- Email: [email protected]
The microbiome, parturition, and timing of birth
46
Abstracts / Journal of Reproductive Immunology 115 (2016) 39–55
S9 Immune uterine profiling increases the live birth rate of patients with previous repeated embryos implantation failures after IVF/ICSI N. Lédée Hopital Saint Louis, Matricelab Innove, INSERM U 976, Paris, France Hopital des bluets, Centre d’assistance médicale à la procreation, Paris, France Implantation is the main limiting factor of success in assisted reproductive medicine. 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 immune mechanism involved. 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 deregulation at the next embryo transfer (ET). The analyzed group was paired to the closest patient included in the IVF program at the time of ET according to biological criteria (age, ART method, number of oocytes, stage and number of transferred Embryo) and constituted the non-analyzed group (193 patients). Live birth rate (LBR) was significantly higher in the analyzed group versus non-analyzed group (27.5% versus 16.6%, p-0.03) and miscarriage rate per initiated pregnancy significantly decreased (respectively 17% versus 41%, p = 0.003). The uterine immune profile may guide physicians through an effective understanding that optimizes the conditions of uterine receptivity. A 65% relative increase of the LBR was observed in patients who benefitted of the immune uterine profiling. http://dx.doi.org/10.1016/j.jri.2016.04.138 OP6 Elevated levels of Tc17 and NK17 cells in early pregnancy are associated with spontaneous abortion in women with a history of recurrent pregnancy loss N. Mansouri-Attia ∗ , D.R. Ritsick, M. Campbell, K. Carlson, A. Vidali, J. Braverman Braverman Reproductive Immunology PC, Woodbury, New York, USA Dysregulated responses of the maternal immune system to the semi-allogenic conceptus have been implicated in immunological rejection resulting in infertility and recurrent pregnancy loss (RPL). This study aimed to examine associations of several immune cell types with spontaneous abortion (SA) in a cohort of women being treated for RPL. Retrospective analysis of immunological data from 205 clinical pregnancies in women being treated for RPL was conducted. These pregnancies resulted in 165 continuing pregnancies (P, 81.5%) and 40 pregnancies (19.5%) resulting in (SA). Testing was performed on peripheral blood of patients drawn prior to pregnancy (T0), at 4–5 (T1), 6–9 (T2), 10–13 (T3), 14–17 (T4) and 18–20 weeks gestation (T5). Flow cytometry was used to detect levels of CD4+ T (Th) cells, CD8+ T (Tc) cells, NKT cells, and NK cells expressing IL-4, IL-10, IL17, TNF␣, and IFN␥. No statistically significant differences in levels of any individual CD4+ T cell or NKT cell subtype were observed between the two groups. In addition, no bias toward a Th1 shift
was observed in the SA group. Rather the data show a significant increase in levels of Tc17 and NK17 cells at T2 in the (SA) compared with the (P) group. These data show a strong association of elevated levels of Tc17 and NK17 cells in the first trimester with (SA) in RPL patients and further studies of the role of these cells in early pregnancy may provide novel insights on immune mechanisms involved in pregnancy failure. http://dx.doi.org/10.1016/j.jri.2016.04.139 S10 Repeated implantation failure: A new potential treatment option A. Makrigiannakis University of Iraklion, Crete, Greece The immunological relationship between mother and conceptus still remains a mystery, although the recent advances in molecular biology have lighten a lot of the parameters that participate in feto-maternal cross-talk during implantation. The atypical expression of major histocompatibility complex (MHC), the specific role of some hormones and cytokines, as well as the modified function of cellular constituents of the feto-maternal interface, represent substantive parameters of fetomaternal immunotolerance during implantation. However the implantation process is currently considered the most important limiting factor for the establishment of a viable pregnancy and the fertility physician is often called upon to perform the unpleasant task of counselling an infertile couple after repeated implantation failure (RIF). Aetiology is often not clear and treatment options are indistinct. Some of these include hysteroscopic treatment, myomectomy, preimplantation genetic diagnosis for aneuploidy screening (PGS), assisted hatching, blastocyst transfer, zygote intra-Fallopian transfer (ZIFT), salpingectomy of hydrosalpinges and immunological treatment. Since some of these remedies have not been proven to be effective (the evidence behind some of these is robust), assisted reproduction programmes should resist offering treatment options that are not evidence based at least until well designed randomized studies show the value of what are today considered as empirical treatments. http://dx.doi.org/10.1016/j.jri.2016.04.140
Session 5: Impact of the microbiome and pathogens in reproduction S11 The microbiome, parturition, and timing of birth A.L. Prince Baylor College of Medicine, Houston, TX, USA Work from the laboratory of Dr. Kjersti Aagaard has focused on the role of the microbiome in pregnancy and how this is affiliated with the offspring microbiome. Upon characterizing the microbiome during pregnancy, we demonstrated the presence of a placental microbiome in healthy, human subjects. Taking these studies further, we have found that the placental microbiome differentiates by term versus preterm birth; additionally, the preterm placental microbiome can be further differentiated in association with gestational weight gain. Recently, we have characterized the placental membrane microbiome, which is altered in association with gestational age and histological chorioamnionitis. Altogether,
Abstracts / Journal of Reproductive Immunology 115 (2016) 39–55
these recent studies suggest that inflammation is associated with alterations in the placental microbiome. With this in mind, we have examined associations between the offspring microbiome and inflammatory factors, such as a maternal high fat diet. In these studies, we utilized a non-human primate model, which demonstrated that the fetus harbors its own unique microbiome. Also, a maternal high fat diet is associated with alterations in the fetal and juvenile offspring microbiome, and these alterations are present at three years of age. Our data indicate that a maternal high fat diet during gestation and lactation alters the offspring microbiome persistently. Overall, our work suggests that inflammatory factors are associated with alterations in the placental and offspring microbiomes. http://dx.doi.org/10.1016/j.jri.2016.04.141 OP7 Influenza A virus infection during pregnancy: Reduced CD8 T cell responses significantly impair the immune reaction against influenza A virus infection in pregnant mice A. Hierweger ∗ , G. Engels, G. Gabriel, H.-W. Mittrücker, P.C. Arck Department of Obstetrics and Fetal Medicine, Laboratory for Experimental Feto-Maternal Medicine & Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany During the 2009 H1N1 influenza A virus (IAV) pandemic, pregnant women showed higher influenza related morbidity and mortality. Thus, it is of great clinical interest to investigate underlying altered immune responses leading to increased risk for pregnant women. We previously developed an IAV mouse model where Balb/cmated C57BL/6 dams were infected with a 2009 pandemic H1N1 IAV strain, showing increased mortality and morbidity that corresponds to clinical observations in humans. Using this model, we could now show that the total number of leukocytes in the lung was reduced in infected pregnant mice compared to infected, non-pregnant mice. Thus, we detected a significantly reduced expression of the chemokines Cxcl10 and Cxcl9 in lungs of pregnant, infected mice. Based on these findings, we next assessed the migration of CD8+ T cells, isolated from congenic CD90.1+ infected, non-pregnant C57BL/6 donors and adoptively transferred them into infected wild type (CD90.2+) pregnant C57BL/6 mice. In pregnant animals we observed significantly reduced frequencies of migrated CD8+ T cells as well as migrated CD8+ T cells bearing the cognate Cxcl10 receptor, CXCR3. These observations strongly support an impaired Cxcl10dependent recruitment of leukocytes to the lung during pregnancy, similar as recently shown for the feto-maternal interface. Whilst bearing benefits for fetal tolerance, these migration constrains are associated with severe disadvantages for maternal health. http://dx.doi.org/10.1016/j.jri.2016.04.142
47
S12 Modulation of neonatal host defense against infection S. Merani 1 , P. Kent 1 , G. Xu 1 , L. Xu 1 , S. Elahi 1,2,∗ 1 Departments of Dentistry, University of Alberta, Canada 2 Medical Microbiology and Immunology, University of Alberta, Canada
Neonates are highly susceptible to disseminated infections, which are often fatal. The mechanisms underlying the susceptibility of neonates to infections and the molecular basis for the transition of immunologic function from fetal to postnatal life has remained a mystery. While most vaccines do induce protective immunity in older children and adults, their efficacy in the very young often requires further manipulation and optimization. Recently, we have reported that CD71+ erythroid cells are physiologically enriched in neonatal mice and human cord blood with distinctive immune suppressive properties. Our most recent data indicate that CD71+ cells impair neonatal adaptive immune responses by suppressing T cell functions in vitro and in vivo. In addition, CD71+ erythroid cells by expression of ligands for inhibitory receptors and production of regulatory cytokines could play an essential role in immune regulation. More interestingly, CD71+ cell mediated susceptibility to infection is counterbalanced by protection against aberrant immune cell activation in the intestine where brisk postnatal colonization with commensal microbes occurs. CD71+ cells quench excessive upregulation of TLRs induced by abrupt commensal colonization after parturition. Our data provide new insight into the role of this novel subset of “suppressor cells” in immunopathogenesis of neonatal infections and immune regulation early in life. http://dx.doi.org/10.1016/j.jri.2016.04.143 OP8 Abnormal vaginal microbiota may be associated with poor reproductive outcomes: A prospective study in IVF patients T. Haahr Aarhus University and The Fertility Clinic Skive, Skive Regional Hospital, Denmark Background: Bacterial vaginosis (BV) has a prevalence of approximately 19% in the infertile population. BV is often subclinical, characterized by a dysbiosis with anaerobic and facultative anaerobic bacteria, such as Gardnerella (g) vaginalis and Atopobium (a) vaginae. Few studies have been conducted in infertile women, and some have suggested a negative impact on fecundity in the presence of BV. Methods: A cohort of 130 infertile patients attending for in vitro fertilization (IVF) treatment was prospectively enrolled. Gram stained slides were assessed according to Nugent’s criteria. PCR primers were specific for four common Lactobacillus spp., G. vaginalis and A. vaginae. Results: The prevalence of BV defined by Nugent score was 21% (27/130), whereas the prevalence of an abnormal vaginal microbiota was 28% (36/130) defined by qPCR with high concentrations of G. vaginalis and/or A. vaginae. The qPCR diagnostic approach had a sensitivity and specificity of 93% and 93% for Nugent-defined BV. Eighty-four patients completed IVF treatment. The overall clinical pregnancy rate was 35% (29/84). Interestingly, only 9% (2/22) with qPCR defined abnormal vaginal microbiota obtained a clinical pregnancy (P = 0.004).
Abstracts / Journal of Reproductive Immunology 115 (2016) 39–55
S9 Immune uterine profiling increases the live birth rate of patients with previous repeated embryos implantation failures after IVF/ICSI N. Lédée Hopital Saint Louis, Matricelab Innove, INSERM U 976, Paris, France Hopital des bluets, Centre d’assistance médicale à la procreation, Paris, France Implantation is the main limiting factor of success in assisted reproductive medicine. 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 immune mechanism involved. 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 deregulation at the next embryo transfer (ET). The analyzed group was paired to the closest patient included in the IVF program at the time of ET according to biological criteria (age, ART method, number of oocytes, stage and number of transferred Embryo) and constituted the non-analyzed group (193 patients). Live birth rate (LBR) was significantly higher in the analyzed group versus non-analyzed group (27.5% versus 16.6%, p-0.03) and miscarriage rate per initiated pregnancy significantly decreased (respectively 17% versus 41%, p = 0.003). The uterine immune profile may guide physicians through an effective understanding that optimizes the conditions of uterine receptivity. A 65% relative increase of the LBR was observed in patients who benefitted of the immune uterine profiling. http://dx.doi.org/10.1016/j.jri.2016.04.138 OP6 Elevated levels of Tc17 and NK17 cells in early pregnancy are associated with spontaneous abortion in women with a history of recurrent pregnancy loss N. Mansouri-Attia ∗ , D.R. Ritsick, M. Campbell, K. Carlson, A. Vidali, J. Braverman Braverman Reproductive Immunology PC, Woodbury, New York, USA Dysregulated responses of the maternal immune system to the semi-allogenic conceptus have been implicated in immunological rejection resulting in infertility and recurrent pregnancy loss (RPL). This study aimed to examine associations of several immune cell types with spontaneous abortion (SA) in a cohort of women being treated for RPL. Retrospective analysis of immunological data from 205 clinical pregnancies in women being treated for RPL was conducted. These pregnancies resulted in 165 continuing pregnancies (P, 81.5%) and 40 pregnancies (19.5%) resulting in (SA). Testing was performed on peripheral blood of patients drawn prior to pregnancy (T0), at 4–5 (T1), 6–9 (T2), 10–13 (T3), 14–17 (T4) and 18–20 weeks gestation (T5). Flow cytometry was used to detect levels of CD4+ T (Th) cells, CD8+ T (Tc) cells, NKT cells, and NK cells expressing IL-4, IL-10, IL17, TNF␣, and IFN␥. No statistically significant differences in levels of any individual CD4+ T cell or NKT cell subtype were observed between the two groups. In addition, no bias toward a Th1 shift
was observed in the SA group. Rather the data show a significant increase in levels of Tc17 and NK17 cells at T2 in the (SA) compared with the (P) group. These data show a strong association of elevated levels of Tc17 and NK17 cells in the first trimester with (SA) in RPL patients and further studies of the role of these cells in early pregnancy may provide novel insights on immune mechanisms involved in pregnancy failure. http://dx.doi.org/10.1016/j.jri.2016.04.139 S10 Repeated implantation failure: A new potential treatment option A. Makrigiannakis University of Iraklion, Crete, Greece The immunological relationship between mother and conceptus still remains a mystery, although the recent advances in molecular biology have lighten a lot of the parameters that participate in feto-maternal cross-talk during implantation. The atypical expression of major histocompatibility complex (MHC), the specific role of some hormones and cytokines, as well as the modified function of cellular constituents of the feto-maternal interface, represent substantive parameters of fetomaternal immunotolerance during implantation. However the implantation process is currently considered the most important limiting factor for the establishment of a viable pregnancy and the fertility physician is often called upon to perform the unpleasant task of counselling an infertile couple after repeated implantation failure (RIF). Aetiology is often not clear and treatment options are indistinct. Some of these include hysteroscopic treatment, myomectomy, preimplantation genetic diagnosis for aneuploidy screening (PGS), assisted hatching, blastocyst transfer, zygote intra-Fallopian transfer (ZIFT), salpingectomy of hydrosalpinges and immunological treatment. Since some of these remedies have not been proven to be effective (the evidence behind some of these is robust), assisted reproduction programmes should resist offering treatment options that are not evidence based at least until well designed randomized studies show the value of what are today considered as empirical treatments. http://dx.doi.org/10.1016/j.jri.2016.04.140
Session 5: Impact of the microbiome and pathogens in reproduction S11 The microbiome, parturition, and timing of birth A.L. Prince Baylor College of Medicine, Houston, TX, USA Work from the laboratory of Dr. Kjersti Aagaard has focused on the role of the microbiome in pregnancy and how this is affiliated with the offspring microbiome. Upon characterizing the microbiome during pregnancy, we demonstrated the presence of a placental microbiome in healthy, human subjects. Taking these studies further, we have found that the placental microbiome differentiates by term versus preterm birth; additionally, the preterm placental microbiome can be further differentiated in association with gestational weight gain. Recently, we have characterized the placental membrane microbiome, which is altered in association with gestational age and histological chorioamnionitis. Altogether,
Abstracts / Journal of Reproductive Immunology 115 (2016) 39–55
these recent studies suggest that inflammation is associated with alterations in the placental microbiome. With this in mind, we have examined associations between the offspring microbiome and inflammatory factors, such as a maternal high fat diet. In these studies, we utilized a non-human primate model, which demonstrated that the fetus harbors its own unique microbiome. Also, a maternal high fat diet is associated with alterations in the fetal and juvenile offspring microbiome, and these alterations are present at three years of age. Our data indicate that a maternal high fat diet during gestation and lactation alters the offspring microbiome persistently. Overall, our work suggests that inflammatory factors are associated with alterations in the placental and offspring microbiomes. http://dx.doi.org/10.1016/j.jri.2016.04.141 OP7 Influenza A virus infection during pregnancy: Reduced CD8 T cell responses significantly impair the immune reaction against influenza A virus infection in pregnant mice A. Hierweger ∗ , G. Engels, G. Gabriel, H.-W. Mittrücker, P.C. Arck Department of Obstetrics and Fetal Medicine, Laboratory for Experimental Feto-Maternal Medicine & Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany During the 2009 H1N1 influenza A virus (IAV) pandemic, pregnant women showed higher influenza related morbidity and mortality. Thus, it is of great clinical interest to investigate underlying altered immune responses leading to increased risk for pregnant women. We previously developed an IAV mouse model where Balb/cmated C57BL/6 dams were infected with a 2009 pandemic H1N1 IAV strain, showing increased mortality and morbidity that corresponds to clinical observations in humans. Using this model, we could now show that the total number of leukocytes in the lung was reduced in infected pregnant mice compared to infected, non-pregnant mice. Thus, we detected a significantly reduced expression of the chemokines Cxcl10 and Cxcl9 in lungs of pregnant, infected mice. Based on these findings, we next assessed the migration of CD8+ T cells, isolated from congenic CD90.1+ infected, non-pregnant C57BL/6 donors and adoptively transferred them into infected wild type (CD90.2+) pregnant C57BL/6 mice. In pregnant animals we observed significantly reduced frequencies of migrated CD8+ T cells as well as migrated CD8+ T cells bearing the cognate Cxcl10 receptor, CXCR3. These observations strongly support an impaired Cxcl10dependent recruitment of leukocytes to the lung during pregnancy, similar as recently shown for the feto-maternal interface. Whilst bearing benefits for fetal tolerance, these migration constrains are associated with severe disadvantages for maternal health. http://dx.doi.org/10.1016/j.jri.2016.04.142
47
S12 Modulation of neonatal host defense against infection S. Merani 1 , P. Kent 1 , G. Xu 1 , L. Xu 1 , S. Elahi 1,2,∗ 1 Departments of Dentistry, University of Alberta, Canada 2 Medical Microbiology and Immunology, University of Alberta, Canada
Neonates are highly susceptible to disseminated infections, which are often fatal. The mechanisms underlying the susceptibility of neonates to infections and the molecular basis for the transition of immunologic function from fetal to postnatal life has remained a mystery. While most vaccines do induce protective immunity in older children and adults, their efficacy in the very young often requires further manipulation and optimization. Recently, we have reported that CD71+ erythroid cells are physiologically enriched in neonatal mice and human cord blood with distinctive immune suppressive properties. Our most recent data indicate that CD71+ cells impair neonatal adaptive immune responses by suppressing T cell functions in vitro and in vivo. In addition, CD71+ erythroid cells by expression of ligands for inhibitory receptors and production of regulatory cytokines could play an essential role in immune regulation. More interestingly, CD71+ cell mediated susceptibility to infection is counterbalanced by protection against aberrant immune cell activation in the intestine where brisk postnatal colonization with commensal microbes occurs. CD71+ cells quench excessive upregulation of TLRs induced by abrupt commensal colonization after parturition. Our data provide new insight into the role of this novel subset of “suppressor cells” in immunopathogenesis of neonatal infections and immune regulation early in life. http://dx.doi.org/10.1016/j.jri.2016.04.143 OP8 Abnormal vaginal microbiota may be associated with poor reproductive outcomes: A prospective study in IVF patients T. Haahr Aarhus University and The Fertility Clinic Skive, Skive Regional Hospital, Denmark Background: Bacterial vaginosis (BV) has a prevalence of approximately 19% in the infertile population. BV is often subclinical, characterized by a dysbiosis with anaerobic and facultative anaerobic bacteria, such as Gardnerella (g) vaginalis and Atopobium (a) vaginae. Few studies have been conducted in infertile women, and some have suggested a negative impact on fecundity in the presence of BV. Methods: A cohort of 130 infertile patients attending for in vitro fertilization (IVF) treatment was prospectively enrolled. Gram stained slides were assessed according to Nugent’s criteria. PCR primers were specific for four common Lactobacillus spp., G. vaginalis and A. vaginae. Results: The prevalence of BV defined by Nugent score was 21% (27/130), whereas the prevalence of an abnormal vaginal microbiota was 28% (36/130) defined by qPCR with high concentrations of G. vaginalis and/or A. vaginae. The qPCR diagnostic approach had a sensitivity and specificity of 93% and 93% for Nugent-defined BV. Eighty-four patients completed IVF treatment. The overall clinical pregnancy rate was 35% (29/84). Interestingly, only 9% (2/22) with qPCR defined abnormal vaginal microbiota obtained a clinical pregnancy (P = 0.004).