Association of alpha globin gene copy number with exhaled nitric oxide in a cross-sectional study of healthy Black adults.

INTRODUCTION: The genetic determinants of fractional exhalation of nitric oxide (FeNO), a marker of lung inflammation, are understudied in Black individuals. Alpha globin (HBA) restricts nitric oxide signalling in arterial endothelial cells via interactions with nitric oxide synthase; however, its role in regulating the release of NO from respiratory epithelium is less well understood. We hypothesised that an HBA gene deletion, common among Black individuals, would be associated with higher FeNO. METHODS: Healthy Black adults were enrolled at four study sites in North Carolina from 2005 to 2008. FeNO was measured in triplicate using a nitric oxide analyzer. The -3.7 kb HBA gene deletion was genotyped using droplet digital PCR on genomic DNA. The association of FeNO with HBA copy number was evaluated using multivariable linear regression employing a linear effect of HBA copy number and adjusting for age, sex and serum immunoglobulin-E levels. Post-hoc analysis employing a recessive mode of inheritance was performed. RESULTS: 895 individuals were in enrolled in the study and 720 consented for future genetic research; 643 had complete data and were included in this analysis. Median (25th, 75th) FeNO was 20 (13, 31) ppb. HBA genotypes were: 30 (4.7%) -a/-a, 197 (30.6%) -a/aa, 405 (63%) aa/aa and 8 (1.2%) aa/aaa. Subjects were 35% male with median age 20 (19, 22) years. Multivariable linear regression analysis revealed no association between FeNO and HBA copy number (ß=-0.005 (95% CI -0.042 to 0.033), p=0.81). In the post-hoc sensitivity analysis, homozygosity for the HBA gene deletion was associated with higher FeNO (ß=0.107 (95% CI 0.003 to 0.212); p=0.045). CONCLUSION: We found no association between HBA copy number and FeNO using a prespecified additive genetic model. However, a post hoc recessive genetic model found FeNO to be higher among subjects homozygous for the HBA deletion.

Regulatory Variant rs2535629 in ITIH3 Intron Confers Schizophrenia Risk By Regulating CTCF Binding and SFMBT1 Expression.

Genome-wide association studies have identified 3p21.1 as a robust risk locus for schizophrenia. However, the underlying molecular mechanisms remain elusive. Here a functional regulatory variant (rs2535629) is identified that disrupts CTCF binding at 3p21.1. It is confirmed that rs2535629 is also significantly associated with schizophrenia in Chinese population and the regulatory effect of rs2535629 is validated. Expression quantitative trait loci analysis indicates that rs2535629 is associated with the expression of three distal genes (GLT8D1, SFMBT1, and NEK4) in the human brain, and CRISPR-Cas9-mediated genome editing confirmed the regulatory effect of rs2535629 on GLT8D1, SFMBT1, and NEK4. Interestingly, differential expression analysis of GLT8D1, SFMBT1, and NEK4 suggested that rs2535629 may confer schizophrenia risk by regulating SFMBT1 expression. It is further demonstrated that Sfmbt1 regulates neurodevelopment and dendritic spine density, two key pathological characteristics of schizophrenia. Transcriptome analysis also support the potential role of Sfmbt1 in schizophrenia pathogenesis. The study identifies rs2535629 as a plausibly causal regulatory variant at the 3p21.1 risk locus and demonstrates the regulatory mechanism and biological effect of this functional variant, indicating that this functional variant confers schizophrenia risk by altering CTCF binding and regulating expression of SFMBT1, a distal gene which plays important roles in neurodevelopment and synaptic morphogenesis.

Inner retinal injury in experimental glaucoma is prevented upon AAV mediated Shp2 silencing in a caveolin dependent manner.

SH2 domain containing tyrosine phosphatase 2 (Shp2; PTPN11) regulates several intracellular pathways downstream of multiple growth factor receptors. Our studies implicate that Shp2 interacts with Caveolin-1 (Cav-1) protein in retinal ganglion cells (RGCs) and negatively regulates BDNF/TrkB signaling. This study aimed to investigate the mechanisms underlying the protective effects of shp2 silencing in the RGCs in glaucomatous conditions. Methods: Shp2 was silenced in the Cav-1 deficient mice and the age matched wildtype littermates using adeno-associated viral (AAV) constructs. Shp2 expression modulation was performed in an acute and a chronic mouse model of experimental glaucoma. AAV2 expressing Shp2 eGFP-shRNA under a strong synthetic CAG promoter was administered intravitreally in the animals' eyes. The contralateral eye received AAV-eGFP-scramble-shRNA as control. Animals with Shp2 downregulation were subjected to either microbead injections or acute ocular hypertension experimental paradigm. Changes in inner retinal function were evaluated by measuring positive scotopic threshold response (pSTR) while structural and biochemical alterations were evaluated through H&E staining, western blotting and immunohistochemical analysis of the retinal tissues. Results: A greater loss of pSTR amplitudes was observed in the WT mice compared to Cav-1-/- retinas in both the models. Silencing of Shp2 phosphatase imparted protection against inner retinal function loss in chronic glaucoma model in WT mice. The functional rescue also translated to structural preservation of ganglion cell layer in the chronic glaucoma condition in WT mice which was not evident in Cav-1-/- mice retinas. Conclusions: This study indicates that protective effects of Shp2 ablation under chronic experimental glaucoma conditions are dependent on Cav-1 in the retina, suggesting in vivo interactions between the two proteins.

Contextual cues from cancer cells govern cancer-associated fibroblast heterogeneity.

Cancer cells function as primary architects of the tumor microenvironment. However, the molecular features of cancer cells that govern stromal cell phenotypes remain unclear. Here, we show that cancer-associated fibroblast (CAF) heterogeneity is driven by lung adenocarcinoma (LUAD) cells at either end of the epithelial-to-mesenchymal transition (EMT) spectrum. LUAD cells that have high expression of the EMT-activating transcription factor ZEB1 reprogram CAFs through a ZEB1-dependent secretory program and direct CAFs to the tips of invasive projections through a ZEB1-driven CAF repulsion process. The EMT, in turn, sensitizes LUAD cells to pro-metastatic signals from CAFs. Thus, CAFs respond to contextual cues from LUAD cells to promote metastasis.

Pan-cancer analysis identifies ITIH1 as a novel prognostic indicator for hepatocellular carcinoma.

Although a previous pan-cancer study has reported the expression patterns of ITIHs in various tumors, their analyses have been restricted to limited cancer types. We thus comprehensively analyzed the expression profiles and clinical significances of ITIHs in a broader spectrum of cancers from TCGA. Our results showed that ITIHs were primarily down-regulated in tested cancers. The ITIH members were associated with either survival advantage or disadvantage, depending on the cancer type tested and the genes queried. Importantly, we for the first time demonstrated that ITIH1 had substantially decreased expression in liver hepatocellular carcinoma (LIHC) compared with corresponding normal tissue, and its down-regulation adversely impacted patient outcome. Moreover, ITIH1 expression was consistently declining during the progression of LIHC. Further analysis revealed that ITIH1 may be involved in cellular metabolic processes. Our findings established ITIH1 as a potential diagnostic and prognostic biomarker for LIHC, which awaits future experimental validation.

Knockout of the radical scavenger α1-microglobulin in mice results in defective bikunin synthesis, endoplasmic reticulum stress and increased body weight.

α1-microglobulin (A1M) is a ubiquitous protein with reductase and radical- and heme-binding properties. The protein is mainly expressed in the liver and encoded by the α1-microglobulin-bikunin precursor (AMBP) gene together with the plasma proteinase inhibitor bikunin. The AMBP polypeptide is translated, glycosylated and the C-terminal bikunin part linked via a chondroitin sulfate glycosaminoglycan chain to one or two heavy chains in the endoplasmic reticulum (ER) and Golgi compartments. After proteolytic cleavage, the A1M protein and complexed bikunin parts are secreted separately. The complete physiological role of A1M, and the reason for the co-synthesis with bikunin, are both still unknown. The aim of this work was to develop an A1M knockout (A1M-KO) mouse model lacking expression of A1M, but with a preserved bikunin expression, and to study the phenotypic traits in these mice, with a focus on hepatic endoplasmic reticulum (ER) function. The bikunin expression was increased in the A1M-KO mouse livers, while the bikunin levels in plasma were decreased, indicating a defective biosynthesis of bikunin. The A1M-KO livers also showed an increased expression of transducers of the unfolded protein response (UPR), indicating an increased ER-stress in the livers. At twelve months of age, the A1M-KO mice also displayed an increased body weight, and an increased liver weight and lipid accumulation. Moreover, the KO mice showed an increased expression of endogenous antioxidants in the liver, but not in the kidneys. Together, these results suggest a physiological role of A1M as a regulator of the intracellular redox environment and more specifically the ER folding and posttranslational modification processes, particularly in the liver.

Novel Neuroprotective Agents to Treat Neonatal Hypoxic-Ischemic Encephalopathy: Inter-Alpha Inhibitor Proteins.

Perinatal hypoxia-ischemia (HI) is a major cause of brain injury and mortality in neonates. Hypoxic-ischemic encephalopathy (HIE) predisposes infants to long-term cognitive deficits that influence their quality of life and place a large burden on society. The only approved treatment to protect the brain after HI is therapeutic hypothermia, which has limited effectiveness, a narrow therapeutic time window, and is not considered safe for treatment of premature infants. Alternative or adjunctive therapies are needed to improve outcomes of full-term and premature infants after exposure to HI. Inter-alpha inhibitor proteins (IAIPs) are immunomodulatory molecules that are proposed to limit the progression of neonatal inflammatory conditions, such as sepsis. Inflammation exacerbates neonatal HIE and suggests that IAIPs could attenuate HI-related brain injury and improve cognitive outcomes associated with HIE. Recent studies have shown that intraperitoneal treatment with IAIPs can decrease neuronal and non-neuronal cell death, attenuate glial responses and leukocyte invasion, and provide long-term behavioral benefits in neonatal rat models of HI-related brain injury. The present review summarizes these findings and outlines the remaining experimental analyses necessary to determine the clinical applicability of this promising neuroprotective treatment for neonatal HI-related brain injury.

The Role of α1-Microglobulin (A1M) in Erythropoiesis and Erythrocyte Homeostasis-Therapeutic Opportunities in Hemolytic Conditions.

α1-microglobulin (A1M) is a small protein present in vertebrates including humans. It has several physiologically relevant properties, including binding of heme and radicals as well as enzymatic reduction, that are used in the protection of cells and tissue. Research has revealed that A1M can ameliorate heme and ROS-induced injuries in cell cultures, organs, explants and animal models. Recently, it was shown that A1M could reduce hemolysis in vitro, observed with several different types of insults and sources of RBCs. In addition, in a recently published study, it was observed that mice lacking A1M (A1M-KO) developed a macrocytic anemia phenotype. Altogether, this suggests that A1M may have a role in RBC development, stability and turnover. This opens up the possibility of utilizing A1M for therapeutic purposes in pathological conditions involving erythropoietic and hemolytic abnormalities. Here, we provide an overview of A1M and its potential therapeutic effect in the context of the following erythropoietic and hemolytic conditions: Diamond-Blackfan anemia (DBA), 5q-minus myelodysplastic syndrome (5q-MDS), blood transfusions (including storage), intraventricular hemorrhage (IVH), preeclampsia (PE) and atherosclerosis.

A functional missense variant in ITIH3 affects protein expression and neurodevelopment and confers schizophrenia risk in the Han Chinese population.

The Psychiatric Genomics Consortium (PGC) has recently identified 10 potential functional coding variants for schizophrenia. However, how these coding variants confer schizophrenia risk remains largely unknown. Here, we investigate the associations between eight potential functional coding variants identified by PGC and schizophrenia in a large Han Chinese sample (n = 4022 cases and 9270 controls). Among the eight tested single nucelotide polymorphisms (SNPs), rs3617 (a missense variant, p.K315Q in the ITIH3 gene) showed genome-wide significant association with schizophrenia in the Han Chinese population (P = 8.36 × 10-16), with the same risk allele as in PGC. Interestingly, rs3617 is located in a genomic region that is highly evolutionarily conserved, and its schizophrenia risk allele (C allele) was associated with lower ITIH3 mRNA and protein expression. Intriguingly, mouse neural stem cells stably overexpressing ITIH3 with different alleles of rs3617 exhibited significant differences in proliferation, migration, and differentiation, suggesting the impact of rs3617 on neurodevelopment. Subsequent transcriptome analysis found that the differentially expressed genes in neural stem cells stably overexpressing different alleles of rs3617 were significantly enriched in schizophrenia-related pathways, including cell adhesion, synapse assembly, MAPK and PI3K-AKT pathways. Our study provides convergent lines of evidence suggesting that rs3617 in ITIH3 likely affects protein function and neurodevelopment and thereby confers risk of schizophrenia.

Integrative analyses indicate an association between ITIH3 polymorphisms with autism spectrum disorder.

It is challenge to pinpoint the functional variants among numerous genetic variants. Investigating the spatial dynamics of the human brain transcriptome for genes and exploring the expression quantitative trait loci data may provide the potential direction to identify the functional variants among autism spectrum disorders (ASD) patients. In order to explore the association of ITIH3 with ASD, the present study included three components: identifying the spatial-temporal expression of ITIH3 in the developing human brain using the expression data from the Allen Institute for Brain Science; examining the cis-acting regulatory effect of SNPs on the ITIH3 expression using UK Brain Expression Consortium database; validating the effect of identified SNPs using a case-control study with samples of 602 cases and 604 controls. The public expression data showed that ITIH3 may have a role in the development of human brain and suggested a cis-eQTL effect for rs2535629 and rs3617 on ITIH3 in the hippocampus. Genetic analysis of the above two SNPs suggested that the over-dominant model of rs2535629 was significantly associated with decreased risk of ASD. Convergent lines of evidence supported ITIH3 rs25352629 as a susceptibility variant for ASD.

Towards coeliac-safe bread.

Gluten-free foods cannot substitute for products made from wheat flour. When wheat products are digested, the remaining peptides can trigger an autoimmune disease in 1% of the North American and European population, called coeliac disease. Because wheat proteins are encoded by a large gene family, it has been impossible to use conventional breeding to select wheat varieties that are coeliac-safe. However, one can test the properties of protein variants by expressing single genes in coeliac-safe cereals like maize. One source of protein that can be considered as coeliac-safe and has bread-making properties is teff (Eragrostis tef), a grain consumed in Ethiopia. Here, we show that teff α-globulin3 (Etglo3) forms storage vacuoles in maize that are morphologically similar to those of wheat. Using transmission electron microscopy, immunogold labelling shows that Etglo3 is almost exclusively deposited in the storage vacuole as electron-dense aggregates. Of maize seed storage proteins, 27-kDa γ-zein is co-deposited with Etglo3. Etglo3 polymerizes via intermolecular disulphide bonds in maize, similar to wheat HMW glutenins under non-reducing conditions. Crossing maize Etglo3 transgenic lines with α-, ß- and γ-zein RNA interference (RNAi) lines reveals that Etglo3 accumulation is only dramatically reduced in γ-zein RNAi background. This suggests that Etglo3 and 27-kDa γ-zein together cause storage vacuole formation and behave similar to the interactions of glutenins and gliadins in wheat. Therefore, expression of teff α-globulins in maize presents a major step in the development of a coeliac-safe grain with bread-making properties.

Plasma extracellular vesicle long RNA profiling identifies a diagnostic signature for the detection of pancreatic ductal adenocarcinoma.

OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is difficult to diagnose at resectable stage. Recent studies have suggested that extracellular vesicles (EVs) contain long RNAs. The aim of this study was to develop a diagnostic (d-)signature for the detection of PDAC based on EV long RNA (exLR) profiling. DESIGN: We conducted a case-control study with 501 participants, including 284 patients with PDAC, 100 patients with chronic pancreatitis (CP) and 117 healthy subjects. The exLR profile of plasma samples was analysed by exLR sequencing. The d-signature was identified using a support vector machine algorithm and a training cohort (n=188) and was validated using an internal validation cohort (n=135) and an external validation cohort (n=178). RESULTS: We developed a d-signature that comprised eight exLRs, including FGA, KRT19, HIST1H2BK, ITIH2, MARCH2, CLDN1, MAL2 and TIMP1, for PDAC detection. The d-signature showed high accuracy, with an area under the receiver operating characteristic curve (AUC) of 0.960, 0.950 and 0.936 in the training, internal validation and external validation cohort, respectively. The d-signature was able to identify resectable stage I/II cancer with an AUC of 0.949 in the combined three cohorts. In addition, the d-signature showed superior performance to carbohydrate antigen 19-9 in distinguishing PDAC from CP (AUC 0.931 vs 0.873, p=0.028). CONCLUSION: This study is the first to characterise the plasma exLR profile in PDAC and to report an exLR signature for the detection of pancreatic cancer. This signature may improve the prognosis of patients who would have otherwise missed the curative treatment window.

Galnt11 regulates kidney function by glycosylating the endocytosis receptor megalin to modulate ligand binding.

Chronic kidney disease (CKD) affects more than 20 million Americans and ∼10% of the population worldwide. Genome-wide association studies (GWAS) of kidney functional decline have identified genes associated with CKD, but the precise mechanisms by which they influence kidney function remained largely unexplored. Here, we examine the role of 1 GWAS-identified gene by creating mice deficient for Galnt11, which encodes a member of the enzyme family that initiates protein O-glycosylation, an essential posttranslational modification known to influence protein function and stability. We find that Galnt11-deficient mice display low-molecular-weight proteinuria and have specific defects in proximal tubule-mediated resorption of vitamin D binding protein, α1-microglobulin, and retinol binding protein. Moreover, we identify the endocytic receptor megalin (LRP2) as a direct target of Galnt11 in vivo. Megalin in Galnt11-deficient mice displays reduced ligand binding and undergoes age-related loss within the kidney. Differential mass spectrometry revealed specific sites of Galnt11-mediated glycosylation within mouse kidney megalin/LRP2 that are known to be involved in ligand binding, suggesting that O-glycosylation directly influences the ability to bind ligands. In support of this, recombinant megalin containing these sites displayed reduced albumin binding in cells deficient for Galnt11 Our results provide insight into the association between GALNT11 and CKD, and identify a role for Galnt11 in proper kidney function.

Overproduction of inter-α-trypsin inhibitor heavy chain 1 after loss of Gα13 in liver exacerbates systemic insulin resistance in mice.

The impact of liver disease on whole-body glucose homeostasis is largely attributed to dysregulated release of secretory proteins in response to metabolic stress. The molecular cues linking liver to whole-body glucose metabolism remain elusive. We found that expression of G protein α-13 (Gα13) was decreased in the liver of mice and humans with diabetes. Liver-specific deletion of the Gna13 gene in mice resulted in systemic glucose intolerance. Comparative secretome analysis identified inter-α-trypsin inhibitor heavy chain 1 (ITIH1) as a protein secreted by liver that was responsible for systemic insulin resistance in Gna13-deficient mice. Liver expression of ITIH1 positively correlated with surrogate markers for diabetes in patients with impaired glucose tolerance or overt diabetes. Mechanistically, a decrease in hepatic Gα13 caused ITIH1 oversecretion by liver through induction of O-GlcNAc transferase expression, facilitating ITIH1 deposition on the hyaluronan surrounding mouse adipose tissue and skeletal muscle. Neutralization of secreted ITIH1 ameliorated glucose intolerance in obese mice. Our findings demonstrate systemic insulin resistance in mice resulting from liver-secreted ITIH1 downstream of Gα13 and its reversal by ITIH1 neutralization.

Matrix-degrading protease ADAMTS-5 cleaves inter-α-inhibitor and releases active heavy chain 2 in synovial fluids from arthritic patients.

Destruction of the cartilage matrix in joints is an important feature of arthritis. Proteolytic degradation of cartilage glycoproteins can contribute to the loss of matrix integrity. Human inter-α-inhibitor (IαI), which stabilizes the extracellular matrix, is composed of the light-chain serine proteinase inhibitor bikunin and two homologous heavy chains (HC1 and HC2) covalently linked through chondroitin 4-sulfate. Inflammation promotes the transfer of HCs from chondroitin 4-sulfate to hyaluronan by tumor necrosis factor-stimulated gene-6 protein (TSG-6). This reaction generates a covalent complex between the heavy chains and hyaluronan that can promote leukocyte invasion. This study demonstrates that both IαI and the HC-hyaluronan complex are substrates for the extracellular matrix proteases ADAMTS-5 and matrix metalloprotease (MMP) -3, -7, and -13. The major cleavage sites for all four proteases are found in the C terminus of HC2. ADAMTS-5 and MMP-7 displayed the highest activity toward HC2. ADAMTS-5 degradation products were identified in mass spectrometric analysis of 29 of 33 arthropathic patients, indicating that ADAMTS-5 cleavage occurs in synovial fluid in arthritis. After cleavage, free HC2, together with TSG-6, is able to catalyze the transfer of heavy chains to hyaluronan. The release of extracellular matrix bound HC2 is likely to increase the mobility of the HC2/TSG-6 catalytic unit and consequently increase the rate of the HC transfer reaction. Ultimately, ADAMTS-5 cleavage of HC2 could alter the physiological and mechanical properties of the extracellular matrix and contribute to the progression of arthritis.

The Effect of AMBP SNPs, Their Haplotypes, and Gene-Environment Interactions on the Risk of Atherothrombotic Stroke Among the Chinese Population.

Background/Objectives: Ischemic stroke (IS) is a severe and complex disorder with high morbidity and mortality rates and it has been associated with both environmental and genetic predisposing factors. The purpose of this study was to evaluate the association of the alpha-1-microglobulin/bikunin precursor (AMBP) gene polymorphisms with IS and any possible interactions between specific AMBP alleles and traditional risk factors among a Han Chinese cohort. Materials and Methods: We conducted a candidate gene study designed to characterize nine (9) single nucleotide polymorphisms (SNPs) of the AMBP gene among 195 patients with atherothrombotic stroke (ATS) (a major subtype of IS) and 184 nonstroke controls. Allelic and genotypic frequency differences were evaluated using a logistic regression model. False discovery rate (FDR) correction for multiple comparisons was used. The interactional analyses were performed using the multifactor dimensionality reduction test. Results: We found an association between the rs2567698 CC genotype (odds ratio [OR], 95% confidence interval [CI]: 2.176, 1.159-4.086) and the T allele (OR, 95% CI: 0.654, 0.446-0.960) with risk of ATS in men. However, these associations did not survive FDR correction. In haplotype analyses, the GCCCCCCCC haplotype had a higher frequency (OR, 95% CI: 2.191, 1.048-4.580) in ATS in the ≥45 years of age subgroup, whereas the GCCTCCCCC haplotype decreased the risk for ATS (OR, 95% CI: 0.543, 0.345-0.853) in men. In addition, we also found interactions for ATS risk between SNPs in the AMBP gene and modifiable risk factors for ATS, including: rs11788411 and hypertension in the overall population and women; rs2251680 and hypertension in subjects aged 45 years and older, as well as the interaction among hypertension and the rs2567698 and rs10817564 genotypes in men. Conclusion: Our results show a possible association between AMBP SNP haplotypes and gene-environment interactions with ATS susceptibility in a Han Chinese cohort.

rA1M-035, a Physicochemically Improved Human Recombinant α1-Microglobulin, Has Therapeutic Effects in Rhabdomyolysis-Induced Acute Kidney Injury.

AIMS: Human α1-microglobulin (A1M) is an endogenous reductase and radical- and heme-binding protein with physiological antioxidant protective functions. Recombinant human A1M (rA1M) has been shown to have therapeutic properties in animal models of preeclampsia, a pregnancy disease associated with oxidative stress. Recombinant A1M, however, lacks glycosylation, and shows lower solubility and stability than A1M purified from human plasma. The aims of this work were to (i) use site-directed mutagenesis to improve the physicochemical properties of rA1M, (ii) demonstrate that the physicochemically improved rA1M displays full in vitro cell protective effects as recombinant wild-type A1M (rA1M-wt), and (iii) show its therapeutic potential in vivo against acute kidney injury (AKI), another disease associated with oxidative stress. RESULTS: A novel recombinant A1M-variant (rA1M-035) with three amino acid substitutions was constructed, successfully expressed, and purified. rA1M-035 had improved solubility and stability compared with rA1M-wt, and showed intact in vitro heme-binding, reductase, antioxidation, and cell protective activities. Both rA1M-035 and rA1M-wt showed, for the first time, potential in vivo protective effects on kidneys using a mouse rhabdomyolysis glycerol injection model of AKI. INNOVATION: A novel recombinant A1M-variant, rA1M-035, was engineered. This protein showed improved solubility and stability compared with rA1M-wt, full in vitro functional activity, and potential protection against AKI in an in vivo rhabdomyolysis mouse model. CONCLUSION: The new rA1M-035 is a better drug candidate than rA1M-wt for treatment of AKI and preeclampsia in human patients.

Inter-α-inhibitor deficiency in the mouse is associated with alterations in anxiety-like behavior, exploration and social approach.

In recent years, several genome-wide association studies have identified candidate regions for genetic susceptibility in major mood disorders. Most notable are regions in a locus in chromosome 3p21, encompassing the genes NEK4-ITIH1-ITIH3-ITIH4. Three of these genes represent heavy chains of the composite protein inter-α-inhibitor (IαI). In order to further establish associations of these genes with mood disorders, we evaluated behavioral phenotypes in mice deficient in either Ambp/bikunin, which is necessary for functional ITIH1 and ITIH3 complexes, or in Itih4, the gene encoding the heavy chain Itih4. We found that loss of Itih4 had no effect on the behaviors tested, but loss of Ambp/bikunin led to increased anxiety-like behavior in the light/dark and open field tests and reduced exploratory activity in the elevated plus maze, light/dark preference and open field tests. Ambp/bikunin knockout mice also exhibited a sex-dependent exaggeration of acoustic startle responses, alterations in social approach during a three-chamber choice test, and an elevated fear conditioning response. These results provide experimental support for the role of ITIH1/ITIH3 in the development of mood disorders.

Underexpression of α-1-microglobulin/bikunin precursor predicts a poor prognosis in oral squamous cell carcinoma.

In the present study, in order to identify novel diagnostic biomarkers for the malignant behavior of oral squamous cell carcinoma (OSCC), we determined the proteomic profiles of several OSCC cell lines and keratinocytes by two-dimensional fluorescence difference gel electrophoresis and liquid chromatography tandem mass spectrometry. The protein expression level of α-1-microglobulin/bikunin precursor (AMBP) was found to be significantly lower in the OSCC cell lines than in the keratinocytes, and a significant decrease in AMBP mRNA expression was confirmed in the OSCC cell lines by RT-qPCR. To investigate the biological function of AMBP in OSCC, the cells were transiently transfected with an AMBP overexpression vector; the AMBP-overexpressing cells exhibited a significantly decreased invasion and migration in comparison to the mock-transfected control cells, although no significant changes in cell proliferation were observed. Immunohistochemistry revealed that the underexpression of AMBP was significantly associated with a high metastatic potential to cervical lymph nodes and a poor overall survival. Thus, the expression of AMBP is an independent predictive factor of cervical lymph node metastasis and a prognostic factor of overall survival, and it is involved in both cell invasion and metastasis in cervical lymph nodes in OSCC.

ITIH3 and ITIH4 polymorphisms and depressive symptoms during pregnancy in Japan: the Kyushu Okinawa Maternal and Child Health Study.

ITIH3 and ITIH4 are involved in the stabilization of the extracellular matrix. Several genome-wide association studies and case-control studies regarding psychiatric disorders have identified ITIH3 and ITIH4 single nucleotide polymorphisms (SNPs). The present case-control study examined the relationship between ITIH3 SNPs rs2535629 and rs736408 and ITIH4 SNPs rs3821831 and rs2239547 and depressive symptoms during pregnancy in Japan. Cases comprised 273 women with depressive symptoms during pregnancy defined as a Center for Epidemiological Studies Depression Scale (CES-D) score ≥ 16. Control subjects comprised 1176 women without depressive symptoms during pregnancy, according to the CES-D criteria, who had not been diagnosed with depression by a doctor. Adjustment was made for age, gestation at baseline, region of residence, the presence of children, family structure, smoking, employment, and education. Compared with the TT genotype of ITIH4 SNP rs2239547, the CC genotype was significantly related to a reduced risk of depressive symptoms during pregnancy: the adjusted odds ratio (95% CI) was 0.84 (0.63-1.11) for the TC genotype and 0.57 (0.36-0.91) for the CC genotype. ITIH3 SNPs rs2535629 and rs736408 and ITIH4 SNP rs3821831 were not related to depressive symptoms during pregnancy. The GCCT haplotype of rs2535629, rs736408, rs3821831, and rs2239547 was significantly positively associated with depressive symptoms during pregnancy. A significant interaction was found between rs2239547 and the presence of children. This is the first study to show significant associations of ITIH4 SNP rs2239547 and the GCCT haplotype with depressive symptoms during pregnancy. The effect of the presence of children might depend on rs2239547.