Maternal immune responses may interfere with offspring allergy development as maternal immunization may suppress IgE development, while maternal allergy may promote allergy. Therefore, we investigated the effect of two different maternal treatments on airway allergy in female and male offspring.
Pregnant mice were immunized (IMM) with ovalbumin (OVA) or immunized and airway-challenged (IMM+AI). At different ages, airway allergy to OVA was induced in offspring by intranasal sensitization.
Maternal IgG1 was found at higher levels in IMM+AI than in IMM offspring. After sensitization, the suppression of OVA-specific IgE and IgG1 was complete in juvenile offspring but waned with age concurrently with maternal IgG1 levels. Cytokine secretion, lung inflammation, and B cell priming were not suppressed although IgE responses were.
High compared with low levels of maternal IgG1 were associated with lower TH2 antibody production after adult offspring were re-exposed to OVA. Thus, offspring allergy-related responses appeared to be shaped by maternal antibody levels.
Earlier studies from our group have established that about 47% cases of autoimmune ovarian failure are due to presence of autoantibodies to Heat Shock Protein 90 (HSP90). However, there are no reports correlating pathological effects of HSP90 autoantibodies leading to ovarian failure.
Antibodies to HSP90 in female mouse model were generated by active immunization with an immunodominant peptide of HSP90, followed by detailed analysis of several reproductive parameters.
Estrous cyclicity remains unchanged; however, there was a significant drop in the fertility index due to an increase in pre- and post-implantation loss, associated with an increased incidence of degenerated eggs and embryos. The ovaries showed an increase in the number of empty and degenerated follicles and extensive granulosa cell deaths, which was reflected by the decrease in the levels of Nobox and Gja1 gene expression.
This study underlines a critical role played by HSP90 in ovarian folliculogenesis and highlights the implications of the presence of anti-HSP90 antibodies in infertile women.
Specific pro-inflammatory cytokine profiles in plasma may characterize women with recurrent miscarriage (RM) but the dynamics of the cytokine profiles with progressing pregnancy is largely unknown.
Plasma was repeatedly sampled in the first trimester from 47 RM patients. The concentrations of five cytokines including tumour necrosis factor alpha (TNF-α) were measured. TNF-α levels were correlated to carriage of five TNFA promoter polymorphisms.
TNF-α levels increased (P = 0.014) with progressing pregnancy, with higher levels in secondary than primary RM (P = 0.042) but with no significant impact on outcome. Carriage of TNFA -863C and TNFA -1031T was associated with higher TNF-α levels, and the former was found more often in secondary than primary RM (P < 0.02).
Plasma TNF-α levels increase during early pregnancy in RM women regardless of outcome, but are higher in secondary than primary RM, which may be partly genetically determined.
Dysregulated natural killer (NK) immunity and T-cell immunity are associated with recurrent pregnancy loss (RPL). We aim to define clinically relevant NK and T-cell parameters for RPL and determine their cutoff values.
Ninety-five women with RPL (>3) including 42 idiopathic and 53 known-etiology RPL, and 29 age-matched fertile controls were enrolled. Peripheral blood immunophenotype, NK cell cytotoxicity (NKC), and T-helper (Th) 1 and Th2 cytokine producing cell ratios (Th1/Th2) were measured using flowcytometry. The cutoff values were determined using Youden's J with likelihood ratio (LR) >2.
Natural killer cell percentage and NKC, TNF-α+ Th1 cells, and TNF-α/IL-10 producing Th1/Th2 cell ratio were significantly higher in idiopathic RPL than controls. By the area under the curve (AUC) analysis, NK cell percentage (AUC = 0.691), NKC (AUC = 0.649), TNF-α+ Th1 cells (AUC = 0.681) and Th1/Th2 cell ratio (AUC = 0.660) were highly specific for RPL. The cutoff values for NK cell percentage, NKC (E:T cell ratio 25:1), and TNF-α/IL-10 producing Th1/Th2 cell ratio are 16.1, 23.8, and 36.2%, respectively. Seventy-six percent of idiopathic RPL showed at least one of more immune abnormalities by these criteria.
Differences in NK cell percentages, NKC and Th1/Th2 cell ratio differentiated RPL from fertile controls.
Endometriosis affects 5–10% of women and is characterized by the growth of endometrial tissue outside of the uterus. Establishing new blood supply is a fundamental requirement for endometriosis lesion growth. Endothelial progenitor cells (EPCs), recruited by stromal cell–derived factor-1 (SDF-1), contribute to neoangiogenesis in endometriotic lesions. We hypothesized that SDF-1 is central to the neoangiogenesis and survival of endometriotic lesions, and blocking of SDF-1 will reduce vascularization of lesions in a mouse model.
Using immunohistochemistry, we evaluated SDF-1 and CD34+ EPCs in human endometriotic lesions and normal endometrium samples. EPCs were co-localized using CD34 and VEGFR2. Effects of SDF-1 blocking on endometriotic lesion survival were assessed in BALB/c-Rag2−/−/IL2rγ−/− mice engrafted with human endometrium and treated with SDF-1-blocking antibody or an isotype control. Weekly blood samples from experimental mice were analyzed for cytokines and EPCs.
SDF-1 and CD34+ EPCs were abundant in human endometriotic lesions compared with eutopic endometrium. In our mouse model, SDF-1-blocking antibody reduced CD31+ microvessels compared with isotype control.
Blocking SDF-1 reduces neovascularization and survival of lesions in a mouse model of endometriosis.
Aim of our study was to investigate whether TIFI, a syntetic peptide able to compete with anti-phospholipid antibodies (aPL) in the binding to endothelium, may restore aPL-inhibited endometrial angiogenesis.
The protective role of TIFI was evaluated on: i) aPL–inhibited of human endometrial endothelial cells (HEEC) angiogenesis in vitro; ii) aPL–inhibited vascular endothelial growth factor (VEGF) and metalloproteases (MMPs) expression; iii) aPL-inhibited Nuclear Factor-κB (NF-κB) and Extracellular signal–Regulated Kinase (ERK) activation and (iv) angiogenesis in vivo.
TIFI restores in a dose-dependent manner: i) aPL-mediated inhibition of HEEC angiogenesis in vitro and in vivo (P < 0.05), ii) VEGF (P < 0.001) and MMP–2 (P < 0.05) expression and iii) NF-κB DNA binding and ERK–1/2 activation (P < 0.05) inhibited by aPL.
Our results show for the first time the protective effects of TIFI, as represented by its ability to interfere with aPL mediated anti-angiogenic activity.
Peripheral counts of CD16+ NK cells have been well characterized in reproductive failure. However, not enough case-control clinical studies have been conducted to establish normal or abnormal CD16+/− values in the endometrium.
Peripheral and endometrial NK cell counts by FACS, IL-6, and VEGF cytokines levels by ELISA were characterized in fertile women and unexplained infertility patients with implantation failures (UI-IF) during implantation window. ROC and correlation analysis were performed.
Receiver Operating Characteristic(ROC) analysis revealed endometrial CD16+ NK cells, IL-6, and VEGF as good diagnostic parameters for unexplained infertility. Almost half of UI-FI patients showed increased total and CD16+ NK cell counts correlating with decreased levels of endometrial IL-6 and VEGF. No correlation was found with peripheral blood values.
Increased CD16+ NK cells were associated with IL-6 and VEGF deficiency in a high proportion of UI-IF patients. Testing for these immunomarkers could be a potential tool in infertility diagnosis.
The type I interferon (IFN), IFN-tau (τ), is the primary embryonic signal for pregnancy maintenance in ruminants. This study determined the effects of heat shock upon IFN-τ (IFNT) gene expression by bovine blastocysts in vitro.
In vitro-produced blastocyst-stage embryos were exposed to 42°C for 4 hr, and mRNA for heat-shock protein 70 (HSP70) and IFNT quantified.
Heat shock increased both HSP70 and IFNT expression. There was a significant correlation between HSP70 and IFNT transcript levels irrespective of whether a blastocyst had been exposed to heat shock or not.
The increase in IFNT as a result of heat shock suggests that a proportion of the variation in IFNT expression observed in blastocyst-stage embryos is a response to stress.
Human endometrial endothelial cell (HEEC) innate immunity remains poorly characterized. Based on their direct contact with the circulation, HEECs are uniquely positioned to be exposed to viral infections. This study evaluated the innate immune response generated by HEECs after exposure to the TLR3 agonist, Poly(I:C) and the TLR8 agonist, viral ssRNA.
HEECs were treated with or without Poly(I:C) or ssRNA. Culture supernatants were measured for cytokines by multiplex analysis. RNA was analyzed by qRT-PCR for type I interferons and antiviral factors.
Treatment of HEECs with Poly(I:C) rapidly upregulated the secretion of IL-2, IL-6, IL-8, IFN-γ, G-CSF, GM-CSF, MCP-1, MIP-1β, RANTES, and GRO-α after 12 hr, while ssRNA treatment induced the slower secretion of IL-6, IL-8, IFN-γ, G-CSF, VEGF, and GRO-α after 24 hr. Both viral components induced HEEC IFN-α and IFN-β expression. While treatment with Poly(I:C) induced APOBEC3G and OAS expression, treatment with ssRNA upregulated APOBEC3G and M×A mRNA.
Our findings demonstrate that HEECs can differentially sense and respond to viral components by generating distinct inflammatory and antiviral immune responses, indicating that these cells likely play an active role in the immune protection of the uterus toward viral infections.
It is unknown whether viral replication or viral components that subvert innate responses in other cells, specifically the virion host shutoff (VHS) protein, play a role in determining primary genital epithelial cell (GEC) innate antiviral responses.
Cultures of primary female GECs were exposed to wildtype (WT), VHS-deleted (vhsB), or UV-inactivated HSV-2. Antiviral pathway induction was evaluated by measuring nuclear factor-κB (NFκB) translocation by immunofluorescent microscopy. Proinflammatory cytokines, chemokines, and interferon (IFN) were measured by Luminex or ELISA. Biological activity of IFN-β was evaluated via VSV-GFP bioassay, by blocking secreted IFN-β with neutralizing antibodies and by measuring interferon-stimulated genes by RT-PCR.
Proinflammatory cytokines and chemokines were upregulated in primary GECs in response to replication-competent HSV-2, but suppressed in the presence of the VHS protein. In contrast, upregulation of IFN-β depended on viral replication, but was not affected by VHS. However, the IFN-β produced was biologically active and reduced the viral burden.
Viral factors such as replication and the presence of the VHS protein play important roles in regulating innate antiviral responses against HSV-2 from primary GECs.
Macrophage function has many implications in a variety of diseases. Understanding their biology becomes imperative when trying to elucidate immune cell interactions with their environment, and in vitro cell lines allow researchers to manipulate these interactions. A common cell line used is THP-1, a promyeloid cell line suggestive to outside factors, and therefore sensitive to culture conditions. In this study, we describe how culture conditions can alter THP-1 morphology and in turn affect their response to differentiation stimuli.
THP-1 cells were cultured in two conditions and treated with phorbol 12-myristate 13-acetate (PMA) or MCSF. CD14 surface expression was determined by flow cytometry and cytokine/chemokine production determined by multiplex analysis.
Culture conditions of THP-1 affect their response to PMA. Highly confluent THP-1 cells differentiate into a heterogeneous population responsive to PMA as seen by an increase in CD14 expression. However, these cells, cultured in low confluence, remain as a homogenous population and do not gain CD14. Additionally, there are major differences in the constitutive cytokine profile.
We demonstrate that the culture conditions of THP-1 cells can alter their response PMA. This suggests that culture techniques may account for the discrepancy in the literature of both basal THP-1 phenotype and their response to PMA.
Uterine leiomyomata are the most common reproductive tumor in women, and their cause is not known.
Plasma samples from 155 women (74 with and 81 without ultrasound-confirmed leiomyoma) from a new study of leiomyoma risk factors in the Detroit, Michigan area, were examined for any cross-sectional associations between commonly examined cytokines and leiomyoma presence.
Associations varied by season of sample collection defined a priori as winter (December–February) and non-winter seasons. In the winter months, interleukin (IL)13 and IL17 were positively and IP10 was inversely associated with having a leiomyoma. In the non-winter samples, VEGF, G-CSF, and IP10 were positively associated and Monocyte chemotactic protein-1, IL13, and IL17 were inversely associated with having a leiomyoma. Associations were not changed by adjustment for age or BMI.
These data suggest that new insight into leiomyoma formation may be acquired through investigation of the immune system.
Chronic histiocytic intervillositis (CHIV) is a rare type of placental pathology that is associated with reproductive loss at all gestational ages. The aim of the study was to investigate the relationship between the severity of CHIV and the outcome of pregnancy and to compare the immune response between CHIV patients and controls to explore an immunological origin of CHIV.
Microscopic slides were reviewed and scored according to a previously published grading system in 30 pregnancies of 22 CHIV patients. Partner-specific mixed lymphocyte reactions, cytotoxic T-lymphocyte precursor frequencies (CTLpf), and anti-HLA antibodies were determined in four patients and seven controls.
Higher CHIV scores are associated with worse pregnancy outcome. CHIV patients demonstrated a higher CTLpf against their partner compared to non-complicated pregnancies (P = 0.03). The CTLpf was extremely high in 75% of the patients. Antipaternal HLA antibodies were only present in 75% of the CHIV patients compared to none of the controls (P = 0.02).
CHIV scores seem to be associated with the severity of adverse pregnancy outcome. High antipaternal cellular (T-cell) and humoral (B-cell) response to partner-specific CTLpf and the presence of anti-HLA antibodies directed to the partner suggest an immunologic origin of CHIV.
The tumor microenvironment is made up of tissue that is responsible for the growth and progression of the tumor as well as its ability to initiate metastases. The cancer cells on the front of the tumor together with the macrophages and fibroblasts help to constitute the aggressive phenotype of the tumor. The presence of this aggressive phenotype is indicated by the local infiltration of cancer cells and by the development of lymph node metastases. In cases of uterine cancer, the extent of the local and distant spread of the disease is crucial for determining the type of therapeutic strategy to be applied – surgery alone, surgery followed by radio-chemotherapy, or radio-chemotherapy alone. In the interest of trying to improve the patient's quality of life, different studies supporting the therapeutic model of surgery alone have been conducted. While the cancer cells on the tumor front together with the macrophages and the fibroblasts help to constitute the aggressive phenotype of the tumor, metallothionein (MT) has been shown to have both pro-proliferative and anti-apoptotic activities and to participate in microenvironment remodeling. The aim of the current study was to determine the levels of MT immunoreactivity in the uterine cervical cancer cells as well as in the stromal fibroblasts and macrophages of the tumor microenvironment with respect to the depth of the local invasion and the extent of the distant metastases, so that its potential predictive value as a therapeutic strategy for cervical cancer can be ascertained.
We determined the levels of immunoreactivity of MT in a total of 57 carcinoma tissue samples (in the tumor front, in its central part, and in the macrophages and fibroblasts present within the tumor microenvironment). The patients from whom the samples derived were divided into groups with respect to the presence of lymph node metastases (distant spread) and to the depth of invasion (local spread) in relation to the FIGO stage.
Metallothionein immunoreactivity was observed in uterine cervical cancer cells; it was also identified in the fibroblasts and macrophages found within the microenvironments of the tumors of patients suffering from the disease. The MT immunoreactivity level significantly increased within the whole cancer nest in relation to the FIGO stage (intensity of the local spread of the disease). Similarly, the infiltration of MT-positive CAFs and TAMs statistically significantly increased in relation to the FIGO stage.
The level of MT immunoreactivity found in the fibroblasts and macrophages within the tumor microenvironment seems to be indicative of the intensity of the remodeled cervical tumor microenvironment, and this in turn may be related to the local advancement of the disease. Moreover, it appears that the intensity of the metallothionein immunoreactivity in the immunoreactivity profile of the cervical tumor may be linked to both the depth of the local invasion and the extent of the distant advancement of the disease.
Magnesium sulfate (MgSO4) exposure reduces the risk of cerebral palsy. As neonatal inflammatory cytokine levels strongly correlate with neurologic outcome, we hypothesize that MgSO4 decreases inflammatory cytokine production.
We assessed the effect of MgSO4 on cellular magnesium levels, cytokine production, and release within THP-1 and cord blood mononuclear cells.
MgSO4 exposure increased intracellular magnesium levels, reducing the frequency of THP-1 cells producing IL-1β, IL-8, and TNF-α following LPS stimulation. Significant reductions in the frequency of neonatal monocytes producing TNF-α (48%) and IL-6 (37%) were seen following LPS stimulation, and MgSO4 also significantly decreased the frequency of monocytes producing TNF-α (35%) under basal conditions. Decreased cytokine production was confirmed via ELISA, demonstrating a sustained effect and dose response. Magnesium also reduced cytokine production following stimulation with TLR ligands representing obstetrical infections (group B Streptococcus and Mycoplasma) and in preterm neonatal monocytes.
MgSO4 increases intracellular magnesium, reducing inflammatory cytokine production and release, potentially elucidating the mechanism by which MgSO4 prevents cerebral palsy, eclampsia, and preterm birth.
This study aimed to comprehensively describe inflammatory responses to trivalent influenza virus vaccine (TIV) among pregnant women and determine whether responses differ compared to non-pregnancy.
Twenty-eight pregnant and 28 non-pregnant women were vaccinated. Serum cytokines were measured at baseline, and 1, 2, and 3 days post-vaccination. Anti-influenza antibody titers were measured at baseline and 1 month post-vaccination.
Overall, following vaccination, tumor necrosis factor (TNF)-α and interleukin(IL)-6 increased significantly, peaking at 1 day post-vaccination (P's < 0.001). Pregnant versus non-pregnant women showed no differences in IL-6, TNF-α, or IL-1β responses. Pregnant women showed no change in IL-8 and increases in migration inhibitory factor (MIF), while non-pregnant showed decreases in both. Pregnancy did not significantly alter antibody responses.
Inflammatory responses to TIV are mild, transient, and generally similar in pregnant and non-pregnant women. Given the variability evidenced, vaccination may provide a useful model for studying individual differences in inflammatory response propensity.
Preterm birth is frequently caused by intrauterine infection and inflammation. Recent studies have demonstrated that carbon monoxide (CO), which is produced endogenously, has potent anti-inflammatory properties. Whether or not CO can prevent infection-mediated preterm birth is unknown.
Mice were assigned to one of four groups: sham infection, sham infection + CO, infection, or infection + CO. Infections were established by intra-uterine injection of Escherichia coli on day 14 of pregnancy. Animals received daily i.p. injections of 1 mL CO-saturated lactated ringers solution (LRS) or LRS alone beginning on the morning of surgery. Gestational age at delivery and litter characteristics was noted. In second experiment, animals were sacrificed 24 hrs post-surgery and tissues were harvested for cytokine analyses.
Escherichia coli intrauterine infection increased the number of animals delivering preterm. This effect was significantly ameliorated by CO-LRS. CO-treatment also increased litter size and weights of the surviving offspring. Cytokines in the amniotic fluid and the placenta were increased by E. coli exposure, but CO had no detectible effect on E. coli-stimulated cytokine production. No effects of CO were detected in sham-infected animals.
Supplemental CO improves pregnancy outcome after intrauterine infection and may function at a point downstream of, or through pathways independent of, induction of proinflammatory cytokines.
Plasminogen activator inhibitor type 1 (PAI-1) regulates fibrinolysis, and the common promoter region variants −675G/A (4G/5G) and −844G/A are associated with increased thrombotic risk. Despite evidence linking altered fibrinolysis with adverse pregnancy events, including idiopathic recurrent pregnancy loss (RPL), the contribution of PAI-1 variants to RPL risk remains controversial. We investigated the association between the PAI-1 −844G/A and 4G/5G (−675G/A) variants with altered risk of RPL.
This was a case–control study involving 304 women with confirmed RPL and 371 age- and ethnically matched control women. PAI-1 genotyping was performed by PCR single-specific primer −675 (G/A) and real-time PCR (−844G/A) analysis.
Minor allele frequency (MAF) of 4G/5G (P < 0.001), but not −844G/A (P = 0.507), was higher in RPL cases. PAI-1 4G/5G single-nucleotide polymorphism (SNP) was significantly associated with RPL under additive, dominant, and recessive genetic models; no association of −844G/A with RPL was seen irrespective of the genetic model tested. Taking common −844G/5G haplotype as reference (OR = 1.00), multivariate analysis confirmed the association of 4G-containing −844A/4G (P < 0.001) and −844G/4G (P = 0.011) haplotypes with increased RPL risk.
4G/5G, but not −844G/A, PAI-1 variant is associated with an increased risk of RPL.
The immune modulatory effect of heme oxygenase-1 (HO-1) is well documented in studies about sepsis and transplantation. This work evaluates the influence of HO-1 on the innate immune response during pregnancy.
Human first-trimester trophoblasts derived from normal pregnancies or spontaneous abortions were analyzed for their basal HO-1, BCL-associated athanogene-1 (Bag-1), and cytokine production before and after LPS treatment. In vivo, pregnant Hmox1+/+ and Hmox1+/− female mice were treated with LPS, and the production of Bag-1 was evaluated.
Human trophoblasts up-regulated the expression of both HO-1 and pro-inflammatory cytokines after LPS treatment, whereas the basal level of HO-1 was higher in normal pregnancies. In vivo, HO-1 deficiency provoked diminished Bag-1 level upon LPS treatment.
HO-1 deficiency causes an inflammatory immune reaction and diminished expression of protective molecules in trophoblasts. Thus, HO-1 emerges as one important modulator of innate immune responses in pregnancy.
Thyroid disease is one of the most common endocrine conditions affecting women during reproductive age. A link between thyroid and assisted reproduction outcome is debated.
Serum TSH levels, number and scoring of oocytes and embryos, and number of clinical pregnancies were retrospectively recorded in 164 women undergoing assisted reproduction technologies (ART) at an University–based fertility center, to evaluate the outcome of the first steps of assisted reproduction (ovarian stimulation, oocyte pickup and fertilization, embryo transfer and implantation) in relation to thyroid function and autoimmunity.
No significant relationship was found between TSH and all parameters, except clinical pregnancy rate (22.3% in TSH ≤ 2.5 group versus 8.9% in TSH > 2.5 mUI/L group; P = 0.045). No pregnancy occurred in women with anti-thyroperoxidase autoantibodies, while pregnancy occurred in 23.9% of cycles without autoimmunity (P = 0.02).
Further studies must be conducted in order to shed light on the link between infertility and thyroid dysfunction.
During pregnancy, antibodies are induced that target the paternal human leukocyte antigens of the semi-allogeneic fetus. The level and presence of these antibodies have been reported increased as well as decreased for a variety of pregnancy complications; the clinical relevance and consequences of these antibodies are not very clear. Therefore, the objective of this review is to determine whether the presence of antipaternal antibodies influences pregnancy outcome.
We performed a systematic search of studies that described the effect of antipaternal antibodies on pregnancy complications. The primary outcome was the risk ratio for HLA class I and class II antibodies on pregnancy complications. Furthermore, we calculated the risk for first- and third-trimester complications.
The seventeen studies that were selected for meta-analysis showed high level of statistical and clinical heterogeneity. In the meta-analysis, we found no significant effect of HLA class I or class II antibodies on pregnancy outcome.
No consistent conclusions can be drawn from the meta-analysis. Discrepancies in the meta-analysis are the result of different screening techniques, varying time points of screening, and use of incorrect control groups. Furthermore, more detailed analyses of the characteristics and specificity of the antibodies involved are essential.
To explore whether cervical carcinomas cells-derived thymic stromal lymphopoietin (TSLP) modulates the biologic behavior of vascular endothelial cells and herein participates in the angiogenesis in the cervical cancer pathogenesis.
We analyzed expression of TSLP and its receptor (TSLPR) in cervical cancer cells by immunohistochemistry, ELISA, and flow cytometry, respectively. We further investigated the effects of TSLP on the proliferation, apoptosis, activation, and angiogenesis in vitro of human umbilical vein endothelial cells (HUVECs).
It has been found that the cervical cancer cells translate TSLP and endothelial cells express TSLPR in cervical cancer tissues. Both HeLa and CaSki cells secret TSLP in a time-dependent manner, and the ratio of TSLPR-positive HUVECs is about 30%. It has been showed that recombinant human TSLP (rhTSLP) can significantly increase Ki67 and CD62E expression in HUVECs and interleukin-6 (IL-6) levels from HeLa and CaSki cells; on the contrary, anti-human TSLP or TSLPR neutralizing antibody down-regulates the expression of Ki67, angiogenesis-relative molecules CD62E, and CD105 in HUVECs cocultured with HeLa or CasKi cells and inhibits IL-6 secretion from HeLa and CaSki cells. Moreover, both rhTSLP and endogenous TSLP from HeLa or CaSki cells obviously stimulate the proliferation, activation, and angiogenesis, but not influence the apoptosis of HUVECs in vitro.
This study has demonstrated that TSLP secreted by cervical carcinomas cells is involved in the angiogenesis of cervical cancer in a paracrine manner.
The renin–angiotensin system is associated with angiogenesis, tissue remodeling, prenatal development, and Th2 cytokine production. The purpose of this study was to evaluate whether polymorphisms in angiotensin I-converting enzyme [ACE insertion/deletion (I/D)], angiotensinogen (AGT M235T), and angiotensin II type 1 receptor (AT1R 1166A>C) affect the prevalence of spontaneously aborted fetuses (SAFs).
One hundred and ninety-eight SAFs were <20 weeks of gestational age. The control subjects were 103 healthy children and 640 adults collected from a convenience sample. Polymerase chain reaction and restriction fragment length polymorphism analysis were performed to identify the ACE I/D, AGT M235T, and AT1R 1166A>C genotypes.
II/MM/AA, II/MT/AA, and II/TT/AC of ACE/AGT/AT1R were significantly different from controls. In particular, the statistical significance of the II/MM/AA genotype remained strong in chromosomally normal SAFs.
Our data suggest that the II/MM/AA of ACE/AGT/AT1R is a possible predisposing factor for spontaneous abortion.
Natural killer (NK) cells play a key role in embryo implantation and pregnancy success, whereas blood and uterine NK expansions have been involved in the pathophysiology of reproductive failure (RF). Our main goal was to design in a large observational study a tree-model decision for interpretation of risk factors for RF.
A hierarchical multivariate decision model based on a classification and regression tree was developed. NK and NKT-like cell subsets were analyzed by flow cytometry.
By multivariate analysis, blood NK cells expansion was an independent risk factor for RF (both recurrent miscarriages and implantation failures). We propose a new decision-tree model for the risk interpretation of women with RF based on a combination of main risk factors.
Women with age above 35 years and >13% CD56+CD16+ NK cells showed the highest risk of further pregnancy loss (100%).
To determine the presence of antinuclear antibodies (ANAs) in follicular fluid (FF) and embryos as to their potential impact on IVF outcome.
A total of 100 women presenting to IVF/ICSI program were recruited in this study, wherein 50 women sero-positive for ANA served as study group (ANA+ group) and 50 women seronegative for ANA served as control group (ANA- group), and patients were age-matched in the two groups. ANA level in serum and FF were examined by ELISA, and immunofluorescence assay was employed to detect IgG within embryos.
In ANA+ group, 36 women showed positive ANA both in serum and FF on the day of ovum pick-up (OPU), wherein 34 exhibited fluorescence of IgG in embryos. All the 50 women in control group showed negative results in serum and FF ANA on OPU day and in embryo immunofluorescence. Serum ANA level positively correlated with FF ANA level. IVF outcomes were significantly poorer in ANA+ group, mainly reflecting the less number of high-quality embryos, reduced rates of pregnancy, clinical pregnancy, and implantation.
These data demonstrated the existence of ANA in FF and embryos, which may exert detrimental impact on IVF outcome.
Pre-eclampsia (PE), a pregnancy complication of unknown etiology, is a major cause of maternal and fetal mortality and morbidity. Previous studies have described placental genes that are up-regulated in expression in PE, but few studies have addressed placental gene suppression in this syndrome.
Gene profiling and quantitative reverse transcription PCR (qRTPCR) analyses were used to identify genes down-regulated in placentas from women with severe preterm PE compared with gestational age-matched normotensive controls with spontaneous preterm birth (sPTB). Western blotting and immunohistochemistry were used to evaluate levels and patterns of cell type–specific protein expression in PE and sPTB group placentas.
Levels of macrophage marker [folate receptor (FR)-β, CD163, and CD68] mRNA and FR-β protein were significantly down-regulated in PE group placentas compared with the sPTB group. Numbers of Hofbauer cells (HBCs, fetal macrophages) and FR-β protein in these cells were reduced in PE group placentas.
Severe PE is associated with decreased placental expression of FR-β and a reduction in the number of HBCs. Reduced placental macrophage function is likely to play a key role in the pathophysiology of PE.
Inflammation and genital infections promote the increase in leukocytes, pro-inflammatory cytokines, and oxygen reactive species, impairing sperm functions such as motility, capacitation, and acrosome reaction. All these functions are primarily regulated by cytoplasmic concentration of Ca2+ ([Ca2+]cyto). This study evaluated the effect of tumor necrosis factor (TNF)-α on the [Ca2+]cyto and its regulation in human sperm.
Sperm loaded with fura-2 were incubated with or without TNF-α (0–500 pg/mL) from 0 to 120 min. After incubation, the basal [Ca2+]cyto and membrane permeability to Ca2+ were evaluated by spectrofluorometry, before and after Ca2+ addition to the extracellular medium.
Without TNF-α, the addition of Ca2+ promotes an transitory increase in [Ca2+]cyto in the spermatozoa, that returns in a few minutes to a basal level, indicating calcium regulation activation. TNF-α decreases the Ca2+ permeation and increases the basal level of [Ca2+]cyto after a Ca2+ pulse (P < 0.04); affecting calcium regulation in a way that is time and concentration dependent. TNF-α effect was partially prevented by the addition of an antioxidant (butylated hydroxytoluene) (P < 0.03).
Tumor necrosis factor-α decreases membrane permeability to Ca2+ and affects Ca2+ regulation in sperm cells in vitro, probably via lipid peroxidation, which may explain the decrease in sperm fertilizing capacity during inflammatory and infectious processes.
The decidual NK (dNK) cell is called on to support placental growth by providing an array of growth factors that directly transform the spiral artery and direct trophoblast invasion. Successful transformation is dependent upon adequate stimulation paradoxically stimulating the cell for placental support rather than cytotoxicity. With the identification of its supportive role, the presence of an intact cytotoxic mechanism has been confusing. Investigators have found that the cell remains fully capable of cytotoxic responses particularly in response to pathogen-specific signals. We postulate a dual threshold model where moderate stimulation results in release of stimulatory factors supporting placentation while intense stimulation, particularly triggered through pathogen-specific receptors, restores the cell to its protective, cytotoxic, role. Individual dNK cells mature attaining the capacity to respond to the delivery of cognate signals. The process, known as ‘licensing’ tunes responsiveness to the degree to which stochastically selected inhibitory receptors block cytotoxic response to self. A changing licensing milieu within the decidua may result in altered and unsuitable receptor expression. We postulate that a heterogeneous population of dNK cells where cells inappropriately licensed for the milieu contributes to pathology.
]]>The object of the study was to investigate the costimulatory phenotype of spleen antigen-presenting cells (APCs) in mice after mating and the influence of costimulatory signal blocking on pregnancy outcome, cytokine production, and Treg cell concentration.
The levels of CD40, CD80, and CD86 on spleen APCs at day 0.5 and 3.5 after mating (C57BL/6Jx DBA/2J and C57BL/6JxBalb/c) were assessed by flow cytometry and RT-PCR. Blocking antibodies against costimulatory molecules were given i.p. at day 3.5 after mating. Pregnancy outcomes, blood cytokine (ELISA), and spleen Treg (flow cytometry) concentrations were examined at day 10.5 after mating.
Differential expression of costimulatory molecules on spleen APCs of mated v. pseudopregnant mice was observed mainly at day 3.5 after conception. Administration of anti-CD86 antibody lowered pregnancy outcome. Cytokine expression was modulated after administration of anti-CD86, anti-CD80 and anti-CD40 antibodies, whereas only anti-CD40 antibody changed the concentration of Treg lymphocytes and the level of Foxp3 protein expression.
Costimulatory phenotype of female spleen APCs is distinctly modulated after mating. Alteration of costimulatory signal derived from APCs during pre-implantation period of pregnancy may have an adverse effect on pregnancy outcome and the tolerogenic immune response.
IL-22 has important functions at mucosal surfaces, including the induction of antimicrobial peptides and maintenance of epithelium. However, IL-22 has not been investigated in the genital tract during TV infection.
Women who visited an STD clinic and women from a cohort with frequent Trichomoniasis were studied. IL-22, IL-17, and antimicrobial peptides were measured in cervicovaginal lavage by ELISA.
In women visiting the STD clinic, those without STDs (n = 10) had a median IL-22 of 0 pg/mL, while women with infections (n = 30) had 27 pg/mL (P = 0.04). In the cohort, women with Trichomoniasis (n = 19) had significantly higher IL-22 than women with no infections (n = 21, 74 versus 0 pg/mL, P = 0.0001). IL-17 was also significantly increased in Trichomoniasis, and there was a correlation between IL-22 and IL-17 (P = 0.001).
IL-22 is increased in STDs generally and in Trichomoniasis specifically suggesting an antimicrobial response of the mucosa and an epithelial repair process induced by the STDs.
This study compares the frequencies of plasminogen activator inhibitor-1 (-675) 4G/5G polymorphism and its relationship with eight antiphospholipid antibodies (aPLs) in serum of 157 patients with repeated pregnancy loss (RPL).
PAI-1 (-675) 4G/5G polymorphism was determined using standard PCR-RFLP method. Enzyme-linked immunosorbent assay was used for the detection of aPLs against ph-serine, ph-ethanolamine, ph-inositol, ph-DL-glycerol, phosphatidic acid, annexin V, cardiolipin, and beta2-GPI. Allelic frequency and distribution of genotypes were calculated. The prevalence of the risk conferring 4G allele and 4G/4G homozygous genotype in patients and controls was compared, and the correlation between aPLs positivity and PAI-1 4G/4G genotype was tested by chi-square test.
Statistically highly significant correlation between RPL and PAI-1 (-675) 4G/4G genotype was found. No correlation between PAI-1 (-675) 4G/5G polymorphism and the presence of antiphospholipid antibodies in RPL patients was observed.
PAI-1 (-675) 4G/4G homozygous genotype increases the risk of RPL independently from the aPLs positivity.
To overcome availability of the purified native zona pellucida (ZP) glycoproteins for immunocontraception, porcine ZP3, and ZP4 were expressed in E. coli.
Purified recombinant proteins were characterized by SDS-PAGE and Western blot, and immunogenicity and contraceptive efficacy determined in FvB/J female mice.
Purified ZP3, ZP3 with promiscuous T-cell epitope of tetanus toxoid, ZP4 and ZP4 incorporating promiscuous T-cell epitope of bovine RNase revealed ~44-, ~49-, ~53-, and ~55-kDa bands by SDS-PAGE and Western blot, respectively. Immunization of female mice with recombinant proteins elicited high antibody titers as well as T-cell responses. Immune sera recognized mouse oocyte ZP and also inhibited in vitro fertilization. Immunized mice showed significant decrease in fertility. Recombinant proteins were able to recall memory antibody response in female mice primed with porcine native ZP.
Availability of recombinant porcine proteins will be useful in the development of contraceptive vaccine.
Successful embryonic implantation requires an appropriate communication network between the embryo and its near environment within the implantation site. Herein, we examined whether human chorionic gonadotropin (hCG), the major embryonic signal, targets endothelial cells and regulate their responsiveness to interleukin 1 (IL1), one of the earliest signals released by embryonic cells.
Human microvascular endothelial cell proliferation and migration following exposure to various concentrations of hCG and/or IL1B for different time periods were analyzed by BrdU incorporation and wound healing assays. The expression of soluble (s) and membrane-bound (mb) IL1 receptors (IL1Rs), IL1R antagonist (IL1RN), luteinizing hormone/choriogonadotropin receptor (LHCGR), and IL8 was determined by real-time PCR, Western blot, and ELISA.
Cell proliferation and migration increased in response to IL1B and further in the presence of hCG. IL1B up-regulated both the signaling IL1R1 and the inhibitory IL1R2, while adding hCG further increased IL1R1 and significantly downregulated IL1R2. This translated into an increased secretion of IL8, which was inhibited in cells where IL1R2 was overexpressed.
These findings reveal a new mechanism by which hCG may target endothelial cells to directly stimulate angiogenesis and favor embryonic growth.
Dairy cattle suffer from high percentages of early embryonic loss, and therefore, it is critical to study the function of the uterus at this time. We hypothesize that the antiviral protein, myxovirus resistance (MX)1, regulates secretion in uterine glandular cells during early pregnancy.
Uterine epithelial cells were used to study uterine function, in vitro. Sucrose gradients, Western blotting, and transmission electron microscopy were used to isolate and identify exosomes. Immunofluorescence and ceramide inhibitors were used for the characterization of exosomes.
Myxovirus resistance1 was associated with exosomes and protected from proteases, indicating it was inside exosomes. MX1 partially colocalized with exosomal protein CD63, and a ceramide inhibitor reduced numbers of MX1-associated exosomes.
This study is the first to characterize MX1-associated exosomes, and we postulate that MX1 regulates secretion in epithelial cells by playing a role in exosome formation or trafficking.
The American Society for Reproductive Immunology's Second Clinical Reproductive Immunology Symposium: ‘Using Reproductive Immunology to Improve Clinical Practices’ was held on October 27th to 28th 2012, in New Haven, Connecticut. The objective of this symposium was to disseminate up-to-date clinical reproductive immunology information and how it relates to current clinical practices, diagnostics, and therapeutics for women's reproductive and pregnancy-related issues, with a view to bridging the distance between clinicians and translational researchers. It provided a common platform to physicians and physician scientists to facilitate the application of research outcomes into current medical practice. This symposium highlighted the current status of health issues during pregnancy including preeclampsia, preterm birth, maternal obesity and diabetes, autoimmunity and infection. In addition, a session was dedicated toward demystifying laboratory techniques of reproductive immunology for clinicians. This symposium was attended by approximately 50 delegates along with the 16 invited speakers and included 17 presentations that addressed key issues in the immunology of maternal–fetal well-being. This report will summarize the highlights of the meeting.
]]>Regulatory T cells (Tregs) have been identified as immunomodulatory cells, which induce tolerance. Evidence from numerous recent studies implicates these cells as critical for maternal tolerance to the fetal-derived placenta, proper functioning of which is required for normal gestation and growth and development of the fetus in utero. This review focuses on the recent studies of Tregs at the maternal–fetal interface in pregnancy complications, as well as their identification in specific inflammatory lesions in the placenta.
]]>Antithrombin (AT) replacement has been described in patients with hereditary AT deficiency undergoing delivery; however, the kinetics of AT replacement in preeclampsia is not adequately understood. Therefore, the Prospective Randomized Evaluation of the Safety and Efficacy of Recombinant Antithrombin in Very Preterm Preeclampsia (PRESERVE-1) study has been proposed.
Sixty women aged ≥18 years at 24 0/7–28 0/7 weeks' gestation and with hypertension and proteinuria will be enrolled and randomly assigned to receive recombinant human AT or placebo until fetal and/or maternal indications cause cessation of expectant management or until 34 0/7 weeks' gestation. The primary endpoint is the increase in gestational age from randomization to delivery. Safety assessments and laboratory assays will also be performed.
PRESERVE-1 study enrollment will begin during the second half of 2013.
The PRESERVE-1 study will provide further insight into the pharmacokinetic activity and safety of AT therapy in preeclampsia.
Gestational diabetes mellitus (GDM) is an inflammatory condition that involves unbalanced cytokine production. We carried out a systematic review on the relationship between GDM and maternal circulating levels of cytokines in the 2nd/3rd trimesters.
Three electronic databases (MEDLINE, EMBASE and LILACS), were searched. Duplicate study selection, extraction and quality assessment was performed.
Twenty-two studies with 1982 participants reporting levels of 9 cytokines (IL-1B, IL-2, IL-6, IL-10, IL-13, IL-18, IFN-G, TGF-B and TNF-A) were included. Most studies differed considerably in selection criteria, sampling and assay methods and in reporting their results. Consequently, only two studies could be pooled: TNF-A concentration was slightly higher in GDM than in control patients, although not significant (WMD = 0.45, 95% CI −0.34–1.23).
New studies with well-defined, more homogeneous methodological parameters are needed to detect whether there are significant differences in circulating levels of cytokines in patients with GDM.
Medical problems of most importance to reproductive health of women differ to some extent between the developed world and resource-disadvantaged countries. Nevertheless, many share a common link in microbial involvement.
A review of the peer-reviewed literature on microbiota, probiotics, and reproductive health.
Indigenous and probiotic lactobacilli express properties antagonistic to pathogens, but complementary to host immunity. These organisms are associated with conception, reducing the risk of infection, as well as potentially lowering the risk of a number of complications of pregnancy that otherwise lead to maternal and infant mortality and morbidity.
The ability to manipulate the microbiome and to improve immunity through probiotics holds much promise. The lack of improvements over the past 40 years in managing urogenital infections in women is incomprehensible. Support for innovative diagnostic and treatment options is needed, including testing and implementing probiotic therapies, especially for women with poor access to healthcare and good nutrition.
This represents an overview of the use of animal models to study the adverse pregnancy outcomes seen in humans. The purpose is to entice clinicians to utilize some of this information to seek out the literature and have more meaningful and profitable discussions with their academic colleagues and enhance transdisciplinary research in reproductive health.
]]>APS causes adverse pregnancy outcome, especially in younger patients with SLE or prior thromboses. LAC is the best predictor of adverse outcome. Prior conclusions on the efficacy of heparin for prevention of poor outcome may be suspect. New therapies are under evaluation in animal models but how to introduce them into the clinic is unclear.
Systemic lupus erythematosus (SLE) is a chronic multisystem autoimmune disorder that predominantly affects women of reproductive age. As clinical outcomes improve, pregnancy in these women is becoming more common. Although epidemiological data have documented an improvement in the prognosis of pregnancy in these women over recent years, they are still at significantly increased risk of pregnancy complications, such as miscarriage, stillbirth, pre-eclampsia and impaired foetal growth. The pathogenesis of SLE involves marked immune dysfunction, and in particular, the function of immunosuppressive elements of the immune system is impaired, including regulatory T-cell function. Because regulatory T cells are likely to be the key cell-modulating feto-maternal tolerance, this review overviews the possibility that regulatory T-cell impairments contribute to pregnancy pathology in women with SLE and contribute to the clinical challenge of managing these women during pregnancy.
]]>Antiphospholipid antibodies (aPL) have been associated with recurrent pregnancy losses (RPL) and other obstetrical complications. The diagnostic criteria for the classical antiphospholipid antibody syndrome (APS) have been utilized for the detection of obstetrical APS in women with RPL. However, laboratory findings and immunopathology of obstetrical APS are significantly different from those of classical APS. In addition, many women with RPL who have positive aPL do not have symptoms consistent with the current APS criteria. The induction of a proinflammatory immune response from trophoblasts and complement activation by aPL rather than thromboembolic changes has been reported as a major immunopathological feature of obstetrical APS. Heparin treatment has been reported to be effective in prevention of early pregnancy loss with APS but not for the late pregnancy loss or complications. The complex effects of heparin may explain the limited efficacy of heparin treatment in RPL. New diagnostic criteria for obstetrical APS are needed urgently, and new therapeutic approaches should be explored further.
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