Case-control Genetic Association Study Research Articles

RPA3-UMAD1 rs12702634 and rheumatoid arthritis-associated interstitial lung disease in European ancestry

Abstract Objective Recently, a genome-wide association identified an association between rheumatoid arthritis (RA)-associated interstitial lung disease (ILD) and RPA3-UMAD1 rs12702634 in the Japanese population, especially for patients with a usual interstitial pneumonia (UIP) pattern. We aimed to replicate this association in a European population and test for interaction with MUC5B rs35705950. Methods In this genetic case-control association study, cases with RA with ILD and controls with RA and no ILD were included from France, the US, and the Netherlands. Only cases and controls from European genetic ancestries determined by principal component analysis were included in the analyses. RA was defined by 1987 ACR or 2010 ACR/EULAR criteria and ILD by chest high-resolution CT scan, except in the control dataset from The Netherlands where the absence of ILD was determined by chart review. Patients were genotyped for RPA3-UMAD1 rs12702634 and MUC5B rs35705950. Associations were tested using logistic regression adjusted for sex, age at RA onset, age at ILD onset or at certified absence of ILD, tobacco smoking status, and country of origin. Results Among the 883 patients included, 322 were RA-ILD cases (36.5%). MUC5B rs35705950 was strongly associated with RA-ILD in all datasets: combined adjusted OR = 2.9, 95%CI [2.1–3.9], P = 1.1x1 0 −11. No association between RPA3-UMAD1 rs12702634 and RA-ILD was observed: combined OR = 1.2 95%CI [0.8–1.6], p = 0.31. No interaction was found between RPA3-UMAD1 rs12702634 and MUC5B rs35705950 (p = 0.70). Conclusion Our findings did not support a contribution of RPA3-UMAD1 rs12702634 to the overall RA-ILD susceptibility in the European population.

The Arg/Arg genotype of leptin receptor gene Gln223Arg polymorphism may be an independent risk factor for nonalcoholic fatty liver disease.

Given that obesity and insulin resistance play key roles in the pathogenesis of nonalcoholic fatty liver disease (NAFLD) and the connection between leptin and these metabolic diseases, the association between NAFLD and a leptin receptor gene (LEPR) polymorphism was examined. In this genetic case-control association study, 144 biopsy-proven NAFLD patients and 144 controls were genotyped for the LEPR gene Gln223Arg (rs1137101) polymorphism using the polymerase chain reaction-restriction fragment length polymorphism method. The distributions of genotypes and alleles of Gln223Arg variant were in accordance with the Hardy-Weinberg equilibrium in the study groups (P>.05). Multivariate logistic regression analysis showed that the LEPR Gln223Arg Arg/Arg genotype was an independent risk factor for NAFLD; the Arg/Arg genotype, compared with the Gln/Gln genotype, was associated with a 2.09-fold increased risk for NAFLD (P=.036, odds ratio=2.09 [95% CI = 1.31-5.95]). We found that the LEPR Gln223Arg Arg/Arg genotype was independently associated with a more than 2-fold rise in biopsy-proven NAFLD risk. Our findings, however, need to be corroborated by further studies.

Association of Tenascin-C Gene Polymorphisms with Risk of Acute Coronary Syndrome in South Indian Population: A Case-Control Genetic Association Study.

Background: The extracellular matrix (ECM) glycoprotein changes are associated with the pathogenesis and complications of atherosclerosis, leading to acute coronary syndrome (ACS). Tenascin-C (TNC), an ECM protein, has been implemented in the pathogenesis, diagnosis, and prognosis of patients with cardiovascular disease. Aim: The study aimed to compare the genetic variants of the TNC gene (rs13321, rs2104772, and rs12347433) between South Indians with ACS and healthy participants. Materials and Methods: This case-control study recruited 150 ACS patients as cases and 150 healthy participants as controls. TNC genotyping was performed using TaqMan 5'-exonuclease allele discrimination assay. Serum TNC levels were measured by enzyme-linked immunosorbent assay. Results: Serum TNC levels were significantly higher in cases compared with controls. No significant difference was observed in allele and genotype frequencies of rs13321, rs2104772, and rs12347433 between cases and controls, which was confirmed by dominant, recessive, codominant, and homozygotic genetic models. The patients with heterozygous genotypes of rs13321, rs2104772, and rs12347433 had significantly lower serum TNC levels than patients with respective homozygous genotypes. Haplotype analyses revealed that the C-T-A haplotype in the block of rs13321-rs12347433-rs2104772 was associated with lower ACS risk (OR = 0.33, 95% CI: 0.15 - 0.75; p = 0.005). Also, the C-T-T and G-T-A haplotypes of the TNC gene were associated with higher and lower serum TNC levels, respectively. Conclusion: Our study demonstrated no genetic association between single nucleotide polymorphisms of the TNC gene and ACS risk; however, the C-T-A haplotype of the TNC gene might be associated with reduced ACS risk in South Indians.

Common Variants rs429358 and rs7412 in APOE Gene Are Not Associated with POAG in a Saudi Cohort.

Adult-onset glaucoma, an age-related neurodegenerative disease, is very prevalent among the elderly Arabs of Saudi origin. This study investigated the association between apolipoprotein E (APOE) gene variants (rs429358 and rs7412) and primary open-angle glaucoma (POAG) in Arabs of Saudi origin. A case-control genetic association study involving 179 POAG patients and 251 controls utilized Sanger sequencing to genotype APOE gene variants. The allele frequencies and genotype distributions for rs429358 and rs7412 did not show significant associations with POAG. The haplotype analysis revealed apoε3 (87.6% and 87.4%) as the most prevalent, followed by ε4 (2.8% and 3.6%) and ε2 (9.6% and 8.9%) in the controls and POAG patients, respectively. Although the ε2/ε3 genotype and ε2-carriers displayed a more than two-fold increased risk, statistical significance was not reached. Notably, these polymorphisms did not affect clinical markers, such as intraocular pressure and cup/disc ratio. The logistic regression analysis demonstrated no significant influence of age, sex, rs429358, or rs7412 polymorphisms on POAG. In conclusion, within the Saudi cohort, APOE variants (rs429358 and rs7412) do not appear to be associated with POAG and are not substantial risk factors for its development. However, additional population-based studies are required to validate these findings.

Genetic associations of central serous chorioretinopathy subtypes, neovascular age-related macular degeneration, and polypoidal choroidal vasculopathy

PurposeTo evaluate the genetic associations of different subtypes of central serous chorioretinopathy (CSCR), neovascular age-related macular degeneration (nAMD), and polypoidal choroidal vasculopathy (PCV). DesignA case-control genetic association study. MethodsThis study enrolled 217 CSCR, 341 nAMD, 288 PCV patients, and 1380 controls. The CSCR patients were classified into those with focal or diffuse leakage, with or without pigment epithelial detachment (PED), and with or without macular neovascularization (MNV). Associations between 11 variants from 8 genes, ADAMTS9, ANGPT2, ARMS2, CFH, NR3C2, PGF, TNFRSF10A and VIPR2, and diseases/subtypes were analyzed by logistic regression analysis adjusted for age and sex, and inter-phenotype comparison by heterogeneity test. ResultsThe CFH rs800292-A conferred a protective effect for CSCR with MNV (OR=0.44, P = 0.002) and a risk effect for CSCR without MNV (OR=1.31, P = 0.023). CSCR patients carrying rs800292-G had a 3.23-fold of increased risk towards developing secondary MNV (P = 1.45 ×10-4). CFH rs3753394, rs800292 and rs1329428 showed similar effects among CSCR with MNV, nAMD and PCV, but opposite effects on CSCR without MNV. TNFRSF10A rs13278062-T was associated with overall CSCR but not with CSCR subtypes, nAMD or PCV. Moreover, CFH and ARMS2 SNPs showed heterogeneous effects in CSCR without MNV against CSCR with MNV, nAMD and PCV. ConclusionsGenetic associations of CSCR with MNV resembled nAMD and PCV compared to CSCR without MNV, indicating differential genetic effects on neovascularization and choroidopathy. Further investigation of the functional roles of CFH, ARMS2, and TNFRSF10A in CSCR, nAMD and PCV should help elucidate the mechanisms of these maculopathies.

Insulin-like growth factor binding protein 3 promoter variant (rs2854744) is associated with nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease and a growing global epidemic. In NAFLD, liver fat surpasses 5% of hepatocytes without the secondary causes of lipid accumulation or excessive alcohol consumption. Given the link between NAFLD and insulin resistance, the possible association between the rs2854744 (-202 G>T) promoter polymorphism of insulin-like growth factor binding protein 3 (IGFBP3) gene and NAFLD was investigated in this study. In this genetic case-control association study, the IGFBP3 rs2854744 genotypes of 315 unrelated individuals, including 156 patients with biopsy-proven NAFLD and 159 controls, were determined using polymerase chain reaction/restriction fragment length polymorphism analyses. The "GT+TT" genotype of the IGFBP3 rs2854744 polymorphism, compared with the "GG" genotype, was associated with a 2.7-fold increased risk of NAFLD after adjustment for confounding factors (P = 0.009; odds ratio [OR] = 2.71; 95% confidence interval [CI] = 1.19-3.18). Additionally, the IGFBP3 rs2854744 "T" allele, in comparison with the "G" allele, was significantly overrepresented in NAFLD patients than the controls (P = 0.008; OR = 1.85; 95%CI = 1.23-2.94). Our findings first indicated that the IGFBP3 rs2854744 "GT+TT" genotype is a marker of increased NAFLD susceptibility; however, it needs to be supported by further investigations in other populations.

Schizophrenia plausible protective effect of microRNA-137 is potentially related to estrogen and prolactin in female patients.

Schizophrenia (SCZ) is a serious chronic mental disorder. Our previous case-control genetic association study has shown that microRNA-137 (miR-137) may only protect females against SCZ. Since estrogen, an important female sex hormone, exerts neuroprotective effects, the relationship between estrogen and miR-137 in the pathophysiology of SCZ was further studied in this study. Genotyping of single-nucleotide polymorphism rs1625579 of miR-137 gene in 1,004 SCZ patients and 896 healthy controls was conducted using the iMLDR assay. The effect of estradiol (E2) on the miR-137 expression was evaluated on the human mammary adenocarcinoma cell line (MCF-7) and the mouse hippocampal neuron cell line (HT22). The relationships between serum E2, prolactin (PRL), and peripheral blood miR-137 were investigated in 41 SCZ patients and 43 healthy controls. The miR-137 and other reference miRNAs were detected by real-time fluorescent quantitative reverse transcription-PCR. Based on the well-known SNP rs1625579, the distributions of protective genotypes and alleles of the miR-137 gene were not different between patients and healthy controls but were marginally significantly lower in female patients. E2 upregulated the expression of miR-137 to 2.83 and 1.81 times in MCF-7 and HT22 cells, respectively. Both serum E2 and blood miR-137 were significantly decreased or downregulated in SCZ patients, but they lacked expected positive correlations with each other in both patients and controls. When stratified by sex, blood miR-137 was negatively correlated with serum E2 in female patients. On the other hand, serum PRL was significantly increased in SCZ patients, and the female patients had the highest serum PRL level and a negative correlation between serum PRL and blood miR-137. The plausible SCZ-protective effect of miR-137 may be female specific, of which the underlying mechanism may be that E2 upregulates the expression of miR-137. This protective mechanism may also be abrogated by elevated PRL in female patients. These preliminary findings suggest a new genetic/environmental interaction mechanism for E2/miR-137 to protect normal females against SCZ and a novel E2/PRL/miR-137-related pathophysiology of female SCZ, implying some new antipsychotic ways for female patients in future.

Clinical and genetic determinants of severe acute pancreatitis: A genetic association study in the UK Biobank.

The clinical severity of acute pancreatitis is unpredictable, ranging from self-limiting disease to life-threatening inflammation. The determinants of severe acute pancreatitis (SAP) are unclear. We aim to identify clinical variables and single nucleotide polymorphisms (SNP) associated with SAP. We used UK Biobank data to conduct a case-control clinical and genetic association study. Pancreatitis patients were identified through national hospital and mortality records across the United Kingdom. Clinical covariates and SAP were analyzed for associations. Genotyped data that included 35 SNPs were assessed for independent associations with SAP and SNP to SNP interaction. A total of 665 patients with SAP and 3304 non-SAP patients were identified. Male sex and older age increased odds of developing SAP (odds ratio [OR] 1.48; 95% confiden interval [CI] 1.24-1.78, P<0.0001) and (OR 1.23; 95% CI 1.17-1.29), P<0.0001), respectively. SAP was associated with diabetes (OR 1.46; 95% CI 1.15-1.86, P=0.002), chronic kidney disease (OR 1.74; 95% CI 1.26-2.42, P=0.001), and cardiovascular disease (OR 2.00; 95% CI 1.54-2.61, P=0.0001). A significant association was established between IL-10 rs3024498 and SAP (OR 1.24; 95% CI 1.09-1.41, P=0.0014). Epistasis analysis revealed that the odds of SAP was greater by an interaction between TLR 5 rs5744174 and Factor V rs6025 (ORinteraction 7.53; P=6.64×10-5 ). This study reports clinical risk factors for SAP. We also show evidence for an interaction between rs5744174 and rs6025 as determinants for SAP in addition to rs3024498 independently altering the severity of acute pancreatitis.

Contribution of Somatic Ras/Raf/Mitogen-Activated Protein Kinase Variants in the Hippocampus in Drug-Resistant Mesial Temporal Lobe Epilepsy

Mesial temporal lobe epilepsy (MTLE) is the most common focal epilepsy subtype and is often refractory to antiseizure medications. While most patients with MTLE do not have pathogenic germline genetic variants, the contribution of postzygotic (ie, somatic) variants in the brain is unknown. To test the association between pathogenic somatic variants in the hippocampus and MTLE. This case-control genetic association study analyzed the DNA derived from hippocampal tissue of neurosurgically treated patients with MTLE and age-matched and sex-matched neurotypical controls. Participants treated at level 4 epilepsy centers were enrolled from 1988 through 2019, and clinical data were collected retrospectively. Whole-exome and gene-panel sequencing (each genomic region sequenced more than 500 times on average) were used to identify candidate pathogenic somatic variants. A subset of novel variants was functionally evaluated using cellular and molecular assays. Patients with nonlesional and lesional (mesial temporal sclerosis, focal cortical dysplasia, and low-grade epilepsy-associated tumors) drug-resistant MTLE who underwent anterior medial temporal lobectomy were eligible. All patients with available frozen tissue and appropriate consents were included. Control brain tissue was obtained from neurotypical donors at brain banks. Data were analyzed from June 2020 to August 2022. Drug-resistant MTLE. Presence and abundance of pathogenic somatic variants in the hippocampus vs the unaffected temporal neocortex. Of 105 included patients with MTLE, 53 (50.5%) were female, and the median (IQR) age was 32 (26-44) years; of 30 neurotypical controls, 11 (36.7%) were female, and the median (IQR) age was 37 (18-53) years. Eleven pathogenic somatic variants enriched in the hippocampus relative to the unaffected temporal neocortex (median [IQR] variant allele frequency, 1.92 [1.5-2.7] vs 0.3 [0-0.9]; P = .01) were detected in patients with MTLE but not in controls. Ten of these variants were in PTPN11, SOS1, KRAS, BRAF, and NF1, all predicted to constitutively activate Ras/Raf/mitogen-activated protein kinase (MAPK) signaling. Immunohistochemical studies of variant-positive hippocampal tissue demonstrated increased Erk1/2 phosphorylation, indicative of Ras/Raf/MAPK activation, predominantly in glial cells. Molecular assays showed abnormal liquid-liquid phase separation for the PTPN11 variants as a possible dominant gain-of-function mechanism. Hippocampal somatic variants, particularly those activating Ras/Raf/MAPK signaling, may contribute to the pathogenesis of sporadic, drug-resistant MTLE. These findings may provide a novel genetic mechanism and highlight new therapeutic targets for this common indication for epilepsy surgery.

Multi-Polymorphism Analysis Reveals Joint Effects in Males With Chronic Central Serous Chorioretinopathy.

Central serous chorioretinopathy (CSCR) is a leading cause of central vision impairment in the working-age population with male predilection. Knowledge about the genetic basis of CSCR and its male predilection remained limited. This study aimed to evaluate the association patterns of multiple gene variants in chronic CSCR (cCSCR) in Chinese patients. This case-control genetic association study included 531 patients with cCSCR and 2383 controls from two independent Chinese cohorts. Nine single-nucleotide polymorphisms (SNPs) of six genes, namely CFH, NR3C2, GATA5, VIPR2, TNFRSF10A, and ARMS2, were genotyped in all subjects. The main outcome measures were the association of individual single-nucleotide polymorphism (SNP) with cCSCR, the sex-stratification effects of individual SNP, and joint effects of different SNPs on cCSCR. Association results in the two cohorts were consistent with low heterogeneities. In the combined analysis, SNPs CFH rs800292 (odds ratio [OR] = 1.25, P = 0.0020), CFH rs1329428 (OR = 1.23, P = 0.0037), and TNFRSF10A rs13278062 (OR = 1.43, P = 0.0014) were significantly associated with cCSCR. In stratification analysis by sex, 3SNPs in CFH, rs3753394, rs800292, and rs1329428, were associated with cCSCR in male patients, but not in female patients. Joint analysis revealed that subjects homozygous for the risk alleles of CFH rs800292 and TNFRSF10A rs13278062 had over 4-fold of increased risk of cCSCR when compared with subjects homozygous for the non-risk alleles (OR = 4.06, P = 2.30 × 10-5). This study revealed main and joint effects of SNPs in CFH and TNFRSF10A on cCSCR, and suggested CFH as a potential genetic factor underlying the male predilection of cCSCR. Further replication in other study populations is needed.

A visfatin gene promoter polymorphism (rs1319501) is associated with susceptibility to nonalcoholic fatty liver disease.

Background: Considering the role of visfatin in nonalcoholic fatty liver disease (NAFLD), a growing global epidemic, this article explores the potential association between the visfatin gene (NAMPT) and NAFLD. Methods: We used the PCR-restriction fragment length polymorphism method to genotype the rs1319501 promoter variant of the NAMPT gene in 154 patients with biopsy-proven NAFLD and 158 controls in this case-control genetic association study. Results: The 'CC+TC' genotype of NAMPT rs1319501 in comparison to the 'TT' genotype occurred less frequently in the cases with NAFLD than the controls, and the difference remained significant after adjustment for confounding factors (p=0.029; odds ratio =0.55; 95% CI=0.31-0.82). Conclusion: This study showed, for the first time, that the carriers of the NAMPT rs1319501 'CC+TC' genotype had a 45% decreased risk for NAFLD.

Application of an in silico approach identifies a genetic locus within ITGB2, and its interactions with HSPG2 and FGF9, to be associated with anterior cruciate ligament rupture risk

ABSTRACT We developed a Biomedical Knowledge Graph model that is phenotype and biological function-aware through integrating knowledge from multiple domains in a Neo4j, graph database. All known human genes were assessed through the model to identify potential new risk genes for anterior cruciate ligament (ACL) ruptures and Achilles tendinopathy (AT). Genes were prioritised and explored in a case–control study comparing participants with ACL ruptures (ACL-R), including a sub-group with non-contact mechanism injuries (ACL-NON), to uninjured control individuals (CON). After gene filtering, 3376 genes, including 411 genes identified through previous whole exome sequencing, were found to be potentially linked to AT and ACL ruptures. Four variants were prioritised: HSPG2:rs2291826A/G, HSPG2:rs2291827G/A, ITGB2:rs2230528C/T and FGF9:rs2274296C/T. The rs2230528 CC genotype was over-represented in the CON group compared to ACL-R (p < 0.001) and ACL-NON (p < 0.001) and the TT genotype and T allele were over-represented in the ACL-R group and ACL-NON compared to CON (p < 0.001) group. Several significant differences in distributions were noted for the gene-gene interactions: (HSPG2:rs2291826, rs2291827 and ITGB2:rs2230528) and (ITGB2:rs2230528 and FGF9:rs2297429). This study substantiates the efficiency of using a prior knowledge-driven in silico approach to identify candidate genes linked to tendon and ACL injuries. Our biomedical knowledge graph identified and, with further testing, highlighted novel associations of the ITGB2 gene which has not been explored in a genetic case control association study, with ACL rupture risk. We thus recommend a multistep approach including bioinformatics in conjunction with next generation sequencing technology to improve the discovery potential of genomics technologies in musculoskeletal soft tissue injuries. Highlights A biomedical knowledge graph was modelled for musculoskeletal soft tissue injuries to efficiently identify candidate genes for genetic susceptibility analyses. The biomedical knowledge graph and sequencing data identified potential biologically relevant variants to explore susceptibility to common tendon and ligament injuries. Specifically genetic variants within the ITGB2 and FGF9 genes were associated with ACL risk. Novel allele combinations (HSPG2-ITGB2 and ITGB2-FGF9) showcase the potential effect of ITGB2 in influencing risk of ACL rupture.

Identification of Novel Genetic Risk Factors for Focal Segmental Glomerulosclerosis in Children: Results From the Chronic Kidney Disease in Children (CKiD) Cohort

Focal segmental glomerulosclerosis (FSGS) is a major cause of pediatric nephrotic syndrome, and African Americans exhibit an increased risk for developing FSGS compared with other populations. Predisposing genetic factors have previously been described in adults. Here we performed genomic screening of primary FSGS in a pediatric African American population. Prospective cohort with case-control genetic association study design. 140 African American children with chronic kidney disease from the Chronic Kidney Disease in Children (CKiD) cohort, including 32 cases with FSGS. Over 680,000 common single-nucleotide polymorphisms (SNPs) were tested for association. We also ran a pathway enrichment analysis and a human leucocyte antigen (HLA)-focused association study. Primary biopsy-proven pediatric FSGS. Multivariate logistic regression models. The genome-wide association study revealed 169 SNPs from 14 independent loci significantly associated with FSGS (false discovery rate [FDR]<5%). We observed notable signals for genetic variants within the APOL1 (P=8.6×10-7; OR, 25.8 [95% CI, 7.1-94.0]), ALMS1 (P=1.3×10-7; 13.0% in FSGS cases vs 0% in controls), and FGFR4 (P=4.3×10-6; OR, 24.8 [95% CI, 6.3-97.7]) genes, all of which had previously been associated with adult FSGS, kidney function, or chronic kidney disease. We also highlighted novel, functionally relevant genes, including GRB2 (which encodes a slit diaphragm protein promoting podocyte structure through actin polymerization) and ITGB1 (which is linked to renal injuries). Our results suggest a major role for immune responses and antigen presentation in pediatric FSGS through (1) associations with SNPs in PTPRJ (or CD148, P=3.5×10-7), which plays a role in T-cell receptor signaling, (2) HLA-DRB1∗11:01 association (P=6.1×10-3; OR, 4.5 [95% CI, 1.5-13.0]), and (3) signaling pathway enrichment (P=1.3×10-6). Sample size and no independent replication cohort with genomic data readily available. Our genetic study has identified functionally relevant risk factors and the importance of immune regulation for pediatric primary FSGS, which contributes to a better description of its molecular pathophysiological mechanisms. We assessed the genetic risk factors for primary focal segmental glomerulosclerosis (FSGS) by simultaneously testing over 680,000 genetic markers spread across the genome in 140 children, including 32 with FSGS lesions. Fourteen independent genetic regions were significantly associated with pediatric FSGS, including APOL1 and ALMS1-NAT8, which were previously found to be associated with FSGS and chronic kidney diseases in adults. Novel genes with relevant biological functions were also highlighted, such as GRB2 and FGFR4, which play a role in the kidney filtration barrier and in kidney cell differentiation, respectively. Finally, we revealed the importance of immune regulation in pediatric FSGS through associations involving cell surface proteins presenting antigens to the immune system and interacting with T-cell receptors.

한국인 투척 선수군에서 CKMM 유전자의 rs8111989(NcoI, A/G) 다형성에 대한 분포 양상

In order to explore the useful genetic marker(s) influencing the athletic performance of throwers, we performed a kind of case-control study genetic association study using the rs8111989(NcoI, A/G) polymorphism in the muscle-specific creatine kinase(CKMM) gene as a candidate gene. A total of 160 unrelated subjects including controls(n=70) and thrower (n=90) were recruited in this study, and total genomic DNA isolated from their saliva. The analysis of A/G polymorphisms in CKMM gene from their total genomic DNA was performed by using SNaPshot method in automated machine. By the result of genotype analysis, there was the significant differences in both genotype and allele frequencies of the A/G polymorphism in CKMM gene between controls and throwers, respectively(p.05). Especially, the frequencies of GG genotype and G allele in this genetic polymorphism showed the significantly higher frequencies in throwers than in controls, respectively. Thus, this genetic polymorphism may be one of useful genetic markers influencing the athletic performance in throwers. However, because athletic performance indicate the polygenic and multifactorial trait, the further studies using other genetic polymorphisms in more candidate genes will be required.

Association of Single Nucleotide Polymorphisms from Angiogenesis-Related Genes, ANGPT2, TLR2 and TLR9, with Spontaneous Preterm Labor

In this study, we hypothesized that the changes localized at angiopoietin-2 (ANGPT2), granulocyte-macrophage colony-stimulating factor (CSF2), fms-related tyrosine kinase 1 (FLT1) and toll-like receptor (TLR) 2, TLR6 and TLR9 genes were associated with spontaneous preterm labor (PTL), as well as with possible genetic alterations on PTL-related coagulation. This case-control genetic association study aimed to identify single nucleotide polymorphisms (SNPs) for the aforementioned genes, which are correlated with genetic risk or protection against PTL in Polish women. The study was conducted in 320 patients treated between 2016 and 2020, including 160 women with PTL and 160 term controls in labor. We found that ANGPT2 rs3020221 AA homozygotes were significantly less common in PTL cases than in controls, especially after adjusting for activated partial thromboplastin time (APTT) and platelet (PLT) parameters. TC heterozygotes for TLR2 rs3804099 were associated with PTL after correcting for anemia, vaginal bleeding, and history of threatened miscarriage or PTL. TC and CC genotypes in TLR9 rs187084 were significantly less common in women with PTL, compared to the controls, after adjusting for bleeding and gestational diabetes. For the first time, it was shown that three polymorphisms—ANGPT2 rs3020221, TLR2 rs3804099 and TLR9 rs187084 —were significantly associated with PTL, adjusted by pregnancy development influencing factors.

P-538 KATNAL1 polymorphisms confer susceptibility to severe phenotypes of male infertility in a large European cohort

Abstract Study question What is the contribution of the common variation of KATNAL1 to the development of severe spermatogenic failure (SPGF) in a phenotypically well-characterized cohort? Summary answer An allelic combination of KATNAL1 single-nucleotide polymorphisms (SNPs) increases the risk to develop SPGF, likely by altering the expression and splicing pattern of the gene. What is known already Spermatogenesis is a process that requires an exhaustive control of gene expression, and subtle alterations of its molecular regulatory network can lead to male infertility. The aetiology of most SPGF cases remains unknown, and increasing evidence clearly suggests that the idiopathic form of SPGF represents a complex trait, in which genetic susceptibility is conferred by the accumulation of risk alleles in genetically predisposed men. In this regard, previous studies reported that rare genetic mutations and polymorphisms in the KATNAL1 locus lead to male infertility through the disruption of microtubule remodelling and premature germ cell exfoliation. Study design, size, duration We designed a case-control genetic association study including three SNPs (rs2077011, rs7338931, and rs2149971) in the 3' and 5' regulatory regions of KATNAL1, which tagged the common genetic variability in the region. The allele frequencies in the study population, composed of 715 infertile men diagnosed with idiopathic SPGF, were compared to those observed in a control group comprising 1058 fertile men from Spain and Portugal. Participants/materials, setting, methods The SPGF group comprised 210 men with severe oligospermia (SO) and 505 with non-obstructive azoospermia (NOA). The latter were phenotypically characterised according to the histological examination of testis biopsies and its outcome (Sertoli cell-only syndrome, SCO; maturation arrest, MA; hypospermatogenesis, HS; and testicular sperm extraction, TESE). After genotyping, case-control association analyses by logistic regression were conducted. In silico functional characterization of risk variants was also carried out using public multiomic databases and bioinformatic tools. Main results and the role of chance Significant genetic associations with different SPGF patterns and/or TESE outcome were observed even after correction for multiple testing when independent SNP models were tested. However, in all cases, the haplotype model including the three risk alleles (rs2077011*C | rs7338931*T | rs2149971*A) was the most informative. This haplotype was specifically over-represented in the SPGF group (P = 3.45E-02, OR = 2.33), which includes all infertile men, and in the NOA group (P = 8.22E-03, OR = 2.97). In addition, subtype-specific associations were observed with the most severe subgroups, namely MA (P = 2.44E-02, OR = 5.00), SCO (P = 4.03E-03, OR = 5.16), and unsuccessful TESE outcome (P = 2.22E-04, OR = 6.13), which indicates the relevant role of KATNAL1 in spermatogenesis development. We prioritized the most likely causal variant/s based on in silico analyses addressing the possible functional implication of the tagged variants. We observed that an alteration of the KATNAL1 splicing pattern, by favouring the overrepresentation of a short non-functional transcript isoform in the testis, might be the cause behind the observed genetic association. The analysis of the testis transcriptome at the single cell level showed that KATNAL1 transcripts were mostly presented in spermatocytes and early spermatids at puberty, which correlates with its effect of premature exfoliation and loss of the germ cells. Limitations, reasons for caution Although a previous low-powered study reported suggestive associations between KATNAL1 and SPGF, additional genetic association studies in independent populations should be conducted to confirm our findings. Moreover, the statistical power for the overall analysis was appropriate, but the subphenotype analyses were performed with reduced power due to smaller study groups. Wider implications of the findings Our results suggest a relevant role of the common genetic variation of KATNAL1 in the susceptibility to develop the most severe histological phenotypes of NOA (i.e., SCO and MA). Studies like the one presented here may definitively help to develop future non-invasive molecular markers of TESE success. Trial registration number Not Applicable

Analysis of parent-of-origin effects for secondary phenotypes using case-control mother-child pair data.

The detection of parent-of-origin effects (POEs) has become a research focus in genetic association studies since POEs play an important role in explaining the heritability of many complex human disorders. Genetic association studies are commonly conducted based on case-control designs. Case-control genetic association studies often collect additional information on secondary phenotypes other than the case-control status. Various statistical methods have been proposed to analyze the secondary phenotypes, but no methods are specifically tailored for identifying POEs of offspring genes on the secondary phenotypes. The parental origin information may not be determined unambiguously using the genotypes of the test locus for some families, and ignoring such families would lose considerable information. In this article, we focus on case-control mother-child pair design that has been widely used for studying human early life growth and development, and propose a robust and efficient retrospective likelihood method to detect POEs for the secondary phenotypes using multilocus genotypes. The proposed method fully utilizes the information from multilocus genotypes, Hardy-Weinberg equilibrium (HWE), Mendelian inheritance law, and conditional independence between child genotype and maternal covariate given maternal genotype. Large sample properties, including consistency and asymptotic normality, are established for our proposed statistical method. The finite sample performance of our method are demonstrated through extensive simulation studies and application to the Danish National Birth Cohort data.

Haplotype phasing of a bipolar disorder pedigree revealed rare multiple mutations of SPOCD1 gene in the 1p36–35 susceptibility locus

BackgroundThe etiology of bipolar disorder (BD) is poorly understood. Considering the complexity of BD, pedigree-based sequencing studies focusing on haplotypes at specific loci may be practical to discover high-impact risk variants. This study comprehensively examined the haplotype sequence at 1p36–35 BD and recurrent depressive disorder (RDD) susceptibility loci. MethodsWe surveyed BD families in Okinawa, Japan. We performed linkage analysis and determined the phased sequence of the affected haplotype using whole genome sequencing. We filtered rare missense variants on the haplotype. For validation, we conducted a case-control genetic association study on approximately 3000 Japanese subjects. ResultsWe identified a three-generation multiplex pedigree with BD and RDD. Strikingly, we identified a significant linkage with mood disorders (logarithm of odds [LOD] = 3.61) at 1p36–35, supported in other ancestry studies. Finally, we determined the entire sequence of the 6.4-Mb haplotype shared by all affected subjects. Moreover, we found a rare triplet of missense variants in the SPOCD1 gene on the haplotype. Notably, despite the rare frequency, one heterozygote with multiple SPOCD1 variants was identified in an independent set of 88 BD type I genotyping samples. LimitationsThe 1p36–35 sequence was obtained from only a single pedigree. The replicate sample was small. Short-read sequencing might miss structural variants. A polygenic risk score was not analyzed. ConclusionThe 1p36–35 haplotype sequence may be valuable for future BD variant studies. In particular, SPOCD1 is a promising candidate gene and should be validated.

Concussion-Associated Gene Variant COMT rs4680 Is Associated With Elite Rugby Athlete Status.

Concussions are common match injuries in elite rugby, and reports exist of reduced cognitive function and long-term health consequences that can interrupt or end a playing career and produce continued ill health. The aim of this study was to investigate the association between elite rugby status and 8 concussion-associated risk polymorphisms. We hypothesized that concussion-associated risk genotypes and alleles would be underrepresented in elite rugby athletes compared with nonathletes. A case-control genetic association study. Institutional (university). Elite White male rugby athletes [n = 668, mean (SD) height 1.85 (0.07) m, mass 102 (12) kg, and age 29 (7) years] and 1015 nonathlete White men and women (48% men). Genotype was the independent variable, obtained by PCR of genomic DNA using TaqMan probes. Elite athlete status with groups compared using χ 2 and odds ratio (OR). The COMT rs4680 Met/Met (AA) genotype, Met allele possession, and Met allele frequency were lower in rugby athletes (24.8%, 74.6%, and 49.7%, respectively) than nonathletes (30.2%, 77.6%, and 54.0%; P < 0.05). The Val/Val (GG) genotype was more common in elite rugby athletes than nonathletes (OR 1.39, 95% confidence interval 1.04-1.86). No other polymorphism was associated with elite athlete status. Elite rugby athlete status is associated with COMT rs4680 genotype that, acting pleiotropically, could affect stress resilience and behavioral traits during competition, concussion risk, and/or recovery from concussion. Consequently, assessing COMT rs4680 genotype might aid future individualized management of concussion risk among athletes.

Modeling Secondary Phenotypes Conditional on Genotypes in Case–Control Studies

Traditional case–control genetic association studies examine relationships between case–control status and one or more covariates. It is becoming increasingly common to study secondary phenotypes and their association with the original covariates. The Orofacial Pain: Prospective Evaluation and Risk Assessment (OPPERA) project, a study of temporomandibular disorders (TMD), motivates this work. Numerous measures of interest are collected at enrollment, such as the number of comorbid pain conditions from which a participant suffers. Examining the potential genetic basis of these measures is of secondary interest. Assessing these associations is statistically challenging, as participants do not form a random sample from the population of interest. Standard methods may be biased and lack coverage and power. We propose a general method for the analysis of arbitrary phenotypes utilizing inverse probability weighting and bootstrapping for standard error estimation. The method may be applied to the complicated association tests used in next-generation sequencing studies, such as analyses of haplotypes with ambiguous phase. Simulation studies show that our method performs as well as competing methods when they are applicable and yield promising results for outcome types, such as time-to-event, to which other methods may not apply. The method is applied to the OPPERA baseline case–control genetic study.