Background:Genome-wide association studies (GWAS) have identified hundreds of loci underlying adult-onset asthma (AOA) and childhood-onset asthma (COA). However, the causal variants, regulatory elements, and effector genes at these loci are largely unknown. Methods:We performed heritability enrichment analysis to determine relevant cell types for AOA and COA, respectively. Next, we fine-mapped putative causal variants at AOA and COA loci. To improve the resolution of fine-mapping, we integrated ATAC-seq data in blood and lung cell types to annotate variants in candidate cis-regulatory elements (CREs). We then computationally prioritized candidate CREs underlying asthma risk, experimentally assessed their enhancer activity by massively parallel reporter assay (MPRA) in bronchial epithelial cells (BECs) and further validated a subset by luciferase assays. Combining chromatin interaction data and expression quantitative trait loci, we nominated genes targeted by candidate CREs and prioritized effector genes for AOA and COA. Results:Heritability enrichment analysis suggested a shared role of immune cells in the development of both AOA and COA while highlighting the distinct contribution of lung structural cells in COA. Functional fine-mapping uncovered 21 and 67 credible sets for AOA and COA, respectively, with only 16% shared between the two. Notably, one-third of the loci contained multiple credible sets. Our CRE prioritization strategy nominated 62 and 169 candidate CREs for AOA and COA, respectively. Over 60% of these candidate CREs showed open chromatin in multiple cell lineages, suggesting their potential pleiotropic effects in different cell types. Furthermore, COA candidate CREs were enriched for enhancers experimentally validated by MPRA in BECs. The prioritized effector genes included many genes involved in immune and inflammatory responses. Notably, multiple genes, including TNFSF4, a drug target undergoing clinical trials, were supported by two independent GWAS signals, indicating widespread allelic heterogeneity. Four out of six selected candidate CREs demonstrated allele-specific regulatory properties in luciferase assays in BECs. Conclusions:We present a comprehensive characterization of causal variants, regulatory elements, and effector genes underlying AOA and COA genetics. Our results supported a distinct genetic basis between AOA and COA and highlighted regulatory complexity at many GWAS loci marked by both extensive pleiotropy and allelic heterogeneity.

The mechanisms that underlie the timing of labor in humans are largely unknown. In most pregnancies, labor is initiated at term (≥ 37 weeks gestation), but in a signifiicant number of women spontaneous labor occurs preterm and is associated with increased perinatal mortality and morbidity. The objective of this study was to characterize the cells at the maternal–fetal interface (MFI) in term and preterm pregnancies in both the laboring and non-laboring state in Black women, who have among the highest preterm birth rates in the U.S. Using mass cytometry to obtain high-dimensional single-cell resolution, we identified 31 cell populations at the MFI, including 25 immune cell types and six non-immune cell types. Among the immune cells, maternal PD1 + CD8 T cell subsets were less abundant in term laboring compared to term non-laboring women. Among the non-immune cells, PD-L1 + maternal (stromal) and fetal (extravillous trophoblast) cells were less abundant in preterm laboring compared to term laboring women. Consistent with these observations, the expression of CD274 , the gene encoding PD-L1, was significantly depressed and less responsive to fetal signaling molecules in cultured mesenchymal stromal cells from the decidua of preterm compared to term women. Overall, these results suggest that the PD1/PD-L1 pathway at the MFI may perturb the delicate balance between immune tolerance and rejection and contribute to the onset of spontaneous preterm labor.

Sex-specific differences in prevalence are well documented for many common, complex diseases, especially for immune-mediated diseases, yet the precise mechanisms through which factors associated with biological sex exert their effects throughout life are not well understood. We interrogated sex-specific transcriptional responses of peripheral blood leukocytes (PBLs) to innate immune stimulation by lipopolysaccharide (LPS) in 46 male and 66 female members of the Hutterite community, who practice a communal lifestyle. We identified 1217 autosomal and 54 X-linked genes with sex-specific responses to LPS, as well as 71 autosomal and one X-linked sex-specific expression quantitative trait loci (eQTLs). Despite a similar proportion of the 15 HLA genes responding to LPS compared to all expressed autosomal genes, there was a significant over-representation of genes with sex by treatment interactions among HLA genes. We also observed an enrichment of sex-specific differentially expressed genes in response to LPS for X-linked genes compared to autosomal genes, suggesting that HLA and X-linked genes may disproportionately contribute to sex disparities in risk for immune-mediated diseases.

Human trophoblast stem cells (hTSC) can be isolated from first trimester placenta but not from term placenta. Here we demonstrate that villous cytotrophoblasts (vCTB) from term placenta can be reprogrammed into induced trophoblastic stem-like cells (iTSC) by introducing sets of transcription factors. The iTSCs express TSC markers such as GATA3, TEAD4 and ELF5, and are multipotent, validated by their differentiation into both extravillous trophoblasts (EVT) and syncytiotrophoblasts (STB) in vitro and in vivo. The iTSC can be passaged indefinitely in vitro without slowing of growth. The transcriptome profile of these cells closely resembles the profile of hTSC isolated from first trimester placentae but different from the term placental vCTB from which they originated. The ability to reprogram cells from term placenta into iTSC will allow study of early gestation events which impact placental function later in gestation, including preeclampsia and spontaneous preterm birth.

There is a life-long relationship between rhinovirus (RV) infection and the development and clinical manifestations of asthma. In this study we demonstrate that cultured primary bronchial epithelial cells from adults with asthma (n = 9) show different transcriptional and chromatin responses to RV infection compared to those without asthma (n = 9). Both the number and magnitude of transcriptional and chromatin responses to RV were muted in cells from asthma cases compared to controls. Pathway analysis of the transcriptionally responsive genes revealed enrichments of apoptotic pathways in controls but inflammatory pathways in asthma cases. Using promoter capture Hi-C we tethered regions of RV-responsive chromatin to RV-responsive genes and showed enrichment of these regions and genes at asthma GWAS loci. Taken together, our studies indicate a delayed or prolonged inflammatory state in cells from asthma cases and highlight genes that may contribute to genetic risk for asthma.

Genome-wide association studies (GWASs) have become a standard tool for dissecting genetic contributions to disease risk. However, these studies typically require extraordinarily large sample sizes to be adequately powered. Strategies that incorporate functional information alongside genetic associations have proved successful in increasing GWAS power. Following this paradigm, we present the results of 20 different genetic association studies for quantitative traits related to complex diseases, conducted in the Hutterites of South Dakota. To boost the power of these association studies, we collected RNA-sequencing data from lymphoblastoid cell lines for 431 Hutterite individuals. We then used Sherlock, a tool that integrates GWAS and expression quantitative trait locus (eQTL) data, to identify weak GWAS signals that are also supported by eQTL data. Using this approach, we found novel associations with quantitative phenotypes related to cardiovascular disease, including carotid intima-media thickness, left atrial volume index, monocyte count and serum YKL-40 levels.

The non-classical major histocompatibility complex molecule, human leukocyte antigen (HLA)-G, is thought to contribute to maternal immune tolerance and successful placentation during pregnancy. Genetic polymorphisms in HLA-G are known to influence expression levels as well as the relative expression of individual protein isoforms. As diminished or aberrant HLA-G expression patterns may contribute to the development of certain pregnancy complications, we sought to investigate the association between functional HLA-G polymorphisms and the risk of pre-eclampsia (PE) in African-American women. The association between maternal and fetal genotype at six HLA-G polymorphisms and risk of PE was assessed in 372 pregnancies (314 normotensive; 58 pre-eclamptic). We observed an elevated risk of PE (P = 0.00027) in pregnancies where the mother carried the 1597ΔC allele, a null allele that abolishes expression of full-length HLA-G isoforms. Furthermore, the frequency of the maternal 1597ΔC allele was highest in the subset of pre-eclamptic pregnancies that were delivered preterm, suggesting an association between the null allele and the severity of PE. We then replicated the association between higher maternal 1597ΔC allele frequency and increased severity of PE (P = 0.038) in an independent sample of 533 African-American women. Finally, to investigate the mechanistic basis of this association, we measured circulating soluble HLA-G (sHLA-G) concentrations in maternal serum collected during pregnancy in 51 healthy, normotensive African-American control women and found significantly lower levels in women carrying the 1597ΔC allele (P = 0.012). These results demonstrate that maternal HLA-G genotype is significantly associated with risk of PE in African-American women and is predictive of circulating sHLA-G levels during pregnancy.

Recent genome-wide association studies (GWAS) have identified a number of novel genetic associations with complex human diseases. In spite of these successes, results from GWAS generally explain only a small proportion of disease heritability, an observation termed the 'missing heritability problem'. Several sources for the missing heritability have been proposed, including the contribution of many common variants with small individual effect sizes, which cannot be reliably found using the standard GWAS approach. The goal of our study was to explore a complimentary approach, which combines GWAS results with functional data in order to identify novel genetic associations with small effect sizes. To do so, we conducted a GWAS for lymphocyte count, a physiologic quantitative trait associated with asthma, in 462 Hutterites. In parallel, we performed a genome-wide gene expression study in lymphoblastoid cell lines from 96 Hutterites. We found significant support for genetic associations using the GWAS data when we considered variants near the 193 genes whose expression levels across individuals were most correlated with lymphocyte counts. Interestingly, these variants are also enriched with signatures of an association with asthma susceptibility, an observation we were able to replicate. The associated loci include genes previously implicated in asthma susceptibility as well as novel candidate genes enriched for functions related to T cell receptor signaling and adenosine triphosphate synthesis. Our results, therefore, establish a new set of asthma susceptibility candidate genes. More generally, our observations support the notion that many loci of small effects influence variation in lymphocyte count and asthma susceptibility.

HLA-G is a nonclassical, class I major histocompatibility complex (MHC) gene that exhibits immunomodulatory properties and likely plays a role in the maintenance of successful pregnancy. In this study, we investigated the role of HLA-G polymorphisms on risk for recurrent pregnancy loss (RPL) and on circulating levels of soluble (s)HLA-G in Iraqi women. DNA and plasma were obtained from blood samples collected at 9-12 weeks gestation from 50 women with RPL and 50 healthy pregnant women in Basrah province, Iraq. As measured by ELISA, median sHLA-G levels were significantly lower in the RPL cases compared to healthy controls (21.4 vs. 38.8 U/ml, respectively; P=0.025), and decreased with increasing maternal age (P=0.0051). However, HLA-G allele and haplotype frequencies did not differ significantly between cases and controls (P values ≥0.12 for all tests). In contrast, homozygosity for the C allele (CC) at a tri-allelic promoter polymorphism, -725C/G/T, was associated with lower concentrations of sHLA-G compared to the CG or CT genotypes (median levels 21.1 vs. 40.1 vs. 42.6 U/ml, respectively; P=0.0089). These results demonstrate that HLA-G genotype influences circulating sHLA-G levels during pregnancy but is not significantly associated with risk of RPL.

The semiallogenic fetus is tolerated by the maternal immune system through control of innate and adaptive immune responses. Trophoblast cells secrete nanometer scale membranous particles called exosomes, which have been implicated in modulation of the local and systemic maternal immune system. Here we investigate the possibility that exosomes secreted from the first trimester and term placenta carry HLA-G and B7 family immunomodulators. Confocal microscopy of placental sections revealed intracellular co-localization of B7-H1 with CD63, suggesting that B7-H1 associates with subcellular vesicles that give rise to exosomes. First trimester and term placental explants were then cultured for 24 h. B7H-1 (CD274), B7-H3 (CD276) and HLA-G5 were abundant in pelleted supernatants of these cultures that contained microparticles and exosomes; the latter, however, was observed only in first trimester pellets and was nearly undetectable in term explant-derived pellets. Further purification of exosomes by sucrose density fractionation confirmed the association of these proteins specifically with exosomes. Finally, culture of purified trophoblast cells in the presence or absence of EGF suggested that despite the absence of HLA-G5 association with term explant-derived exosomes, it is present in exosomes secreted from mononuclear cytotrophoblast cells. Further, differentiation of cytotrophoblast cells reduced the presence of HLA-G5 in secreted exosomes. Together, the results suggest that the immunomodulatory proteins HLA-G5, B7-H1 and B7-H3, are secreted from early and term placenta, and have important implications in the mechanisms by which trophoblast immunomodulators modify the maternal immunological environment.

Preeclampsia occurs more frequently in women of African ancestry. The cause of this hypertensive complication is unclear, but placental oxidative stress may play a role. Because mitochondria are the major sites of oxidative phosphorylation, we hypothesized that placentas of preeclamptic pregnancies harbor mitochondrial DNA (mtDNA) mutations. Next-generation sequencing of placental mtDNA in African American preeclamptics (N = 30) and controls (N = 38) from Chicago revealed significant excesses in preeclamptics of nonsynonymous substitutions in protein-coding genes and mitochondrially encoded nicotinamide adenine dinucleotide dehydrogenase 5 gene and an increase in the substitution rate (P = .0001). Moreover, 88% of preeclamptics and 53% of controls carried at least one nonsynonymous substitution (P = .005; odds ratio [OR] = 6.36, 95% confidence interval [CI]: 1.5-39.1). These results were not replicated in a sample of African American preeclamptics (N = 162) and controls (N = 171) from Detroit. Differences in study design and heterogeneity may account for this lack of replication. Nonsynonymous substitutions in mtDNA may be risk factors for preeclampsia in some African American women, but additional studies are required to establish this relationship.

Plasma lipoprotein(a) (Lp[a]) level is an independent risk factor of cardiovascular disease that is under strong genetic control. We conducted a genome-wide association study of plasma Lp(a) in 386 members of a founder population that adheres to a communal lifestyle, proscribes cigarette smoking, and prepares and eats meals communally. We identified associations with 77 single nucleotide polymorphisms (SNPs) spanning 12.5 Mb on chromosome 6q26-q27 that met criteria for genome-wide significance (P <or= 1.3 x 10(-7)) and were within or flanking nine genes, including LPA. We show that variation in at least six genes in addition to LPA are significantly associated with Lp(a) levels independent of each other and of the kringle IV repeat polymorphism in the LPA gene. One novel SNP in intron 37 of the LPA gene was also associated with Lp(a) levels and carotid artery disease number in unrelated Caucasians (P = 7.3 x 10(-12) and 0.024, respectively), also independent of kringle IV number. This study suggests a complex genetic architecture of Lp(a) levels that may involve multiple loci on chromosome 6q26-q27.

BACKGROUND:HLA-G is a non-classical HLA with important immunomodulatory roles in pregnancy. A polymorphism in the promoter region, -725G, was previously associated with sporadic miscarriage in women who were unselected with respect to reproductive history. In this study, the transcription levels of different HLA-G promoter haplotypes were examined to determine whether the miscarriage-associated -725G allele influences transcription.METHODS:Five naturally occurring promoter haplotypes and three variant haplotypes created by site-directed mutagenesis were sub-cloned into luciferase expression vectors and transfected into JEG-3 cells. Expression levels of these eight haplotypes were examined in cultured cells before and after treatment with interferon-beta (IFN-beta), cytosine-5-DNA methyltransferase (M. SssI) and 5-aza-2'-deoxycytidine. Differences in expression levels between haplotypes were determined by analysis of variance (ANOVA).RESULT:Promoter haplotypes with the miscarriage-associated -725G allele were expressed at significantly higher levels in all culture conditions compared with otherwise identical haplotypes that had a -725C or -725T allele.CONCLUSION:Variation in the HLA-G promoter region influences transcription rates. Contrary to expectations, increased expression of HLA-G may be disadvantageous in some pregnancies.

Asthma affects nearly 14 million people worldwide and has been steadily increasing in frequency for the past 50 years. Although environmental factors clearly influence the onset, progression, and severity of this disease, family and twin studies indicate that genetic variation also influences susceptibility. Linkage of asthma and related phenotypes to chromosome 6p21 has been reported in seven genome screens, making it the most replicated region of the genome. However, because many genes with individually small effects are likely to contribute to risk, identification of asthma susceptibility loci has been challenging. In this study, we present evidence from four independent samples in support of HLA-G as a novel asthma and bronchial hyperresponsiveness susceptibility gene in the human leukocyte antigen region on chromosome 6p21, and we speculate that this gene might contribute to risk for other inflammatory diseases that show linkage to this region.

The HLA-G gene is primarily expressed in placental cells that invade the maternal decidua during pregnancy. This gene encodes multiple isoforms that fulfill a variety of functions at the maternal-fetal interface throughout gestation. Recently, a null allele for the most abundant HLA-G isoform was associated with recurrent miscarriage in two independent studies, suggesting that reduced levels of the HLA-G1 protein may compromise successful pregnancy. We initiated the present study to determine whether other polymorphisms that could affect expression levels of HLA-G were associated with fetal loss in women participating in a 15-year prospective study of pregnancy outcome. We genotyped these subjects for 18 single-nucleotide polymorphisms in the 1,300 bp upstream of exon 1, 13 of which were identified as part of this study, as well as for an insertion/deletion (in/del) polymorphism in the 3' untranslated region. The 18 SNPs defined eight unique haplotypes. One polymorphism, -725C/G, was associated with fetal loss, with an increased risk for miscarriage in couples in which both partners carried the -725G allele, compared with couples not carrying this allele (odds ratio 2.76, 95% confidence interval 1.08-7.09; P=.035). Further, the G at nucleotide -725 creates a CpG dinucleotide, and we demonstrate that this CpG site is methylated on -725G alleles. Overall, this study identified extraordinary levels of variation in the 5'-upstream regulatory region of HLA-G and provides evidence for an association between a promoter-region SNP and fetal loss rates, further attesting to the novel features and critical role of this gene in pregnancy.

OBJECTIVE:As part of a study to determine population-based frequencies of CFTR mutations in an ethnically diverse, midwestern cystic fibrosis (CF) population, clinical histories were studied in 119 CF patients.METHODOLOGY:We sought to examine the association between genotype as characterized by the delta F508 and 11 other commonly occurring mutations and clinical parameters including age at diagnosis, clinical presentation, sweat chloride level, chest roentgenogram score, clinical scores, pulmonary function test results, percent weight for height, and presence of associated CF complications.RESULTS:Age at diagnosis of CF was significantly associated with homozygosity for delta F508 (mean age at diagnosis +/- SE: 1.7 +/- 0.3 years for delta F508/delta F508 vs 3.9 +/- 0.9 years for delta F508/other and other/other; P = .03). No other age-adjusted clinical parameter was significantly associated with delta F508 or any other genotype.CONCLUSION:These data suggest that in this sample of CF patients, delta F508 genotype is not predictive of disease severity. The lack of association between disease severity and genotype in this ethnically diverse sample may reflect the presence of more severe undetected mutations in our sample, or the effects of modifying genes at other, non-CF loci.

Maternal-fetal histocompatibility for alleles at HLA class II loci, HLA-DQA1 and HLA-DQB1, was examined in 40 abortuses and 31 liveborn children of 68 couples with a history of idiopathic recurrent spontaneous abortion (RSAB) who underwent leukocyte immunization prior to the index pregnancy. Significantly more couples with RSAB shared two HLA-DQA1 alleles as compared with fertile control couples (0.18 vs. 0.03, respectively; P = 0.031). There were no differences in HLA sharing between couples with RSAB who experienced a repeat abortion in the index pregnancy as compared with couples with RSAB who were delivered of a liveborn child. Non-significant deficits of abortuses who were compatible for alleles at the HLA-DQA1 (6 observed vs. 8.5 expected; P = 0.225) and the HLA-DQB1 (7 observed vs. 9.2 expected; P = 0.254) loci were observed. A significant deficit of HLA-DQA1 compatible liveborn children was observed (1 observed vs. 5.5 expected; P = 0.0069). The overall deficit of HLA-DQA1 compatible fetuses (7 observed vs. 14.0 expected; P = 0.0018) after approximately 8 weeks gestation suggests that HLA-DQA1 compatible fetuses may be aborted early in pregnancy, prior to the time when fetal tissue can be recovered for genetic studies.

Journal Article HindIII polymorphism in the BCL6 gene Get access Beverly W.Baron, Beverly W.Baron * * To whom correspondence should be addressed Search for other works by this author on: Oxford Academic PubMed Google Scholar Christine Billstrand, Christine Billstrand 1Department of Obelectrics & Gynecology, University of ChicagoChicago, IL 60637, USA Search for other works by this author on: Oxford Academic PubMed Google Scholar Luisa Madronero, Luisa Madronero Search for other works by this author on: Oxford Academic PubMed Google Scholar Timothy W. McKeithan Timothy W. McKeithan Search for other works by this author on: Oxford Academic PubMed Google Scholar Human Molecular Genetics, Volume 2, Issue 9, September 1993, Page 1513, https://doi.org/10.1093/hmg/2.9.1513 Published: 01 September 1993