Accelerated epigenetic aging in alcohol dependence.

Alcohol dependence poses a global health threat associated with aging and reduced life expectancy. Recently, aging research through deoxyribonucleic acid (DNA) methylation has gained attention. New epigenetic clocks have been developed; however, no study has investigated GrimAge components, GrimAge2 components and DunedinPACE in patients with alcohol dependence. In this study, we aimed to perform epigenetic clock analysis to evaluate epigenetic age acceleration and DNA methylation-based age-predictive components in patients with alcohol dependence and controls. We utilized publicly available DNA methylation data (GSE98876) for our analysis. Additionally, we compared the values of the same items before and after the patients underwent a treatment program. The dataset comprised 23 controls and 24 patients. We observed that DunedinPACE accelerated more in patients with alcohol dependence. AgeAccelGrim and AgeAccelGrim2 decelerated more after the treatment program than before, and beta-2-microglobulin and Cystatin C decreased after the treatment program than before. These findings are crucial as they affect the cranial nerve area, potentially contributing to cognitive dysfunction and psychiatric symptoms in patients with alcohol dependence.

Epigenome-wide association study on methamphetamine dependence.

Repeated abuse of methamphetamine (METH) can cause dependence, repeated relapse of psychotic symptoms, compulsive drug-seeking behaviour, and various neurological symptoms. These long-term biological changes may be associated with epigenetic mechanisms; however, the association between METH use and epigenetic mechanisms has been poorly investigated. Thus, we performed an epigenome-wide association study of METH dependence using genomic DNA extracted from the blood samples of 24 patients with METH dependence and 24 normal controls. All participants were of Japanese descent. We tested the association between METH dependence and DNA methylation using linear regression analysis. We found epigenome-wide significant associations at four CpG sites, one of which occurred in the CNOT1 gene and another in the PUM1 gene. We especially noted the CNOT1 and PUM1 genes as well as several other genes that indicated some degree of association with METH dependence. Among the relatively enriched Gene Ontology terms, we were interested in terms of mRNA metabolism, respirasome, and excitatory extracellular ligand-gated ion channel activity. Among the relatively enriched Kyoto Encyclopedia of Genes and Genome pathways, we noted pathways of several neurological diseases. Our results indicate that genetic changes akin to those in other psychiatric or neurodegenerative disorders may also occur via epigenetic mechanisms in patients with METH dependence.

Association study of a single nucleotide polymorphism in the hypoxia response element of the macrophage migration inhibitory factor gene promoter with suicide completers in the Japanese population.

BACKGROUND: More than 800 000 people die by suicide annually. The heritability of suicide is 30%-50%. We focused on the hypoxia response element (HRE), which promotes the expression of macrophage migration inhibitory factor (MIF) via the hypoxia-inducible factor (HIF) pathway, important in neurogenesis and neuroprotection. We examined a genetic polymorphism of rs17004038, a single-nucleotide polymorphism (SNP), in suicide completers and controls. METHODS: The study population included 1336 suicide completers and 814 unrelated healthy controls. All participants were Japanese. We obtained peripheral blood, extracted DNA, and genotyped the patients for SNP rs17004038 (C > A). RESULTS: No significant differences were observed between the two groups in either the allele or genotype analyses. Subgroup analyses by sex, age (<40 or ≥40), and suicide method (violent or nonviolent suicide) were performed with similar results. CONCLUSION: No association was observed between SNP rs17004038 and suicide completion. Although it is challenging to collect a large number of samples from suicide completers, further MIF-related genetic studies, including those of rs17004038, are necessary with larger sample sizes.

Accelerated epigenetic aging and decreased natural killer cells based on DNA methylation in patients with untreated major depressive disorder.

Major depressive disorder (MDD) is known to cause significant disability. Genome-wide DNA methylation (DNAm) profiles can be used to estimate biological aging and as epigenetic clocks. However, information on epigenetic clocks reported in MDD patients is inconsistent. Since antidepressants are likely confounders, we evaluated biological aging using various DNAm-based predictors in patients with MDD who had never received depression medication. A publicly available dataset consisting of whole blood samples from untreated MDD patients (n = 40) and controls (n = 40) was used. We analyzed five epigenetic clocks (HorvathAge, HannumAge, SkinBloodAge, PhenoAge, and GrimAge), DNAm-based telomere length (DNAmTL), and DNAm-based age-related plasma proteins (GrimAge components), as well as DNAm-based white blood cell composition. The results indicate that patients with untreated MDD were significantly associated with epigenetic aging acceleration in HannumAge and GrimAge. Furthermore, a decrease in natural killer cells, based on DNAm, was observed in patients with untreated MDD.

Association of two variable number of tandem repeats in the monoamine oxidase A gene promoter with suicide completion: The present study and meta-analysis.

BACKGROUND: One potential cause of suicide is serotonergic dysfunction. Sex differences have been reported to modulate the effects of serotonergic polymorphisms. Monoamine oxidase A (MAOA) is an enzyme that degrades serotonin and is located on the X chromosome. A previous study indicated that the upstream (u) variable number of tandem repeat (VNTR) in the MAOA gene promoter may be associated with suicide. However, a meta-analysis showed that this polymorphism may not be related to suicide. According to a recent study, compared with the uVNTR, the distal (d)VNTR and the haplotypes of the two VNTRs modulate MAOA expression. METHODS: We examined the two VNTRs in the MAOA gene promoter in 1007 subjects who committed suicide and 844 healthy controls. We analyzed the two VNTRs using fluorescence-based polymerase chain reaction assays. We conducted a meta-analysis for the two VNTRs to update it. RESULTS: Our results demonstrated that neither the genotype-based associations nor allele/haplotype frequencies of the two VNTRs were significantly associated with suicide. In the meta-analysis, we did not indicate relationships between uVNTR and suicide nor did we identify articles analyzing dVNTR in suicide. CONCLUSION: Overall, we did not find a relationship between the two VNTRs in the MAOA promoter and suicide completion; thus, warranting further studies are required.

Epigenetic clock analysis reveals increased plasma cystatin C levels based on DNA methylation in major depressive disorder.

Major depressive disorder (MDD) is a common mental illness and a major public health concern worldwide. Depression is associated with epigenetic changes that regulate gene expression, and analyzing these changes may help elucidate the pathophysiology of MDD. Genome-wide DNA methylation (DNAm) profiles can function as 'epigenetic clocks' that can help estimate biological aging. Here, we assessed biological aging in patients with MDD using various DNAm-based indicators of epigenetic aging. We used a publicly available dataset containing data obtained from the whole blood samples of MDD patients (n = 489) and controls (n = 210). We analyzed five epigenetic clocks (HorvathAge, HannumAge, SkinBloodAge, PhenoAge, and GrimAge) and DNAm-based telomere length (DNAmTL). We also investigated seven DNAm-based age-predictive plasma proteins (including cystatin C) and smoking status, which are components of GrimAge. Following adjustment for confounding factors such as age and sex, patients with MDD showed no significant difference in epigenetic clocks and DNAmTL. However, DNAm-based plasma cystatin C levels were significantly higher in patients with MDD than controls. Our findings revealed specific DNAm changes predicting plasma cystatin C levels in MDD. These findings may help elucidate the pathophysiology of MDD, leading to the development of new biomarkers and medications.

Psychological Distress among University Staff before and during the COVID-19 Pandemic.

(1) Background: The COVID-19 pandemic has distressed many populations worldwide, and since its beginning, many institutes have performed cross-sectional studies to assess mental health. We longitudinally examined psychological distress and depressive symptoms among university staff in Japan from 2019 to 2021, before and during the COVID-19 pandemic.; (2) Methods: Participants were teachers and hospital staff working at institutions related to Kobe University, who completed the Brief Job Stress Questionnaire (BJSQ) from 2019 to 2021. This study used the definition recommended by the guideline to identify high-stress. We analyzed the relationship between those who identified as having high-stress before versus during the COVID-19 pandemic using logistic regression analysis (adjusted for age, sex, and occupation).; (3) Results: Results showed that Stress Reaction scores increased slightly in 2020 and significantly in 2021. Time and other factors had a synergistic effect on mental health. The increase in Stress Reaction was significantly associated with females and nurses over the three years. Those with high-stress in 2019 had approximately twenty-fold odds ratios (OR) of having high-stress in 2020 and 2021.; (4) Conclusions: The long-term COVID-19 pandemic may disturb university staff's mental health. Those who originally experienced high levels of stress were vulnerable to the negative effects of the COVID-19 pandemic.

Psychological symptoms and related risk factors among healthcare workers and medical students during the early phase of the COVID-19 pandemic in Japan.

Aim: The aim of this study was to investigate the mental health status of healthcare workers and medical students during the early phase of the COVID-19 pandemic. Methods: An online questionnaire was administered to 637 students and 3189 healthcare workers from May to July, 2020. The patient healthcare questionnaire-9 (PHQ-9) and state anxiety (A-State) of the state-trait anxiety inventory-form (STAI) were used to assess depression and anxiety symptoms, respectively. Individuals were categorized into severe (15 or higher) depression and severe (50-51 or higher) anxiety groups. Results: Healthcare workers and those taking care of COVID-19 patients had a higher risk of severe depression (PHQ-9 scores >15) than other comparison groups. Students and men also had a higher risk of severe anxiety (STAI > 50-51). Multivariable logistic regression analysis showed that healthcare workers had a fivefold higher risk of developing severe depression symptoms (adjusted odds ratio [OR] = 4.99, confidence interval [CI] 2.24-5.97, P-value < 0.001) and those taking care of COVID-19 patients had 2.8-fold higher risk of developing severe depression symptoms (OR 2.75, CI 1.36-5.53, P-value = 0.005). Conclusion: Both medical students and healthcare workers have been experiencing depression and anxiety symptoms during the first wave of the pandemic. Our findings showed a high rate of severe anxiety symptoms in medical students and a high rate of severe depression symptoms in healthcare workers. Those who treated COVID-19 patients were at greater risk of developing major depressive disorder than those who treated non-COVID-19 patients.

miR-19b is elevated in peripheral blood of schizophrenic patients and attenuates proliferation of hippocampal neural progenitor cells.

MicroRNAs (miRNAs) have been investigated in neurodevelopmental and psychiatric disorders including schizophrenia (SZ). Previous studies showed miRNAs dysregulation in postmortem brain tissues and peripheral blood of SZ patients. These suggest that miRNAs may play a role in the pathophysiology of SZ and be a potential biomarker of SZ. Previous studies also showed that miRNAs regulated neurogenesis and that neurogenesis was involved in the pathophysiology of SZ. In addition, a recent study showed that miR-19a and 19b, enriched in neural progenitor cells (NPC) in adult hippocampus, were increased in human NPC derived from induced pluripotent stem cell derived from SZ patients. However, it remains unclear whether the levels of miR-19a and 19b are altered in peripheral blood of SZ patients and how miR-19a and 19b affects neurogenesis. To elucidate them, first we examined the levels of miR-19a and 19b in peripheral blood of SZ patients with quantitative RT-PCR and showed that the level of miR-19b, but not miR-19a, was significantly higher (miR-19a: p = 0.5733, miR-19b: p = 0.0038) in peripheral blood of SZ patients (N = 22) than that of healthy controls (N = 19). Next, we examined the involvement of miR-19b in proliferation and survival of mouse neonatal mice hippocampus-derived NPC with BrdU assay and TUNEL assay. The silencing of miR-19b significantly increased proliferation (N = 5, p = 0.0139), but not survival (N = 5, p = 0.9571), of neonatal mice hippocampus-derived NPC. These results suggest that the level of miR-19b in peripheral blood is a potential biomarker of schizophrenia and that the higher level of miR-19b may increase the vulnerability of SZ via attenuating proliferation of hippocampal NPC.

Major depressive disorder-associated SIRT1 locus affects the risk for suicide in women after middle age.

A recent genome-wide association study (GWAS) for major depressive disorder (MDD) in Chinese women identified a single-nucleotide polymorphism (SNP), rs12415800, near the Sirtuin1 (SIRT1) gene as one of the top candidate loci. However, no study has shown a genetic association between SIRT1 and completed suicide, which is one of the most serious outcomes of MDD. In this study, 778 suicide completers and 760 controls in a Japanese population were genotyped for two SNPs in strong linkage disequilibrium (rs12415800 and rs4746720 in 3'UTR). We found significant associations between both SNPs and completed suicide among women aged ≥50 years. Additional analysis using postmortem brain tissues (10 suicide brains and 13 non-suicide brains) revealed the following: while SIRT1 gene expression in the prefrontal cortex did not differ between suicide and non-suicide brains, DNAJC12 gene expression, potentially implicated by the SNPs genotyped here, was significantly decreased in suicide brains (p = 0.003). In conclusion, regarding the genetic association of SIRT1 with MDD that was previously identified in women by the Chinese GWAS, we successfully validated our results using a female suicidal cohort in the same Asian population with the same direction of allelic effect.

Epigenetic clock analysis of blood samples from Japanese schizophrenia patients.

The accelerated aging hypothesis of schizophrenia (SCZ) has been proposed. DNA methylation profiles were developed for determining "epigenetic age." Here, we assessed intrinsic and extrinsic epigenetic age acceleration (IEAA and EEAA, respectively) in SCZ. We examined two independent cohorts of Japanese ancestry. The first cohort consisted of 80 patients with SCZ under long-term or repeated hospitalization and 40 controls, with the economical DNA pooling technique. The second cohort consisted of 24 medication-free patients with SCZ and 23 controls. Blood of SCZ subjects exhibited decreased EEAA in the first cohort (p = 0.0162), but not in the second cohort. IEAA did not differ in either cohort. We performed replication analyses using publicly available datasets from European ancestry (three blood and one brain datasets). One blood dataset showed increased EEAA in SCZ (p = 0.0228). Overall, our results provide evidence for decreased EEAA in SCZ associated with hospitalization in the Japanese population.

Mitochondrial DNA copy number of peripheral blood in bipolar disorder: The present study and a meta-analysis.

Numerous evidence indicated mitochondrial abnormalities in the pathophysiology of bipolar disorder (BD); however, it remains unclear whether aberrant mitochondrial DNA (mtDNA) copy number (cn) occur in BD due to the conflicting results in previous studies. Here, peripheral blood mtDNAcn in 69 BD patients and 54 controls were analysed via qPCR. BD patients had significantly lower mtDNAcn compared to controls (regardless of their BD type [BD I or II]). Meta-analysis for all previous BD-mtDNAcn studies combining our results with previously published studies failed to identify any significant association. Meanwhile, Asian-specific meta-analysis remarkably revealed lower mtDNAcn in BD patients.

A cross-sectional study exploring useful indicators for low bone mineral density in male alcoholic patients.

BACKGROUND: Alcohol dependence induces low bone mineral density (BMD), predicting osteoporosis, while low and moderate alcohol consumption may even increase BMD. In recent years, undercarboxylated osteocalcin (ucOC) and tartrate-resistant acid phosphatase-5b (TRACP-5b), bone turnover markers, have gained special interest as useful indicators of low BMD. However, it remains unclear whether other alcohol-related variables (eg, duration of abstinence and continuous drinking) are linked to aberrant BMD. In addition, no previous study has investigated whether ucOC or TRACP-5b is clinically useful to predict low BMD not only in the general population, but also in alcohol-dependent subjects. PATIENTS AND METHODS: We recruited 275 male alcohol-dependent subjects and collected information about their drinking habits, comorbid diseases, smoking history and walking exercise behavior. BMD in each subject was determined by ultrasonography. Serum liver enzymes (AST, ALT, ALP, ChE, γ-GTP and LDH), ucOC and TRACP-5b were measured in all subjects. T-scores were calculated according to BMD for all subjects. RESULTS: The mean T-scores of our subjects were negatively shifted compared to the general population (-0.75±1.36 SD). We divided our subjects into a normal BMD group (n=137) and a low BMD group (n=138) according to their T-scores (T-score ≥-1 SD, normal BMD; T-score <-1 SD, low BMD). Multivariate logistic regression analysis showed that body mass index (BMI) was negatively associated with low BMD (95% CI: 0.75-0.90). By contrast, long abstinence period (95% CI: 1.40-4.21), smoking (95% CI: 1.30-5.56), hypertension (95% CI: 1.04-3.76), lactate dehydrogenase (LDH) (95% CI: 1.00-1.01) and ucOC (95% CI: 1.04-1.22) were positively associated with low BMD. CONCLUSION: In alcohol-dependent males, smoking habits and higher ucOC are associated with low BMD. Our study suggests that smoking cessation may prevent lower BMD, and ucOC may predict lower BMD in alcohol-dependent individuals.

Aberrant telomere length and mitochondrial DNA copy number in suicide completers.

Short telomere length (TL) occurs in individuals under psychological stress, and with various psychiatric diseases. Recent studies have also reported mitochondrial DNA copy number (mtDNAcn) alterations under several neuropsychiatric conditions. However, no study has examined whether aberrant TL or mtDNAcn occur in completed suicide, one of the most serious outcomes of mental illnesses. TL and mtDNAcn in post-mortem samples from 528 suicide completers without severe physical illness (508 peripheral bloods; 20 brains) and 560 samples from control subjects (peripheral bloods from 535 healthy individuals; 25 post-mortem brains) were analysed by quantitative polymerase chain reaction. Suicide completers had significantly shorter TL and higher mtDNAcn of peripheral bloods with sex/age-dependent differences (shorter TL was more remarkably in female/young suicides; higher mtDNAcn more so in male/elderly suicides). The normal age-related decline of TL and mtDNAcn were significantly altered in suicide completers. Furthermore, shorter TL and lower mtDNAcn of post-mortem prefrontal cortex were seen in suicide completers compared to controls. This study shows the first association of aberrant telomeres and mtDNA content with suicide completion. Our results indicate that further research on telomere shortening and mitochondrial dysfunction may help elucidate the molecular underpinnings of suicide-related pathophysiology.

Association study of MIF promoter polymorphisms with suicide completers in the Japanese population.

BACKGROUND: Numerous studies suggest that inflammation plays a key role in suicidal behavior. Macrophage migration inhibitory factor (MIF), a proinflammatory cytokine, has received increasing attention in depression research. However, no study has investigated whether MIF has genetic involvement in completed suicide. In this study, we sought to explore the relationship between two functional polymorphisms on the MIF gene promoter (MIF-794CATT5-8 microsatellite and MIF-173G/C single-nucleotide polymorphism [SNP]) and completed suicide by using one of the largest samples of suicide completers ever reported. METHODS: The subjects comprised 602 suicide completers and 728 healthy controls. We genotyped MIF-794CATT5-8 microsatellite by polymerase chain reaction-based size discrimination assay and MIF-173G/C SNP by TaqMan® SNP genotyping assay. The allele-, genotype-, or haplotype-based association analyses between the suicide completers and the controls were carried out with the χ2 test, the Cochran-Armitage trend test, or Fisher's exact test. RESULTS: Analyses of allele or genotype frequency distributions of the polymorphisms studied here did not reveal any significant differences between the suicide completers and the controls. Haplotype analysis also revealed no association with completed suicide. CONCLUSION: To our knowledge, this is the first study that has examined the genetic association between MIF and completed suicide. Our results suggest that the effects of MIF-794CATT5-8 microsatellite and MIF-173G/C SNP on the MIF gene promoter might not contribute to the genetic risk of completed suicide in the Japanese population.

The cell cycle-related genes as biomarkers for schizophrenia.

BACKGROUND: Recent studies suggest that genomic abnormalities such as single nucleotide polymorphisms (SNPs) and copy number variations (CNVs) may elevate the risk of schizophrenia. Such genomic abnormalities often occur during chromosomal DNA replication in the S phase of cell cycle. In addition, several studies showed that abnormal expressions of several cell cycle-related genes are associated with schizophrenia. Therefore, here we compared mRNA expression levels of cell cycle-related genes in peripheral blood cells between patients with schizophrenia and healthy controls. METHOD: mRNA expression levels of cell cycle-related genes in peripheral blood cells from patients with schizophrenia and healthy controls were measured with quantitative reverse transcription polymerase chain reaction (Q-RT-PCR). The discovery, replication and intervention studies with Q-RT-PCR were performed as follows: discovery (40 cases and 20 controls), replication (82 cases and 74 controls) and intervention (22 cases and 18 controls). RESULT: Nine genes were identified in the discovery and replication stages as schizophrenia-associated genes. Moreover, the combination of mRNA expression levels of CDK4, MCM7 and POLD4 was identified as a potential biomarker for schizophrenia with multivariate logistic regression analysis. The intervention stage revealed that the mRNA expression levels of these three genes were significantly decreased in the acute state of schizophrenia, and CDK4 was significantly recovered in the remission state of schizophrenia. CONCLUSION: The combination of mRNA expression levels of three cell cycle-related genes such as CDK4, MCM7 and POLD4 is expected to be a candidate for useful biomarkers for schizophrenia. Especially, the mRNA expression changes of CDK4 may be potential as both trait and state markers for schizophrenia.

The serum level of NX-DCP-R, but not DCP, is not increased in alcoholic liver disease without hepatocellular carcinoma.

BACKGROUND AND AIM: Alcoholic liver disease (ALD) is the most common cause of hepatocellular carcinoma (HCC) worldwide. Des-gamma-carboxy prothrombin (DCP) is elevated in many patients with HCC, but also in severe alcoholics without HCC. We aimed to clarify whether the DCP/NX-DCP ratio (NX-DCP-R) could have a high specificity in ALD patients without HCC. METHODS: We performed a prospective cohort study on a total of 703 consecutive outpatients of liver diseases including severe alcoholics and healthy volunteers, who underwent blood biochemical examinations at Kobe University Hospital. Serum DCP was measured by electrochemiluminescence immunoassay (ECLIA) using a monoclonal antibody, MU-3. A novel parameter, serum NX-DCP, which represents predominantly DCP caused by reduced vitamin K availability, was also measured by ECLIA using monoclonal antibodies P-16 and P-11. The diagnostic accuracy of DCP and NX-DCP-R in patients with and without excessive alcohol intake was statistically examined. RESULTS: DCP was significantly higher in alcoholics than in non-alcoholics (p= 0.005), whereas the NX-DCP-R did not differ between alcoholics and non-alcoholics (p= 0.375). DCP was significantly increased in the serum of each patient with alcoholic hepatitis and alcoholic cirrhosis (p< 0.05), whereas the NX-DCP-R was not increased (p> 0.05). CONCLUSIONS: NX-DCP-R, but not DCP, was not increased in alcoholics without HCC. As for negative screening for HCC, the specificity of the NX-DCP-R in alcoholics without HCC was better than that of DCP in alcoholics without HCC, and so could be a useful negative screening tool for HCC in millions of alcoholics worldwide.

A decrease in protein level and a missense polymorphism of KIF17 are associated with schizophrenia.

It has been shown that the dysfunction of N-methyl-d-asparate (NMDA) receptors-mediated neurotransmission plays a role in the pathophysiology of schizophrenia. Especially, GluN2B, a subunit of NMDA receptors, associated trafficking complex is altered in the prefrontal cortex of schizophrenia. The kinesin superfamily motor protein 17 (KIF17) is known as a transporter of NR2B.Previous studies showed that a structural variant of KIF17 gene is associated with a schizophrenic phenotype. Therefore, here we investigated KIF17 levels in postmortem prefrontal cortex in schizophrenia and the association of a missense polymorphism (Ile341Val) in KIF17 with schizophrenia. The protein expression of KIF17 in schizophrenic postmortem brains was significantly lower than that in controls. Next, the association of missense polymorphisms (rs631375, rs13375609, rs522496 and rs2296225) of KIF17 gene in 567 schizophrenia and 710 healthy subjects was examined. Both genotypic distribution and allelic frequency of rs2296225 polymorphism were significantly different between the chronic schizophrenia subjects and controls. However, our findings described above were not replicated with the independent subjects (555 schizophrenia and 814 healthy controls). Furthermore, the two alleles of rs2296225 polymorphism did not affect the mRNA expression of KIF17. These results suggest that the dysfunction of KIF17 might be involved in the pathophysiology of schizophrenia.