Kininogen-1 Research Articles

Variants in the Atherogenic ALOX5AP, THBD, and KNG1 Genes Potentiate the Risk of Ischemic Stroke via a Genetic Main Effect and Epistatic Interactions in a Chinese Population

Ischemic stroke (IS) is a multifactorial disease that displays a strong genetic predisposition. However, the genetic architecture of IS has yet to be fully elucidated. It was hypothesized that epistasis between genes in multiple atherothrombotic pathways may play a vital role in determining the susceptibility to IS. The aim of the present study was to investigate the contributions of the hypothesized genetic factors to IS and the interactions between these genetic factors in a Chinese population. In this study, 351 cases with IS and 417 control subjects from a Chinese population were genotyped for single-nucleotide polymorphisms (SNPs) in 12 genes hypothesized to be involved in atherosclerosis, coagulation, and related pathways. We examined SNP main effects and epistatic interactions between these polymorphic loci. rs710446 of the KNG1 gene was associated with IS susceptibility based on an additive genetic model (rs710446: P = .012; odds ratio [OR], 1.247; 95% confidence interval [CI], 1.050-1.481) after adjusting for covariates. Furthermore, an epistatic interaction between the ALOX5AP, THBD, and KNG1 gene was also identified in association with stroke susceptibility (P < .001 after 1000 permutations). Based on the chi-squared test, the OR of the high-risk combination of the three-locus model increased the risk of IS by 2.53-fold (95% CI, 1.60-4.01; P < .0001). Our findings support the association of the epistatic interactions of ALOX5AP, THBD, and KNG1 and present novel evidence for the main effect of KNG1 gene on IS susceptibility, suggesting a modulation of stroke risk by a genetic main effect and gene-gene interactions.

Hyperexpression of Inflammatory Genes in Placental Tissue in Patients with Hemoglobin SC Disease

Introduction: Pregnant women with sickle cell disease (SCD) are at higher risk of obstetrical complications, such as preeclampsia and preterm delivery. The placental circulation is susceptible to vaso-occlusion, which may contribute to fibrosis, necrosis and infarction of the placental villi. Restricted uteroplacental circulation is associated with low birth weight and fetal growth restriction. The mechanisms by which SCD affects placental physiology are largely unknown, and the chronic inflammatory state may be involved in this pathological process. The aim of this study was to evaluate the expression of mediators of inflammatory response in the placenta of pregnant women with SCD.Methods: This is an ongoing case control study that has recruited 4 pregnant patients with hemoglobin SC disease and 4 healthy control pregnant women paired for gestational age and route of delivery. Placental tissue was collected for total RNA extraction and synthesis of cDNA. Samples were run on a qPCR Array (Inflammatory Response and Autoimmunity RT2 Profiler, Qiagen), which includes 84 genes. Gene expression levels were normalized to housekeeping genes (ACTB, B2M, and RPLP0), and will be further validated by RT-qPCR.Results: Two patients underwent cesarean delivery and one patient delivered preterm. The analysis of placental gene expression showed that 16 genes were upregulated. The top 5 most upregulated genes were CCL24 (18.1 fold), TLR9 (8.6 fold), inducible nitric oxide synthase NOS2 (6.8 fold), IL17A (5.4 fold), and CCL16 (4.8 fold). Upregulated genes also included other chemokines and their receptors (CCL21, CCL22, CXCL3, and CCR7), interleukins (IL1RAP, IL5, and IL9), another toll-like receptor (TLR3), transcription factor BCL6, CD40 ligand (CD40LG), and kininogen 1 (KNG1). Interestingly, the two genes found to be downregulated were CRP (C reactive protein) and IFNG (interferon gamma).Conclusion: This is the first study of differential gene expression in placenta of patients with SCD. Despite patients with hemoglobin SC disease being clinically less severe than homozygous sickle cell anemia, we have found that placental tissue in this subset of patients displays increased expression of several genes involved in inflammatory response. Our data support local inflammatory changes in the placenta of these patients that might explain the development of complications in SCD pregnancies. Although the association between obstetric complications and SCD has been long described, our data should contribute to better define the mechanisms affecting the placental physiology that lead to complications, such as preeclampsia. DisclosuresNo relevant conflicts of interest to declare.

Kininogen 1 and insulin‐like growth factor binding protein 6: candidate serum biomarkers of proliferative vitreoretinopathy

BackgroundThe aim was to validate whether kininogen 1 (KNG1) or insulin‐like growth factor binding protein 6 (IGFBP‐6) are serum biomarkers of proliferative vitreoretinopathy (PVR).MethodsSamples from vitreous and corresponding serum samples were collected from patients with PVR. The donor vitreous samples and serum samples from healthy volunteers and volunteers who had undergone vitrectomies for other conditions were used as controls. The samples were subsequently analysed using Western blotting (WB) and enzyme‐linked immunosorbent assay.ResultsThe Western blotting outcomes indicated both IGFBP‐6 and KNG1 could be specifically detected in the vitreous and serum samples of patients with PVR. The concentrations of KNG1 and IGFBP‐6 were significantly higher in both vitreous and serum samples from patients with severe PVR than in the samples from patients with moderate PVR. The serum concentrations of KNG1 or IGFBP‐6 had decreased by the post‐vitrectomy examinations. The receiver operating characteristic (ROC) analyses when the concentrations of IGFBP‐6 or KNG1 were greater than 181.4-pg/ml or 441.75-ng/ml, respectively, predicted severe PVR with both a sensitivity and specificity of over 70 per cent. When the concentrations of IGFBP‐6 or KNG1 were greater than 98.5-pg/ml or 88.5-ng/ml, respectively, they predicted the PVR prognosis with both a sensitivity and specificity of 80 per cent.ConclusionsKNG1 and IGFBP‐6 may be candidate serum biomarkers of PVR.

Proteomics research about effect of zinc finger protein 139 knockdown with RNA interference on serum of orthotopic transplantation nude mouse with gastric cancer

Objective To observe the effect of inhibiting zinc finger protein 139 (ZNF139) on serum proteins in orthotopic transplantation nude mice model with gastric cancer,and to investigate the mechanism for ZNF139 involved in progression of gastric cancer.Methods Small interfering RNA (siRNA) recombinant plasmid targeting ZNF139 gene was constructed and imported into gastric cancer cell line SGC7901.After screened stably transfected cells by G418,gastric cancer cells were injected into nude mice subcutaneously.Transplanted tumor tissue was constructed through repeatedly subcutaneously inoculating and passaging between nude mice,and 6th-generation subcutaneous tumor was pasted with OB biological glue orthotopically on the stomach wall of BALB/c nude mice.The proteins in nude mice serum samples were separated by fluorescence two-dimensional difference gel electrophoresis (2D-DIGE) and the differential proteins were identified by liquid chromatography-mass spectrometry (LC-MS).Parts of the proteins were identified by Western blotting.Results The tumor-forming rate in orthopotic implantation was 100％ (15/15).ZNF139 was inhibited obviously,and the tumor growth of orthotopic transplantation nude mice model was obviously slower in siRNA-ZNF139 group.Five differential proteins were identified in orthotopic transplantation nude mice serum samples.Results showed that apolipoprotein E (APOE) and the kininogen 1 (KNG1) were down-regulated in gastric cancer orthotopic mouse serum,and mTERF was upregulated after siRNA-ZNF139 interference.Results of Western blotting were consistent with proteomics.Conclusion ZNF139 in gastric cancer may up-regulate serum levels of APOE and KNG1,while down-regulate mTERF to participate in the progression of gastric cancer. Key words: Orthotopic transplantation of gastric cancer; Zinc finger protein 139; Proteomics

Peptidomic Identification of Serum Peptides Diagnosing Preeclampsia

We sought to identify serological markers capable of diagnosing preeclampsia (PE). We performed serum peptide analysis (liquid chromatography mass spectrometry) of 62 unique samples from 31 PE patients and 31 healthy pregnant controls, with two-thirds used as a training set and the other third as a testing set. Differential serum peptide profiling identified 52 significant serum peptides, and a 19-peptide panel collectively discriminating PE in training sets (n = 21 PE, n = 21 control; specificity = 85.7% and sensitivity = 100%) and testing sets (n = 10 PE, n = 10 control; specificity = 80% and sensitivity = 100%). The panel peptides were derived from 6 different protein precursors: 13 from fibrinogen alpha (FGA), 1 from alpha-1-antitrypsin (A1AT), 1 from apolipoprotein L1 (APO-L1), 1 from inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4), 2 from kininogen-1 (KNG1), and 1 from thymosin beta-4 (TMSB4). We concluded that serum peptides can accurately discriminate active PE. Measurement of a 19-peptide panel could be performed quickly and in a quantitative mass spectrometric platform available in clinical laboratories. This serum peptide panel quantification could provide clinical utility in predicting PE or differential diagnosis of PE from confounding chronic hypertension.

Abstract MP56: A Gene-centric Association Study of Activated Partial Thromoplastin Time in European Americans and African Americans: The ARIC Study

Introduction: Activated partial thromboplastin time (aPTT) is commonly used to screen for coagulation factor deficiencies. Shorter aPTT is also a risk marker for incident and recurrent venous thromboembolism (VTE). Genetic factors influencing aPTT are not well understood. aPTT was associated with common genetic variants of coagulation factors V (F5), XI (F11), XII (F12), KNG1, HRG, and ABO in previously reported genome-wide association studies (GWAS) that were conducted in individuals of European ancestry; no data have been reported in other race groups. Hypothesis: The present study aimed to identify aPTT-related gene variants in European Americans (EAs) and African Americans (AAs). Methods: We conducted a large-scale candidate gene study for aPTT in 9,719 EAs and 2,799 AAs from the Atherosclerosis Risk in Communities (ARIC) study. Subjects on anticoagulants were excluded. Nearly 50,000 single nucleotide polymorphisms (SNPs) located in 2,100 candidate genes were genotyped by the Candidate gene Association Resource (CARe) gene chip. The association between each SNP and aPTT was assessed with an additive genetic model using linear regression adjusted for age, sex, and field center. We additionally adjusted for principal components in AAs to account for potential population stratification. P-value for significant threshold was set at 2x10-6 after accounting for multiple testing. Results: In EAs, fifty-five SNPs from F5, HRG, KNG1, F11, F12, and ABO genes exceeded the significant p-value threshold. The signals in HRG, KNG1, F11, F12, and ABO genes replicated the previously reported GWAS findings. The top variant in F5 identified in EAs was only weakly associated with the previously reported GWAS variant (rs2239852, p=1.89x10-08 and r2=0.02 with rs9332701 reported in the previously reported GWAS). In AAs, twenty-seven SNPs from the HRG, KNG1, F12, and ABO genes were significantly associated with aPTT. The top signals from the HRG (rs9898, p=1.19x10-27) and KNG1 genes ( rs710446 , p=8.41x10-42) replicated the previously reported signals in EAs with similar effect size and direction of association, but the top signals in the F12 and ABO genes were weakly associated with the previously reported variants in EAs (rs1801020 in F12: p=1.01x10-84 and r2=0.12 with rs2545801, and rs8176722 in ABO: p=1.62x10-29 and r2=0.26 with rs687621 , respectively). Conclusions: Our study replicated the previously reported associations of aPTT with HRG, KNG1, F11, F12, and ABO genes in EAs and with HRG and KNG1 in AAs. The signals from F5 identified in EAs and from F12 and ABO identified in AAs may represent new genetic variants for aPTT.

A gene-centric analysis of activated partial thromboplastin time and activated protein C resistance using the HumanCVD focused genotyping array

Activated partial thromboplastin time (aPTT) is an important routine measure of intrinsic blood coagulation. Addition of activated protein C (APC) to the aPTT test to produce a ratio, provides one measure of APC resistance. The associations of some genetic mutations (eg, factor V Leiden) with these measures are established, but associations of other genetic variations remain to be established. The objective of this work was to test for association between genetic variants and blood coagulation using a high-density genotyping array. Genetic association with aPTT and APC resistance was analysed using a focused genotyping array that tests approximately 50 000 single-nucleotide polymorphisms (SNPs) in nearly 2000 cardiovascular candidate genes, including coagulation pathway genes. Analyses were conducted on 2544 European origin women from the British Women's Heart and Health Study. We confirm associations with aPTT at the coagulation factor XII (F12)/G protein-coupled receptor kinase 6 (GRK6) and kininogen 1 (KNG1)/histidine-rich glycoprotein (HRG) loci, and identify novel SNPs at the ABO locus and novel locus kallikrein B (KLKB1)/F11. In addition, we confirm association between APC resistance and factor V Leiden mutation, and identify novel SNP associations with APC resistance in the HRG and F5/solute carrier family 19 member 2 (SLC19A2) regions. In conclusion, variation at several genetic loci influences intrinsic blood coagulation as measured by both aPTT and APC resistance.

Kininogen-1 and insulin-like growth factor binding protein-6 as serum biomarkers for proliferative vitreoretinopathy

To evaluated the predictive potential of kininogen-1 and insulin-like growth factor binding protein-6 (IGFBP-6) for PVR. Vitreous and serum samples were obtained from 24 PVR patients. Vitreous from 8 donated normal eyes, and serum samples from 20 healthy volunteers served as control. Patients who underwent vitrectomy with C(3)F(8) gas tamponade (n = 15) and silicone tamponade (n = 8) and patients who experienced recurrent retinal detachment after scleral buckling surgery (n = 8) were recruited for serum tests as well. Western blot analysis was employed to detect the presence of kininogen-1 and IGFBP-6. The protein concentration was measured by using enzyme linked immunosorbent assay (ELISA) analysis. All date were analyzed with the SPSS 3.0 for Windows (only-way analysis of variance and t test). Western blot analysis displayed that except that IGFBP-6 was absent in 2 PVR vitreous, both kininogen-1 and IGFBP-6 were otherwise found in all PVR vitreous and serum samples. Neither kininogen-1 nor IGFBP-6 can be detected in normal vitreous or serum samples. Protein expression was more intensive in severe PVR vitreous than in mild PVR vitreous, which was confirmed by a significantly higher concentration of each protein in sever PVR vitreous. The ELISA outcomes documented that kininogen-1 concentration in vitreous were significantly higher in severe PVR patients than those in mild PVR (281.0 ± 63.0 & 237.5 ± 32.1) µg/L (t = 5.44, P < 0.05). Kininogen-1 was about 2 times higher in serum than in vitreous (443.3 ± 190.1) µg/L (t = 5.27, P < 0.05). At 6 months after vitrectomy with gas tamponade in 15 patients, their kininogen-1 level in serum was significantly lower than that of preoperation (81.9 ± 18.6 & 443.3 ± 190.1) µg/L (t = 5.26, P < 0.05) and encircling failure group was (116.8 ± 45.1) µg/L, it was higher than that of normal and silicone tamponade groups (t = 3.95, 4.34;P < 0.05). Similarly, IGFBP-6 concentration in vitreous were significantly higher in severe PVR patients than those in mild PVR (352.9 ± 64.4 & 283.9 ± 69.9) ng/L (t = 5.08, P < 0.05) and its level in serum was (185.3 ± 34.9) ng/L and lower than that of in vitreous(t = 7.95, P < 0.05). At 6 months after vitrectomy with gas tamponade in 15 patients, their IGFBP-6 level in serum decreased comparing that of preoperation (65.4 ± 31.8) ng/L (t = 11.10, P < 0.05) and encircling failure group was (109.2 ± 6.6) ng/L, it was higher than that of normal and silicone tamponade groups (t = 3.16, 2.77; P < 0.05). Kininogen-1 and IGFBP-6 are presented in serum and vitreous in PVR patients. The strength of protein expression is related to the severity of PVR. These results suggested that kininogen-1 and IGFBP-6 can be biomarkers for severe PVR.