Placental Complement Activation in Fetal and Neonatal Alloimmune Thrombocytopenia: An Observational Study

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a disease that causes thrombocytopenia and a risk of bleeding in the (unborn) child that result from maternal alloantibodies directed against fetal, paternally inherited, human platelet antigens (HPA). It is hypothesized that these alloantibodies can also bind to the placenta, causing placental damage. This study aims to explore signs of antibody-mediated placental damage in FNAIT. We performed a retrospective study that included pregnant women, their newborns, and placentas. It comprised 23 FNAIT cases, of which nine were newly diagnosed (14 samples) and 14 were antenatally treated with intravenous immune globulins (IVIg) (21 samples), and 20 controls, of which 10 had anti-HLA-class I antibodies. Clinical information was collected from medical records. Placental samples were stained for complement activation markers (C1q, C4d, SC5b-9, and mannose-binding lectin) using immunohistochemistry. Histopathology was examined according to the Amsterdam criteria. A higher degree of C4d deposition was present in the newly diagnosed FNAIT cases (10/14 samples), as compared to the IVIg-treated FNAIT cases (2/21 samples, p = 0.002) and anti-HLA-negative controls (3/20 samples, p = 0.006). A histopathological examination showed delayed maturation in four (44%) placentas in the newly diagnosed FNAIT cases, five (36%) in the IVIg-treated FNAIT cases, and one in the controls (NS). C4d deposition at the syncytiotrophoblast was present in combination with low-grade villitis of unknown etiology in three newly diagnosed FNAIT cases that were born SGA. We conclude that a higher degree of classical pathway-induced complement activation is present in placentas from pregnancies with untreated FNAIT. This may affect placental function and fetal growth

Prevalence and predictors of depression and well-being after hysterectomy: an observational study

OBJECTIVES: To assess risk and predictive factors for depression and well-being, 3 and 12 months after elective hysterectomy. Secondary objectives were to assess the incidence of depression, level of well-being, and feelings of femininity. STUDY DESIGN: A prospective multicenter cohort study was performed among 419 women, undergoing hysterectomy for benign indication. Data were collected in the week prior to surgery, and in the per- and postoperative period up to the fourth postoperative day and 3 and 12 months after surgery. Sociodemographic variables, baseline health status, psychosocial predictors, and surgery data were assessed. Outcome measures were Center for Epidemiological Studies-Depression scale (CES-D, range 0-60), the 12-item well-being questionnaire energy and positive well-being subscales (range 0-12), and feelings of femininity. Predictor analyses were performed using linear mixed model analyses. RESULTS: Levels of depression, energy, and positive well-being after hysterectomy were predicted by their corresponding baseline levels (estimate 0.62 p<0.001, 0.39 p<0.001, 0.37 p<0.001, respectively) and baseline pain (0.31 p=0.003, -0.09 p=0.026, -0.10 p=0.008). Postoperative infection reported at 12 months affected CES-D and energy level. Several other gynaecological, psychosocial, or perioperative factors were also predictive for one of the outcomes. Prevalence of depression at baseline, 3 and 12 months was 24%, 19%, and 21%, respectively. In general, well-being scores were slightly higher 3 and 12 months after hysterectomy than at baseline. Feelings of femininity were not negatively affected in 92% of the patients. CONCLUSIONS: Preoperative psychosocial status, perioperative pain, and postoperative infection were found as predictors of psychological outcome after hysterectomy. In the majority of patients we observed small but significant improvements with regard to postoperative depression and well-being, while feelings of femininity were unaffected

HIP (HPA-screening in pregnancy) study: protocol of a nationwide, prospective and observational study to assess incidence and natural history of fetal/neonatal alloimmune thrombocytopenia and identifying pregnancies at risk

INTRODUCTION: Fetal and neonatal alloimmune thrombocytopenia (FNAIT) may lead to severe fetal or neonatal bleeding and/or perinatal death. Maternal alloantibodies, targeted against fetal human platelet antigens (HPAs), can result thrombocytopenia and bleeding complications. In pregnancies with known immunisation, fetal bleeding can be prevented by weekly maternal intravenous immunoglobulin infusions. Without population-based screening, immunisation is only detected after birth of an affected infant. Affected cases that might have been prevented, when timely identified through population-based screening. Implementation is hampered by the lack of knowledge on incidence, natural history and identification of pregnancies at high risk of bleeding. We designed a study aimed to obtain this missing knowledge. METHODS AND ANALYSIS: The HIP (HPA-screening in pregnancy) study is a nationwide, prospective and observational cohort study aimed to assess incidence and natural history of FNAIT as well as identifying pregnancies at high risk for developing bleeding complications. For logistic reasons, we invite rhesus D-negative or rhesus c-negative pregnant women, who take part in the Dutch population-based prenatal screening programme for erythrocyte immunisation, to participate in our study. Serological HPA-1a typing is performed and a luminex-based multiplex assay will be performed for the detection of anti-HPA-1a antibodies. Results will not be communicated to patients or caregivers. Clinical data of HPA-1a negative women and an HPA-1a positive control group will be collected after birth. Samples of HPA-1a immunised pregnancies with and without signs of bleeding will be compared with identify parameters for identification of pregnancies at high risk for bleeding complications. ETHICS AND DISSEMINATION: Ethical approval for this study has been obtained from the Medical Ethical Committee Leiden-The Hague-Delft (P16.002). Study enrolment began in March 2017. All pregnant women have to give informed consent for testing according to the protocol. Results of the study will be disseminated through congresses and publication in relevant peer-reviewed journals. TRIAL REGISTRATION NUMBER: NCT04067375

RHC and RHc genotyping in different ethnic groups

BACKGROUND: RH genotyping assays are mainly based on research in whites. These assays may not be reliable in a multiracial society because of the genetic variation in RH among ethnic groups. STUDY DESIGN AND METHODS: Five groups from different ethnic backgrounds were serologically typed for C and c and were genotyped on nucleotide C48 and intron 2 for RHC and RHc on nucleotides C178 and C307. RESULTS: RHc genotyping with both methods proved to be reliable. RHC genotyping on C48 is not reliable because of a 48G>C mutation in the RHce allele (false-positive prediction of C). This mutation was found in every ethnic group and does not affect c or e expression. RHC genotyping on intron 2 is unreliable because of r's (Cdes) alleles (a false-negative prediction of C). This allele was found in whites and blacks from Curaçao and South Africa. Reactions of r's cells with anti-C are weaker, but no negative reactions with various MoAbs were found. A new method (RHC/c/hex3-intron 4/exon 7 multiplex PCRs) was developed based on intron 2 and r's hybrid exon 3 characteristics (RHC) and C307 (RHc). CONCLUSIONS: Reliable RHC and RHc genotyping is possible in different ethnic groups with the RHC/c/hex3-intron 4/exon 7 multiplex PCR approac

RHD(T201R, F223V) cluster analysis in five different ethnic groups and serologic characterization of a new Ethiopian variant DARE, the DIII type 6, and the RHD(F223V)

BACKGROUND: The RHD phylogeny in humans shows four main clusters of which three are predominantly observed in (African) black persons. Each of the African clusters is characterized by specific amino acid substitutions relative to the Eurasian RHD allele. RH phylogeny defines the framework for identification of clinically relevant aberrant alleles. This study focuses on the weak D type 4 cluster (characterized by RHD(T201R, F223V) (602C>G 667T>G)) in five ethnic groups. STUDY DESIGN AND METHODS: A total of 1702 samples were screened for the presence of 602C>G and 667T>G by sequence-specific polymerase chain reaction (PCR-SSP). Eighty samples were assigned to the weak D type 4 cluster and were molecularly characterized by PCR-SSP and RHD sequencing. Antigens of aberrant alleles were characterized with monoclonal anti-D according to the 37-epitope model when possible. RESULTS: Five new aberrant alleles, DIII type 6, DIII type 7, DARE, RHD(T201R, F223V) (without 819G>A), and RHD(F223V), were identified and DIII type 6, DARE, and RHD(F223V) were serologically characterized with monoclonal anti-D. Both the DARE and RHD(F223V) showed epitope loss. It is postulated that the 1136C>T nucleotide substitution (characteristic for the DAU allele cluster) is present on the DVa(KOU) allele. CONCLUSION: Identification of the new variant alleles refines the phylogeny of RHD in humans. The proposed DVa(KOU) allele with 1136C>T (DVa(KOU)T379M) is probably caused by conversion of the DAU0 allele and the DVa(KOU) allele, forming a phylogenetic link between the DV allele and the DAU cluster. By describing the RHD(F223V) (602C>G) and RHD(T201R, F223V) (602C>G and 667T>G) alleles formal proof is given for the origin of the non-Eurasian cluste

Clinical characteristics of human platelet antigen (HPA)-1a and HPA-5b alloimmunised pregnancies and the association between platelet HPA-5b antibodies and symptomatic fetal neonatal alloimmune thrombocytopenia

Fetal neonatal alloimmune thrombocytopenia (FNAIT) is caused by maternal alloantibodies directed against the human platelet antigens (mostly HPA-1a or HPA-5b) of the (unborn) child and can lead to severe bleeding. Anti-HPA-1a-mediated FNAIT shows a severe clinical outcome more often than anti-HPA-5b-mediated FNAIT. Given the relatively high prevalence of anti-HPA-5b in pregnant women, the detection of anti-HPA-5b in FNAIT-suspected cases may in some cases be an incidental finding. Therefore we investigated the frequency of anti-HPA-5b-associated severe bleeding in FNAIT. We performed a retrospective nationwide cohort study in cases with clinical suspicion of FNAIT. HPA antibody screening was performed using monoclonal antibody-specific immobilisation of platelet antigens. Parents and neonates were typed for the cognate antigen. Clinical data were collected by a structured questionnaire. In 1 864 suspected FNAIT cases, 161 cases (8·6%) had anti-HPA-1a and 60 (3·2%) had anti-HPA-5b. The proportion of cases with severe bleeding did not differ between the cases with anti-HPA-1a (14/129; 11%) and anti-HPA-5b (4/40; 10%). In multigravida pregnant women with a FNAIT-suspected child, 100% (81/81) of anti-HPA-1a cases and 79% (38/48) of anti-HPA-5b cases were HPA-incompatible, whereas 86% and 52% respectively were expected, based on the HPA allele distribution. We conclude that anti-HPA-5b can be associated with severe neonatal bleeding symptoms. A prospective study is needed for true assessment of the natural history of anti-HPA-5b mediated FNAIT

Long-term neurodevelopmental outcome in children after antenatal intravenous immune globulin treatment in fetal and neonatal alloimmune thrombocytopenia

Background: Children with fetal and neonatal alloimmune thrombocytopenia face increased risk of intracranial hemorrhage potentially leading to developmental impairment. To prevent intracranial hemorrhage, pregnant women with alloantibodies against fetal platelets are often treated with intravenous immunoglobulin. Intravenous immunoglobulin seems effective in vastly reducing the risk of fetal or neonatal bleeding complications. However, information on long-term neurodevelopment of these children is lacking. Objective: This study aimed to evaluate long-term neurodevelopmental outcome in children with fetal and neonatal alloimmune thrombocytopenia who were treated with intravenous immunoglobulin antenatally. Study Design: An observational cohort study was performed, including children of mothers treated with intravenous immunoglobulin during pregnancy because a previous child was diagnosed with fetal and neonatal alloimmune thrombocytopenia. Children were invited for a follow-up assessment including standardized cognitive and neurologic tests. The parents were asked to complete a behavioral questionnaire and school performance reports. The primary outcome was severe neurodevelopmental impairment, defined as severe cognitive impairment (intelligence quotient <70), cerebral palsy with Gross Motor Function Classification System Level ≥3, bilateral blindness, and/or bilateral deafness (requiring amplification). The secondary outcome was mild to moderate neurodevelopmental impairment, defined as either mild to moderate cognitive impairment (intelligence quotient <85), cerebral palsy with Gross Motor Function Classification System Level ≤2, minor neurologic dysfunction, vision loss, and/or hearing loss. Results: Between 2003 and 2017, 51 children were live-born after antenatal intravenous immunoglobulin treatment. One family moved abroad and was therefore not eligible for inclusion. In total, 82% (41/50) of the eligible cases were included for neurodevelopmental assessment at a median age of 9 years and 8 months. Severe neurodevelopmental impairment was not detected. The incidence of mild to moderate neurodevelopmental impairment was 14% (6/41; 95% confidence interval, 6%–29%). The children's mean cognitive score, behavioral scores, and academic achievement were not different from those observed in the Dutch norm groups. Neuroimaging was performed in 90% (37/41) of cases. Severe intracranial hemorrhage was diagnosed in 2 cases (5%), one antenatally before the start of intravenous immunoglobulin and the other case 1 day after birth. Both cases had a normal neurodevelopmental outcome. Conclusion: The risk of neurodevelopmental impairment in children whose mothers were treated for fetal and neonatal alloimmune thrombocytopenia with antenatal intravenous immunoglobulin is comparable to that reported in the general population

A multicentre double-blinded randomized controlled trial on the efficacy of laser-assisted hatching in patients with repeated implantation failure undergoing IVF or ICSI

STUDY QUESTION: Does assisted hatching increase the cumulative live birth rate in subfertile couples with repeated implantation failure? SUMMARY ANSWER: This study showed no evidence of effect for assisted hatching as an add-on in subfertile couples with repeated implantation failure. WHAT IS KNOWN ALREADY: The efficacy of assisted hatching, with regard to the live birth rate has not been convincingly demonstrated in randomized trials nor meta-analyses. It is suggested though that especially poor prognosis women, e.g. women with repeated implantation failure, might benefit most from assisted hatching. STUDY DESIGN, SIZE, DURATION: The study was designed as a double-blinded, multicentre randomized controlled superiority trial. In order to demonstrate a statistically significant absolute increase in live birth rate of 10% after assisted hatching, 294 participants needed to be included per treatment arm, being a total of 588 subfertile couples. Participants were included and randomized from November 2012 until November 2017, 297 were allocated to the assisted hatching arm of the study and 295 to the control arm. Block randomization in blocks of 20 participants was applied and randomization was concealed from participants, treating physicians, and laboratory staff involved in the embryo transfer procedure. Ovarian hyperstimulation, oocyte retrieval, laboratory procedures, embryo selection for transfer and cryopreservation, the transfer itself, and luteal support were performed according to local protocols and were identical in both the intervention and control arm of the study with the exception of the assisted hatching procedure which was only performed in the intervention group. The laboratory staff performing the assisted hatching procedure was not involved in the embryo transfer itself. PARTICIPANTS/MATERIALS, SETTING, METHODS: Participants were eligible for inclusion in the study after having had either at least two consecutive fresh IVF or ICSI embryo transfers, including the transfer of frozen and thawed embryos originating from those fresh cycles, and which did not result in a pregnancy or as having had at least one fresh IVF or ICSI transfer and at least two frozen embryo transfers with embryos originating from that fresh cycle which did not result in a pregnancy. The study was performed at the laboratory sites of three tertiary referral hospitals and two university medical centres in the Netherlands. MAIN RESULTS AND THE ROLE OF CHANCE: The cumulative live birth rate per started cycle, including the transfer of fresh and subsequent frozen/thawed embryos if applicable, resulted in 77 live births in the assisted hatching group (n?=?297, 25.9%) and 68 live births in the control group (n?=?295, 23.1%). This proved to be statistically not significantly different (relative risk: 1.125, 95% CI: 0.847 to 1.494, P?=?0.416). LIMITATIONS, REASONS FOR CAUTION: There was a small cohort of subfertile couples that after not achieving an ongoing pregnancy, still had cryopreserved embryos in storage at the endpoint of the trial, i.e. 1 year after the last randomization. It cannot be excluded that the future transfer of these frozen/thawed embryos increases the cumulative live birth rate in either or both study arms. Next, at the start of this study, there was no international consensus on the definition of repeated implantation failure. Therefore, it cannot be excluded that assisted hatching might be effective in higher order repeated implantation failures. WIDER IMPLICATIONS OF THE FINDINGS: This study demonstrated no evidence of a statistically significant effect for assisted hatching by increasing live birth rates in subfertile couples with repeated implantation failure, i.e. the couples which, based on meta-analyses, are suggested to benefit most from assisted hatching. It is therefore suggested that assisted hatching should only be offered if information on the absence of evidence of effect is provided, at no extra costs and preferably only in the setting of a clinical trial taking cost-effectiveness into account. STUDY FUNDING/COMPETING INTEREST(S): None. TRIAL REGISTRATION NUMBER: Netherlands Trial Register (NTR 3387, NL 3235, https://www.clinicaltrialregister.nl/nl/trial/26138). TRIAL REGISTRATION DATE: 6 April 2012. DATE OF FIRST PATIENT’S ENROLMENT: 28 November 2012