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As part of a drug discovery program, 4-pyrazin-2-yl-1H-pyrrole-2-carboxamides were accessed along with a number of bicyclic analogues. Routes to these compounds were largely absent from the scientific literature. The synthesis of a 4-(pyrazin-2-yl)-1H-pyrrole-2-carboxamide and several fused bicyclic analogues all using standard procedures (SNAr, borylation, C–C cross couplings, hydrolysis, amide bond formation, cyclisation, halogenation, and alkylation) from readily available starting materials is reported. The synthetic sequences range from 4–12 steps per final compound, with yields of isolated intermediates ranging from 20 to ∼100%.

This review presents an account of the palladium-catalyzed functionalizations of alkenes and alkynes developed at the Laboratory of Catalysis and Organic Synthesis (LCSO). Starting from the intramolecular oxy- and aminoalkynylation of alkenes, tethered methods were then developed to functionalize allylic amines and alcohols, as well as propargylic amines. Finally, a new dynamic kinetic asymmetric transformation was developed based on the use of a ‘one-arm’ Trost-type ligand, giving access to enantiopure amino alcohols. Each section is a personal account by the researcher(s) who performed the work.1 Introduction,2 Oxy- and Aminoalkynylation of Olefins,3 In Situ Tethering Strategies for the Synthesis of Vicinal Amino Alcohols and Diamines,4 Carboamination of Allylic Alcohols,5 Carbooxygenation of Propargylic Amines,6 Enantioselective Carboetherification/Hydrogenation via a Catalytically Formed Chiral Auxiliary,7 Conclusion

Chiral auxiliaries and asymmetric catalysis are the workhorses of enantioselective transformations, but they still remain limited in terms of either efficiency or generality. Herein, we present an alternative strategy for controlling the stereoselectivity of chemical reactions. Asymmetric catalysis is used to install a transient chiral auxiliary starting from achiral precursors, which then directs diastereoselective reactions. We apply this strategy to a palladium-catalyzed carboetherification/ hydrogenation sequence on propargylic amines, providing fast access to enantioenriched chiral amino alcohols, important building blocks for medicinal chemistry and drug discovery. All stereoisomers of the product could be accessed by the choice of ligand and substituent on the propargylic amine, leading to a stereodivergent process.

PURPOSE: BRAF(V600E), a major driver of thyroid cancer, evaluated in the context of thyroid hormones and human relaxin. METHODS: Immunohistochemical expression of BRAF(V600E), TSH, TSH receptor (TSHR), T4, T3 receptor (T3R), RLNH2, and its receptor, RXFP1, were evaluated in thyroid tumors from a retrospective U.S. population of 481 cancer cases diagnosed in 1983-2004. RESULTS: BRAF(V600E) was expressed in 52% of all thyroid tumors; expression of other markers ranged from 25% for T4 to 98% for RLNH2. Tumors predominantly exhibited hypothyroid-like conditions characterized by elevated TSH and TSHR and reduced T4. BRAF(V600E) prevalence was significantly higher in tumors expressing TSH, TSHR, T3R, and RXFP1 and lower in tumors expressing T4. The proportion of BRAF(V600E) mutation in classic papillary tumors significantly increased from 56% to 72% over the 21-year period of diagnoses while expression of RXFP1, TSH, TSHR, and T3R decreased in non-tumor. Racial/ethnic differences were observed in thyroid hormone marker expression. Non-tumor expression of TSH, TSHR, and T3R were each associated with shorter overall survival but did not remain significant after adjustment for demographic and clinical factors. CONCLUSIONS: Our study provides the first evidence of the potential interaction of BRAF(V600E) mutation, relaxin, and thyroid hormones in thyroid carcinogenesis. Moreover, our results suggest that hypothyroidism, influenced by RLNH2 activity, may underlie the development of the majority of thyroid cancers and mediate the role of BRAF(V600E) in thyroid carcinogenesis. BRAF(V600E) mutation is increasing in papillary thyroid cancers and may be contributing to the rising incidence of this malignancy

Construction projects are characterised by supply chains with multiple contracts and significant transaction costs. An example of these costs is to be found in the management of project delays. The operationalisation and measurement of transaction costs, especially in the construction context, has hitherto proved difficult. The work reported here is concerned with defining and measuring the resources required for the management of contractual disputes relating to project delays. Its main argument is that concepts from transaction cost economics (TCE) theory (bounded rationality, uncertainty, information asymmetry, and opportunistic behaviour) present serious problems for transaction efficiency. This is exemplified in forensic analysis of construction project delays. Data from twelve case studies are analysed, to reveal that up to 90% of time spent on delay analysis was concerned with searching for and validating information which could, ostensibly, be automatically and reliably captured using digital technologies. This research forms part of a wider study that considers the implications of the identified inefficiencies and makes a case for the exploitation of advances in information technology on the more efficient resolution (or even avoidance) of contractual disputes. It concludes that there is a prima facie case for this, and therefore for the reduction in the transaction costs that relate to the management of construction project delays.

Construction delay claims are a leading cause for disputes in the sector. There are two primary aspects to such disagreements: legal and factual. The focus of this paper is the latter. It identifies, evaluates and ranks the factors that impact the analysis of evidence concerning construction delays. The research method is two-fold, involving content analysis of twenty case studies, that identifies the reasons for difference in expert opinion (or factors), and testing of these findings in a survey questionnaire designed to examine the impact and comprehensiveness of those factors. This method provides a more rigorous assessment than previous studies and therefore a more precise list of variables. It is concluded that the factors can be grouped into two categories: (i) materials and documents and (ii) matters of interpretation. Although both categories are important, the deficiencies in the former are often exploited by the parties and/or legal professionals to generate conflicting results in terms of extension of time entitlements. Incorrect, incomplete, undisclosed and/or unagreed records and procedures are used to generate ambiguity and create opportunities to perpetuate claims and disagreements. The work presented here provides additional support to the proposition that opportunistic behaviour is at the core of factual delay disputes and further evidence that tailored contractual delay protocols such as delay analysis clauses can be an effective aspect of a broader solution.

Background Alcohol is a possible risk factor for abdominal aortic aneurysm (AAA), but evidence from individual studies is weak and inconsistent. Existing narrative reviews suggest the possibility of non-linear associations. The aim here was to quantify any association using a systematic literature review, followed by dose–response meta-analysis of prospective studies. Methods MEDLINE, Embase and Web of Science were searched systematically to January 2017 for relevant prospective studies of alcohol consumption and AAA risk. Summary estimates of highest versus lowest levels of consumption, and linear and non-linear dose–response curves were quantified using random-effects models. Results Eleven relevant cohorts were identified describing results from 3580 individuals with among 473 092 participants. Data were extracted from ten cohorts for meta-analyses of high versus low levels of alcohol consumption (risk ratio for AAA 0·93, 95 per cent c.i. 0·78 to 1·11; P = 0·4, I2 = 47 per cent). The linear dose–response risk ratio for AAA, derived from 11 cohorts, was 1·00 (0·97 to 1·04) per 8 g alcohol per day (P = 0·9, I2 = 73 per cent). Non-linear dose–response results showed a tick-shaped curve with lower risk up to 2 units/day, but increasing risk beyond that (P = 0·05). The increase in risk beyond 2 units/day was stronger in men than in women. Conclusion Although the linear dose–response analysis revealed little evidence of an association between alcohol consumption and AAA risk, a tick-shaped trend in the association was observed. This non-linear dose–response analysis revealed reduced risks for alcohol consumption below 2 units/day, masking increased risks for 2 or more units/day.

Background: The combination of pepsinogen, gastrin-17 and anti-H. pylori antibodies serological assays (panel test) is a non-invasive tool for the diagnosis of atrophic gastritis. However, the diagnostic reliability of this test is still uncertain. Aim: To assess the diagnostic performance of the serum panel test for the diagnosis of atrophic gastritis. Methods: Medline via PubMed, Embase, Scopus, Cochrane Library databases and abstracts of international conferences proceedings were searched from January 1995 to December 2016 using the primary keywords “pepsinogens,” “gastrin,” “atrophic gastritis,” “gastric precancerous lesions.” Studies were included if they assessed the accuracy of the serum panel test for the diagnosis of atrophic gastritis using histology according to the updated Sydney System as reference standard. Results: Twenty studies with a total of 4241 subjects assessed the performance of serum panel test for the diagnosis of atrophic gastritis regardless of the site in the stomach. The summary sensitivity was 74.7% (95% confidence interval (CI), 62.0-84.3) and the specificity was 95.6% (95%CI, 92.6-97.4). With a prevalence of atrophic gastritis of 27% (median prevalence across the studies), the negative predictive value was 91%. Few studies with small sample size assessed the performance of the test in detecting the site of atrophic gastritis. Conclusions: The combination of pepsinogen, gastrin-17 and anti-H. pylori antibodies serological assays appears to be a reliable tool for the diagnosis of atrophic gastritis. This test may be used for screening subjects or populations at high risk of gastric cancer for atrophic gastritis; however, a cost-effectiveness analysis is needed.

Relaxin, a systemic and placental hormone, has potential roles in fetoplacental growth. Human placenta expresses two RLN genes, RLNH1 and RLNH2. Maternal obesity is common and is associated with abnormal fetal growth. Our aims were to relate systemic and cord blood RLNH2, placental RLNs and their receptor (RXFP1) with fetoplacental growth in context of maternal body mass index, and associations with insulin-like growth factor 2 (IGF2) and vascular endothelial growth factor A (VEGFA) in the same placentas. Systemic, cord blood and placental samples were collected prior to term labor, divided by prepregnancy body mass index: underweight/normal (N = 25) and overweight/obese (N = 44). Blood RLNH2 was measured by ELISA; placental RLNH2, RLNH1, RXFP1, IGF2 and VEGFA were measured by quantitative immunohistochemistry and mRNAs were measured by quantitative reverse transcription PCR. Birthweight increased with systemic RLNH2 only in underweight/normal women (P = 0.036). Syncytiotrophoblast RLNH2 was increased in overweight/obese patients (P = 0.017) and was associated with placental weight in all subjects (P = 0.038). RLNH1 had no associations with birthweight or placental weight, but was associated with increased trophoblast and endothelial IGF2 and VEGFA, due to female fetal sex. Thus, while systemic RLNH2 may be involved in birthweight regulation in underweight/normal women, placental RLNH2 in all subjects may be involved in placental weight. A strong association of trophoblast IGF2 with birthweight and placental weight in overweight/obese women suggests its importance. However, an association of only RLNH1 with placental IGF2 and VEGFA was dependent upon female fetal sex. These results suggest that both systemic and placental RLNs may be associated with fetoplacental growth.

Visfatin/nicotinamide phosphoribosyltransferase (Nampt), an enzyme involved in energy metabolism and sirtuins, SIRT1 and SIRT3, which are NAD-dependent deacetylases, are critical for cellular function. All three either regulate or are regulated by intracellular NAD+ levels and therefore available cellular energy, important for placental cell survival and successful pregnancy. This study investigates whether these protective proteins are involved in the placental pathophysiology of pre-eclampsia (PE) and if they are associated with 8-oxo-deoxyguanosine (8OHdG), a marker of oxidative damage or with placental telomere length.Maternal blood and placental samples were collected from 31 patients with PE and 30 controls between 31 and 40 weeks gestation. Quantitative immunohistochemistry was performed on placental specimens for visfatin/Nampt, SIRT1, SIRT3, and nuclear 8OHdG. Plasma visfatin was measured by ELISA and telomere length by Southern blot analysis of telomere restriction fragments.Visfatin/Nampt and SIRT1 in syncytiotrophoblast decreased in PE compared to controls (p 0.0001, p = 0.004 respectively). SIRT3 decreased in PE most significantly at preterm (p = 0.002). 8OHdG was only significantly lower in preterm controls compared to term controls (p = 0.01) and correlated with SIRT1 in all samples (r = 0.27). Telomere length was not different in PE and controls.Decreased visfatin/Nampt, SIRT1 and SIRT3 in syncytiotrophoblast in PE suggests a lack of placental reserve in metabolic energy efficiency, increased inflammation, and lower resistance to environmental stressors. However, there was little effect on nuclear function, or evidence of genomic DNA damage, which would lead to cellular senescence and death.

Fluorination is commonly employed to optimize bioactivity and pharmaco-kinetic properties of drug candidates. Aliphatic fluorination often reduces the lipophilicity (log P), but polyfluoroalkylation typically increases lipophilicity. Hence, identification of polyfluorinated motifs that nonetheless lead to similar or even reduced lipophilicities is of interest to expand the arsenal of medicinal chemistry tools in tackling properties such as compound metabolic stability or off-target selectivity. We show that changing a CF

This study was designed to show whether placental relaxin (RLN), its receptor (RXFP1), or insulin-like peptide 4 (INSL4) might have altered expression in patients with placenta accreta. The baseline expression of their genes through gestation (n = 34) was quantitated in the placental basal plate (BP) and villous trophoblast (TR), and compared to their expression in placenta accreta (n = 6). The proteins were also immunolocalized and quantitated in the accreta tissues. The messenger RNAs (mRNAs) of matrix metalloproteinase 9, -2, and tissue inhibitors of matrix metalloproteinase (TIMP)-1 were also measured. Results demonstrated that the BP and TR expressed low levels of RLN/RXFP1 and INSL4 through gestation. In accreta, increased RLN gene and protein in BP were associated with antepartum bleeding whereas INSL4 expression decreased throughout the TR. There were no changes in mRNAs for MMPs, but TIMP-1 was increased only in the invasive TR.

Intrauterine infection frequently leads to preterm birth (PTB), with the pathophysiology involving activation of the innate immune system and its associated inflammatory response. The choriodecidua produces relaxin (RLN) and elevated levels are associated with preterm premature rupture of the fetal membranes. However, it is not increased in bacterially-mediated PTB, but may act as an endogenous sterile inflammatory mediator. Elevated systemic RLN levels from the corpus luteum are also associated with PTB, but the mechanism is unknown. In clinical obstetrics, intrauterine inflammation or infection can coexist with elevated RLN. Therefore, in this study, we further characterized the effects of RLN alone or together with an inflammatory mediator on the production of IL1B, CSF2 (GM-CSF), IL6, IL8 and TNF, from chorionic cytotrophoblasts (CyT), decidual fibroblasts (DF) and stromal cells (DSC), using interleukin-1 beta (IL1B) to mimic sterile inflammation or lipopolysaccharide (LPS) for bacterial infection. Endogenous differences between the cells showed that the CyT expressed more RLN, its receptor RXFP1 and the RXFP1 splice variant D. CyT also showed the most robust cAMP response to RLN with increased IL6 secreted after 4 h, preceded by increased transcription at 1 h, likely due to activation of RXFP1 and cAMP. When all cell types were treated with IL1B and RLN, RLN augmented secretion of IL6 and IL8 from CyT and DF, but not DSC. Similarly, RLN augmented LPS-induced IL6 secretion from CyT and DF. Despite the structural similarity between TLR4 and RXFP1, blocking TLR4 in CyT had no effect on RLN-induced IL6 secretion, suggesting specific activation of RXFP1. Thus, we have shown that in the presence of a low level of intrauterine inflammation/infection, elevated RLN could act on the CyT and DF to augment the inflammatory response, contributing to the pathophysiology of PTB. Summary RLN augments the inflammatory responses induced by IL1B or LPS in chorionic cytotrophoblasts and decidual fibroblasts.

Relaxin (RLN) is a systemic hormone from the corpus luteum, and its levels remain low during normal human gestation. Indeed, elevation of circulating RLN has long been associated with preterm birth, for which there has been no physiological explanation. Recent studies have shown that RLN suppresses endotoxin-induced cytokine secretion from THP-1 monocytic cells by acting on the glucocorticoid receptor (GR), but its effects on primary macrophages are unknown. Therefore, in the present study, we examined the effects of RLN on cytokine secretion from primary decidual macrophages (DMs) obtained at term before labor. Unlike THP-1 cells, RLN had no effects on the cytokine responses induced by either lipopolysaccharide (LPS) or interleukin (IL) 1B, mimicking infection-induced or sterile inflammation, respectively. However, RLN alone for 4 h significantly decreased (P < 0.05) colony-stimulating factor 2 (CSF2; also known as granulocyte-macrophage colony-stimulating factor) and IL8 but for 24 h significantly increased IL6 (P < 0.01). We show that DMs express both the RLN receptor (RXFP1) and the GR. RLN suppression of CSF2 and IL8 was sensitive to the GR-antagonist mifepristone (RU-486). However, RLN activation of RXFP1 induced a dose-dependent cAMP response, which when mimicked by forskolin also caused significantly increased (P < 0.05) secretion of IL6. Thus, RLN may be anti-inflammatory in DMs via activation of the GR but proinflammatory via activation of RXFP1 and cAMP. In summary, we have shown that RLN targeting DMs may modulate proinflammatory cytokine secretion at the maternal-fetal interface and contribute to the localized inflammatory response associated with parturition in women.

We present a 18 m W fiber-coupled single-mode super-luminescent diode with 85 nm bandwidth for application in optical coherence tomography (OCT). First, we describe the effect of quantum dot (QD) growth temperature on optical spectrum and gain, highlighting the need for the optimization of epitaxy for broadband applications. Then, by incorporating this improved material into a multicontact device, we show how bandwidth and power can be controlled. We then go on to show how the spectral shape influences the autocorrelation function, which exhibits a coherence length of < 11 mu m, and relative noise is found to be 10 dB lower than that of a thermal source. Finally, we apply the optimum device to OCT of in vivo skin and show the improvement that can be made with higher power, wider bandwidth, and lower noise, respectively.

Size-related diet shifts are important characteristics of fish trophodynamics. Here, body size–related changes in muscle δ15N and δ13C of four coral reef fishes, Acanthurus nigrofuscus (herbivore), Chaetodon lunulatus (corallivore), Chromis xanthura (planktivore) and Plectropomus leopardus (piscivore) were investigated at two locations in the Solomon Islands. All four species occupied distinct isotopic niches and the concurrent δ13C′ values of C. xanthura and P. leopardus suggested a common planktonic production source. Size-related shifts in δ15N, and thus trophic level, were observed in C. xanthura, C. lunulatus and P. leopardus, and these trends varied between location, indicating spatial differences in trophic ecology. A literature review of tropical fishes revealed that positive δ15N-size trends are common while negative δ15N-size trends are rare. Size-δ15N trends fall into approximately equal groups representing size-based feeding within a food chain, and that associated with a basal resource shift and occurs in conjunction with changes in production source, indicated by δ13C. The review also revealed large scale differences in isotope-size trends and this, combined with small scale location differences noted earlier, highlights a high degree of plasticity in the reef fishes studied. This suggests that trophic size analysis of reef fishes would provide a productive avenue to identify species potentially vulnerable to reef impacts as a result of constrained trophic behaviour.

This paper details the development of broadband sources at 1050 nm for optical coherence tomography applications. A method for obtaining a broad and smooth emission and gain spectrum for 1050 nm quantum dot (QD) layers is presented. The design, fabrication, and operating characteristics of multicontact superluminescent diodes are then set out, and the operating characteristics of this device, incorporating the broadband QD material, are described. It is shown that this device allows the tuning of the emission spectrum peak position, peak shape, emission bandwidth, and power, which are advantageous for imaging applications. A simplified device, utilizing a single current source, is achieved by incorporating a resistor network. The operation of the multicontact device under various drive topologies is discussed.

In normal pregnancy, the fetal membranes become increasingly distended towards term and in multifetal gestations they become over-distended. Apoptosis of the amniotic epithelium increases with advancing gestation and may contribute to fetal membrane weakening and rupture. The effects of chronic static stretching for 36 h have been investigated using primary amniotic epithelial cells. Pre-B cell colony-enhancing factor (PBEF) is a stretch-responsive cytokine and expression of its gene, intracellular and secreted protein were all significantly increased by 4 h and its secretion sustained over 36 h, contrasting with the rapid increase and decline in expression of IL-8. Increased expression of SIRT1 and decreased p53 paralleled the changes in PBEF, are known to be responsive to PBEF, and contribute to cell survival. Distension had no effects on proliferation or necrosis but protected the cells from apoptosis, knocking-down PBEF with antisense probes abrogated this protective effect. There was increased immunostaining of PBEF in the compact layer of the amnion in multifetal tissues and significantly fewer apoptotic amniotic epithelial cells. These results show that chronic stretching of the amniotic epithelial cells increases PBEF expression, which protects them from apoptosis.

A general method for the synthesis of functionalized pyridazinylboronic acids/esters is described involving a directed ortho metalation (DoM)--boronation protocol (Schemes 1 and 2). A comprehensive study of the reactivity of the C-B bond in palladium-catalyzed cross-couplings with aryl/heteroaryl halides is presented. Aryl-/heteroarylpyridazines are thereby obtained in synthetically viable yields (typically 40-75%) although in some cases competing protodeboronation has been observed. A series of pyridazin-3(2H)-one derivatives, including 4,6-diaryl/heteroaryl derivatives, have been obtained from the corresponding 3-methoxypyridazines in straightforward procedures (Schemes 3 and 4). Several X-ray crystal structures of aryl-/heteroarylpyridazines and derived pyridazin-3(2H)-one derivatives are reported. These multi-ring systems are of considerable interest in contemporary N-heterocyclic chemistry.

The human fetal membranes are complex tissues that perform many important functions during gestation. The extracellular matrix provides their strength to withstand the forces directed from the fetus and myometrium. Relaxin is a collagenolytic hormone that causes increased production of the matrix metalloproteinases. Its expression from the decidua is increased in patients with preterm premature rupture of the membranes, and its leucine-rich G receptor 7 is upregulated at preterm. The authors previously showed that relaxin is not involved in the infection-mediated cytokine response, but in the absence of infection, it causes increased secretion of both interleukin -6 and interleukin-8 from the membranes. In this article, the authors propose that relaxin is one of a number of sterile stimuli capable of causing increased proinflammatory cytokines, similar to but less robust than the effects of infection. These probably represent distinct inflammatory pathways involving different intracellular signaling events, which can result in either preterm premature rupture of the membranes or preterm labor. The current challenge is to fully understand these pathways and to clarify their similarities and differences.

A localized intrauterine inflammatory response is often associated with the initiation of normal human parturition, whereas infection causes a similar but more florid response initiating preterm labor. Pre-B-cell colony-enhancing factor (PBEF) is expressed in the human fetal membranes and is up-regulated by labor, severe infection and inflammatory stimuli. The aim of this study was to determine the involvement of the transcription factors NF-kappaB and AP-1 in the response of PBEF to an inflammatory stimulus and compare it with IL-8. The results showed that this treatment of amniotic epithelial-like cells (WISH) and primary amniotic epithelial cells increased expression of PBEF and IL-8, but IL-8 responded 100-fold more than PBEF. IL-1beta treatment together with a panel of NF-kappaB and AP-1 inhibitors demonstrated the involvement of these transcription factors in the up-regulation of PBEF. These data show that an inflammatory stimulus in the fetal membranes inducing NF-kappaB and AP-1 would up-regulate PBEF as well as IL-8.

Safety issues related to thermal cook-off are important for handling and storing explosive devices. Violence of an event as a function of confinement is important for prediction of collateral events. There are major issues which require an understanding of the following events: (1) transit to detonation of a pressure wave from a cook-off event, (2) sensitivity of changes in HMX-based explosives with thermally induced phase transitions, and (3) the potential danger of neighboring explosive devices being affected by a cook-off reaction. Results of cook-off events of known size, confinement, and thermal history allow for development and/or calibration of computer models for calculating events that are difficult to measure experimentally.

Visfatin is both a systemic adipocytokine and the cytosolic enzyme, nicotinamide phosphoribosyl transferase (Nampt). This is a longevity protein, which extends the lifespan of human cells by activating sirtuin 1 (SIRT1). In this study, we sought a role for these proteins in obese pregnant women, who experience more postterm deliveries. Thus, 78 women (26 lean, 24 overweight, and 28 obese) were recruited and maternal blood and placental tissue collected prior to term labor. Plasma levels were measured by enzyme-linked immunosorbent assay and quantitative immunohistochemistry used for placenta. We confirmed maternal plasma interleukin 6 increased according to prepregnancy body mass index (BMI; P < .0001) and showed a linear relationship between BMI and syncytiotrophoblast visfatin/Nampt (P = .021) but not with its levels in maternal plasma. Both systemic and placental visfatin/Nampt were significantly associated with placental SIRT1 levels (P = .028 and .017). Thus, higher visfatin/Nampt may prevent a labor-associated decrease in SIRT1 leading to postterm delivery in obesity.

This study aimed to gain new insights into both systemic and placental leptin and its receptors, with reference to the maternal prepregnancy body mass index (BMI). Thus, 84 women (29 lean, 24 overweight, and 31 obese) were recruited and maternal, cord blood, and placental tissues collected prior to term labor. Plasma levels were measured by enzyme-linked immunosorbent assay and for placenta, immunohistochemistry and messenger RNAs (mRNAs) were quantitated. We confirmed that maternal leptin increased linearly as the soluble receptor decreased with BMI (P = .001). Fetal leptin increased with maternal BMI (P = .02) and birth weight (P = .006) and was higher in female infants (P < .001). Placental mRNA levels of leptin and its receptors showed no change in BMI. However, we show a significant (P = .043) linear increase in leptin in the placental vascular endothelial cells with maternal obesity, while leptin in syncytiotrophoblast showed no statistical change. Leptin receptors localized to syncytiotrophoblast and intravillous macrophages and were unchanged with BMI.

The relaxin receptor has been recently described as a leucine-rich repeat G-protein coupled receptor and designated as LGR7. A closely related receptor, LGR8, is co-expressed by some cells. This study explored the expression of the genes for these receptors in the human fetal membranes and placenta by RT-PCR and the LGR7 protein by immunolocalization. The results showed that LGR7 was well expressed in the fetal membranes, with significantly more in the decidua (p < 0.05) than in the amnion. On the other hand, relatively low levels were expressed in the placenta. The major splice variant of LGR7 was undetectable in either the placenta or fetal membranes. Expression of LGR8 was also below the level of detectability in either tissue. Immunostaining for LGR7 was conducted with antisera to both its endomain and ectodomain, in order to seek evidence for a solubilized ectodomain. However, similar staining patterns were obtained with both antisera, with predominant staining in the cells of the amniotic epithelium, chorionic cytotrophoblast and decidua. Full-thickness fetal membranes from preterm deliveries, before and after labor or after preterm premature rupture of the membrane (PPROM) and labor were collected. In addition, membranes at term, both before and after spontaneous labor were used for analysis of LGR7 gene expression. There was significantly greater LGR7 expressed (p = 0.01) in the preterm period compared to term, indicating a potentially important role for relaxin at this time. There was a marginal decline in LGR7 gene expression after labor and delivery both at preterm and term, which did not reach significance. Immunostaining patterns showed less inter-patient variability than did gene expression, with more intense staining for LGR7 after labor and delivery.

Improved methods for selecting cultivars for their ability to grow well on low P soils will improve productivity and minimise pollution. Estimates are required of the relative values for different genotypes of physiologically meaningful parameters that define yield response to increasing levels of plant available soil P. The object of this study was to devise ways of obtaining these estimates from conventional trials that include numerous genotypes but only two P treatments. The parameters were estimated as coefficients of a modified Michaelis–Menten equation, namely A, the maximum yield that can be obtained with ample P, Km, the concentration of plant available P in the rooting medium at which yield is half the maximum, and B, the gradient of yield against available P as it approaches the origin. Two novel methods of estimation were devised. They both require inputs of yield and its variance at each of two concentrations of available P, but these must be within certain limits. Estimates of parameter values for 12 Brassica oleracea genotypes made by these methods from two P-levels in each of two experiments with six P levels were compared with the values obtained by conventional fitting to the data from all six P levels. According to regression and ranking analyses, the relative values of A obtained for the different genotypes by both two P-level methods were similar to those from the six P-level method and both were reproducible. None of the methods however detected any reproducible differences in Km. Application of one of the two P-level methods to published data for genotypes of other species indicated that there was little intra-species variation in Km within some data sets on maize, beans and leguminous cover crops but much within other data sets. Sensitivity and algebraic analyses are presented to define the experimental conditions required for successful use of two P-level methods. It is concluded that, provided these experimental conditions are met, measurements at two P levels can give almost as much information about intra-species differences in P response parameters as measurements at six P levels. The procedure may also be useful for interpreting other responses of a “diminishing returns” type such as those to other nutrients or plant spacing.

Early placental insulin-like protein (INSL4 or EPIL) is a member of the insulin superfamily of hormones, which is highly expressed in the placenta. We have confirmed this at term and shown it to be expressed by the maternal decidua. Although an abundance of locally acting growth factors are produced within the uterus during pregnancy, we hypothesized that INSL4 plays an important role in fetal and placental growth. We have demonstrated with cell lines and primary cells that it has a growth-inhibitory effect by causing apoptosis and loss of cell viability. We used primary amniotic epithelial cells for flow cytometry to show that INSL4 caused apoptosis, which was dose-related and significant (P < 0.05) at 50 ng/ml. This was confirmed by measurement of the nuclear matrix protein in the media. In comparison, relaxin treatment (up to 200 ng/ml) had no effect on apoptosis. The addition of INSL4 (3-30 ng/ml) also caused a loss of cell viability, although it had no effect on the numbers of cells at different phases of the cell cycle. Placental apoptosis is an important process in both normal placental development and in fetal growth restriction. Therefore, an in vivo clinical correlate was sought in fraternal twins exhibiting discordant growth. Expression of the INSL4 gene was doubled in the placenta of the growth-restricted twin compared to the normally grown sibling, suggesting that it may be linked to a higher level of apoptosis and loss of cell viability and, therefore, that it may contribute to fetal growth restriction.

Better criteria for selecting cultivars for their ability to grow well on low-P soils could reduce waste of fertilizer P. The objective of this study was to quantify and determine the reproducibility of two crop parameters (A and K m ) responsible for differences in the response of 12 Brassica oleracea L. genotypes to P supply. Modified Michaelis-Menten equations were fitted to shoot dry weight responses to P supply for each genotype in each of three glasshouse and three field experiments. One of the fitted parameters (A) defined the maximum shoot dry weight that could be obtained with ample P, and the other (K m ) defined the efficiency of root P-uptake as the concentration of extractable soil P in the rooting medium at which growth was half the maximum. The equations fitted the data well. Ranking and regression analyses showed that intergenotypic differences in A were considerable, and reproducible, but that differences in K m were small. Thus, B. oleracea genotypes yielding most on a P-sufficient soil will also yield most on a P-deficient soil. However, both the maximum yield (A) and the efficiency of root-P uptake (K m ) vary considerably between different vegetable crops. Thus, a graphical procedure to select crops for highest yields on low-P soils, on the basis of a comparison of A and K m values, is described.

The purpose of this study was to determine whether pre-B-cell colony-enhancing factor is a secreted cytokine in the human amnion and to study its chemotaxic and antiapoptotic properties.Pre-B-cell colony-enhancing factor secretion was studied from amniotic epithelial-like WISH cells and primary amniotic epithelial cells that were seeded on squares of immobilon-P membrane and stimulated with lipopolysaccharide or tumor necrosis factor-alpha, respectively. The pre-B-cell colony-enhancing factor protein was detected both intracellularly and after secretion, as bound to the membrane, by immunostaining and densitometry. Medium and cell lysates that were obtained from WISH cells that were treated with lipopolysaccharide alone or together with a pre-B-cell colony-enhancing factor antisense oligonucleotide to block pre-B-cell colony-enhancing factor translation were also analyzed for secreted pre-B-cell colony-enhancing factor by Western blotting and densitometry. A chemotaxic effect of pre-B-cell colony-enhancing factor on human neutrophils was compared with the chemoattractants interleukin-8 and N-Formyl-Met-Leu-Phe methyl ester in a rapid fluorescence-based neutrophil migration assay. Apoptosis was induced in primary amniotic epithelial cells and fibroblasts by actinomycin D (1 microg/mL); the antiapoptotic effects of pre-B-cell colony-enhancing factor on early apoptosis were measured by the annexin V assay, and the late effects were determined by measurement of nuclear matrix protein in the media.Treatment of amnion cells that adhered to immobilon-P membrane to induce the secretion of pre-B-cell colony-enhancing factor showed significantly (P.05) more pre-B-cell colony-enhancing factor protein surrounding the cells compared with the controls. Although the addition of lipopolysaccharide to cultured WISH cells caused the secretion of pre-B-cell colony-enhancing factor into the medium, co-treatment with an antisense oligonucleotide to pre-B-cell colony-enhancing factor obliterated it. Analysis of the cell lysates showed no significant change, which suggests that most of the pre-B-cell colony-enhancing factor protein had been secreted. No significant chemotaxic effects of pre-B-cell colony-enhancing factor were observed; however, pre-B-cell colony-enhancing factor treatment (100 ng/mL), together with actinomycin D, cancelled the early induction of apoptosis, although there was a dose-dependent and significant late antiapoptotic effect on primary amnion epithelial cells (P.001) and fibroblasts (P.01).Pre-B-cell colony-enhancing factor is a secreted protein from amniotic epithelial cells. Although it had no chemotaxic effects, it was antiapoptotic for both amniotic epithelial cells and fibroblasts and may protect these cells against apoptosis that is induced by chronic distension, labor, or infection.

Relaxin, a peptide hormone important to the outcome of human pregnancy is expressed in a tissue specific manner as two genes known as relaxins H1 and H2, in addition to a third human relaxin H3, expressed primarily in the brain. The H1 and H2 genes are highly homologous, differentially expressed in reproductive tissues and appear to activate the same receptor, but their regulation is poorly understood. Based upon the known physiology of these hormones and the response elements in their 5'- and 3'-flanking regions, the possibility that progesterone and/or the glucocorticoids might influence their differential expression was therefore investigated. The changes in the mRNA levels of the relaxin genes in response to either medroxyprogesterone acetate (MPA) or dexamethasone (Dex) were analyzed by RT-PCR using a choriocarcinoma cell line (JAR) as a model system, because the expression of these genes in any primary human cell type is too low for such a study. The addition of 0.5 microM MPA to JAR cells, significantly upregulated the mRNA of only the relaxin H2, while the addition of 0.5 microM Dex significantly upregulated the mRNAs for both the relaxins, after 6h of treatment. Promoter assays indicated an early activation of transcription (1 h), which by 6 h had decreased. Progesterone and/or glucocorticoids could exert their effects via the GRE motif found on the 5'-flanking region of the relaxin genes. The H1-GRE differs from the H2-GRE by a single nucleotide, which may affect H1-GRE binding to the progesterone receptor (PR) but not the glucocorticoid receptor (GR). The antiprogestin RU486 inhibited the binding of the GR to both H1-GRE and H2-GRE, while it enhanced the binding of the PR to these GREs. As determined by gel shift assays, this GRE motif could bind to both the PR and GR and was therefore considered to be functional. Thus, both progesterone and glucocorticoids are capable of differentially regulating the expression of the two human relaxin genes in a model system.

A silicon-detector-based neutral particle analyzer array was used to study fast ion distributions in the Large Helical Device for different plasma heating conditions. GNET code simulations of the measurements are needed for accurate interpretation of the data.

The study was conducted to determine whether relaxin has a proliferative effect on amniotic epithelial cells and to show that this effect is caused by its stimulation of the insulin-like growth factor-II (IGF-II) gene.Immunolocalization and Northern analysis were used to confirm the expression of IGF-II by the fetal cells in the membranes. Human amniotic epithelial (WISH) cells were treated with doses of IGF-II or human relaxin and their proliferative effects measured. The mechanism of the effect of relaxin on cellular proliferation was studied with the use of an IGF-II-blocking antibody and Northern analysis for IGF-II gene expression after treatment with relaxin. An in vivo correlate was sought by quantitation of relaxin gene expression in 10 fetal membranes from women with normally grown and large for gestational age infants.The amniotic epithelial and cytotrophoblast cells of the fetal membranes expressed IGF-II, as did the amniotic epithelial-like (WISH) cell line. Treatment of WISH cells with IGF-II or relaxin caused a significant (P.03) and dose-related increase in WISH cell proliferation over 5 days. The concurrent treatment with a blocking antibody to IGF-II significantly decreased the proliferative response to IGF-II (P.002) and relaxin (P.002). Treatment with relaxin caused a significant increase (P.003) in the transcription of IGF-II in 24 hours. In fetal membranes, the levels of relaxin gene expression correlated with fetal membrane surface area (r = 0.76) and was significantly greater (P.008) in the membranes from macrosomic infants (4020-4729 g) compared with those normally grown (2855-3830 g).IGF-II and relaxin both caused the proliferation of WISH cells. Concurrent treatment with an IGF-II-blocking antibody abrogated the proliferative effects of both hormones. Relaxin increased the transcription of IGF-II, and its expression levels in the fetal membranes correlated with the membrane surface area as well as neonatal birth weight. These data suggest that relaxin is a growth factor for the fetal membranes.

Objective: Our purpose was to determine whether pre-B-cell colony-enhancing factor (PBEF) is expressed in the human fetal membranes during normal gestation and parturition in the absence of infection and to show its effects on the expression of interleukin (IL)-6 and IL-8. Study Design: PBEF was immunolocalized in the fetal membranes from early pregnancy, at preterm, and at term. Its expression was quantitated by Northern analysis in separated uninfected amnion, chorion, decidua, and placenta of patients at term before labor and in full-thickness membranes before and after spontaneous labor at preterm and at term. Amnion-like epithelial (WISH) cells and fetal membrane explants were treated with recombinant PBEF (rhPBEF), and the expression of IL-6 and IL-8 was quantitated. Results: PBEF was immunolocalized throughout gestation in the amniotic epithelium and mesenchymal cells as well as the chorionic cytotrophoblast and parietal decidua. Northern analysis showed significantly more (P

Preterm birth (PTB) is a global problem with a high incidence in the developing world. Relaxin (RLN) has classically been associated with parturition, but its role(s) in the human have been difficult to determine. For the first time, we bring together the systemic (ovarian) and autocrine/paracrine (intrauterine) sources of RLN, in an attempt to understand how RLN contributes to PTB in women.

Objective: To compare the clinical failure rates of brackets bonded using a prototype hydrophilic primer, designed to be insensitive to moisture, with brackets bonded with a conventional primer.Design: Single centre randomised controlled clinical study. Thirty-three patients were bonded using a split mouth technique: randomly allocating the hydrophilic primer to one side of the mouth and a conventional primer to the other.Setting: Hospital orthodontic department, Bradford, UK.Subjects: Orthodontic patients requiring fixed appliancesMain outcome measures: The site and time to bond failure was recorded for each bracket that failed over 6 months.Results: Using survival analysis, there was an increased risk of bracket failure when bonded with the hydrophilic primer compared with the conventional primer (hazard ratio = 2•2, 95% confidence interval: 1•1 to 4•5, P = 0•01).Conclusions: This hydrophilic primer cannot be recommended for routine clinical use.

Pre-B-cell colony enhancing factor (PBEF) was first isolated from an activated peripheral blood lymphocyte cDNA library and was found to be involved in the maturation of B-cell precursors. It was subsequently identified as one of the genes upregulated by distending the human fetal membranes in vitro. Here we report on the genomic organization of this gene, which is composed of 11 exons and 10 introns, spanning 34.7 kb of genomic DNA. Neither the gene nor the protein has any homology with other cytokines in any currently available database. The use of two promoters (proximal and distal) may result in differential, tissue specific expression of the PBEF transcripts. The 5'-flanking region lacks the classical sequence motif that would place it with the hematopoietic cytokines; however, it has several putative regulatory elements, suggesting that this gene may be chemically and mechanically responsive to inducers of transcription. The three PBEF mRNA transcripts were observed in both normal and infected human fetal membranes but were significantly upregulated (P