Analysis of FCGR3A-p.176Val variants in women with recurrent pregnancy loss and the association with CD16a expression and anti-HLA antibody status.

Natural Killer (NK) cells have been implicated in recurrent pregnancy loss (RPL). The p.Val176Phe (or Val158Phe) Single Nucleotide Polymorphism (SNP) in the FCGR3A gene encoding the FcγRIIIA or CD16a receptor has been associated with an enhanced affinity for IgG and stronger NK-mediated antibody-dependent cellular cytotoxicity. We hypothesized that the presence of at least one p.176Val variant associates with RPL and increased CD16a expression and alloantibodies e.g., against paternal human leukocyte antigen (HLA). In 50 women with RPL, we studied frequencies of the p.Val176Phe FCGR3A polymorphisms. Additionally, CD16a expression and anti-HLA antibody status were analyzed by flowcytometry and Luminex Single Antigens. In woman with RPL, frequencies were: 20% (VV), 42% (VF) and 38% (FF). This was comparable to frequencies from the European population in the NCBI SNP database and in an independent Dutch cohort of healthy women. NK cells from RPL women with a VV (22,575 [18731-24607]) and VF (24,294 [20157-26637]) polymorphism showed a higher expression of the CD16a receptor than NK cells from RPL women with FF (17,367 [13257-19730]). No difference in frequencies of the FCGR3A-p.176 SNP were detected when comparing women with or without class I and class II anti-HLA antibodies. Our study does not provide strong evidence for an association between the p.Val176Phe FCGR3A SNP and RPL.

Natural killer cell profiles in recurrent pregnancy loss: Increased expression and positive associations with TACTILE and LILRB1.

PROBLEM: NK cells are important for healthy pregnancy and aberrant phenotypes or effector functions have been associated with RPL. We compared expression of a broad panel of NK cell receptors, including immune checkpoint receptors, and investigated their clinical association with RPL as this might improve patient stratification and prediction of RPL. METHOD OF STUDY: Peripheral blood mononuclear cells were isolated from 52 women with RPL and from 2 women with an uncomplicated pregnancy for flowcytometric analysis and plasma was used to determine anti-CMV IgG antibodies. RESULTS: Between RPL and controls, we observed no difference in frequencies of T-, NKT or NK cells, in CD56dimCD16+ or CD56brightCD16- NK cell subsets or in the expression of KIRs, NKG2A, NKG2C, NKG2D, NKp30, NKp44, NKp46 or DNAM1. NK cells from women with RPL had a higher expression of LILRB1 and TACTILE and this was associated with the number of losses. The immune checkpoint receptors PD1, TIM3 and LAG3 were not expressed on peripheral blood NK cells. In RPL patients, there was a large variation in NKG2C expression and higher levels could be explained by CMV seropositivity. CONCLUSION: Our study identified LILRB1 and TACTILE as NK cell receptors associated with RPL. Moreover, we provide first support for the potential role of CMV in RPL via its impact on the NK cell compartment. Thereby our study could guide future studies to confirm the clinical association of LILRB1, TACTILE and NKG2C with RPL in a larger cohort and to explore their functional relevance in reproductive success.

Intravenous immunoglobulins improve live birth rate among women with underlying immune conditions and recurrent pregnancy loss: a systematic review and meta-analysis.

Intravenous immunoglobulin (IVIG) is increasingly used as a treatment for recurrent pregnancy loss (RPL) despite lack of clear evidence on efficacy. Recent data suggest IVIG might be more effective in a subgroup of women with an aberrant immunological profile. Therefore, a systematic review and meta-analysis of studies on the effectiveness of IVIG treatment on pregnancy outcome among women with RPL and underlying immunological conditions (e.g., elevated NK cell percentage, elevated Th1/Th2 ratio, diagnosis with autoimmune disorders) was conducted. Eight non-randomized controlled trials, including 478 women (intervention: 284; control: 194), met eligibility criteria. Meta-analysis showed that treatment with IVIG was associated with a two-fold increase in live birth rate (RR 1.98, 95% CI 1.44-2.73, P < 0.0001). The effect of IVIG was particularly marked in the subgroup of studies including patients based on presence of elevated (> 12%) NK-cell percentage (RR 2.32, 95% CI 1.77-3.02, P < 0.0001) and when starting intervention prior to or during cycle of conception (RR 4.47, 95% CI 1.53-13.05, P = 0.006). In conclusion, treatment with IVIG may improve live birth rate in women with RPL and underlying immune conditions. However, these results should be interpreted with caution as studies are limited by low number of participants and the non-randomized design, which represent seriously biases. Future randomized controlled trials in women with RPL and underlying immune conditions are needed before using IVIG in a clinical setting.

Preconceptional evaluation of women with recurrent pregnancy loss: the additional value of assessing vascular and metabolic status.

BACKGROUND: A majority of recurrent pregnancy loss cases (RPL) remains unexplained. We hypothesized that complications in vascular and metabolic status may guide towards underlying problems that also predispose to RPL and that the number of pregnancy losses is related. METHODS: A retrospective study in 123 women with either a history of low-order RPL (2-3 pregnancy losses) or high-order RPL (≥ 4 pregnancy losses) and 20 women with a history of uncomplicated pregnancy (controls) was performed. Vascular status was assessed by measuring hemodynamic parameters, determining abnormal parameters and analyzing their contribution to the circulatory risk profile (CRP). In a similar way, metabolic status was assessed. Metabolic parameters were measured, used to determine abnormal parameters and analyzed for their contribution to the metabolic syndrome (MetS). RESULTS: No major differences were observed in vascular or metabolic parameters between women with RPL and controls. There was no relation with the number of pregnancy losses. However, when analyzing the presence of abnormal constituents, more than 80% of women with RPL had at least one abnormal constituent of the CRP. While only 27% had one or more abnormal constituent of the MetS. CONCLUSIONS: The presence of abnormal circulatory factors prior to pregnancy, and to lesser extent constituents of the metabolic syndrome, may predispose to RPL and offer new insights to its pathophysiology.

Annexin A1 as Neuroprotective Determinant for Blood-Brain Barrier Integrity in Neonatal Hypoxic-Ischemic Encephalopathy.

Blood-brain barrier (BBB) disruption is associated with hypoxia-ischemia (HI) induced brain injury and life-long neurological pathologies. Treatment options are limited. Recently, we found that mesenchymal stem/stromal cell derived extracellular vesicles (MSC-EVs) protected the brain in ovine fetuses exposed to HI. We hypothesized that Annexin A1 (ANXA1), present in MSC-EVs, contributed to their therapeutic potential by targeting the ANXA1/Formyl peptide receptor (FPR), thereby preventing loss of the BBB integrity. Cerebral ANXA1 expression and leakage of albumin into the fetal ovine brain parenchyma after HI were analyzed by immunohistochemistry. For mechanistic insights, barrier integrity of primary fetal endothelial cells was assessed after oxygen-glucose deprivation (OGD) followed by treatment with MSC-EVs or human recombinant ANXA1 in the presence or absence of FPR inhibitors. Our study revealed that BBB integrity was compromised after HI which was improved by MSC-EVs containing ANXA1. Treatment with these MSC-EVs or ANXA1 improved BBB integrity after OGD, an effect abolished by FPR inhibitors. Furthermore, endogenous ANXA1 was depleted within 24 h after induction of HI in cerebovasculature and ependyma and upregulated 72 h after HI in microglia. Targeting ANXA1/FPR with ANXA1 in the immature brain has great potential in preventing BBB loss and concomitant brain injury following HI.