values, respectively. We also explored the roles of P2X receptors in hormone-induced and inflammation-induced preterm delivery models. In the former, mifepristone caused the P2X4 and P2X7 mRNA levels to increase to 2.1 and 4.1 times the control values, respectively. In the latter, lipopolysaccharide (LPS) caused the mRNA levels of P2X4 and P2X7 to increase dramatically to 7.4 and 18.6 times the control values, respectively. These findings suggest that increased P2X4 and P2X7 receptor expression in pregnant rats is related to uterine contraction leading to term and preterm delivery.

The Role of IGF-1 and Ghrelin in the Compensation of Intrauterine Growth Restriction

Mon, 16 Nov 2009 19:08:50 PST

The role of insulin-like growth factor 1 (IGF-1) and ghrelin in intrauterine growth restricted (IUGR) neonates in comparison to appropriate for gestational age (AGA) ones was investigated. Levels of IGF-1/insulin-like growth factor binding protein 3 (IGFBP3), ghrelin, insulin, and cortisol were determined in 20 singleton, full-term IUGR and 20 respective AGA neonates at birth (umbilical cord-UC), on days 1 (d1) and 4 (d4) postnatally. The ratio of IGF-1 to birth weight was higher in IUGR than in AGA in both UC (18.2 ± 1.2 vs14.4 ± 0.9, P = .05) and d1 (9.6 ± 0.5 vs 6.8 ± 0.3, P = .05). A significant positive correlation was found between IGF-1 and ghrelin levels and a negative one between IGFBP3 and ghrelin only in IUGR. In both groups, fetal IGF-1 levels negatively correlated with fetal cortisol levels. Intrauterine growth restricted neonates demonstrate a relative IGF-1 resistance in an attempt to drive energy toward survival on the expense of growth. The observed correlations between ghrelin and IGF-1/IGFBP3 postnatally indicate that ghrelin might play a role in the compensation of intrauterine undernutrition, promoting postnatal growth.

Do Alterations in Placental 11{beta}-Hydroxysteroid Dehydrogenase (11{beta}HSD) Activities Explain Differences in Fetal Hypothalamic-Pituitary-Adrenal (HPA) Function Following Periconceptional Undernutrition or Twinning in Sheep?

Mon, 16 Nov 2009 19:08:50 PST

Periconceptional undernutrition (UN) in sheep accelerates fetal hypothalamic-pituitary-adrenal (HPA) axis activation, resulting in preterm birth. In contrast, twin conception suppresses fetal HPA function and delays prepartum HPA activation. We hypothesized that these dissimilar effects on fetal HPA activity result from different influences of maternal glucocorticoid (GC) on maturation of the fetal HPA axis, mediated via different activities of placental 11β-hydroxysteroid dehydrogenase (11βHSD) isozymes. We examined the effects of twinning and maternal periconceptional UN from 60 days before until 30 days after mating on the ontogeny of placental 11βHSD-1 and -2 enzyme activities. At day 85 of gestation, placental 11βHSD-2 activity was lower in UN than in normally nourished (N) fetuses (P < .05) and was higher in twins than in singletons (P < .05). Furthermore, placental 11βHSD-1 activity was not different between nutritional groups but was higher in twins than in singletons (P = .01). At day 85, fetal plasma cortisol (P < .001) and cortisone (P < .001) concentrations were lower in UN than in N fetuses, but the cortisol to cortisone ratio was higher in UN than in N fetuses (P = .01). There was no effect of fetus number on plasma cortisol or cortisone concentrations or on the ratio of cortisol to cortisone at day 85. Therefore, periconceptional UN and twinning may result in the alterations of placental 11βHSD isozyme activities at particular times during gestation. Changes in these activities during critical periods of fetal development could affect transplacental transfer or placental generation of GCs that reach the fetus, potentially influencing the timing of activation of the fetal HPA axis, fetal maturation, and hence the development and health later in life.

Proteinuria in Women With Preeclampsia: Understanding the Dialogue Between 2 Neighbors

Henao, D. E. — Thu, 08 Oct 2009 15:34:26 PDT

Review: Human Endogenous Retroviruses and the Placenta

Sugimoto, J., Schust, D. J. — Thu, 08 Oct 2009 15:34:26 PDT

Up to 8% of the human genome is of retroviral origin. These stably integrated retroviral sequences that characterize the human endogenous retrovirus (HERV) arose from retroviral infections that occurred more than 25 million years ago. The host and the retrovirus have subsequently coevolved as retrovirally derived genetic material is propagated in a Mendelian fashion. Although most HERV sequences are silenced, several have been described that are functional. The effects of some HERV-derived products are linked to human disease; others appear essential to human organ function. The human placenta, unique in its active expression of retroviral sequences that are not expressed in other tissues, may hold the key to an improved understanding of the functional significance of HERVs. In this review, we discuss the contribution of retroelements, particularly HERVs, to placental function and dysfunction. We describe fusogenic and immunosuppressive HERV activities and emphasize epigenetic regulation of retroelement expression.

Postpartum Outcome of Cervical Intraepithelial Neoplasia in Pregnant Women Determined by Route of Delivery

Thu, 08 Oct 2009 15:34:26 PDT

Cervical intraepithelial neoplasia (CIN) has its highest incidence during women’s reproductive years. During 2 sequential 7-year periods, 1994 to 2000 and 2001 to 2007, 3695 and 3894 deliveries were performed, respectively, at Osaka University Hospital. CIN was detected in 21 cases (0.57%) during 1994-2000 and in 43 cases (1.1%) during 2001-2007. By comparison, cervical intraepithelial neoplasia—complicated pregnancies increased significantly in the latter period (P = .015 by Fisher exact test, Odds ratio = 1.95; 95%CI: 1.16-3.30). We observed CIN regression in 34 (76%) of 45 cases of vaginal delivery and in 6 (50%) of 12 cases of cesarean delivery, indicating that the outcome of an initially diagnosed CIN and the delivery routes appeared not to be significantly related. However, a different result was obtained when only those patients whose CIN lesions persisted until the delivery were analyzed. Among the 35 such cases in the vaginal delivery group, 24 cases (69%) regressed after the delivery; in 8 such cases from the cesarean delivery group, only 2 cases (25%) regressed afterward. Our study clearly shows that pregnancy complicated with CIN is increasing rapidly in Japan. We also find that there is a significantly more frequent postpartum regression of biopsy-proven CIN lesions following a vaginal delivery compared to cesarean section (P = .042 by Fisher exact test, Odds ratio = 6.55; 95% CI: 1.13-37.8).

Imidazole-Based Erythrocyte Markers of Oxidative Stress in Preeclampsia--An NMR Investigation

Turner, E., Brewster, J. A., Simpson, N. A. B., Walker, J. J., Fisher, J. — Thu, 08 Oct 2009 15:34:26 PDT

Using ¹H-nuclear magnetic resonance (NMR) spectroscopy and statistical models, we sought to identify ‘‘biomarkers’’ present in erythrocytes that would distinguish between women with normal pregnancy and those suffering from preeclampsia, and investigate possible links with previously identified plasma ‘‘markers.’’ Erythrocytes from 22 normotensive pregnant women and 15 preeclamptics were analyzed by ¹H Carr-Purcell-Meiboom-Gill (CPMG) NMR. Multivariate analysis and logistic regression were applied to differentiate between the 2 groups of patients, and used to develop a diagnostic model based on the concentrations of the constituents identified as being influential. Significantly higher concentrations of alanine (P < .001), glycine (P = .025), and ergothioneine (P = .049) were found in erythrocytes from preeclamptic patients. Discriminant analysis and regression of NMR data permitted 100% accurate diagnosis of the health status of new patients. Chemically related imidazole-based molecules, histidine and ergothioneine, are important in the classification process and the etiology of preeclampsia (PE).

The Effect of Progesterone on Myometrial Contractility, Potassium Channels, and Tocolytic Efficacy

Anderson, L., Martin, W., Higgins, C., Nelson, S. M., Norman, J. E. — Thu, 08 Oct 2009 15:34:26 PDT

Objectives: Recent clinical trials have demonstrated a beneficial effect of supplementation with progesterone to prevent preterm labor. We aimed to determine the effects of progesterone treatment in vitro and in vivo and 17-hydroxyprogesterone caproate (17OHPC) in vitro on myometrial contractions. Methods: Myometrial strips were taken from women undergoing cesarean delivery at term. We also obtained myometrial biopsies from women participating in a clinical trial of progesterone to prevent preterm labor in twins (STOPPIT). After establishment of spontaneous contractions, strips were exposed to progesterone or 17OHPC. Separate strips were exposed to oxytocin and tocolytics alone and in combination with progesterone. Potassium channel blockers were added in conjunction with progesterone. STOPPIT samples were used to compare the effects of in vivo progesterone and placebo. We measured amplitude, frequency and activity integral of contractions. Results: Maximum inhibition of contraction amplitude was 93 ± 2% and 67 ± 14% for progesterone at 30 µM and vehicle (70% ethanol), respectively, P < 0.05. 17OHPC did not exert an inhibitory effect. Water soluble progesterone exerted a maximal inhibitory effect on amplitude of contractions of 82 ± 10% at 100 µM, P < 0.05. The inhibitory effect of progesterone was unaffected by potassium channel blockers. There was no difference between in vivo placebo and progesterone-treated groups in either amplitude or frequency of contractions, nor was there any difference in the response to oxytocin or the tocolytic drugs. Conclusions: Progesterone exerts rapid inhibition of the amplitude of myometrial contractions in vitro but 17OHPC does not. The action of progesterone does not appear to operate via potassium channels nor does it enhance the activity of certain tocolytic drugs.

Loss of Proliferative Capacity in a Retroviral Immortalized Human Uterine Smooth Muscle Cell Line Derived From Leiomyoma Is Restored by hTERT Overexpression

Chang, B., Myatt, L., Cui, X.-L. — Thu, 08 Oct 2009 15:34:26 PDT

Overexpression of human telomerase reverse transcriptase (hTERT) has facilitated establishing in vitro model systems for biological research. The plasmid containing hTERT gene was stably transfected into ULTR cells, a retroviral transformed human uterine leiomyomatous smooth-muscle cell line. Cells that express hTERT, termed as ULTR-hT, shared the morphological characteristics of the parental proliferative ULTR cells. They expressed a set of smooth-muscle-specific genes and had increased proliferation rate and prolonged lifespan. Quantitative real-time polymerase chain reaction (PCR) analysis revealed a correlation of proliferation rates of ULTR-hT clonal cells with the level of hTERT expression. ULTRhT cells also preserved expression of estrogen, progesterone, and oxytocin receptor genes, confirming a myometrial phenotype. Expression of angiotensin II receptors and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase isoforms were also preserved. Our finding suggests that ULTR-hT cells can be a useful in vitro model for studying human myometrium differentiation both in pregnancy and pathological growth.

Local Uteroplacental Influences are Responsible for the Induction of Uterine Artery Myogenic Tone during Rat Pregnancy

Gokina, N. I., Kuzina, O. Y., Fuller, R., Osol, G. — Thu, 08 Oct 2009 15:34:26 PDT

Uterine artery constrictor responses to elevation of intraluminal pressure (myogenic tone) are considerably enhanced in late pregnant rats, although the underlying causes remain unknown. A single uterine horn ligation model was used to differentiate local from systemic influences, and to test the hypothesis that factors associated with the site of placentation, rather than systemic hormonal changes, are primarily involved in the induction of this adaptive process. Radial uterine arteries were dissected from the gravid and nongravid uterine horns of late pregnant rats, cannulated, and pressurized. Changes in arterial diameter and smooth muscle [Ca²⁺]_i in response to the elevation of intraluminal pressure were studied using intact and endothelium-denuded arteries loaded with the ratiometric Ca²⁺-sensitive dye fura-2. Elevations of pressure from 10 to 60 and 100 mm Hg resulted in passive arterial distention of arteries from nongravid horns with a minor change in [Ca²⁺]_i. In contrast, arteries from gravid horns developed myogenic tone associated with a significant elevation in [Ca²⁺]_i. Synchronous oscillations in [Ca²⁺]_i and lumen diameter were frequently observed in vessels from gravid horns. Endothelial denudation augmented tone in the gravid horn but did not uncover myogenic tone in vessels from the nongravid horn. In summary, pregnancy-associated uterine artery myogenic behavior is due to an upregulation of calcium-handling mechanisms, occurs independently of the endothelium, and is induced by local uteroplacental influences.

Plasma From Women With Preeclampsia Inhibits Trophoblast Invasion

Harris, L.K., Clancy, O.H., Myers, J.E., Baker, P.N. — Thu, 08 Oct 2009 15:34:26 PDT

To assess whether plasma from women with preeclampsia altered trophoblast invasion, SGHPL-4 extravillous trophoblasts were treated with pooled plasma from women with preeclampsia (PE-P; 10%) or with plasma from healthy pregnant controls (C-P). PE-P significantly inhibited SGHPL-4 invasion through Matrigel-coated transwells (P < .01), reduced mitochondrial dehydrogenase activity (P < .01), and increased apoptosis (P < .05); however, invading cells were no more susceptible to PE-P-induced apoptosis than their static counterparts. C-P did not alter rates of invasion, proliferation, or apoptosis. The bioactivity of PE-P was retained after removal of the 6 most abundant plasma proteins using an immunodepletion column (P < .05). Fractionation of PE-P demonstrated that the reduction in invasion was predominantly mediated by factors >100 kd in size. The authors conclude that plasma from women with preeclampsia contains multiple factors that inhibit invasion. These factors do not specifically target invading cells, but instead may reduce the number of cells available to invade.

Relationship Between Prepregnancy and Early Pregnancy Uterine Blood Flow and Resistance Index

Hale, S. A., Schonberg, A., Badger, G. J., Bernstein, I. M. — Thu, 08 Oct 2009 15:34:26 PDT

We evaluated the relationship between prepregnancy and early pregnancy uterine blood flow (UBF) and resistance index (RI). Nineteen nulliparous participants were studied during cycle day 8 + 4, and early pregnancy (13.4 + 1.6 weeks). Color Doppler ultrasound of both uterine arteries and maternal heart was performed to calculate uterine RI, volumetric UBF, and cardiac output (CO), respectively. We observed a strong negative association of uterine RI with prepregnancy UBF (r = —.82, P < .001) that weakened, but remained significant in early pregnancy (r =—.48, P = .04). Prepregnancy uterine index (UBF/CO) was significantly associated with early pregnancy uterine index; r = .48, P = .04). There was also a trend associating prepregnancy and early pregnancy volumetric UBF (r = .44, P = .068). Prepregnancy UBF may be a determinant of early pregnancy UBF and UBF may have independent value as a predictor of adverse pregnancy outcome.

Pharmacokinetic Comparison Between Systemic and Local Chemotherapy by Carboplatin in Dogs

Chen, C., Wang, W., Zhou, H., Huang, J., Liu, P., Song, T., Sun, M. — Thu, 08 Oct 2009 15:34:26 PDT

The aim of the current study was to compare pharmacokinetics of local chemotherapy by pelvic intra-arterial administration with intravenous injection in dogs. A total of 18 female dogs (weight: 10-15 kg) were randomly divided into 3 groups: the peripheral vein administration (group A, n = 6), the abdominal aorta administration (group B, n = 6), and the internal iliac artery administration (group C, n = 6). Carboplatin at a dose of 1.2 mg/kg was administered by infusing into the arteries or the vein. For analysis, plasma and uterine tissue samples were collected at different times following infusion. The peak local concentration of platinum in the uterus of dogs in group C was significantly higher than those of groups A and B (P < .05). The area under the tissue concentration—time curve (AUC) of uterine tissues was significantly higher in dogs of group C compared to those of the other 2 groups (P < .05). There was no significant difference in the AUC of the uterine tissues of dogs between groups A and B (P > .05). The peak concentration of platinum in plasma was significantly higher in group A compared to those of the other 2 administration routes (P < .05). We observed the pharmacokinetic advantages of local chemotherapy by internal iliac artery perfusion with the chemotherapeutic agent, carboplatin, to the uterus, thereby leading to a high-drug concentration that may be more effective in treating cervical cancer.

Correlation of Human Leukocyte Antigen-G (HLA-G) Expression and Disease Progression in Epithelial Ovarian Cancer

Thu, 08 Oct 2009 15:34:26 PDT

Human leukocyte antigen-G (HLA-G) expression has been reported to be relevant to cancer development and immune tolerance. The purpose of this study was to investigate the correlation between HLA-G expression and disease progression and to assess the use of HLA-G expression as a prognostic immunomarker in epithelial ovarian carcinoma. Human leukocyte antigen-G expression in 41 ovarian cancer tissues and 8 normal ovarian tissues was analyzed using immunohistochemistry and Western blot assay. Quantitative real-time reverse transcription polymerase chain reaction (RT-PCR) was used for HLA-G messenger RNA (mRNA) expression. Human leukocyte antigen-G mRNA and protein levels were significantly greater in advanced ovarian cancer tissues than in normal or early stage ovarian cancer tissues (P < .05 and P < .05, respectively). Patients with HLA-G expression had a significantly worse prognosis. There is a significant correlation between HLA-G immunoreactivity and patient survival in univariate analysis (P = .04). Our data was consistent with the concept that HLA-G expression might play a pivotal role in the development and disease progression of epithelial ovarian cancer.

Reviews: Adipocytokines in Normal and Complicated Pregnancies

Briana, D. D., Malamitsi-Puchner, A. — Wed, 16 Sep 2009 16:55:56 PDT

Human pregnancy is characterized by insulin resistance, traditionally attributed to the effects of placental hormones. Normal pregnancy—induced insulin resistance is further enhanced in pregnancy complications, associated with disturbed placental function, such as gestational diabetes mellitus, preeclampsia, and intrauterine growth restriction. Compelling evidence suggests that these pregnancy disorders are associated with future development of maternal metabolic syndrome. However, the pathogenetic mechanisms underlying the association between abnormal placental development, insulin resistance, and maternal metabolic syndrome are not fully understood. A large body of evidence has recently supported the role of adipose tissue in the regulation of insulin resistance in both nonpregnant and pregnant participants. In this respect, adipocytokines, which are adipocyte-derived hormones, have been implicated in the regulation of maternal metabolism and gestational insulin resistance. Adipocytokines, including leptin, adiponectin, tumor necrosis factor , interleukin 6, as well as the newly discovered resistin, visfatin, and apelin, are also known to be produced within the intrauterine environment. However, data concerning the pattern of adipocytokines secretion in normal and complicated pregnancies are still limited and partially contradictory. Given the importance of adipose tissue and its hormones in terms of adequate metabolic control and energy homeostasis, we present a review of published data related to the role of adipocytokines in pregnancy, especially in relation to pregnancy complications. Focus will be placed on the functions and other potential roles of the novel adipocytokines resistin, visfatin, and apelin.

Metformin Does Not Improve the Reproductive or Metabolic Profile in Women With Polycystic Ovary Syndrome (PCOS)

Wed, 16 Sep 2009 16:55:56 PDT

To determine whether metformin, when given to women with polycystic ovary syndrome (PCOS), promotes folliculogenesis by prompting a drop in free sex steroids resulting in a compensatory follicle stimulating hormone (FSH) rise, we conducted a randomized, double-blind, placebo-controlled crossover clinical trial. Eight mid-reproductive age PCOS participants with mean obese body mass index (BMI) and normal glucose tolerance received 8 weeks of metformin, given in a step-up fashion to a maximum dose of 2000 mg daily or placebo with daily urine sampling, 4-6 weeks washout, and crossover to the remaining arm for 8 weeks. To confirm the effects of metformin on glucose and other metabolic markers, a hyperinsulinemic, euglycemic 3-dose clamp (physiologic: 30 mU/m² per minute, high: 400 mU/m² per minute) followed each treatment. Urinary FSH, luteinizing hormone (LH), or pregnanediol glucuronide (Pdg) did not differ by treatment. Glucose disposal, endogenous glucose production, BMI, ovulation rates, serum sex steroids, free fatty acids, and lipids did not significantly differ by treatment, despite good evidence for compliance with the protocol. During the clamp, high-dose insulin administration was associated with an acute drop in serum LH. We conclude that short-term, high-dose metformin exerts minimal effects on both metabolic markers and reproductive hormones in a small sample of overall morbidly obese women.

A Major Effect of Simulated Microgravity on Several Stages of Preimplantation Mouse Development is Lethality Associated With Elevated Phosphorylated SAPK/JNK

Wang, Y., Xie, Y., Wygle, D., Shen, H. H., Puscheck, E. E., Rappolee, D. A. — Wed, 16 Sep 2009 16:55:56 PDT

We tested whether microgravity affects mouse development during a period when gravity cues chick and frog embryo development. A rotating vessel developed ~ 0.1% simulated microgravity (MGS) for embryos. Microgravity simulation resulted in blocked cell accumulation in E2.5 embryos. E1.5 and E3.5 embryos showed lesser effects. For E1.5/2.5 embryos, cell accumulation block was followed by lethality at 48 hours after MGS. For E3.5 embryos, MGS blocked development without lethality but with apoptosis. E1.5-3.5 embryos from the rotational control developed lesser effects than MGS embryos. Embryonic stress-activated protein kinase (SAPK) was phosphorylated during MGS and mediated apoptosis. Increased pSAPK suggested that lethality is due to cellular stress induced by MGS, unlike the dysfunctional development after gravitational disorientation in frog and chick embryos. Thus, MGS causes lethality, a novel phenotype not often observed in microgravity or MGS. Embryonic lethality at E2.5 and apoptosis at E3.5 are associated with SAPK function, suggesting that MGS causes a general stress response that immediately affects many aspects of development. In addition, MGS and many aspects of In vitro fertilization/assisted reproductive technologies (IVF/ART) produce nonphysiological, nonevolutionary stresses that are mediated by SAPK, suggesting the primacy of this protein kinase in a wide range of mechanisms mediating negative reproductive outcomes in IVF/ART and potentially in spaceflight.

Expression and Organization of Basement Membranes and Focal Adhesion Proteins in Pregnant Myometrium is Regulated by Uterine Stretch

Shynlova, O., Chow, M., Lye, S. J. — Wed, 16 Sep 2009 16:55:56 PDT

The mechanisms underlying the preparation of the uterus for labor are not fully understood. We have previously found a significant increase in the expression of messenger RNA (mRNAs) encoding extracellular basement membrane (BM) proteins of the smooth muscle cells (SMCs) in late pregnant rat myometrium. At term, the myometrium is stretched by growing fetuses and these mechanical signals are transmitted from extracellular matrix into SMCs through focal adhesions (FA). The aim of this study was to investigate the effect of gravidity on the expression and spatiotemporal distribution of major BM proteins, laminin-2 and collagen IV, as well as typical FA constituents, vinculin and paxillin, in the myometrium during gestation and parturition, using a unilaterally pregnant rat model. We found that the expression of laminin-2 and collagen IV proteins increased significantly with gestational age (P < .05) and was dependent on gravidity whereas vinculin and paxillin proteins were not affected. Near term, BM proteins from gravid horn myometrium demonstrated increased extracellular immunostaining and major rearrangement from sporadic protein distribution to organized, continuous, and regular structures surrounding the plasma membrane of each myocyte. Examination of FA proteins revealed that paxillin was translocated from the cytoplasm to the cell periphery, while vinculin was sequestered specifically to FAs. At labor, BM and FA proteins, organized in similar bead-like structures, were localized on opposing sides of SMC plasma membrane into 2 different compartments. We suggest that these stretch-induced changes facilitate formation of stable cell-matrix adhesions and provide the molecular basis for optimal force transduction during labor contractions.

Decreased Nephrin and GLEPP-1, But Increased VEGF, Flt-1, and Nitrotyrosine, Expressions in Kidney Tissue Sections From Women With Preeclampsia

Zhao, S., Gu, X., Groome, L. J., Wang, Y. — Wed, 16 Sep 2009 16:55:56 PDT

Renal injury is a common pathophysiological feature in women with preeclampsia as evidenced by increased protein leakage (proteinuria) and glomerular injury (glomerular endotheliosis). Recently, podocyturia was found in preeclampsia, suggesting podocyte shedding occurs in this pregnancy disorder. However, podocyte function in preeclampsia is poorly understood. In this study, the authors have examined podocyte-specific protein expressions for nephrin, glomerular epithelial protein 1 (GLEPP-1), and ezrin in kidney biopsy tissue sections from women with preeclampsia. Expressions for vascular endothelial growth factor (VEGF) and its receptor Flt-1 and oxidative stress marker nitrotyrosine and antioxidant CuZn-superoxide dismutase (CuZn-SOD) were also examined. Kidney tissue sections from nonhypertensive and chronic hypertensive participants were stained as controls. The findings were (1) nephrin and GLEPP-1 were mainly expressed in glomerular podocytes; (2) ezrin was expressed in both glomerular podocytes and tubular epithelial cells; (3) compared to tissue sections from nonhypertensive and chronic hypertensive participants, nephrin and GLEPP-1 expressions were much reduced in tissue sections from preeclampsia and ezrin expression was reduced in podocytes; (4) enhanced VEGF, Flt-1, and nitrotyrosine, but reduced CuZn-SOD, expressions were observed in both glomerular podocytes and endothelial cells in tissue sections from preeclampsia; and (5) the expression pattern for nephrin, GLEPP-1, ezrin, VEGF, Flt-1, and CuZn-SOD were similar between tissue sections from nonhypertensive and chronic hypertensive participants. Although the authors could not conclude from this biopsy study whether the podocyte injury is the cause or effect of the preeclampsia phenotype, the data provide compelling evidence that podocyte injury accompanied by altered angiogenesis process and increased oxidative stress occurs in kidney of patients with preeclampsia.

The Relationship of Plasma Volume, Sympathetic Tone, and Proinflammatory Cytokines in Young Healthy Nonpregnant Women

Bernstein, I. M., Damron, D., Schonberg, A. L., Shapiro, R. — Wed, 16 Sep 2009 16:55:56 PDT

Objective. Preeclampsia has been associated with elevated proinflammatory markers, increased sympathetic activity, and decreased plasma volume (PV). We hypothesized that these associations would be identified in women prior to a first pregnancy. Methods. We studied 76 healthy nulligravid participants measuring the proinflammatory markers C-reactive protein (CRP), interleukin-6 (IL-6), and tumor necrosis factor (TNF-). Plasma volume was measured in supine position and corrected for body mass index (BMI). We examined supine plasma levels of epinephrine and norepinephrine and blood pressure response to Valsalva maneuver to quantify sympathetic activation. We then examined the association of PV and sympathetic activity with proinflammatory cytokines with P < .05 accepted for significance. Results. CRP was significantly increased in participants with lowest PV/BMI quartile when compared to middle 2 quartiles and highest quartile (analysis of variance [ANOVA], P = .037). We found no significant association of PV/BMI with either IL-6 or TNF-. Both plasma epinephrine concentration (r = .29, P = .02) and the late phase II (phase II_L) blood pressure response to Valsalva maneuver (r = .44, P < .0001) were associated with serum IL-6 concentrations. Conclusions. Low PV is associated with increased CRP levels and increased sympathetic tone is linked to elevated IL-6 concentration in young nonpregnant women. These findings represent elements of a nonpregnancy phenotype that parallels the findings observed in preeclampsia and in women at risk for ischemic cardiovascular disease. This suggests that the relationships observed during preeclampsia, which have been associated with placental pathology, may predate pregnancy and be independent of placental activity.

Declines in Levels of Hyperpolarization-Activated Cation (HCN) Channels in the Rat Ovary After Cisplatin Exposure

Yeh, J., Beom Su Kim, , Peresie, J., Page, C. — Wed, 16 Sep 2009 16:55:56 PDT

The distribution of the 4 hyperpolarization-activated cation (hyperpolarization-activated, cyclic nucleotide-gated [HCN]) channels (HCN1-4), channels associated with rhythmic electrical bursting, varies in the different cells of the ovary. Cisplatin is a broadly used chemotherapeutic agent that results in ovarian damage. The objective of this study was to test the hypothesis that cisplatin treatment will affect expression of the ovarian HCN channels. Adult female Sprague-Dawley rats were dosed with saline, 4.5 mg/kg cisplatin, or 6.0 mg/kg cisplatin as 2 weekly injections and then were killed 5 days after the second cisplatin dose. The ovaries were studied for HCN1-4 using semiquantitative H score immunohistochemical analysis and using Western blot analysis. By immunohistochemistry, for HCN4, the oocyte-specific staining declined after cisplatin treatment (P < .001). There was a decline in HCN2 intensity after cisplatin in the antral granulosa cells (P < .05) and thecal cells (P < .05). There were no differences found for the immunohistochemical HCN1 and HCN3 H scores after cisplatin. The Western blot analysis revealed that there was a decline in HCN3 levels after cisplatin exposure (P < .05). For HCN1 and HCN2, there was no change in the total ovarian protein levels after cisplatin. Treatment in vivo with cisplatin resulted in decline in the levels of the HCN channels in the rat ovary, with oocytes, antral cells, and thecal cells being specifically affected.

Increased Circulating Levels of the Antiangiogenic Factor Endostatin in Early-Onset But Not Late-Onset Preeclampsia

Wikstrom, A.-K., Larsson, A., Akerud, H., Olovsson, M. — Wed, 16 Sep 2009 16:55:56 PDT

Changes in circulating angiogenic factors seem to play a key role in the pathogenesis of preeclampsia and it seems as if these changes are of greater importance in the pathogenesis of early-onset than of late-onset disease. Endostatin is a potent, broad spectrum antagonist of angiogenesis whose role in preeclampsia is poorly investigated. The aim of this study was to estimate whether circulating endostatin levels are altered in preeclampsia, and whether women with early-onset (before 32 weeks of gestation; n = 16) and late-onset (after 35 weeks of gestation; n = 19) preeclampsia differ in this regard. Women with early-onset, but not of late-onset preeclampsia had higher levels of endostatin than healthy pregnant women in corresponding lengths of gestation. The results of the study support the hypothesis that there is heterogeneity between early- and late-onset preeclampsia, with a stronger association between an altered angiogenic balance and early-onset than late-onset disease.

Clostridium botulinum Toxin A Inhibits Contractility in Pregnant Human Myometrium In Vitro

Burd, I. D., Ness, A., DiMuzio, P., Ren, G.-Y., Tulenko, T. N. — Wed, 16 Sep 2009 16:55:56 PDT

Studies were undertaken to evaluate the effect of Botulinum neurotoxin type-A (BoNTA) preparation on oxytocin-induced contractions of pregnant human myometrium in vitro. Human myometrial tissue was exposed to increasing concentrations (1-50 000 U/mL) of BoNT/A. Isometric contractions were measured using a force displacement transducer. The cumulative effect of BoNT/A on myometrial activity (time to half relaxation [TTR50], frequency, and amplitude) was evaluated. The frequency of myometrial contractions was depressed by 40% from baseline (P < .05) and relaxation time was increased by 30% (P < .05) from baseline within a narrow range of concentrations. There was no significant difference in amplitude. The observed effects were rapidly reversed after complete wash out of the tissue. BoNT/A or its analogues with more specific tissue affinity may be of value as future agents for prevention of unwanted uterine contractile activity associated with preterm labor and fetal surgery.

Angiotensin II Receptor Blocker Candesartan Cilexetil, but Not Hydralazine Hydrochloride, Protects Against Mouse Cardiac Enlargement Resulting From Undernutrition In Utero

Wed, 16 Sep 2009 16:55:56 PDT

Epidemiologic studies have shown that malnutrition in utero is a risk factor for cardiovascular disease (CVD) in adulthood. Recently, we reported a mouse animal model of 30% maternal caloric reduction, in which adult offspring (undernourished [UN] offspring) showed a significant increase in cardiac remodeling-associated parameters, such as cardiac enlargement (CE) and coronary perivascular fibrosis (CPVF), as risk factors for CVD. To investigate the possible involvement of the angiotensin system, an angiotensin II receptor antagonist, candesartan cilexetil, or a nonspecific vasodilator, hydralazine hydrochloride, was administrated via a subcutaneously implanted miniosmotic pump to the UN offspring from 9 to 17 weeks after birth. Administration of candesartan cilexetil, but not hydralazine hydrochloride, significantly protected against CE. While administration of not only candesartan cilexetil but also hydralazine hydrochloride prevented an augmentation of CPVF. The angiotensin system seems to make a critical contribution to the developmental origins of cardiac enlargement.

The ''Toll'' of Labor

Young, S. L. — Fri, 21 Aug 2009 10:40:03 PDT

Review: In Vitro Models for the Study of Early Human Embryo-Endometrium Interactions

Teklenburg, G., Macklon, N. S. — Fri, 21 Aug 2009 10:40:03 PDT

The molecular interactions at the embryo-endometrial interface during the period of blastocyst adhesion and subsequent invasion into the endometrial stroma are not fully understood. Current knowledge is primarily based on evidence from implantation studies in the mouse. The degree to which data derived from animal studies mirror human implantation is limited. The ethical and technical challenges studying implantation in the human can partly be overcome by designing in vitro models of embryo-endometrium interactions. In this review, the principal models in current use are described. Basic models using tissue explants and monolayers are distinguished from complex models using multilayer isolated cells, and embryo-endometrium coculture systems used therapeutically. Although there are limitations to current approaches, a number of research questions that could be addressed using these techniques are identified.

Angiotensin-Converting Enzyme and Adducin-1 Polymorphisms in Women With Preeclampsia and Gestational Hypertension

Fri, 21 Aug 2009 10:40:03 PDT

The angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism and the Adducin-1 (ADD1) G460W nonsense single nucleotide polymorphism (SNP) have previously been associated to hypertension, whereas their association with preeclampsia (PE) and gestational hypertension (GH) is still controversial. We genotyped ACE I/D, ADD1 G460W, and ADD1 S586C polymorphisms in 672 unrelated pregnant women: 204 PE (81/204 mild PE), 56 GH, and 412 controls, evaluating both their single and combined effects on these pathologies. The genotype combination of the 3 polymorphisms was not statistically different in cases versus controls, nor were ACE and ADD1 polymorphisms in GH. Nevertheless, the distribution of ACE genotypes was different in PE. This was confirmed in mild PE, whereas no significance was found in severe PE. This could suggest that different factors may lead to mild and severe PE, with ACE polymorphism playing a more important role in the mild form.

Murine Xenograft Model for Human Uterine Fibroids: An In Vivo Imaging Approach

Suo, G., Sadarangani, A., LaMarca, B., Cowan, B., Wang, J. Y. J. — Fri, 21 Aug 2009 10:40:03 PDT

Uterine fibroids are the most prevalent benign tumors in women of reproductive age. The current knowledge on the fibroid disease mechanism has derived from studies of the Eker rat model where a unique germ line defect in the tuberous sclerosis 2 (Tsc2) tumor suppressor gene leads to the development of leiomyosarcoma, leiomyoma, and renal cancer. To study fibroids of human origin, we sought to establish fibroid xenografts in immune-compromised mice. We determined that lentiviral-mediated transduction of a green fluorescence protein (GFP)-luciferase (LUC) fusion gene and bioluminescence-based whole animal imaging allowed for the monitoring of transplanted fibroid cells in mice. We used this in vivo imaging approach to test a series of transplantation protocols and found that only freshly dissociated fibroid cells, but not the fibroid-derived smooth muscle cells grown in ex vivo cultures, can generate stable xenografts in subcutaneous Matrigel implants. Formation of the fibroid-xenografts requires the implantation of 17βestradiol-releasing pellets in the recipient mice. Furthermore, freshly dissociated myometrial cells do not form xenografts under the experimental conditions. The xenograft protocol developed from this study provides an avenue for investigating the pathogenesis and drug responses of human fibroids.

The Role of Toll-Like Receptors (TLR-2 and -4) and Triggering Receptor Expressed on Myeloid Cells 1 (TREM-1) in Human Term and Preterm Labor

Fri, 21 Aug 2009 10:40:03 PDT

Objective. To define the role of Toll-like receptors (TLR-2, TLR-4) and triggering receptor expressed on myeloid cells-1 (TREM-1) in pregnant human myometrium. Study Design. Expression of TLR-2 and -4 mRNA and protein and TREM-1 mRNA was quantified in human myometrial samples. Subsequently, we investigated the effect of medroxyprogesterone acetate (MPA) as a functional inhibitor of the TLR agonist lipopolysaccharide (LPS). Results. Messenger RNA expression of TLR-2 and -4 was significantly higher in myometrium at term compared with preterm (P = .009 and .016, respectively). Toll-like receptor-2 protein expression was significantly higher during labor (P = .002) compared with nonlaboring samples. Triggering receptor expressed on myeloid cells-1 mRNA expression was increased in both myometrium and cervix after labor at term (P < .05). Medroxyprogesterone acetate significantly suppressed LPS-induced production of interleukin 1b (IL-1b), IL-6, and IL-8 in vitro (P < .05). Conclusion. Toll-like receptors 2and 4 and TREM-1 are expressed in human myometrium and may play a role in the mechanism of labor at term, and their functional effects are inhibited by MPA.

Expression of Angiogenesis-Related Genes in the Cellular Component of the Blood of Preeclamptic Women

Fri, 21 Aug 2009 10:40:03 PDT

The purpose of this study is to assess the changes in the expression of angiogenesis-related genes in the cellular component of the blood from preeclamptic patients. Blood samples were obtained from the preeclampsia (PE) and control participants. Cellular RNA was analyzed by reverse transcription polymerase chain reaction (PCR) to identify any angiogenesis-related genes and thereby assess the mRNA expression among women with PE and controls during weeks 35 to 41 of gestation. Significant differences were observed between PE and controls in all of the angiogenesis-related genes examined. In PE, for the increased expression of transforming growth factor-β1 (TGF-β1), endoglin and fms-like tyrosine kinase-1 (Flt-1); and the reduced expression of vascular endothelial growth factor (VEGF), placental growth factor (PlGF). fms-Like tyrosine kinase-1 and endoglin significantly correlated with the systolic pressure, while VEGF, Flt-1, and endoglin all correlated with proteinuria. An altered expression of angiogenesis-related genes was demonstrated in the cellular component of blood from preeclamptic patients. These findings indicate that this approach may offer an alternative way for evaluating the pathogenesis of PE.

Long-Term Hypoxia Increases Endothelial Nitric Oxide Synthase Expression in the Ovine Fetal Adrenal

Fri, 21 Aug 2009 10:40:03 PDT

This study was designed to test the hypothesis that fetal adrenal nitric oxide synthase (NOS) is elevated in response to long-term hypoxia (LTH). Pregnant ewes were maintained at high altitude (3820 m) for approximately the last 100 days of gestation. Between days 138 and 141 of gestation, adrenal glands were collected from LTH fetuses and age-matched normoxic controls. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western analysis were used to quantify NOS expression, and NOS distribution was examined by immunohistochemistry and double-staining immunofluorescence for endothelial NOS (eNOS) and 17-hydroxylase (CYP17). Neuronal NOS (nNOS) was expressed at very low levels and with no differences between groups. Expression of eNOS was significantly greater in the LTH group compared with control. Neuronal NOS was distributed throughout the cortex while the greatest density of eNOS was observed in the zona fasciculata/reticularis area and eNOS co-localized with CYP17. We conclude that LTH enhances eNOS expression in the inner adrenal cortex which may play a role in regulation of cortisol biosynthesis in the LTH fetus.

Effect of Nicotine Exposure During Pregnancy and Lactation on Maternal, Fetal, and Postnatal Rat IGF-II Profile

Gruslin, A., Cesta, C. E., Bell, M., Qing Qiu, , Petre, M. A., Holloway, A. C. — Fri, 21 Aug 2009 10:40:04 PDT

Smoking during pregnancy has been shown to result in an increased risk of low birth weight. However, the mechanisms underlying this association are poorly understood. The insulin-like growth factor (IGF) system plays a critical role in the regulation of feto-placental growth and development, and abnormal processing of proIGF-II may alter its biological function. Our goal was to investigate the effects of exposure to nicotine on maternal, fetal, and neonatal IGF-II processing. Nulliparous female Wistar rats were randomly assigned to receive saline (vehicle) or nicotine bitartrate (1 mg·kg^—1·d^{— 1}). After mating, dams were euthanized at embryonic days 15, 18, and 21, and fetal body weight was recorded. Serum (fetal and maternal) was collected for determination of the IGF-II profile by Western blot analysis. Nicotine exposure prevented the decrease in maternal IGF-II processing seen in controls with advancing gestation. However, there was no influence of nicotine on fetal levels of IGF-II. Postnatally (postnatal day [PND] 21), pups exposed to nicotine in utero had decreased levels of big IGF-II. Our results show, for the first time, that nicotine exposure prevents the decrease of IGF-II processing in the maternal compartment. This may represent a compensatory mechanism allowing the mother to counteract the negative influence of nicotine on fetal growth and development. Our postnatal findings of suppressed IGF-II may help explain some of the long-term health complications seen in individuals exposed to smoking in utero.

Proteomic Analysis of the Luteal Endometrial Secretome

Scotchie, J. G., Fritz, M. A., Mocanu, M., Lessey, B. A., Young, S. L. — Fri, 21 Aug 2009 10:40:04 PDT

Endometrium attains a secretory architecture in preparation for embryo implantation, but the identity of most endometrial secretory products remains unknown. Our objective was to characterize the endometrial secretome and compare protein expression between prereceptive (luteinizing hormone [LH]+4) and receptive (LH+9) phase endometrium. Endometrial lavage was performed in 11 participants and analyzed by difference gel electrophoresis (DIGE). LH+4 and LH+9 specimens were labeled with cyanine fluorescent dyes Cy3 and Cy5 tags, respectively, and combined. Proteins were separated using 2-dimensional gel electrophoresis, isolated, trypsin-digested, and subjected to mass spectrometry. In all, 152 proteins were identified; 82 were differentially expressed. Most proteins with increased expression on LH+9 functioned in host defense, while proteins with decreased expression had many functions. A total of 14 proteins had changes suggesting altered posttranslational modification. This article describes the first application of proteomic analysis to endometrial secretions, allowing identification of novel endometrial proteins as well as those differentially secreted in prereceptive and receptive phases.

Maternal Protein Deprivation: Changes in Systemic Renin-Angiotensin System of the Mouse Fetus

Goyal, R., Galffy, A., Field, S. A., Gheorghe, C. P., Mittal, A., Longo, L. D. — Fri, 21 Aug 2009 10:40:04 PDT

We tested the hypothesis that maternal protein deprivation during gestation results in changes in expression of the systemic renin-angiotensin system in fetal mice. Fetal weight was decreased significantly as a consequence of 50% maternal protein deprivation during second half of gestation. In fetal liver, angiotensinogen protein expression was reduced significantly despite a significant increase in messenger RNA (mRNA). In fetal kidneys, both mRNA and protein levels of renin were increased significantly. In the lungs, we observed a decrease in both angiotensin-converting enzyme I and II mRNA expression, whereas protein expression of both isoforms was increased significantly. The fetal heart showed significant increases in expression of angiotensin II type 1 (AT-1) and type 2 (AT-2) receptors mRNA. Protein expression of AT-1 receptors increased, while that of AT-2 receptors decreased. We conclude that maternal low-protein diet during gestation leads to significant changes in expression of the systemic renin-angiotensin system in fetal mice and may be important in the genesis of hypertension in the adult.

Chymotrypsin-Like Protease (Chymase) Mediates Endothelial Activation by Factors Derived From Preeclamptic Placentas

Yang Gu, , Chang Liu, , Alexander, J. S., Groome, L. J., Yuping Wang, — Fri, 21 Aug 2009 10:40:04 PDT

Endothelial cells (EC) activation is an important inflammatory phenotypic change in the vascular system in women with preeclampsia (PE). In PE, maternal vessel chymotrypsin-like protease (CLP)/chymase expression was increased. Chymase is an inflammatory protease. In this study, we specifically examined whether placental-derived CLP could induce EC activation and whether EC activation is associated with increased cellular protease expression. Human uterine microvascular endothelial cells (UtMVECs) were used. Endothelial activation was determined by endothelial adhesion molecule P-selectin, E-selectin, inter-cellular adhesion molecule (ICAM), and vascular cell adhesion molecule (VCAM) expressions and by extracellular regulated kinase (ERK) activity. Activation of endogenous CLP/chymase associated with ERK phosphorylation was further examined by CLP/chymase short interfering RNA (siRNA). Our results showed that cells treated with PE placental conditioned medium revealed increased P-selectin, E-selectin, and VCAM-1 expressions and increased ERK phosphorylation. Increased endothelial adhesion molecule expression and phosphorylated ERK (pERK) induction could be attenuated or abolished by depletion of CLP in the conditioned medium or by transfecting ECs with CLP/chymase siRNA. These observations suggest that placental-derived CLP/chymase is responsible for inducing endothelial inflammatory phenotypic changes possibly by upregulation of cell adhesion molecule expressions, activation of cellular protease, and induction of ERK phosphorylation. We speculate that activation of endothelial CLP/chymase may directly relate to the increased inflammatory phenotypic changes in the vascular system in women with PE.