Primipaternities and human birthweights.

OBJECTIVES: To investigate in singleton multiparous pregnancies the effect of having a new father for an index pregnancy on new-borns' birthweights and intrauterine growth restriction. DESIGN: 20 year-observational cohort study (2001-2020). SETTINGS: Centre Hospitalier Universitaire Hospitalier Sud Reunion's maternity (French overseas department, Indian Ocean). MAIN OUTCOMES AND MEASURES: Comparing the 811 multiparas (cases) who had a new partner with the 49,712 who did not (controls), there were no differences concerning maternal age, education, ovulation induction/IVF, previous miscarriages, exams during pregnancies, pre-pregnancy BMI, gestational diabetes, and chronic hypertension. Cases had more previous pregnancies than controls (gravidity 4.2 vs 2.8, p < 0.001), volunteer abortions (OR1.93, p < 0.001), in vitro fecundations (OR 4.34, p < 0.001), were more likely to be unmarried (OR 2.94, p < 0.001) smoker (OR 2.2, p < 0.0001) and consuming alcohol during pregnancy (OR 2.35, p = 0.001). Cases had a much higher risk of preeclampsia than controls (OR 3.94, p < 0.001), especially early-onset preeclampsia (< 34 weeks) with an OR 4.1 (p < 0.001). Controlling for confounding factors (preeclampsia, smoking, alcohol use, early prematurity < 33 weeks, maternal ethnicity), primipaternity was an independent factor for small for gestational age newborns (OR 1.48, p < 0.001). CONCLUSIONS: It has been known for decades that primiparas have lighter babies than multiparas. Primipaternity represents also a risk for lower birth weights. Human birthweight seems to be linked with a "couple habituation" (to paternal genes) which may be not fully established in the first pregnancy of the couple.

Decreased Immune Response in Alexithymic Women: A One-Year Longitudinal Study.

Although previous cross-sectional studies suggested significantly dysregulated immune response in alexithymia, there is a lack of longitudinal studies. We sought to determine the reliability of the reported relationship between alexithymia and decreased immune response in a longitudinal study. Thirty-eight healthy women who had participated in a cross-sectional study were recontacted 1-year later. Of this sample, 26 were finally included: 13 females who had been found to be alexithymic, and 13 females who were classified as non-alexithymic under the 20-item Toronto Alexithymia Scale during the first phase of the study. A year later, they were still healthy women without any psychiatric disorders, their ages now ranging from 19 to 28 years old. Lymphocyte subset counts (CD4, CD8), in vitro production of interleukin 1ß (IL-1ß), IL-2, IL-4, and IL-10 by phytohemagglutinin stimulated peripheral blood lymphocytes, as well as serum cortisol levels, were compared between women with and without alexithymia. One-year later, alexithymic women still had significantly lowered in vitro production of IL-2 and IL-4, with lowered IL-2/IL-10 ratio and a reduced percentage of CD4. This is the first ever published study assessing cytokine production during a follow-up of alexithymics. Although our results should be interpreted with caution due the small sample size, they suggest a sustained reduction in both major type 1 and type 2 cytokines while the former seems to be more affected. The potential long-term health impact, if any, is still to be determined.

Glycan expression in chorionic villi from histocultures of women with early-onset preeclampsia: Immunomodulatory effects on peripheral natural killer cells.

New evidence suggests that glycan expression in placental cells of women with invasive disorders of pregnancy differs from that in normal pregnant women. Hypothesizing that modifications of glycan expression could account for the course of preeclampsia, we established placental villous histocultures and compared glycan expression in women with preeclampsia with that in normal pregnant women and also in syncytialized BeWo cells, and we tested the effect of glycan expression on the functional phenotypes of circulating natural killer (NK) cells. Histocultures of third-trimester placentae from women with preeclampsia and full-term placentae from healthy pregnant women and BeWo choriocarcinoma cells were assessed for the expression of terminal glycans by lectin-binding assays. Circulating NK cells from nonpregnant healthy donors were tested in vitro for their cytotoxic activity and intracellular cytokine content. Histocultures from women with preeclampsia expressed significantly more mannose than did those from healthy pregnant women. Both histocultures and BeWo cells expressed terminal fucose, mannose, sialic acid, and N -acetylgalactosamine, although mean fluorescence intensity (MFI) expression was lower in choriocarcinoma cells than in cells from histocultures. Cocultures of circulating NK cells with K562 target cells resulted in a dose-dependent cytotoxicity effect, but the use of BeWo cells as target reduced cytotoxic activity; this reduction was not affected by syncytialization. Histocultures of placental villous tissue of women with preeclampsia expressed high levels of terminal mannose. We proposethat placental glycans may modulate the functional activity of circulating NK cells in the context of systemic inflammatory response in preeclampsia.

Endometrial Immune Profiling: A Method to Design Personalized Care in Assisted Reproductive Medicine.

Objective: To assess the efficiency of the endometrial immune profiling as a method to design personalized care to enhance the pregnancy rate in a large heterogeneous infertile population. We hypothesized that some reproductive failures could be induced by a uterine immune dysregulation which could be identified and corrected with a targeted plan. Design: Prospective cohort study. Setting: Multicentric study. Intervention(s) and Main outcome measure(s): One thousand and seven hundred thirty-eight infertile patients had an immune profiling on a timed endometrial biopsy between 2012 and 2018. This test documented the absence or the presence of an endometrial immune dysregulation and identified its type. In case of dysregulation, a targeted personalized plan was suggested to the treating clinician aiming to supply the anomaly. One year after the test, the clinician was contacted to provide the outcome of the subsequent embryo transfer with the applied suggested plan. Result(s): After testing, 16.5% of the patients showed no endometrial immune dysregulation, 28% had a local immune under-activation, 45% had a local immune over-activation, and 10.5% had a mixed endometrial immune profile. In patients with a history of repeated implantation failures (RIF) or recurrent miscarriages (RM), the pregnancy rate was significantly higher if an endometrial dysregulation was found and the personalized plan applied, compared to the patients with an apparent balanced immune profile (respectively 37.7 and 56% vs. 26.9 and 24%, p < 0.001). In contrast, in good prognosis IVF (in vitro fertilization) subgroup and patients using donor eggs, this difference was not significant between dysregulated and balanced subgroups, but higher pregnancy rates were observed in absence of dysregulation. For patients with immune over-activation, pregnancy rates were significantly higher for patients who had a test of sensitivity, regarding the type of immunotherapy introduced, when compared to the ones who did not (51 vs. 39.9%, p = 0.012). Conclusion(s): Local endometrial immunity appears to be a new and important parameter able to influence the prognosis of pregnancy. Targeted medical care in case of local immune dysregulation resulted in significantly higher pregnancy rates in RIF and RM patients.

High incidence of early onset preeclampsia is probably the rule and not the exception worldwide. 20th anniversary of the reunion workshop. A summary.

The 11th workshop on Immunology of preeclampsia in Reunion 2018 celebrated its 20th candle In this paper we try to summarize the main tracks of reflections during these two decades. First, of course, the advances in immunology of reproduction in the field of preeclampsia, which was poorly developed 2 decades ago when we first started in 1998. But, this workshop has not been dedicated only to immunology. Second, one of the main reflections has always been, workshop after workshop: "why does preeclampsia exists in humans?" in an evolutionary view, as we have no established natural animal models in the other some 4500 other mammal species. Third, besides the reflections on the biological plausibility of preeclampsia-disease-of-first-pregnancies-at-a-level-of-a-couple (primipaternity rather than primigravidity), i.e. immunology, paternal-maternal conflict, we had to face an apparent conundrum: the human species should have disappeared (almost 40-50% incidence of hypertensive disorders of pregnancy in couples conceiving within the first 4 months of sexual cohabitation). We report then the dialogues we were obliged to have with zoologists who themselves had no clues on our apparent "extravagant sexuality" and strange reproduction (ridiculous low fertility rate of the human female: 25%). Fourth, debates on the main difference between early onset ("rather immunological") and late onset PE ("rather maternal vascular predispositions"). Further, the debate of why high income countries report 90% of their PE being LOP, while other countries describe epidemiologically very high incidences of EOP. Finally, and always present at all workshops, the physiopathology of the reversible systemic maternal vascular inflammation.

A unified (but in fact not fully testable) model of preeclampsia triggering.

In this summary of my presentation in the last Reunion workshop I discuss a few assertions on preeclampsia, then turn on a (not fully testable) model where an embryonic defect in expression of embryo/ placental regulatory proteins results in complement activation, itself responsible for a down regulation of the T regs activity, resulting in a very early lack of complete down regulation of the preimplantation decidual inflammation, causing in the post implantation stage a low grade but chronic inflammatory state.

Intralipid® may represent a new hope for patients with reproductive failures and simultaneously an over-immune endometrial activation.

PROBLEM: Continuous failures to achieve a pregnancy despite effective embryo transfers is extremely distressing for couples. In consequence, many adjuvant therapies to IVF have been proposed to achieve an "ideal" immune environment. We here focus on Intralipid® therapy (IL) reported to have immunosuppressive properties on NK cells. METHOD OF STUDY: 94 patients exhibited an immune profile of endometrial over-immune activation and an history of repeated implantation failures despite multiple embryos transfers (RIF). They received a slow perfusion of Intralipid®. We here report the live birth rate following the procedure at the next embryo transfer. To get new insight on its mechanism of action, a second immune profiling had been performed under Intralipid® before the embryo transfer. RESULTS: The live birth rate of the RIF cohort treated with Intralipid® reached 54% (51/94) at the next embryo transfer. In patients successfully pregnant under Intralipid® who benefitted of a test of sensibility before the embryo transfer, we observed a significant decrease of the three biomarkers used to diagnose the over-immune endometrial activation (CD56 cells; IL-18/TWEAK, IL-14/FN-14). CONCLUSIONS: Double blind placebo versus Intralipid® studies should be conducted. Intralipid® may be an option to explore in RIF patients who exhibit an over-immune activation of uNK cells.

Preeclampsia and the 20th century: "Le siècle des Lumières".

The authors delineate seven quantum leap forward and, or revolutions having occurred during the 20th century in the understanding of the physiopathology of preeclampsia. First the discovery of the inflatable arm band permitting to measure blood pressure in 1896. Second, the discovery that eclamptic (convulsions), and later "pre"eclamptic (proteinuria) women presented hypertension in 1897 and confirmed in 1903, discovery of the hypertensive disorders of pregnancy. Third, the eight major textbooks published all along the 20th century by delineating risk factors of preeclampsia with the concept of "preeclampsia, disease of primigravidae". Fourth, the discovery in the 1970's that human trophoblast implantation was far deeper than in other mammalian species. Fifth, and a major step forward, description at the end of the 1980's that the maternal syndrome in preeclampsia (glomeruloendotheliosis, HELLP syndrome, eclampsia) could be unified in a global endothelial cell inflammation. Sixth, the epidemiological descriptions in the 1970-1990's that indeed preeclampsia was a disease of first pregnancies at the level of a couple ("primipaternity concept"), leading to an explosion in immunological research in the last decade, beginning in 1998. Seventh and finally, in the search for the "factor X" explaining the vascular inflammation in preeclamptic women (inositol phospho glycans P-type were described in 2000, while soluble Flt-1 and S-endoglins have been clearly predicted since 1997). The majority of the seeds or findings have been grounded or realized in the 20th century. Indeed, for preeclampsia, the 20th century has been le "Siècle des Lumières" (the Enlightments).

Impact of prednisone in patients with repeated embryo implantation failures: Beneficial or deleterious?

INTRODUCTION: Corticotherapy is the leading medication worldwide for patients with history of repeated implantation failures (RIF) after IVF/ICSI. Nevertheless, we still do not know its local mechanism of action, hence its precise indication. Our objective is to document the impact of prednisone on the endometrial expression of immune biomarkers (CD56 cells count, IL-18/TWEAK, IL-15/Fn-14 mRNA ratio) at the time of uterine receptivity in a RIF population. MATERIALS AND METHOD: An endometrial biopsy was realized in the mid luteal phase for immune profiling: IL-15/Fn-14 and IL-18/TWEAK mRNA ratios were determined by quantitative RT-PCR and CD56 mobilization per IHC. Fifty-five patients with a RIF history were diagnosed to have local over-immune activation [high IL-18/TWEAK mRNA ratio, and/or high IL-15/Fn-14 mRNA ratio] likely to impair the implantation process. They underwent a second immune profiling with supplementation of prednisone. A paired comparison of the immune profile before and under prednisone was performed in the subset of patients subsequently pregnant under prednisone. FINDING: In 54.5% of the cases, both immune biomarkers were normalized and in 16.5%, only one was normalized under prednisone. In 29% we observed a paradoxical increase of both immune biomarkers. The IL-18/TWEAK mRNA ratio reflecting the Th-1/Th-2 local equilibrium was significantly reduced (0.29 versus 0.10, p = .004), through very significant increase of TWEAK expression, in patients who were subsequently pregnant under prednisone. CONCLUSION: Testing the response to prednisone in a RIF context may be very useful. Less than half of RIF patients with immune deregulation may be prednisone responders and would benefit from its administration.

Historical evolution of ideas on eclampsia/preeclampsia: A proposed optimistic view of preeclampsia.

Eclampsia (together with epilepsy) being the first disease ever written down since the beginning of writings in mankind 5000 years ago, we will make a brief presentation of the different major steps in comprehension of Pre-eclampsia. 1) 1840. Rayer, description of proteinuria in eclampsia, 2) 1897 Vaquez, discovery of gestational hypertension in eclamptic women, 3) In the 1970's, description of the "double" trophoblastic invasion existing only in humans (Brosens & Pijnenborg,), 4) between the 1970's and the 1990's, description of preeclampsia being a couple disease. The "paternity problem" (and therefore irruption of immunology), 5) at the end of the 1980's, a major step forward: Preeclampsia being a global endothelial cell disease (glomeruloendotheliosis, hepatic or cerebral endotheliosis, HELLP, eclampsia), inflammation (J.Roberts.C Redman, R Taylor), 6) End of the 1990's: Consensus for a distinction between early onset preeclampsia EOP and late onset LOP (34 weeks gestation), EOP being rather a problem of implantation of the trophoblast (and the placenta), LOP being rather a pre-existing maternal problem (obesity, diabetes, coagulopathies etc…). LOP is predominant everywhere on this planet, but enormously predominant in developed countries: 90% of cases. This feature is very different in countries where women have their first child very young (88% of world births), where the fatal EOP (early onset) occurs in more than 30% of cases. 7) What could be the common factor which could explain the maternal global endotheliosis in EOP and LOP? Discussion about the inositol phospho glycans P type.

Uterine immune profiling for increasing live birth rate: A one-to-one matched cohort study.

BACKGROUND: Embryo implantation remains the main limiting factor in IVF/ICSI program. Endometrial immune remodeling events begin before implantation and are a vital process for pregnancy, preparing future maternal immune tolerance and regulating the placentation process. METHODS: Between 2012 and 2014, 193 patients (analyzed group) enrolled in our IVF program benefitted of an endometrial immune profiling to determine if their uterus was immunologically ready to accept an embryo and, if not, the specific immune mechanisms involved. Subsequently, they had an effective embryo transfer (ET) with personalization of their treatments if an immune deregulation has been diagnosed. Each analyzed patient was paired to the closest patient included in the IVF program according to biological criteria (age, number of mature oocytes, stage and number of transferred embryo), which had no endometrial immune profiling (193 patients, non-analyzed group). FINDING: 78% of analyzed patients had a uterine immune dysregulation and therefore care personalization. Their corresponding live birth rate (LBR) was twice higher than observed in the matched control group with conventional cares (30.5% versus 16.6%, OR: 2.2 [1.27-3.83] p=0.004) with a simultaneous drastic reduction of miscarriages per initiated pregnancy (17.9% versus 43.2%, OR: 0.29 [0.12-0.71], p=0.005). 22% of analyzed patients had no dysregulation. They did not differ from their matched controls for LBR and miscarriages. CONCLUSION: Uterine immune profiling enables an integrated approach of infertility that includes endometrial immunity as a key factor in planning personalized IVF/ICSI treatments. Personalization of treatment according to the woman's uterine immune balance produced a very significantly higher LBR.

The Uterine Immune Profile May Help Women With Repeated Unexplained Embryo Implantation Failure After In Vitro Fertilization.

LABELED PROBLEM: Embryo implantation remains the main limiting factor in assisted reproductive medicine (20% success rate). METHODS OF STUDY: An endometrial immune profiling was performed among 394 women with the previous history of repeated embryo implantation failures (RIF). The endometrial immune profile documented the ratio of IL-15/Fn-14 mRNA as a biomarker of uNK cell activation/maturation (together with the uNK cell count) and the IL-18/TWEAK mRNA ratio as a biomarker of both angiogenesis and the Th1/Th2 balance. According to their profile, we recommended personalized care to counteract the documented dysregulation and assessed its effects by the live birth rate (LBR) for the next embryo transfer. RESULTS: Endometrial immune profiles appeared to be dysregulated in 81.7% of the RIF patients compared to control. Overactivation was diagnosed in 56.6% and low activation in 25%. The LBR among these dysregulated/treated patients at the first subsequent embryo transfer was 39.8%. CONCLUSION: Endometrial immune profiling may improve our understanding of RIF and subsequent LBR if treated.

An essay of reflection: Why does preeclampsia exist in humans, and why are there such huge geographical differences in epidemiology?

This workshop had four main objectives: (A) Trying to look at the preeclampsia (PE) problem "from the Space Shuttle": why preeclampsia has emerged in humans (a specific human reproductive feature among 4300 mammal species)? (B) Epidemiology: there are major geographical differences concerning early onset PE and late onset PE throughout the world. (C) Vascular: The very promising use of pravastatin in the treatment of the vascular maternal syndrome (based on the metabolism of carbon monoxide (CO), the role of inositol phosphate glycans P-type (IPG-P), a major role in comprehending the insulin resistance phenotype in preeclampsia. (D) Immunology: the specialty of these workshops since their start in 1998; our understanding of the role of the immune system and the regulation of the deep implantation of the human trophoblast (and the obligatory compromises between the fetal/placental unit and the mother) have reached a kind of "maturity," following the pivotal studies exploring the biology of repetitive sperm exposure in the female genital tract. The meeting of people who never meet each other in the course of their normal professional lives (obstetricians, evolutionists, geneticists, immunologists, fundamentalist vascular biologists, epidemiologists, anthropologists, neonatologists, etc.) permitted some fruitful reflections to be made again this year.

Reconsidering the Medawar paradigm placental viviparity existed for eons, even in vertebrates; without a "problem": Why are Tregs important for preeclampsia in great apes?

The classic Medawar paradigm sees viviparity in vertebrates as a "problem". Established in 1953, it was then largely determined by a self-non-self view of the immune system. However, there are alternative models of the immune system, such as the danger model. For these models, pregnancy is neither a problem nor a danger. Supporting this view, we recall that placenta or placental-like-dependent(1) (allo) pregnancy has existed for eons. In fact, it appeared as far back as the time of aquatic colony invertebrates, such as some of the Bryozoa.(2) Since then, convergent evolution has seen placentation appear in a large variety of phyla. These placentae did not seem to cause "immunological problems", even in vertebrates possessing a graft rejection potential. The reappearance of placentae in marsupial and eutherian mammals found placentae confronted with a highly developed adaptive immune system. Two strategies were developed, therefore: short-term only placentation (marsupials) or specialised control of T cell-mediated immunity (Tregs). The problem is likely to be most acute in cases of deep invasive placentation. As an alternative to a restricted view of the Medawar paradigm for preeclampsia, an integrated model putting both inflammation and Tregs into perspective is proposed, somehow embedding the questioning of the initial Medawar paradigm.

Critical Role and Therapeutic Control of the Lectin Pathway of Complement Activation in an Abortion-Prone Mouse Mating.

The abortion-prone mating combination CBA/J × DBA/2 has been recognized as a model of preeclampsia, and complement activation has been implicated in the high rate of pregnancy loss observed in CBA/J mice. We have analyzed the implantation sites collected from DBA/2-mated CBA/J mice for the deposition of the complement recognition molecules using CBA/J mated with BALB/c mice as a control group. MBL-A was observed in the implantation sites of CBA/J × DBA/2 combination in the absence of MBL-C and was undetectable in BALB/c-mated CBA/J mice. Conversely, C1q was present in both mating combinations. Searching for other complement components localized at the implantation sites of CBA/J × DBA/2, we found C4 and C3, but we failed to reveal C1r. These data suggest that complement is activated through the lectin pathway and proceeds to completion of the activation sequence as revealed by C9 deposition. MBL-A was detected as early as 3.5 d of pregnancy, and MBL-A deficiency prevented pregnancy loss in the abortion-prone mating combination. The contribution of the terminal complex to miscarriage was supported by the finding that pregnancy failure was largely inhibited by the administration of neutralizing Ab to C5. Treatment of DBA/2-mated CBA/J mice with Polyman2 that binds to MBL-A with high affinity proved to be highly effective in controlling the activation of the lectin pathway and in preventing fetal loss.

Male fertility and apoptosis in normal spermatogenesis are regulated by vacuolar-ATPase isoform a2.

The a2 isoform of vacuolar-ATPase (ATP6V0A2, referred to as a2V) is required for normal spermatogenesis and maturation of sperm. Treatment of male mice with anti-a2V disturbs the testicular cytokine/chemokine balance and leads to severe deficiencies of spermatogenesis. The aim of the present study was to investigate the role of a2V in male fertility and in the regulation of apoptotic pathways required for normal spermatogenesis in mice. To study the role of a2V single dose of anti-a2V monoclonal antibody or mouse IgG isotype (3µg/animal) was injected i.p. into males on alternate days for 10 days. The expression of sperm maturation-related molecules and pro-apoptotic molecules was measured by real-time PCR or immunohistochemistry in control and anti-a2V-treated testes. The caspase levels and their activity were measured by western blot and fluorometry. We found that the expression of the sperm maturation-related molecules SPAM1, ADAM1, and ADAM2 was significantly decreased in testes from anti-a2V-treated males. The expression of pro-apoptotic molecules (Bax, p53, and p21) and molecules involved in the intrinsic pathway of apoptosis (caspase-9, caspase-3, and PARP), which are crucial for normal spermatogenesis was significantly reduced in testes from anti-a2V-treated males compared with the control. The total ATP level was significantly lower in anti-a2V-treated testes. The data provide novel evidence showing that a2V can regulate the apoptotic pathways, an essential testicular feature, and is necessary for efficient spermatogenesis.

Colony Stimulating Factors 1, 2, 3 and early pregnancy steps: from bench to bedside.

Reproductive immunology applies general immunology principles to specialised targets, reproduction and development. The involvement of colony-stimulating factors (CSFs) in reproduction illustrates this. The CSF family includes CSF-1 or macrophage CSF (M-CSF), CSF-2 or granulocyte macrophage CSF (GM-CSF), and CSF-3 or granulocyte CSF (G-CSF). Each member has a specific localisation and timed expression in the reproductive tract with specific functions involving them in ovulation, embryo implantation, placentation and further embryonic development. They are used in reproductive medicine, either as biomarkers of oocyte quality and competence (follicular G-CSF), or to supplement embryo culture media with human recombinant GM-CSF, or they are used as an innovative therapy by using human recombinant G-CSF for infertile patients. Given fundamental considerations on CSFs and their strong implication in reproduction, this review aimed to detail the current knowledge for each member of the family to improve our understanding of their implication in the maternal-foetal cytokinic dialogue and in possibly preventing reproductive disorders.

Are animal models useful or confusing in understanding the human feto-maternal relationship? A debate.

The proposition "This house agrees that the proper study of man is woman" was debated. For those negating the proposition, the alternative was that "animal models are useful in understanding the human feto-maternal relationship." Evidence for the proposition emphasized molecular and structural differences between the human and animal placenta and placentation. Evidence against the proposition and in favor of the alternative focused on functional and structural homologies, emphasizing that different molecules could be used in humans to achieve similar functional effects seen in animal (e.g., mouse) models. It was agreed that one always needed to test the validity of animal data by studying humans. The advantages and limitations of animal models were discussed.