The effect of uterine fibroids on embryo implantation

Uterine fibroids are common but their role in infertility and effect on embryo implantation is unclear. There is evidence that submucosal fibroids are associated with poor reproductive outcome and that treatment with myomectomy is associated with an improvement in pregnancy rates. Various theories have been proposed to explain this relationship. Fibroids cause a mechanical distortion of the endometrial cavity-their presence may alter gamete and embryo transport (due to blockage of the tubal ostia or by altering uterine contractility and peristalsis) and subsequent embryo implantation (due to compression of the endometrium). They may lead to disruption of the junctional zone within the myometrial layer, affecting general uterine function in the initial stages of embryo invasion and later placentation. Altered vasculature due to the abnormal expression of angiogenic factors by uterine fibroids (such as basic fibroblast growth factor and platelet-derived growth factor) could play a role in a reduced implantation rate in patients with fibroids. Similarly, changes in the endometrium mediated by inflammation and factors involved in the process of fibrosis (such as transforming growth factor) could also have a detrimental effect. In addition, fibroids may affect gene expression pattern in the endometrium (such as HOXA10), disrupting the window of implantation. The supporting evidence for these theories is discussed in this review.     

The M1 muscarinic receptor is present in situ as a ligand-regulated mixture of monomers and oligomeric complexes

The quaternary organization of rhodopsin-like G protein-coupled receptors in native tissues is unknown. To address this we generated mice in which the M₁ muscarinic acetylcholine receptor was replaced with a C-terminally monomeric enhanced green fluorescent protein (mEGFP)–linked variant. Fluorescence imaging of brain slices demonstrated appropriate regional distribution, and using both anti-M₁ and anti–green fluorescent protein antisera the expressed transgene was detected in both cortex and hippocampus only as the full-length polypeptide. M₁-mEGFP was expressed at levels equal to the M₁ receptor in wild-type mice and was expressed throughout cell bodies and projections in cultured neurons from these animals. Signaling and behavioral studies demonstrated M₁-mEGFP was fully active. Application of fluorescence intensity fluctuation spectrometry to regions of interest within M₁-mEGFP–expressing neurons quantified local levels of expression and showed the receptor was present as a mixture of monomers, dimers, and higher-order oligomeric complexes. Treatment with both an agonist and an antagonist ligand promoted monomerization of the M₁-mEGFP receptor. The quaternary organization of a class A G protein-coupled receptor in situ was directly quantified in neurons in this study, which answers the much-debated question of the extent and potential ligand-induced regulation of basal quaternary organization of such a receptor in native tissue when present at endogenous expression levels.

Measuring and understanding the extent and potential significance of quaternary organization of members of the class A (rhodopsin-like) family of G protein-coupled receptors (GPCRs) have both fascinated and frustrated researchers for many years (1, 2). Over time, a wide range of methods have been applied to address this question, and many different GPCRs have been examined. Outcomes have ranged from assertions that such receptors are monomeric and that results consistent with other conclusions reflect either artifacts of the method of measurement or that studies have been performed at nonphysiological levels of expression of the receptor being studied, to those that have suggested rather stable dimeric or tetrameric complexes (1). Only in the case of rhodopsin, the photon receptor expressed at very high levels (in the range of 24,000–30,000 molecules/µm²) in rod outer segments of the eye, have detailed studies been conducted in situ on a class A GPCR. In this example, various studies have shown that rhodopsin is organized as rows of dimers (3, 4). However, to our knowledge, no other GPCR is expressed natively at levels akin to rhodopsin. As such, although a substantial number of studies, generally performed in transfected cell lines or in artificial bilayer systems, have provided evidence that other GPCRs can and do form dimeric and/or higher-order quaternary complexes in a concentration-dependent manner (1, 2), how levels of expression required to observe such complexes relate to expression levels in native cells and tissues has been poorly defined, as is the stability of such complexes and whether they are regulated by ligand binding.Developments in fluorescence fluctuation analysis (FFA) have facilitated efforts to define the oligomeric status of transmembrane receptor proteins (5, 6). Unlike methods based on resonance energy transfer, only a single fluorophore-linked protein is required to be expressed to use FFA. It is, therefore, more practical to use such methods in native cells and tissues if linked to genome-editing approaches and/or the generation of transgenic “knock-in” animal models in which a receptor of interest is replaced with a fluorophore-tagged, modified form of the receptor. Moreover, the recent introduction of fluorescence intensity fluctuation (FIF) spectrometry (7–10) has overcome issues with other methods based on FFA that result in information being compressed due to averaging of oligomer-size data from interrogated regions of interest (RoIs) in which complex mixtures of oligomers of different sizes may be present (7, 8).To define whether the class A M₁ muscarinic acetylcholine receptor is present in hippocampal and cortical neurons as strict monomers or as a range of monomeric, dimeric, and, potentially, oligomeric complexes, we applied FIF spectrometry to images of such neurons isolated from a line of transgenic mice in which we replaced the M₁ receptor with a form of the receptor that includes C-terminally linked monomeric enhanced green fluorescent protein (mEGFP). We first show that both expression levels and function of the introduced M₁-mEGFP construct appear equivalent to the native M₁ receptor in wild-type (WT) mice, using a range of methods and measures ranging from [³H]ligand binding and cell signaling assays to locomotion. We then demonstrate in hippocampal and cortical neurons that in the basal state, the M₁-mEGFP construct is present as a mixture of monomers and dimeric or oligomeric complexes. We also show that the presence of either an agonist or an antagonist ligand promotes monomerization of the receptor. In these studies, we combined analysis of images of a fluorophore-modified receptor in situ with calculation of receptor oligomer complexity. The studies provide a clear and unambiguous answer to a long-standing question that has been the subject of considerable debate (11–13) but that has previously been restricted to studies performed on transfected cell lines. Moreover, these studies are a model for subsequent studies for researchers who plan to explore the topic of dimerization of rhodopsin-family GPCRs.     

Why MSM in Rural South African Communities Should be an HIV Prevention Research Priority

Research into HIV and men who have sex with men’s (MSM) health in South Africa has been largely confined to the metropolitan centres. Only two studies were located making reference to MSM in rural contexts or same-sex behaviors among men in the same. There is growing recognition in South Africa that MSM are not only disproportionately affected by HIV and have been underserved by the country’s national response, but that they contribute significantly to sustaining the high number of new infections recorded each year. We argue that to meet the objectives of the country’s national strategic plan for HIV, STI and TB it is important we know how these behaviours may be contributing to the sustained rural HIV epidemic in the youngest age groups and determine what constitutes appropriate and feasible programmatic response that can be implemented in the country’s public sector health services.     

Magnetic resonance imaging of the normal pituitary gland using ultrashort TE (UTE) pulse sequences (REV 1.0)

Introduction The purpose of this study was to examine the normal pituitary gland in male subjects with ultrashort echo time (TE) pulse sequences, describe its appearance and measure its signal intensity before and after contrast enhancement. Methods Eleven male volunteers (mean age 57.1 years; range 36–81 years) were examined with a fat-suppressed ultrashort TE (= 0.08 ms) pulse sequence. The studies were repeated after the administration of intravenous gadodiamide. The MR scans were examined for gland morphology and signal intensity before and after enhancement. Endocrinological evaluation included baseline pituitary function tests and a glucagon stimulatory test to assess pituitary cortisol and growth hormone reserve. Results High signal intensity was observed in the anterior pituitary relative to the brain in nine of the 11 subjects. These regions involved the whole of the anterior pituitary in three subjects, were localised to one side in two examples and were seen inferiorly in three subjects. Signal intensities relative to the brain increased with age, with a peak around the sixth or seventh decade and decreasing thereafter. Overall, the pituitary function tests were considered to be within normal limits and did not correlate with pituitary gland signal intensity. Conclusion The anterior pituitary shows increased signal intensity in normal subjects when examined with T₁-weighted ultrashort TE pulse sequences. The cause of this increased intensity is unknown, but fibrosis and iron deposition are possible candidates. The variation in signal intensity with age followed the temporal pattern of iron content observed at post mortem. No relationship with endocrine status was observed.     

Graft loss following renal transplantation in Australia: is there a centre effect?

BACKGROUND: Assessment of centre variation in renal transplantation outcome provides an opportunity to examine differences in quality of care between centres. However, differences in outcome may represent differences in patient factors between centres and be biased by sampling variability and inadequate data ascertainment. METHODS: Differences in 12-month graft survival in 1986 primary renal transplant adult recipients from 16 centres in Australia between 1993 and 1998 were examined. Fifteen recipient and donor factors known prior to transplantation were examined to determine factors independently predictive of graft survival. Differences between centres in these factors were examined. Unadjusted and multivariable adjusted outcomes for each centre were compared to the average for all centres. Multivariable hierarchical modelling was employed to account for potential bias due to sampling variability. RESULTS: Factors predictive of reduced 12-month graft survival on multivariable analysis that were significantly different between centres were time on dialysis prior to transplantation, donor age, organ source, and number of human lymphocyte antigen mismatches. Unadjusted 12-month graft survival for all centres was 91.7% and ranged from 83.1 to 96.4%. Although two centres performed significantly lower than average, this poorer outcome was accounted for in one of these two centres after adjusting for factors shown to be independently predictive of outcome. However, multivariable hierarchical modelling failed to identify any centre as performing significantly different to average, with 12-month graft survival ranging from 89.2 to 92.2%. Outcome in patients excluded from the study due to inadequate data ascertainment was significantly worse than patients who were included. CONCLUSIONS: There was no evidence of centre variation after accounting for variation in risk factors predictive of poor outcome between centres, as well as potential bias due to sampling variability. Exclusion of patients due to inadequate data remains an important source of bias in estimating accurate outcomes. Appropriate analytical strategies and consideration of sources of bias are important for the valid identification of centres with poorer outcomes.