Healthcare staff perceptions towards influenza and potential COVID-19 vaccination in the 2020 pandemic context.

The COVID-19 pandemic generated renewed focus on infectious disease transmission in healthcare settings. This study aimed to evaluate staff perceptions towards influenza vaccination in the COVID-19 context. All healthcare workers within a major UK tertiary referral hospital were invited to answer a survey conducted from September 2nd to 13th, 2020. In all, 593 responses were received across a spectrum of roles; 44% reported they were more likely to get an influenza vaccine this year due to COVID-19; however, 10% felt that an influenza vaccine was less important due to social distancing. Additional questions evaluated intention to receive COVID-19 vaccination. There were substantial differences of opinion between staff groups.

Healthcare staff perceptions and misconceptions regarding antibody testing in the United Kingdom: implications for the next steps for antibody screening.

BACKGROUND: Healthcare workers have been at increased risk of exposure, infection and serious complications from COVID-19. Antibody testing has been used to identify staff members who have been previously infected by SARS-CoV-2, and has been rolled out rapidly in the United Kingdom. A number of comment and editorial articles have been published that raise concerns about antibody testing in this context. We present perceptions of National Health Service (NHS) healthcare workers in relation to SARS-CoV-2 antibody testing. METHODS: An electronic survey regarding perceptions towards SARS-CoV-2 antibody testing was distributed to all healthcare workers at a major NHS tertiary hospital following implementation of antibody testing. RESULTS: In total, 560 healthcare workers completed the survey (80% female; 25% of Black and Minority Ethnic background; 58% from frontline clinical staff). Exploring whether they previously had COVID-19 was the primary reported reason for choosing to undergo antibody testing (85.2%). In case of a positive antibody test, 72% reported that they would feel relieved, whilst 48% felt that they would be happier to work in a patient-facing area. Moreover, 12% responded that a positive test would mean "social distancing is less important", with 34% of the responders indicating that in this case they would be both less likely to catch COVID-19 and happier to visit friends/relatives. CONCLUSIONS: NHS staff members primarily seek out SARS-CoV-2 antibody testing for an appropriate reason. Based on our findings and given the lack of definite data regarding the extent of immunity protection from a positive SARS-CoV-2 antibody test, significant concerns may be raised regarding the reported interpretation by healthcare workers of positive antibody test results. This needs to be further explored and addressed to protect NHS staff and patients.

The relationship between serum prolactin concentration and pregnancy outcome in women with unexplained recurrent miscarriage.

The role of prolactin in early pregnancy is controversial. The aim of this study was to evaluate the relationship between serum prolactin concentration and the risk of miscarriage in women with unexplained recurrent miscarriage (RM). A series of 174 women with unexplained RM, who had serum prolactin concentrations measured from January 2000 to September 2009 at the Recurrent Miscarriage Clinic in Royal Hallamshire Hospital in Sheffield, were included in this study. Among the 174 patients with unexplained RM, 40 patients did not conceive during the study period, 9 were lost to follow-up and 125 patients conceived again. Patients who did not conceive were significantly older than those who conceived (p < 0.05, OR: 1.08, 95% CI: 1.03-1.13). Among those who conceived again, the pregnancy outcome data were available for analysis in 109 patients. Those who miscarried were older (p < 0.05, OR: 1.1, 95% CI: 1.01-1.22) and had significantly lower serum prolactin concentrations (p < 0.05, adjusted OR: 0.99, 95% CI: 0.97-0.99) after adjustment has been made for age, than those who had a live birth. Lower basal serum prolactin concentrations were associated with an increased risk of miscarriage in a subsequent pregnancy in women with unexplained RM.

A prospective single-centre evaluation of the intra-individual variability of faecal calprotectin in quiescent Crohn's disease.

BACKGROUND: As a non-invasive marker of gastrointestinal inflammation, faecal calprotectin (FC) is being increasingly used to guide the management of Crohn's disease. It is therefore a concern that studies have shown variability in day to day levels. AIM: To determine the degree of this intrapersonal variability in the context of quiescent Crohn's disease. METHODS: A single-centre prospective study was undertaken in 143 Crohn's disease patients in clinical remission. Three faecal calprotectin levels were analysed from stool samples on consecutive days. Consistency of faecal calprotectin levels was determined by measuring the intraclass correlation (ICC). Due to higher variability at higher faecal calprotectin levels, the ICC was calculated for the log-transformed values. The reliability of detecting a 'case' of active inflammation as defined for specific concentrations of faecal calprotectin was measured by the kappa statistic. RESULTS: Ninety-eight complete sets of results were obtained. The ICC was 0.84 (95% CI: 0.79-0.89), which represents low variability across samples. The kappa statistic for the reliability of detecting a case as defined by an FC level of >50 µg/g was substantial at 0.648 (0.511-0.769). CONCLUSIONS: Day to day variability of faecal calprotectin is low in our cohort of quiescent Crohn's disease patients and the reliability of defining a 'case' is moderately good. These data provide reassurance to clinicians using a single calprotectin sample to inform therapeutic strategies in this cohort.

Congenital heart disease in Cornelia de Lange syndrome: phenotype and genotype analysis.

Congenital heart disease (CHD) has been reported to occur in 14-70% of individuals with Cornelia de Lange syndrome (CdLS, OMIM 122470) and accounts for significant morbidity and mortality when present. Charts from a cohort of 479 patients with CdLS were reviewed for cardiac evaluations, gene testing and information to determine phenotypic severity. Two hundred fifty-nine individuals had either documented structural defects or minor cardiac findings. The presence of CHD was then quantified as a function of mutation status and severity of CdLS: mild, moderate, or severe. Different types of CHD were also evaluated by mutation status to assess for any genotype-phenotype correlation. NIPBL, SMC1A, and SMC3 mutation-positive patients were equally likely to have CHD, although the number of SMC1A and SMC3 mutation-positive patients were small in comparison. Structural CHDs were more likely to be present in individuals with moderate and severe CdLS than in the mild phenotype. This study evaluates the trends of CHD seen in the CdLS population and correlates these findings with genotype.

The correlation of autoantibodies and uNK cells in women with reproductive failure.

There is conflicting evidence on the role of autoimmune disorders in reproductive failure, including recurrent miscarriage (RM) and recurrent implantation failure (RIF), after in vitro fertilisation (IVF). Several commonly studied autoimmune markers in women with reproductive failure include antiphospholipid antibodies (APAs), thyroid peroxidase antibodies (TPA) and uterine natural killer (uNK) cells. However, there have not been any studies that have examined the correlation of these markers in women with reproductive failure. To determine if women who tested positive for autoantibodies (APA and thyroid peroxidase antibodies) have significantly higher uNK cell numbers than women who tested negative for these antibodies, the percentage of stromal cells that stained positive for CD56 was identified by immunocytochemistry in endometrial biopsies from 42 women with unexplained RM (29 women tested negative for autoantibodies and 13 women tested positive for autoantibodies) and 40 women with unexplained RIF (30 women tested negative for autoantibodies and 10 women tested positive for autoantibodies). Biopsies were obtained on days LH+7 to LH+9. There was no significant difference in uNK cell numbers between women with unexplained RM who tested negative and those who tested positive for autoantibodies. Similarly, there was no significant difference in uNK cell numbers between women with unexplained RIF who tested negative and those who tested positive for autoantibodies. In women with reproductive failure the presence of autoantibodies does not appear to affect the numbers of uNK cells in the endometrium around the time of implantation.

Expression of leukaemia inhibitory factor and interleukin 15 in endometrium of women with recurrent implantation failure after IVF; correlation with the number of endometrial natural killer cells.

BACKGROUND: Several studies have suggested that endometrial interleukin 15 (IL-15) and the leukaemia inhibitory factor (LIF) may be important in embryo implantation. IL-15 is postulated to play a role in the control of uterine natural killer (uNK) cell proliferation and function, and uNK cells are also known to play a role in implantation. The aims of this study was to (1) compare endometrial levels of IL-15 and the LIF in women with recurrent implantation failure (RIF) after IVF with those in fertile women (controls) and (2) examine the relation of IL-15 and LIF levels to the uNK cell number. METHODS: We investigated IL-15 and LIF in precisely timed endometrial biopsies (days LH + 7-LH + 9, where the day of the LH surge is LH + 0) obtained from control women (n = 15) and women with RIF (n = 45) by immunohistochemistry. A semi-quantitative analysis was performed by the H-score analysis of staining intensity in the stroma, glandular epithelium and luminal epithelium, separately. We also correlated expression of LIF and IL15 with uNK cell numbers (obtained in an earlier study of the same samples). RESULTS: The quantity of the LIF protein in endometrial glandular epithelium in women with RIF [median and range; 179 (70-365)] was lower (P = 0.01) than in control women [median and range; 247 (120-287)]. In contrast, the level of the IL-15 protein in the stroma in women with RIF [median and range; 90 (0-175)] was higher (P = 0.009) than in control women [median and range; 60 (15-150)]. There was a significant correlation between the uNK cell number and stromal expression of IL-15 (r = 0.427, P = 0.001). No correlation between the LIF expression in any compartment and the uNK cell number was seen. CONCLUSIONS: The results show an altered expression of LIF and IL-15 in the endometrium of women with RIF. Despite the limitation of not identifying uNK cells by phenotypic markers, the correlation between the uNK cell number and the stromal cell IL-15 suggests that IL-15 may play a role in the control of endometrial uNK cell function or proliferation.

Measurements of CD56+ cells in peripheral blood and endometrium by flow cytometry and immunohistochemical staining in situ.

BACKGROUND: CD56+ cells in peripheral blood or the endometrium may be increased in women with reproductive failure. However, the relationship between numbers of peripheral blood CD56+ and endometrial CD56+ cells is uncertain. The aim of this study was (i) to compare the numbers of CD56+ cells in peripheral blood and endometrium in samples taken simultaneously and (ii) to compare measurements by flow cytometry and immunohistochemistry of CD56+ cells in the same endometrial biopsy. METHOD: Endometrial biopsies and blood were obtained from women with recurrent miscarriage (n= 25) on days LH+7-LH+9 of the cycle. The total number of CD56+, CD56+ CD16- and CD56+CD16+ cells in blood was measured by flow cytometry; the number of CD56+ cells in the endometrium was determined by immunohistochemistry. Endometrial samples were also obtained from fertile women (n= 20) and used to measure CD56+ and CD45+ cells, by both flow cytometry and immunostaining. RESULTS: There was no correlation between the numbers of total CD56+, CD56+CD16- or CD56+CD16+ in peripheral blood and the number of endometrial CD56+ cells in the same women. In endometrium from fertile women, a significant correlation was found between the numbers of CD56+ cells measured by flow cytometry and immunohistochemistry (correlation= 0.497, P= 0.026, when expressed as % total cells; correlation= 0.570, P= 0.009 when expressed as % CD45+ cells). CONCLUSIONS: Measurements of CD56+ cells in peripheral blood do not correlate with endometrial CD56+ cell numbers and therefore should not be extrapolated to events in the endometrium. In contrast, measurements of endometrial CD56+ cells by flow cytometry and immunostaining correlate well.

Expression of CXCL12 and CXCR4 in human endometrium; effects of CXCL12 on MMP production by human endometrial cells.

BACKGROUND: Although several studies have suggested that CXCL12 and its receptor, CXCR4, may play a role in embryo implantation, there are limited reports of expression of CXCR4 and CXCL12 in human endometrium. The aim of this study was to investigate CXCL12 and CXCR4 expression in human endometrium and to see if CXCL12 could affect matrix metalloproteinase (MMP) production by endometrial stromal and epithelial cells. METHODS: Quantitative real-time RT-PCR (qRT-PCR) was used to detect the expression of CXCL12 and CXCR4 mRNA in endometrial biopsy samples obtained from fertile women (n = 30). Immunohistochemical analysis was carried out to determine where in the endometrium CXCL12 and CXCR4 were expressed. Primary cell culture followed by qRT-PCR and zymography was used to investigate whether CXCL12 affected MMP-2 and -9 production by endometrial stromal and epithelial cells. RESULTS: Both CXCL12 and CXCR4 were detected in the endometrium. There was no difference in CXCL12 expression at different times in the cycle, but expression of CXCR4 mRNA was significantly higher in the early proliferative (P < 0.01) compared with late proliferative and secretory phases of the cycle. CXCL12 expression was strongest in the epithelial compartment, and weaker in blood vessel walls. CXCR4 immunostaining was strong in the epithelium and blood vessel walls and weaker in the stroma. CXCL12 (10 and 100 ng/ml) had no effect on mRNA expression or activity of MMP-2 or MMP-9 in either stromal or epithelial cells. CONCLUSIONS: The results show that the expression of CXCL12 in human endometrium does not alter during the menstrual cycle, while the endometrial expression of its receptor, CXCR4, is highest in the early proliferative phase. In contrast to its effects in other cells, CXCL12 had no effect on MMP-2 or MMP-9 production by endometrial stromal or epithelial cells.

Characterization of invasive trophoblasts generated from human embryonic stem cells.

BACKGROUND: Abnormal human embryo implantation leads to poor foetal development and miscarriage, or pre-eclampsia. Ethical and practical considerations concerning implantation limit its investigation, and it is often difficult to extrapolate findings in laboratory animals when implantation processes show diverse species differences. Therefore, it is important to develop new in vitro models to study the earliest events of human implantation. The aim of this study was to derive trophoblast cell lines from human embryonic stem cells (hESCs) by a robust protocol and co-culture of these cells with an established endometrial cell culture system to validate a model of trophoblast invasion at implantation. METHODS: Derivation of trophoblast cell lines from hESC lines was established by spontaneous and induced differentiation of embryoid bodies and by initial measurement of hCGß secretion by enzyme-linked immunosorbent assay and their phenotype investigated using gene- and protein-expression markers. Vesicles formed from an aggregating trophoblast were co-cultured with decidualized human endometrial stromal cells in hypoxic (2% oxygen) and normoxic (20% oxygen) environments. RESULTS: Derived villous cytotrophoblast cell (CTB) lines further differentiated to invasive, extra-villous CTBs. Eventually, cells lost their proliferative capacity, with some lines acquiring karyotypic changes, such as a gain in the X chromosome. Cell-invasion assays confirmed that the extra-villous CTBs were invasive and erosion of the endometrial stromal layer occurred, particularly under hypoxic conditions in vitro. CONCLUSIONS: Trophoblast cell lines derived from hESCs differentiate and adapt in vitro and can be used as a model to study the mechanisms of early trophoblast invasion.

A preliminary study comparing the endometrial expression of inhibin, activin and follistatin in women with a history of implantation failure after IVF treatment and a control group.

The aim of this study was to evaluate the expression of activin: beta A and beta B subunit and follistatin in endometrium of women with implantation failure (n = 10) and compare it with a fertile control group (n = 7). Immunohistochemical staining intensity for follistatin in the endometrial glandular epithelium from women with implantation failure were significantly lower than that in control women (P = 0.03). The decreased expression of follistatin in epithelial cells in the endometrium of women with implantation failure after in vitro fertilisation (IVF) may suggest that follistatin may play a role in the implantation process.

Does free androgen index predict subsequent pregnancy outcome in women with recurrent miscarriage?

BACKGROUND: Several studies have investigated plasma androgen levels in women with recurrent miscarriage (RM) with conflicting results on whether an association between hyperandrogenaemia and RM exists. However, none of these studies included sensitive androgen measurements using a large data set. We therefore investigated the free androgen index (FAI) in a large number of women with RM in order to ascertain whether hyperandrogenaemia is a predictor of subsequent pregnancy outcome. METHODS: We studied 571 women who attended the Recurrent Miscarriage Clinic in Sheffield and presented with > or =3 consecutive miscarriages. Serum levels of total testosterone and sex hormone-binding globulin were measured in the early follicular phase and FAI was then deduced. RESULTS: The prevalence of hyperandrogenaemia in RM was 11% and in a subsequent pregnancy, the miscarriage rate was significantly higher in the raised FAI group (miscarriage rates of 68% and 40% for FAI > 5 and FAI < or = 5 respectively, P = 0.002). CONCLUSIONS: An elevated FAI appears to be a prognostic factor for a subsequent miscarriage in women with RM and is a more significant predictor of subsequent miscarriage than an advanced maternal age (> or =40 years) or a high number (> or =6) of previous miscarriages in this study.

Prognostic value of the measurement of uterine natural killer cells in the endometrium of women with recurrent miscarriage.

BACKGROUND: Studies in mice suggest that CD56 + uterine natural killer (uNK) cells play an important role in implantation. Studies in humans have described an increase in the number of uNK cells in the non-pregnant mid-secretory endometrium of women with unexplained recurrent miscarriage (RM). However, the predictive value of uNK cell number in the maintenance of pregnancy is controversial. METHODS: A blind retrospective study was undertaken. The percentage of stromal cells positive for CD56 was identified by immunocytochemistry in endometrial biopsies from 10 normal control women and 87 women with unexplained RM, of whom 51 became pregnant following biopsy. Biopsies were obtained on days LH + 7 to LH + 9. RESULTS: As in previous studies, the number of uNK cells in the 87 women with RM (mean 11.2% range 1.1-41.4%) was significantly higher (P = 0.013) than in the control women (mean 6.2% range 2.2-13.9%). No significance difference in uNK numbers was observed between 19 women who miscarried (mean 9.6% range 1.7-25.0%) and 32 women who had a live birth (mean 13.3% range 1.1-41.4%) in a subsequent pregnancy. CONCLUSIONS: In this study numbers of uNK cells in the peri-implantation endometrium of women with unexplained recurrent miscarriage did not predict subsequent pregnancy outcome.

Impact of high body mass index on endometrial morphology and function in the peri-implantation period in women with recurrent miscarriage.

There is evidence that women with a high body mass index may have a higher risk of miscarriage. It is not known if this is due to an endometrial or embryo defect. The aim of this retrospective study was to examine markers of endometrial function in overweight and obese women with recurrent unexplained miscarriage. A total of 136 women were included in the study and classified according to their body mass index (BMI) into two groups, normal BMI (< 25 kg/m(2), n = 70) and high BMI (> or = 25 kg/m(2), n = 66). Endometrial morphology was examined in all patients. A subgroup of 28 patients was examined for endometrial oestrogen and progesterone receptors in different components of the endometrium, and in a further subgroup of 28 patients, endometrial glandular leukaemia inhibitory factor and leukocyte populations were examined. A modest increase in the BMI (30.4 +/- 0.71 kg/m(2)) does not have a significant impact on endometrial steroid receptors, leukocyte populations or endometrial morphology. However, there was a significant negative correlation between endometrial glandular leukaemia inhibitory factor concentrations and the BMI (r = -0.4, P = 0.02), warranting further investigation in prospective studies that include patients with higher BMI levels.

Mutations in cohesin complex members SMC3 and SMC1A cause a mild variant of cornelia de Lange syndrome with predominant mental retardation.

Mutations in the cohesin regulators NIPBL and ESCO2 are causative of the Cornelia de Lange syndrome (CdLS) and Roberts or SC phocomelia syndrome, respectively. Recently, mutations in the cohesin complex structural component SMC1A have been identified in two probands with features of CdLS. Here, we report the identification of a mutation in the gene encoding the complementary subunit of the cohesin heterodimer, SMC3, and 14 additional SMC1A mutations. All mutations are predicted to retain an open reading frame, and no truncating mutations were identified. Structural analysis of the mutant SMC3 and SMC1A proteins indicate that all are likely to produce functional cohesin complexes, but we posit that they may alter their chromosome binding dynamics. Our data indicate that SMC3 and SMC1A mutations (1) contribute to approximately 5% of cases of CdLS, (2) result in a consistently mild phenotype with absence of major structural anomalies typically associated with CdLS, and (3) in some instances, result in a phenotype that approaches that of apparently nonsyndromic mental retardation.

Effect of laparoscopic ovarian diathermy on circulating inhibin B in women with anovulatory polycystic ovary syndrome.

BACKGROUND: Laparoscopic ovarian diathermy (LOD) frequently induces ovulation in patients with polycystic ovary syndrome (PCOS). The mechanism by which this effect occurs remains largely unexplained. The aim of this study was to measure changes in inhibin B production in response to LOD to see whether this could explain the mechanism of action of LOD. METHODS: This prospective study included 50 anovulatory women with PCOS. All women underwent LOD. Blood samples were collected before and after LOD to measure plasma concentrations of inhibin B, gonadotrophins and androgens. RESULTS: The pre-operative median plasma concentration of inhibin B was 110.0 pg/ml (range 19.0-567.0 pg/ml). There was a statistically significant inverse correlation (r= -0.286; P < 0.05) between body mass index (BMI) and inhibin B. Non-obese women with PCOS (BMI 30 kg/m2; n=13). Following LOD, 39 women ovulated. No statistically significant change of inhibin B after LOD was observed in the overall group of women with PCOS or in the subgroup of non-obese PCOS women with higher pre-operative inhibin B. CONCLUSIONS: The lack of any change of inhibin B after LOD makes it unlikely that this hormone has any role to play in the mechanism of action of LOD.

Cytokine expression in the endometrium of women with implantation failure and recurrent miscarriage.

One potential cause of reproductive failure such as infertility and recurrent miscarriage may be an endometrial defect. Numerous studies in mice have suggested the importance of various different cytokines in successful pregnancy outcome. This article reviews the literature available on the role of T helper cytokines and IL-1, IL-11, LIF, IL-12 and IL-18 in infertility and recurrent miscarriage, with particular emphasis on the role that endometrial cytokines may play. Although there are numerous studies on cytokines in recurrent miscarriage, much less has been reported on their role in infertility with or without failure after IVF. There is also considerable variation in the results obtained from various different studies, which may be due to different populations studied, the different timing of the sample collection, and whether the cytokines were measured in whole tissue or a specific cell population. The presence of complicated networks of cytokines and their overlapping biological activities means that alteration of one cytokine is likely to affect others and this also makes the study of their role in implantation failure very difficult. There is an urgent need to re-examine the role played by various cytokines in reproductive failure through carefully planned and vigorously designed studies and to compare the different types of reproductive failure.

Cytotrophoblast stem cell lines derived from human embryonic stem cells and their capacity to mimic invasive implantation events.

BACKGROUND: An effective embryonic-maternal interaction is crucial for successful human pregnancy. Failure of this process is a major cause of infertility and can lead to placental dysfunction resulting in recurrent miscarriage, fetal retardation and pre-eclampsia. Research is severely constrained by ethical and practical considerations; therefore, we aimed to generate cytotrophoblast stem (CTBS) cell lines from human embryonic stem cells (HESCs). METHOD: Beta-HCG was used as a marker of viable trophoblast cells. In defined culture, embryoid bodies were generated from HESCs and selected for trophoblast enrichment by rounds of cellular aggregation and disaggregation. Distinct CTBS cell lines were isolated and characterized. Spheroid cytotrophoblast bodies were generated and their interaction with luteal-phase endometrial stroma was analysed by real-time image analysis. RESULTS: Three CTBS cell lines were derived, which were maintained in the absence of residual HESCs, fibroblast feeder cells or extracellular matrix. CTBS cells displayed typical cytotrophoblast and syncytiotrophoblast characteristics and exhibited further differentiation to invasive endovascular cell phenotype. One cell line was generated with constitutive expression of enhanced green fluorescent protein (eGFP). Spheroid trophoblast bodies mimicked closely the early invasive stages of implantation when incubated with human endometrial stromal preparations in vitro. CONCLUSION: These human CTBS cell lines are a significant new model for investigating human placentation and may have considerable potential in cell therapy applications.

Expression of interleukin-11 receptor alpha and interleukin-11 protein in the endometrium of normal fertile women and women with recurrent miscarriage.

Interleukin-11 (IL-11) is a member of the IL-6 family of cytokines. Previous studies have suggested that IL-11 may play a role in human endometrial function. In this study, we have used immunocytochemistry to compare endometrial IL-11Ralpha and IL-11 expression in precisely timed peri-implantation biopsies from 9 normal fertile women and 16 recurrent miscarriage (RM) women. Immunocytochemistry was semi-quantified by obtaining an H-score value, which showed increased expression of both IL-11 and IL-11Ralpha in epithelial cells compared to stromal cells in all biopsies. There was a significant (P<0.01) reduction in epithelial cell IL-11, but not stromal cell IL-11, expression in endometrium from RM women compared to normal fertile women. There were no significant differences in expression of IL-11Ralpha protein in both stromal and epithelial cells in endometrium from the two groups of women. This work shows the presence of IL-11 and IL-11Ralpha within the endometrium of RM women during the peri-implantation period. The decreased expression of IL-11 in epithelial endometrium in RM women suggests that this cytokine may play a role in preventing miscarriage.