A feasibility trial of screening women with idiopathic recurrent miscarriage for high uterine natural killer cell density and randomizing to prednisolone or placebo when pregnant.

STUDY QUESTION: Is it feasible to screen women with idiopathic recurrent miscarriage (RM) for high uterine natural killer (uNK) cell density and then randomize them to prednisolone or placebo when pregnant? SUMMARY ANSWER: It was feasible to recruit women with idiopathic RM into a 'screen and treat' trial despite their desire for active medication. WHAT IS KNOWN ALREADY: Clinical trials of immunotherapy in women with idiopathic RM have failed to substantiate efficacy in preventing miscarriage. Preimplantation uNK cell density is higher in women with RM and can be reduced with prednisolone. STUDY DESIGN, SIZE, DURATION: In a pilot RCT, 160 eligible women were screened with an endometrial biopsy and those with high uNK cell density were invited to return when pregnant for randomization to prednisolone (20 mg for 6 weeks, 10 mg for 1 week, 5 mg for 1 week) or identical placebo tablets. Randomization was by random number generation and patients, clinicians and outcome assessors were blinded to allocation. The study size and duration was determined by funding, which was for a feasibility trial, for 2 years, sufficient to screen 150 women and randomize 40 women. The outcome measures were recruitment rate, women's perspectives, compliance, live birth and miscarriage rates and pregnancy complications. PARTICIPANTS/MATERIALS, SETTING, METHODS: The trial was advertised nationally in the UK. Women who attended research clinics run by one consultant (SQ) with three or more consecutive idiopathic miscarriages were included. Women's perspectives of the trial were sought through a questionnaire. The endometrium was sampled 5-9 days after the LH surge, stained using immunohistochemistry for CD56 and the sub-epithelial region analysed with image analysis. Women with a high uNK cell density (>5%) were invited to contact the clinic at 4-6 weeks gestation for randomization. Compliance with medication was assessed using a daily log, and side effects recorded by the women in a diary and on a structured proforma completed in the clinic at the end of the first trimester. All women had ultrasound scans every 2 weeks until 14 weeks' gestation and growth scans at 28 and 34 weeks' gestation in addition to routine antenatal care and a follow-up in person or by telephone 6 weeks after delivery. MAIN RESULTS AND THE ROLE OF CHANCE: Despite the fact that 85% of women said they would prefer the active treatment, the trial recruitment occurred at the planned rate. Eligible women (n = 160) attended the research clinics and had the uNK test, 72 were screen positive and 40 returned when pregnant for randomization. Compliance with medication was reported to be 100%. The active treatment was associated with side effects of insomnia and flushing. The live birth rate was 12/20 (60%) with prednisolone and 8/20 (40%) with placebo (Risk Ratio 1.5, 95% confidence interval (CI) 0.79-2.86, absolute difference 20% CI-10%, +50%), and hence, this was not significant. There were no pregnancy complications or serious adverse fetal outcomes. LIMITATIONS, REASONS FOR CAUTION: This was a feasibility trial so of insufficient size to assess efficacy or safety. There was inconsistency in the start date of the trial medication and this may have affected the outcome in the active treatment group. WIDER IMPLICATIONS OF THE FINDINGS: It was feasible to recruit women with idiopathic RM into a 'screen and treat' trial despite their desire for active medication. Our data also suggest that in future trials the primary outcome measure is live birth rate after 24 weeks gestation.

Prednisolone Trial: Study protocol for a randomised controlled trial of prednisolone for women with idiopathic recurrent miscarriage and raised levels of uterine natural killer (uNK) cells in the endometrium.

BACKGROUND: Idiopathic recurrent miscarriage is defined as 3 consecutive pregnancy losses with no contributing features found on investigations. At present there are no treatments of proven efficacy for idiopathic recurrent miscarriage. Uterine natural killer (uNK) cells, the most predominant leucocyte in the endometrium are adjacent to foetal trophoblast cells and thought to be involved in implantation. The exact mechanisms of how uNK cells affect implantation are not clear but are probably through the regulation of angiogenesis. Multiple studies have shown an association between high density of uterine natural killer cells and recurrent miscarriage. We have shown that prednisolone reduces the number of uNK cells in the endometrium. The question remains as to whether reducing the number of uNK cells improves pregnancy outcome. METHODS: We propose a randomised, double-blind, placebo controlled trial of prednisolone with a pilot phase to assess feasibility of recruitment, integrity of trial procedures, and to generate data to base future power calculations. The primary aim is to investigate whether prednisolone therapy during the first trimester of pregnancy is able to improve live birth rates in patients with idiopathic recurrent miscarriage and raised uNK cells in the endometrium. Secondary outcomes include conception rate, karyotype of miscarriage, miscarriages (first and second trimester), stillbirths, pregnancy complications, gestational age at delivery, congenital abnormality and side effects of steroids. The trial has 2 stages: i) screening of non-pregnant women and ii) randomisation of the pregnant cohort. All patients who fit the inclusion criteria (<40 years old, > or =3 consecutive miscarriages with no cause found and no contraindications to prednisolone therapy) will be asked to consent to an endometrial biopsy in the mid-luteal phase to assess their levels of uNK cells. Women with high levels of uNK cells (> or =5%), will be randomised to either prednisolone or placebo when a pregnancy is confirmed. Follow-up includes 2 weekly ultrasound scans in the first trimester, an anomaly scan at 20 weeks gestation, growth scans at 28 and 34 weeks gestation and a postnatal follow-up at 6 weeks. TRIAL REGISTRATION: Current Controlled Trials ISRCTN28090716.

Uterine natural killer cells and angiogenesis in recurrent reproductive failure.

BACKGROUND: Increased numbers of phenotypically unusual CD56(bright) CD16- uterine natural killer (uNK) cells have been associated with recurrent reproductive failure. uNK cells produce angiogenic growth factors and are potential regulators of decidual angiogenesis in early pregnancy. The final common mechanism for early pregnancy loss is thought to be early onset of the maternal circulation and excessive placental oxidative stress. We tested the hypothesis that increased uNK cells in preimplantation endometrium are associated with altered angiogenesis. METHODS: Women with recurrent reproductive failure (n = 122) were investigated with uterine artery Doppler and endometrial biopsy. Immunohistochemistry was used to identify uNK, endothelial and vascular smooth muscle cells and image analysis was used to assess location, density and differentiation. RESULTS: uNK cell density was positively correlated with the formation of blood (P = 0.005, r = 0.5) and lymphatic vessels (P = 0.0001, r = 0.6), spiral arteriole smooth muscle differentiation (P = 0.01, r = 0.5) and endometrial oedema (P = 0.004). The functional effect of this was a reduced uterine artery resistance to blood flow. CONCLUSIONS: These data suggest that uNK cells may regulate angiogenesis in non-pregnant endometrium. The mechanisms of reproductive failure associated with increased uNK cell density appear to be increased angiogenesis and peri-implantation blood flow, which may lead to early maternal circulation and hence pregnancy failure due to excessive oxidative stress.

CSF cytokine levels in preterm infants may reflect systemic inflammation and are independent of gestation.

BACKGROUND: Among preterm infants, high concentrations of inflammatory mediators in cerebrospinal fluid (CSF) are associated with poor outcome. Previous studies have not indicated whether CSF concentrations of inflammatory mediators are associated with important confounders such as gestational age. AIMS: To examine associations between CSF concentrations of inflammatory mediators and gestational age, maternal features suggestive of inflammation, characteristics of the CSF sample or the presence of a systemic inflammatory response. STUDY DESIGN AND SUBJECTS: Aliquots of CSF obtained during routine investigation of potential sepsis among infants born before 35 weeks gestation were assayed for 17 mediators of inflammation using a fluorescent multi-bead analyser. Other information was collected from routine clinical records. RESULTS: 39 infants were assessed. CSF levels of mediators of inflammation were not correlated with gestational age. CSF red blood cell counts were correlated with CSF concentrations of IL-6, GM-CSF and IL-17 (each p<0.003). CSF lactate was correlated with CSF concentrations of IL-1beta, IL-6, GM-CSF, G-CSF, IFN-gamma and MIP-1beta. CSF concentrations of IL-1beta, IL-6, G-CSF, TNF-alpha and IFN-gamma were higher in infants with a raised CRP within 24 h of delivery (each p<0.003). CONCLUSIONS: CSF concentrations of inflammatory mediators most probably reflect inflammatory pathologies and are not influenced by gestational age. They may also, however, reflect contamination with blood or systemic inflammation. CSF concentrations of inflammatory mediators may not provide a specific indicator of CNS inflammation.

Genetic polymorphisms and retinopathy of prematurity.

PURPOSE: Retinopathy of prematurity (ROP) is a major problem among very preterm survivors of neonatal intensive care. Neovascularization of the retina is prominent in the proliferative stages of ROP and is under the control of several factors such as vascular endothelial growth factor (VEGF). This study was undertaken on the hypothesis that genetic polymorphisms of VEGF, transforming growth factor (TGF)-beta1, and tumor necrosis factor (TNF)-alpha would occur more frequently in preterm infants with progressive ROP than in those with mild or no disease. METHODS: The frequencies of VEGF -634 G-->C, VEGF *936 C-->T, TNF-alpha -308 G-->A, and TGF-beta -509 C-->T were determined in DNA from 91 infants who had received treatment for threshold ROP and 97 comparison infants. RESULTS: The frequencies of the VEGF *936 C-->T, TNF-alpha -308 G-->A and TGF-beta -509 C-->T polymorphisms were similar in both groups. The distribution of alleles at VEGF -634 was significantly different between the two groups (P = 0.03). Homozygotes for the G allele, associated with higher VEGF production were twice as likely to have threshold ROP. CONCLUSIONS: The progression of ROP to threshold ROP in very preterm infants may be influenced by genetic differences in VEGF production. Future efforts at prevention of threshold ROP may be directed toward blocking excess production of VEGF.