William Murk

Endometriosis and Infertility : Epidemiology and Evidence-based Treatments

Endometriosis is an estrogen‐dependent disorder defined as the presence of endometrial tissue outside of the uterine cavity. A leading cause of infertility, endometriosis has a prevalence of 0.5–5% in fertile and 25–40% in infertile women. The optimal choice of management for endometriosis‐associated infertility remains obscure. Removal or suppression of endometrial deposits by medical or surgical means constitutes the basis of endometriosis management. Current evidence indicates that suppressive medical treatment of endometriosis does not benefit fertility and should not be used for this indication alone. Surgery is probably efficacious for all stages of the disease. Controlled ovarian hyperstimulation with intrauterine insemination is recommended in early‐stage and surgically corrected endometriosis when pelvic anatomy is normal. In advanced cases, in vitro fertilization is a treatment of choice, and its success may be augmented with prolonged gonadotropin‐releasing hormone analog treatment. Further randomized clinical trials focusing on diverse etiopathogenic mechanisms and therapeutic innovation are necessary to find more conclusive, evidence‐based answers regarding this enigmatic disease.

Prenatal or Early-Life Exposure to Antibiotics and Risk of Childhood Asthma: A Systematic Review

CONTEXT: The increasing prevalence of childhood asthma has been associated with low microbial exposure as described by the hygiene hypothesis. OBJECTIVE: We sought to evaluate the evidence of association between antibiotic exposure during pregnancy or in the first year of life and risk of childhood asthma. METHODS: PubMed was systematically searched for studies published between 1950 and July 1, 2010. Those that assessed associations between antibiotic exposure during pregnancy or in the first year of life and asthma at ages 0 to 18 years (for pregnancy exposures) or ages 3 to 18 years (for first-year-of-life exposures) were included. Validity was assessed according to study design, age at asthma diagnosis, adjustment for respiratory infections, and consultation rates. RESULTS: For exposure in the first year of life, the pooled odds ratio (OR) for all studies (N = 20) was 1.52 (95% confidence interval [CI]: 1.30–1.77). Retrospective studies had the highest pooled risk estimate for asthma (OR: 2.04 [95% CI: 1.83–2.27]; n = 8) compared with database and prospective studies (OR: 1.25 [95% CI: 1.08–1.45]; n = 12). Risk estimates for studies that adjusted for respiratory infections (pooled OR: 1.16 [95% CI: 1.08–1.25]; n = 5) or later asthma onset (pooled OR for asthma at or after 2 years: OR: 1.16 [95% CI: 1.06–1.25]; n = 3) were weaker but remained significant. For exposure during pregnancy (n = 3 studies), the pooled OR was 1.24 (95% CI: 1.02–1.50). CONCLUSIONS: Antibiotics seem to slightly increase the risk of childhood asthma. Reverse causality and protopathic bias seem to be possible confounders for this relationship.

Matrix Metalloproteinase 9 (MMP9) Expression in Preeclamptic Decidua and MMP9 Induction by Tumor Necrosis Factor Alpha and Interleukin 1 Beta in Human First Trimester Decidual Cells

Extravillous trophoblasts (EVTs) invade human decidua via sequential integrin-mediated binding and proteolysis of basement membrane proteins in the extracellular matrix (ECM). In preeclampsia, shallow EVT invasion impairs spiral artery and arteriole remodeling to reduce uteroplacental blood flow. Excess decidual cell-expressed matrix metalloproteinases (MMPs) 2 and 9, in response to preeclampsia-related interleukin 1 beta (IL1B) and tumor necrosis factor alpha (TNF), may inappropriately degrade these basement membrane proteins and impede EVT invasion. This study found significantly higher immunohistochemical MMP9 levels in decidual cells and adjacent interstitial trophoblasts in placental sections of preeclamptic versus gestational age-matched control women. In contrast, immunostaining for MMP2 and tissue inhibitor of matrix metalloproteinases 1 and 2 (TIMP1 and TIMP2) were similar in preeclamptic and control groups. First-trimester decidual cells were incubated with estradiol (E(2)) or E(2) + medroxyprogesterone acetate (MPA), with or without TNF or IL1B. As measured by ELISA, both cytokines elicited concentration-dependent increases in secreted MMP9 levels that were unaffected by MPA. In contrast, secreted levels of MMP2, TIMP1, and TIMP2 were unchanged in all treatment groups. Substrate gel zymography and Western blotting confirmed that each cytokine increased secreted levels of MMP9 but not MMP2. Similarly, quantitative RT-PCR found that TNF and IL1B enhanced MMP9, but not MMP2, mRNA levels. At the implantation site, inflammatory cytokine-enhanced MMP9 may promote preeclampsia by disrupting the decidual ECM to interfere with normal stepwise EVT invasion.

Spermatozoa Bound to Solid State Hyaluronic Acid Show Chromatin Structure With High DNA Chain Integrity: An Acridine Orange Fluorescence Study

During human spermiogenesis, the elongated spermatids undergo a plasma membrane remodeling step that facilitates formation of the zona pellucida and hyaluronic acid (HA) binding sites. Various biochemical sperm markers indicated that human sperm bound to HA exhibit attributes similar to that of zona pellucida-bound sperm, including minimal DNA fragmentation, normal shape, and low frequency of chromosomal aneuploidies. In this work, we tested the hypothesis that HA-bound sperm would be enhanced in sperm of high DNA chain integrity and green acridine orange fluorescence (AOF) compared with the original sperm in semen. Sperm DNA integrity in semen and in their respective HA-bound sperm fractions was studied in 50 men tested for fertility. In the semen samples, the proportions of sperm with green AOF (high DNA integrity) and red AOF (DNA breaks) were 54.9% ± 2.0% and 45.0% ± 1.9%, whereas in the HA-bound sperm fraction, the respective proportions were 99% and 1.0%, respectively. The data indeed demonstrated that HA shows a high degree of selectivity for sperm with high DNA integrity. These findings are important from the points of view of human sperm DNA integrity, sperm function, and the potential efficacy of HA-mediated sperm selection for intracytoplasmic sperm injection.

Risk analysis of torsion and malignancy for adnexal masses during pregnancy

OBJECTIVE To find the risk factors of torsion and malignancy for adnexal tumors during pregnancy. DESIGN Retrospective, historical cohort study. SETTING University hospital. PATIENT(S) Patients from 1990 to 2004 with adnexal tumors >or=4 cm during pregnancy. INTERVENTION(S) Surgery undertaken antepartum, concurrently with cesarean delivery, or postpartum. MAIN OUTCOME MEASURE(S) Tumor size, progression, pathology, incidence of malignancy, and torsion. RESULT(S) Almost all 213 managements analyzed had good surgical and obstetric outcomes. In 174 patients who were followed through pregnancy with known tumor existence, 14.84% +/- 3.05% encountered tumor torsion. Adnexal masses with sizes between 6 and 8 cm had a significantly higher risk of torsion compared with other sizes (22.41% vs. 9.48%; odds ratio 2.8, 95% confidence interval [CI], [1.1, 6.6]). Sixty percent of the torsion happened between the 10th and 17th weeks of gestation, and only 5.9% happened after 20 weeks. The incidence of malignancy was 3.4%, while that of ovarian cancer was 2.3%. Tumor diameters >or=10 cm at initial diagnosis had a higher risk of malignancy versus smaller sizes (8.77% vs. 0.85%; odds ratio 11.2, 95% CI, [1.3, 97.9]), and tumor growth rates >or=3.5 cm/week also had a significantly higher risk of malignancy versus lower rates (8.33% vs. 0.88%; odds ratio 10.2, 95% CI, [1.0, 101.2]). CONCLUSION(S) Adnexal tumors bearing higher risks for torsion and malignancy should be strongly considered for an aggressive strategy of management during pregnancy.

Involvement of human decidual cell-expressed tissue factor in uterine hemostasis and abruption.

Vascular injury increases access and binding of plasma-derived factor VII to perivascular cell membrane-bound tissue factor (TF). The resulting TF/VIIa complex promotes hemostasis by cleaving pro-thrombin to thrombin leading to the fibrin clot. In human pregnancy, decidual cell-expressed TF prevents decidual hemorrhage (abruption). During placentation, trophoblasts remodel decidual spiral arteries into high conductance vessels. Shallow trophoblast invasion impedes decidual vascular conversion, producing an inadequate uteroplacental blood flow that elicits abruption-related placental ischemia. Thrombin induces several biological effects via cell surface protease activated receptors. In first trimester human DCs thrombin increases synthesis of sFlt-1, which elicits placental ischemia by impeding angiogenesis-related decidual vascular remodeling. During pregnacy, the fibrillar collagen-rich amnion and choriodecidua extracellular matrix (ECM) provides greater than additive tensile strength and structural integrity. Thrombin acts as an autocrine/paracrine mediator that degrades these ECMs by augmenting decidual cell expression of: 1) matrix metalloproteinases and 2) interleukin-8, a key mediator of abruption-associated decidual infiltration of neutrophils, which express several ECM degrading proteases. Among the cell types at the maternal fetal interface at term, TF expression is highest in decidual cells indicating that this TF meets the hemostatic demands of labor and delivery. TF expression in cultured term decidual cells is enhanced by progestin and thrombin suggesting that the maintenance of elevated circulating progesterone provides hemostatic protection and that abruption-generated thrombin acts in an autocrine/paracrine fashion on decidual cells to promote hemostasis via enhanced TF expression.

Extracellularly Signal-Regulated Kinase Activity in the Human Endometrium: Possible Roles in the Pathogenesis of Endometriosis

CONTEXT Endometriosis is an estrogen-dependent disease characterized by the presence of endometrial tissue outside of the uterine cavity, causing pelvic pain and infertility in 10% of reproductive-aged women. It is unclear why ectopic endometrium remains viable in only a subset of women. ERK1/2 plays key intracellular roles in activating cellular survival and differentiation processes. OBJECTIVE We sought to determine ERK1/2 activity in patients with endometriosis and its possible roles in regulating endometrial cell survival. DESIGN ERK1/2 phosphorylation and expression throughout the menstrual cycle were evaluated in vivo in normal and endometriotic human endometrium, and in vitro techniques assessed the steroidal regulation of ERK1/2 and its effect on endometrial cell survival. RESULTS Total ERK1/2 remained constant in normal and endometriotic endometrium throughout the menstrual cycle. Phospho-ERK1/2 was high in the late proliferative and secretory phases in normal endometrium (P < 0.05). In endometriotic glandular cells, there was no cyclical variation in phospho-ERK1/2. In endometriotic stromal cells, there was also a reduction in phospho-ERK1/2 variation, with higher levels in the early-mid secretory phase (P < 0.05). In cultured endometrial stromal cells (ESCs), estrogen plus progesterone increased ERK1/2 phosphorylation within 15 min (P < 0.05). Although estrogen alone did not induce ERK1/2 phosphorylation in normal ESCs, there was a significant response to estrogen in ESCs isolated from eutopic endometriotic endometrium (P < 0.05). ERK1/2 inhibition in ESCs reduced proliferation and increased apoptosis (P < 0.05). CONCLUSION Abnormally high levels of ERK1/2 activity may be involved in endometriosis, possibly by stimulating endometrial cell survival.

Regulation of Interleukin-6 Expression in Human Decidual Cells and Its Potential Role in Chorioamnionitis

Chorioamnionitis frequently precedes both genital tract and placental inflammation and is both a primary cause of maternal morbidity and a major antecedent of preterm premature rupture of the membranes (PPROM) as well as preterm delivery (PTD). In most cases of chorioamnionitis, neutrophils dominate the decidua. In a subset of these cases, a predominance of monocytes is uniquely associated with both neonatal intraventricular hemorrhage and death. The multifunctional cytokine, interleukin-6, promotes local monocyte dominance via several mechanisms. In this study, immunostaining of placental sections revealed significantly higher interleukin-6 HSCOREs in decidual cells (DCs) but not in interstitial trophoblasts, in chorioamnionitis versus gestational age-matched control placentas (P < 0.05). In confluent leukocyte-free term DCs, secreted interleukin-6 levels in incubations with estradiol-17β were increased 2500-fold by IL-1β (P < 0.05). This up-regulation was inhibited by more than 50% in parallel incubations that included medroxyprogesterone acetate (n = 12, P < 0.05). Western blotting data confirmed these enzyme-linked immunosorbent assay results; quantitative RT-PCR findings demonstrated corresponding changes in interleukin-6 mRNA levels. Specific inhibitors of signaling for both nuclear factor-κB activation and p38-mitogen-activated protein kinase, but not for protein kinase C, significantly decreased IL-1β-enhanced interleukin-6 expression levels in cultured DCs. In conclusion, in situ and in vitro results indicate that significantly enhanced interleukin-6 expression levels in DCs during chorioamnionitis could be pivotal in skewing decidual monocyte differentiation to macrophages.

Progestin and thrombin regulate tissue factor expression in human term decidual cells.

CONTEXT Perivascular cell membrane-bound tissue factor (TF) initiates hemostasis via thrombin generation. The identity and potential regulation of TF-expressing cells at the human maternal-fetal interface that confers hemostatic protection during normal and preterm delivery is unclear. OBJECTIVES The objective of the study were to identify TF-expressing cells at the maternal-fetal interface in term and preterm decidual sections by immunohistochemistry and evaluate progestin, thrombin, TNF-alpha, and IL-1beta effects on TF expression by cultured human term decidual cells (DCs). INTERVENTIONS AND MAIN OUTCOME MEASURES Serial placental sections were immunostained for TF. Leukocyte-free term DC monolayers were incubated with 10(-8) M estradiol (E2) or E2 plus 10(-7) M medroxyprogestrone acetate (MPA) +/- thrombin or TNF-alpha or IL-1beta. ELISA and Western blotting assessed TF in cell lysates. Quantitative real-time RT-PCR measured TF mRNA levels. RESULTS Immunolocalized TF in DC membranes in preterm and term placental sections displayed higher Histologic Scores than villous mesenchymal cells (P < 0.05). TF was undetected in interstitial or extravillous trophoblasts. Compared with DCs incubated with E2, MPA and 2.5 U/ml thrombin each doubled TF levels (P < 0.05) and E2 + MPA + thrombin further doubled TF levels (P < 0.05), whereas TNF-alpha and IL-1beta were ineffective. Western blotting confirmed the ELISA results. Quantitative RT-PCR revealed corresponding changes in TF mRNA levels. CONCLUSIONS In human term placental sections, DC-expressed TF exceeds that of other cell types at the maternal-fetal interface and is localized at the cell membranes in which it can bind to factor VII and meet the hemostatic demands of labor and delivery via thrombin formation. Unlike the general concept that TF is constitutive in cells that highly express it, MPA and thrombin significantly enhanced TF expression in term DC monolayers.

Attempted Replication of 50 Reported Asthma Risk Genes Identifies a SNP in RAD50 as Associated with Childhood Atopic Asthma

Objectives: Asthma is a childhood disease that is strongly influenced by genetic factors. We sought to replicate an association between single nucleotide polymorphisms (SNPs) of the top-ranked candidate genes and childhood atopic asthma in Perinatal Risk of Asthma in Infants of Asthmatic Mothers (PRAM) study subjects. Methods: Using data from a systematic literature search and an exploratory genome-wide association study conducted in a subset of the PRAM cohort, we followed a strict procedure to generate a ranked list of candidate genes. SNPs in the top 50 genes were genotyped in the full PRAM cohort (n = 103 cases with doc- tor-diagnosed atopic asthma at age 6, and n = 499 controls). Results: The literature search identified 251 prior risk genes from 469 publications. RAD50 (rs2706347) and PTPRE (rs10830196) revealed crude associations with asthma at a Bonferroni-corrected level of significance (p < 0.0011). IL4R (rs1801275), CCL5 (rs2280788), and TBXA2R (rs4523) revealed nominal significance (p < 0.05). When adjusted for race and gender, only rs2706347 in RAD50 remained significantly associated with asthma. SNPs in frequently replicated asthma risk genes, including TNF, IL13, ADAM33, TGFB1, and MS4A2, revealed no association. Conclusion:RAD50 may be a promising candidate asthma risk gene. Lack of evidence of highly reported polymorphisms in the present study highlights the genetic heterogeneity of asthma and emphasizes the need for robust replication of candidate genes.