Receptor isoforms that mediate estrogen and progestagen action in the female lower urinary tract.

PURPOSE: Bladder symptoms can be ameliorated by sex steroids but to our knowledge the mechanism of action is unknown. Previous studies of steroid receptor expression in the bladder did not indicate receptor subtype expression. We report the distribution of estrogen and progesterone receptor isoforms in the female lower urinary tract. MATERIALS AND METHODS: Prospectively recruited women undergoing routine urogynecological or gynecological surgery provided cold cup biopsy samples from the bladder dome, trigone, and proximal and distal urethra. The samples were immediately frozen or fixed in formalin. After RNA extraction transcripts for estrogen receptor alpha and beta, and progesterone receptor A and B were noted on reverse transcriptase-polymerase chain reaction using isoform specific primers. The precise cellular localization of receptor proteins and their relative levels were assessed by immunochemistry in formalin fixed tissue sections with isoform specific antibodies. RESULTS: Nine premenopausal and 10 postmenopausal women were recruited into the study. Two postmenopausal women on hormone replacement therapy. Estrogen receptor alpha and beta, and progesterone receptor A and B transcripts were detected in whole bladder extracts. Nuclear estrogen receptor alpha immunoreactivity was present in squamous epithelium but absent from transitional epithelium. Estrogen receptor beta immunoreactivity was expressed in squamous and transitional cell epithelium. Nuclear progesterone receptor expression was present in urethral squamous epithelium only. Progesterone receptor expression was greater in premenopausal women and in postmenopausal women on estrogen. CONCLUSIONS: Estrogen receptor alpha and beta genes are transcribed in bladder tissue but only estrogen receptor beta is translated into protein, suggesting that the urothelium responds to endogenous estrogen via estrogen receptor beta. Progesterone receptor expression is confined to urethral squamous epithelium and the major isoform is progesterone receptor A.

The cytoplasmic 60 kDa progesterone receptor isoform predominates in the human amniochorion and placenta at term.

BACKGROUND: The mechanism that initiates human parturition has been proposed to be 'functional progesterone withdrawal' whereby the 116 kDa B-isoform of the progesterone receptor (PR-B) switches in favour of the 94 kDa A-isoform (PR-A) in reproductive tissues. Recently, other PR isoforms, PR-S, PR-C and PR-M generated from the same gene have been identified and partially characterised. METHODS AND RESULTS: Using immunohistochemical, western blotting and RT-PCR techniques, evidence is provided that indicates the major PR isoform present in human term fetal membranes (amnion and chorion) and syncytiotrophoblast of the placenta is neither of the classical nuclear PR-B or PR-A isoforms but is the N-terminally truncated 60 kDa PR-C isoform. Evidence is also provided that this 60 kDa isoform resides in the cytoplasm of the expressing cell types. Data are also presented to show that PR-B, PR-A and PR-S isoforms are essentially absent from the amnion and chorion, whereas PR isoforms A, B, C and S are all present in the decidua, with PR-A being the major isoform. The syncytiotrophoblast of the placenta contains the cytoplasmic 60 kDa isoform, but not isoforms PR-A, PR-B or PR-S. CONCLUSION: The major PR isoform in the amnion, chorion and placenta is a 60 kDa protein that could be PR-C, suggesting that the cytoplasmic isoform has a specific role in extra-embryonic tissues and may be involved in the regulation of human parturition.

Spatio-temporal expression patterns of anandamide-binding receptors in rat implantation sites: evidence for a role of the endocannabinoid system during the period of placental development.

BACKGROUND: Although there is growing evidence that endocannabinoids play a critical role in early pregnancy, there are no studies describing the possible targets for this system after implantation. The endometrial stroma, which undergoes extensive proliferation and differentiation giving rise to the decidua and the trophoblast cells that invade after the initial stages of implantation, are potential targets. Since high anandamide (AEA) levels, the main endocannabinoid, are detrimental to implantation and in order to gain insight into the role of the endocannabinoid system in the development of the fetoplacental unit, the spatio-temporal pattern of expression of the anandamide-binding receptors, CB1, CB2 and the vanilloid receptor (TRPV1), were investigated by quantitative RT-PCR, western blot and immunohistochemistry. METHODS: Rat uterine maternal tissues from different days of pregnancy were used to investigate the expression of CB1, CB2 and vanilloid receptors by quantitative RT-PCR, western blot and immunohistochemistry. RESULTS: The data indicate that all the three receptors were expressed in decidualized cells and placenta. Interestingly, CB1 and CB2 were also expressed in smooth muscle cells of maternal blood vessels and in endovascular trophoblast cells, whereas TRPV1 was mainly expressed in uterine natural killer (uNK) cells and in the longitudinal muscle layer throughout pregnancy. In all tissues, CB2 protein was present at a lower level than CB1. CONCLUSION: These observations support a role for the endocannabinoid system during the period of decidualization and placental development.

Prevention of tamoxifen induced endometrial polyps using a levonorgestrel releasing intrauterine system long-term follow-up of a randomised control trial.

OBJECTIVES: In a RCT, we have previously shown that the levonorgestrel intrauterine system (LNG-IUS, Mirena) produces a decidual response protecting the endometrium at one year follow-up. We here report on the long-term follow-up of this group of women, to test the hypothesis that a LNG-IUS could prevent the pro-proliferative uterine responses of tamoxifen for up to 4.5 years. METHODS: A randomised-controlled trial of postmenopausal women who had taken at least one year of adjuvant tamoxifen therapy. RESULTS: One hundred twenty-two women were recruited. Nine were found to be ineligible after randomisation. The average duration of follow-up was 26.25 months (IQR 14.5-36 months) in the surveillance group and 24.2 months (IQR 13.75-32.5 months) in the LNG-IUS group. Women with LNG-IUS in situ at the time of final assessment had decidualised endometrium, and no polyps. In the surveillance group new polyps arose in 8 cases. There were 3 new polyps in the group initially randomised to LNG-IUS, one in a patient who did not have the device inserted and 2 occurred in patients following the removal of the LNG-IUS. Univariate Cox proportional hazards regression models identified only endometrial thickness at trial entry as a statistically significant variable (HR 1.12, 95% CI 1.02 to 1.22, p=0.01) for the development of polyps. CONCLUSION: This study confirms that LNG-IUS induces benign endometrial changes and prevents endometrial polyps but only during its use in women taking tamoxifen. Endometrial thickness is a risk factor for the development of polyps.

Expression of the endocannabinoid system in human first trimester placenta and its role in trophoblast proliferation.

The endocannabinoid, anandamide, which binds to two major receptor proteins, the cannabinoid receptors (CBs) 1 and 2 (CB1 and CB2), has been shown to play a role in first trimester miscarriage possibly through impairment of the developing trophoblast. Although the precise molecular mechanisms underlying this are unknown, plasma anandamide levels are known to be regulated by the progesterone-induced enzyme, fatty acid amide hydrolase (FAAH). Here, we tested the hypothesis that temporal-spatial expression of FAAH, CB1, and CB2 is regulated during early pregnancy and that anandamide detrimentally alters trophoblast proliferation. Transcripts for CB1, CB2, and FAAH were demonstrated in first trimester trophoblast extracts with only the CB1 transcript being significantly regulated. The significant 4.7-fold increase in expression at wk 10 gestation was reduced to 8.9% of the peak value by wk 12. Transcripts for CB2 showed a similar pattern of expression but were not significantly induced. By contrast, FAAH transcript levels appeared to increase toward the end of the first trimester, but again did not reach significance. These observations were supported by immunohistochemical studies that demonstrated a similar pattern of expression at the protein level, with cellular localization for all three proteins concentrated within the syncytiotrophoblast layer. Anandamide also prevented BeWo trophoblast cell proliferation in a dose-dependent manner, with a 50-60% significant inhibition of cell proliferation with concentrations in excess of 3 mum. This effect was mediated through CB2. Together, these data provide insights into how elevated plasma anandamide levels increase the risk of first trimester miscarriage.

Differential regulation of insulin-like growth factor binding protein-1 and -2 by insulin in the baboon (Papio anubis) endometrium.

BACKGROUND: The purpose of this study was to examine the effect of insulin on expression and synthesis of IGFBP-1 and IGFBP-2 in the baboon endometrium in vitro. METHODS: Baboon endometrial explants collected from cycling, ovariectomized, steroid-treated, simulated-pregnant and pregnant animals were cultured for 48 h in the presence or absence of insulin, with or without estradiol, progesterone and hCG. RESULTS: Insulin clearly inhibited IGFBP-1 production and mRNA expression in a time- and dose-dependent manner, whereas IGFBP-2 synthesis was not significantly affected. The inhibitory effects of insulin on IGFBP-1 were more evident in explants of non-pregnant tissue or tissue away from the implantation site. In the absence of insulin, synthesis of IGFBP-1 was induced in explants with low levels of de novo synthesis whereas IGFBP-2 synthesis was inhibited. This effect was potentiated by steroids and hCG in the explant cultures. CONCLUSION: Insulin differentially regulates endometrial IGFBP-1 and IGFBP-2 secretion in the baboon.

The progesterone receptor in human term amniochorion and placenta is isoform C.

The mechanism that initiates human parturition has been proposed to be functional progesterone withdrawal whereby the 116-kDa B isoform of the progesterone receptor (PR-B) switches in favor of the 94-kDa A isoform (PR-A) in reproductive tissues. Recently other PR isoforms, PR-S, PR-C, and PR-M generated from the same gene have been identified and partially characterized. Using immunohistochemical, Western blotting, and RT-PCR techniques, evidence is provided that the major PR isoform present in human term fetal membranes (amnion and chorion) and syncytiotrophoblast of the placenta is neither of the classical nuclear PR-B or PR-A isoforms but is the N terminally truncated 60-kDa PR-C isoform. Evidence is also provided that the PR-C isoform resides in the cytoplasm of the expressing cell types. Data are also presented to show that PR-B, PR-A, and PR-S isoforms are essentially absent from the amnion and chorion, whereas PR isoforms A, B, C, and S are all present in the decidua, with PR-A being the major isoform. The syncytiotrophoblast of the placenta contains the cytoplasmic PR-C isoform but not PR-A, PR-B, or PR-S. The major PR isoform in the amnion, chorion, and placenta is PR-C, suggesting that the cytoplasmic PR-C isoform has a specific role in extraembryonic tissues and may be involved in the regulation of human parturition.

Changes in tenascin-C isoform expression in invasive and preinvasive breast disease.

Tenascin-C (TN) is an extracellular matrix protein that is expressed at low levels in normal adult tissue but is highly expressed around many tumors including breast carcinoma. TN exists as multiple isoforms generated through alternative splicing, and these isoforms have different effects on cell growth and migration. This study has analyzed in detail the pattern of TN isoform expression in benign, preinvasive, and invasive breast lesions using reverse transcription-PCR and Southern blotting. Significant differences in the profile of TN isoforms were identified. Although all tissues expressed the fully truncated TN, expression of two additional isoforms, one containing exon 16 (TN16) and one containing both exons 14 and 16 (TN14/16), were significantly associated with the invasive phenotype (P < 0.001). A subset of ductal carcinoma in situ (DCIS) cases were also found to express these isoforms, which may be indicative of a high risk of invasion in these lesions. Expression of these isoforms correlated with the presence of TN protein in the stroma in place of or in addition to basement membrane TN. Immunohistochemistry and in situ hybridization confirmed the production of exon 14-containing higher molecular weight isoforms by stromal fibroblasts in malignant tissue and both periductal fibroblasts and residual myoepithelial cells in DCIS. Although no evidence of tumor cell synthesis of TN was detected in the tissues, two highly invasive breast cancer cell lines (MDA-MB 231 and MDA-MB 468) were found to produce TN in contrast with tumor cells with a lower invasive capacity (MCF-7 and T47D). These results demonstrate for the first time that specific TN isoforms are expressed in invasive breast carcinomas and that these isoforms are identified in a subset of DCIS and suggest that detection of TN16 and/or TN14/16 may be used as a predictor for invasion. Functional studies are now essential to establish the effect of these isoforms on tumor behavior and evaluate whether they will provide appropriate targets for therapeutic intervention.

3-dimensional colour power angiography for staging human placental development.

Maldevelopment of placental villous trees and their blood vessels results in impaired fetal growth, which can greatly compromise fetal, neonatal, childhood, and adulthood health. There are no means of directly assessing such maldevelopment. We have applied a new technique of imaging (colour power angiography [CPA]) with 3-dimensional reconstruction to assess directly villous development in human pregnancy in vivo in 20 uncomplicated pregnancies from 13 to 38 weeks' gestation. The chronology of villous trees was much the same in 3-dimensional CPA, scanning electron micrography, and classical histology from controls matched by age. This approach provides a unique opportunity to examine normal placental development directly, and should provide the bases for real-time investigation of placental pathology and a robust method for diagnosis and surveillance during pregnancy.

Urine protein estimation in hypertensive pregnancy: which thresholds and laboratory assay best predict clinical outcome?

OBJECTIVE: To determine what threshold for proteinuria could best predict clinical outcome and whether this threshold could be applied universally to any biochemical assay. DESIGN: A prospective observational study of hypertensive pregnancies referred for further assessment after in a UK University hospital (n=197). Twenty-four hour urine protein was measured by two different assays [benzethonium chloride assay (BCA) and Bradford assay]. The differences between the two assays were calculated from Receiver Operating Characteristic (ROC) curves. Commonly used thresholds for defining preeclampsia (0.3 and 0.5 g/24 hours) were explored for both assays for the prediction of adverse clinical outcomes (severe hypertension, Birthweight<10th percentile, preterm delivery, and a composite biochemical/haematological derangement). RESULTS: The two assays are not equivalent. The prevalence of>300 mg/24 hour proteinuria and, hence, the prevalence of preeclampsia differed between the two assays. ROC curve analysis demonstrates that the two assays are similar in terms of overall performance as predictive tests. However the threshold of 300 mg/24 hours performs poorly as a predictor of clinical risk. Likelihood ratios (LR) for the BCA at the 300 mg/L threshold for each clinical outcome do not achieve statistical significance. At the 500 mg/L threshold, the LR+for the BCA assay does achieve statistical significance for severe hypertension (LR+:1.51 95% CI 0.99-2.28) and for birthweight<10th percentile (LR+:1.72 95% CI 1.11-2.66). For the Bradford assay at the 300 mg/24 hour threshold, the LR+does achieve statistical significance for birthweight<10th percentile (LR+:1.71 95% CI 1.41-4.31). However, at the 500 mg/24 hour threshold, the LR+is significant for severe hypertension (LR+:2.15 95% CI 1.07-4.34), birthweight<10th percentile (LR+:2.79 95% CI 1.4-5.54) and biochemical disease (LR+:2.47 95% CI 1.22-5.01). CONCLUSIONS: This study suggests that thresholds for proteinuria need to be higher (possibly>or=0.5 g/24 hours) and there is the need for a "gold standard" proteinuria assay against which all other measures of quantification can be assessed.

Plasma levels of the endocannabinoid anandamide in women--a potential role in pregnancy maintenance and labor?

Although exposure to exocannabinoids (e.g. marijuana) is associated with adverse pregnancy outcome, little is known about the biochemistry, physiology, and consequences of endocannabinoids in human pregnancy. In these studies, we measured the levels of the endocannabinoid anandamide (N-arachidonoylethanolamine, AEA) by HPLC-mass spectrometry in 77 pregnant and 25 nonpregnant women. The mean +/- sem plasma AEA levels in the first, second, and third trimesters were 0.89 +/- 0.14, 0.44 +/- 0.12, and 0.42 +/- 0.11 nm, respectively. The levels in the first trimester were significantly higher than those in either the second or third trimester. During labor, AEA levels were 3.7 times nonlaboring term levels (2.5 +/- 0.22 vs. 0.68 +/- 0.09 nm, P < 0.0001). During the menstrual cycle, levels in the follicular phase were significantly higher than those in the luteal phase (1.68 +/- 0.16 vs. 0.87 +/- 0.09 nm, P < 0.005). Postmenopausal and luteal-phase levels were similar to those in the first trimester. These findings suggest that successful pregnancy implantation and progression requires low levels of AEA. At term, AEA levels dramatically increase during labor and are affected by the duration of labor, suggesting a role for AEA in normal labor.

Immunohistological localization of insulin-like growth factor binding protein-1 in primary implantation sites and trauma-induced deciduomal tissues of the rhesus monkey.

We have earlier demonstrated that there is a close similarity in the temporo-spatial pattern in the onset of oedema, epithelial-plaque transformation, stromal decidualization and influx of granulated lymphocytes in artificially trauma-induced deciduomal endometrium with such events in maternal endometrium at the primary implantation site during early stages of pregnancy in the rhesus monkey. In the present study, we have immunohistochemically examined whether the pattern of insulin-like growth factor-binding protein 1 (IGFBP-1) protein expression in conceptus tissue and maternal endometrium during lacunae and villous placenta stages of gestation in the rhesus monkey is developmental stage-specific and whether a discernible difference exists in the temporo-spatial characteristics of IGFBP-1 protein expression between conceptus associated implantation-decidualization and trauma induced deciduoma in the rhesus monkey. Trophoblast cells failed to exhibit IGFBP-1 immunopositive staining at any stage of implantation-placentation studied. Epithelial cells in plaque acini, endothelial cells, and vascular smooth muscle also did not show any immunopositive staining for IGFBP-1 in samples of primary implantation sites and trauma-induced deciduoma. Maternal endometrial epithelial and stromal-decidual cells however exhibited a temporal and spatial pattern of IGFBP-1 expression in cell-type specific manner and clear distinctions were observed between conception and deciduoma samples. Our results suggest that IGFBP-1 expression is highly tissue and development-specific and that conceptus-derived signals are necessary to initiate the glandular expression of IGFBP-1 during the early stage of gestation.

Endogenous cannabinoids: metabolism and their role in reproduction.

Over the past two decades a number of endogenous compounds that act as ligands for the cannabinoid receptors has been discovered. In analogy with the "endorphins" these compounds have been called "endocannabinoids". Endocannabinoids have been demonstrated in many mammalian tissues including humans and are widely distributed in the CNS, peripheral nerves, uterus, leukocytes, spleen and testicles. The uterus contains the highest levels of anandamide, the first discovered endocannabinoid, suggesting an important role for this substance in reproduction. Several studies have shown anandamide to be involved in the regulation of implantation and reduced activity of the enzyme that degrades anandamide has been associated with early pregnancy loss in humans. The bulk of the literature concerning endocannabinoids is based upon anandamide related studies; therefore, in this review we focus on the metabolism of anandamide and its role in reproduction.

Inhibition of endometrial carcinoma cell growth using antisense estrogen receptor oligodeoxyribonucleotides.

BACKGROUND: The sex hormone 17 beta-estradiol acts as a mitogen in a number of tissues, including the endometrium, through direct interaction with the estrogen receptor (ER alpha). In the protection of the female breast and endometrium from cancer progression it would be advantageous to inhibit estrogenic action, therefore many estrogen receptor antagonists have been made. However, the most clinically relevant anti-estrogens for breast cancer have a detrimental effect on the endometrium and induce or exacerbate existing endometrial oncogenesis. Specific anti-estrogenic potential that is directed against the endometrial ER alpha theoretically could be achieved using antisense oligodeoxyribonucleotide transfection techniques. MATERIALS AND METHODS: To discover the most effective antisense oligodeoxyribonucleotides against the human ER alpha, a series of oligodeoxyribonucleotides were synthesised and tested in a human endometrial cancer cell line that expresses a functional ER alpha. RESULTS: Transfection with antisense oligodeoxyribonucleotides, directed against different regions of the human ER alpha, significantly inhibited ER alpha protein production without affecting ER beta protein levels. Antisense oligodeoxyribonucleotides directed against the translational start site demonstrated reduced binding of radiolabeled 17 beta-estradiol and a complete inhibition of estrogen-dependent endometrial cancer cell proliferation. The inhibitory effect of the antisense oligodeoxyribonucleotides on ER alpha production and ligand binding was enhanced in the presence of exogenous 17 beta-estradiol. CONCLUSION: The data suggest that antisense oligodeoxyribonucleotides against the human ER alpha have the potential of acting as anti-proliferative agents for estrogen-dependent endometrial cancers and may help in elucidating the relative roles of the two estrogen receptors, ER alpha and ER beta, in cell processes other than proliferation.

Validation of the DCA 2000 microalbumin:creatinine ratio urinanalyzer for its use in pregnancy and preeclampsia.

OBJECTIVE: To determine the accuracy of the DCA 2000 albumin/creatinine ratio urinanalyzer (Bayer Corp., Elkhart, IN) in uncomplicated pregnancy and preeclampsia. METHODS: This was a prospective observational study in a large teaching maternity hospital. Ninety one uncomplicated pregnant women and 100 women referred for assessment of de novo hypertension in pregnancy had albumin concentrations, creatinine concentrations, and albumin/creatinine ratios (ACR) compared between the DCA 2000 and the laboratory gold standard assays (Dade Dimension clinical chemistry autoanalyzer), for both early morning urines (EMU) and 24-hr urine collections. RESULTS: The interassay and intra-assay variability for the DCA 2000 were less than 5.1%. In uncomplicated pregnancy the mean difference in ACR between the DCA 2000 and the laboratory assay was 0.08 mg/mmol (SD 0.28; 95% limits of agreement, -0.47, 0.63) for EMU and 0.06 (SD 0.23; 95% limits of agreement, -0.39, 0.51) for 24-hr samples. In the hypertensive cohort the ACR mean differences were -0.82 (SD 7.13; 95% limits of agreement, -14.79, 13.15) for EMU and -0.76 (SD 4.14; 95% limits of agreement, -8.87, 7.35) for 24-hr samples. The mean differences between assays in the hypertensive group had broader 95% limits of agreement due to greater variability in the samples with high albumin concentrations (>40 mg/L). CONCLUSIONS: The DCA 2000 is accurate for the measurement of albumin creatinine ratios in the uncomplicated pregnant population. In the hypertensive population the DCA 2000 remains accurate though when the albumin concentration is greater than 40 mg/L the 95% limits of agreement are broader. We would recommend that all other automated urinalysis devices be validated by similar protocols to allow meaningful comparisons of accuracy.

Delta9-tetrahydrocannabinol inhibits cytotrophoblast cell proliferation and modulates gene transcription.

Cannabis use in pregnancy is associated with a range of obstetrical conditions. The molecular mechanisms underlying these effects have not been elucidated but are attributed to the actions of delta-9-tetrahydrocannabinol (Delta9-THC). In this study, concentrations of Delta9-THC equivalent to those found in the serum of cannabis users, i.e. approximately 20 microM, inhibited proliferation and activated a restricted tight transcriptional programme in the BeWo trophoblast cell line. Employing genome-wide expression profiling methods, we found that the pattern of gene expression differs from that described in the placenta of patients with fetal growth restriction (FGR), associated with either hypoxia or discordant dichorionic twins, or of patients with pre-eclampsia. It was also dissimilar to the patterns obtained from the transcriptome of other tissues, such as the mouse brain, treated with Delta9-THC. The expression of transcription factors, such as thyroid hormone receptor-beta1 (TRbeta1), and transcriptional co-repressors, such as histone deactylase 3 (HDAC3), was affected by Delta9-THC in a dose-dependent manner, whereby 15 microM Delta9-THC caused a 2.8-fold inhibition of TRbeta1 expression, but a 3.5-fold increase in HDAC3 expression. These data were confirmed by end-point RT-PCR analyses and underpin the observed Delta9-THC-induced inhibition of BeWo cell proliferation. Genes encoding for growth, apoptosis, cell morphology and ion exchange pathways were modulated by 15 microM Delta9-THC. This study may provide insight into the mechanisms underlying the effects of Delta9-THC and cannabis use upon placental development during pregnancy.

Relationship between antenatal inflammation and antenatal infection identified by detection of microbial genes by polymerase chain reaction.

Although antenatal infection is thought to play an important role in the pathogenesis of preterm labor and neonatal diseases, the exact mechanisms are largely unknown. We sought to clarify the relationship between antenatal infection and intrauterine and neonatal inflammation. Samples were obtained from 41 preterm infants of <33 wk gestation delivered to 36 mothers and analyzed for the presence of 16s ribosomal RNA (16s rRNA) genes using PCR and for the proinflammatory cytokines IL-6 and IL-8. In 16 (44%) mother-baby pairings, at least one sample was found to be positive for the presence of 16s rRNA genes. All but one of the positive samples were from mothers presenting with preterm prelabor rupture of membranes (pPROM) or in spontaneous idiopathic preterm labor. A strong association was found between the presence of 16s rRNA genes and chorioamnionitis and with funisitis. A marked increase in IL-6 and IL-8 was noted in all tissues positive for 16s rRNA genes, including placenta, fetal membranes, cord blood serum, and, where samples were available, in bronchoalveolar lavage fluid (BAL) and in amniotic fluid. Interestingly, gastric fluid was always positive for 16s rRNA genes if any other intrauterine or BAL sample was positive, suggesting that this sample may provide an alternative to amniotic fluid to identify antenatal infection. In conclusion, we have found that microbial genes are particularly prevalent in pPROM and spontaneous preterm labor groups and that their presence is strongly associated with a marked intrauterine inflammatory response.

Optimal bedside urinalysis for the detection of proteinuria in hypertensive pregnancy: a study of diagnostic accuracy.

OBJECTIVE: To compare semi-quantitative visual and automated methods of urine testing with fully quantitative point of care urinalysis for the detection of significant proteinuria (0.3 g/24 hours) in pregnancy complicated by hypertension. DESIGN: A prospective comparative study. SETTING: A large teaching maternity hospital. SAMPLE: One hundred and seventy-one pregnant women referred to the obstetric day-care unit for assessment of newly arisen hypertension. METHODS: Early morning urine specimens were tested with four dipstick techniques (Multistix 8SG visual and automated and microalbumin/creatinine ratio visual and automated; Bayer, Elkhart, USA) as well as a fully quantitative measure of the microalbumin creatinine ratio with the DCA 2000 (a point of care assay for albumin; Bayer). These results were compared to a 24-hour urine protein measurement and measures of diagnostic accuracy/prediction are reported. MAIN OUTCOME MEASURES: Significant proteinuria (> or =0.3 g/24 hours) measured by laboratory assay. RESULTS: Automated dipstick urinalysis using the Clinitek 50 has significantly better predictive values for significant proteinuria (LR(+) 4.27, 95% CI 2.78 to 6.56; LR(-) 0.225, 95% CI 0.14 to 0.37) than conventional visual dipstick urinalysis (LR(+) 2.27, 95% CI 1.47 to 3.51; LR(-) 0.635, 95% CI 0.49 to 0.82). Dipstick microalbumin/creatinine ratio testing did not improve overall detection rates with automated or visual testing. Fully quantitative point of care measurement of albumin/creatinine ratio (ACR) was significantly better than any dipstick technique (LR(+) 14.6, 95% CI 6.74 to 31.8; LR(-) 0.069, 95% CI 0.030 to 0.16). CONCLUSIONS: This study confirms that in pregnancy automated dipstick urinalysis is a more accurate screening test for the detection of proteinuria than visual testing. ACR testing can offer a significant improvement over conventional urinalysis if a fully quantitative method of detection is employed that uses pregnancy-specific thresholds. Dipstick assessment of ACR does not improve the detection rate of significant proteinuria.

Mapping of zones of altered morphology and chorionic connective tissue cellular phenotype in human fetal membranes (amniochorion and decidua) overlying the lower uterine pole and cervix before labor at term.

OBJECTIVE: The purpose of this study was to determine the location, frequency, and extent of altered fetal membrane morphology before term labor and its relation to myofibroblast activation in their connective tissue layers. STUDY DESIGN: Fetal membranes that were obtained from 10 women who underwent prelabor cesarean delivery at 38 to 39 weeks of gestation underwent biopsy examination with respect to the internal os of the cervix. The thickness of their constituent layers was measured, and the numbers of alpha-smooth muscle actin immunoreactive cells (ie, marker of myofibroblast activation) within the reticular layer were counted. RESULTS: A region that measured 119+/-21cm(2), that exhibited altered morphology of the fetal membranes from the lower uterine pole, and that was characterized by increased connective tissue thickness and decreased thickness of the cellular layers was demonstrated in all patients. In 8 of 10 patients, this region was centered on the location of the Babcock tissue forceps. Within this region was an area of fetal membranes that exhibited alpha-smooth muscle actin immunoreactivity in the cells of the reticular layer and whose number correlated with parameters of altered morphology. CONCLUSION: All patients before labor at term possess an area of fetal membranes that are located in the lower uterine pole that exhibit altered morphology that is associated with myofibroblastic activation in the chorionic connective tissue.

Urinary microalbumin/creatinine ratios: reference range in uncomplicated pregnancy.

During uncomplicated pregnancy, the development of proteinuria is accepted as a poor prognostic sign and is associated with increasing maternal and perinatal mortality and morbidity. Physiological proteinuria increases with increasing gestation and one of its largest constituents is albumin. The reference range for the (micro)albumin/creatinine ratio (ACR) has not been described for normal pregnancy. This prospective cross-sectional study describes the gestation-specific 95% reference ranges for urinary microalbumin concentration, creatinine concentration and ACR in uncomplicated pregnancy. There is a significant increase ( P =0.016) in the ACR in the third trimester. The mean difference is 0.091 mg of albumin/mmol of creatinine (95% confidence interval, 0.014-0.168). Our results describe the first well-defined gestation-specific 95% reference range for a point-of-care measurement of the ACR. These data are essential if such testing is to be employed in antenatal care.