Integrative analysis of genetics, epigenetics and RNA expression data reveal three susceptibility loci for smoking behavior in Chinese Han population.

Despite numerous studies demonstrate that genetics and epigenetics factors play important roles on smoking behavior, our understanding of their functional relevance and coordinated regulation remains largely unknown. Here we present a multiomics study on smoking behavior for Chinese smoker population with the goal of not only identifying smoking-associated functional variants but also deciphering the pathogenesis and mechanism underlying smoking behavior in this under-studied ethnic population. After whole-genome sequencing analysis of 1329 Chinese Han male samples in discovery phase and OpenArray analysis of 3744 samples in replication phase, we discovered that three novel variants located near FOXP1 (rs7635815), and between DGCR6 and PRODH (rs796774020), and in ARVCF (rs148582811) were significantly associated with smoking behavior. Subsequently cis-mQTL and cis-eQTL analysis indicated that these variants correlated significantly with the differential methylation regions (DMRs) or differential expressed genes (DEGs) located in the regions where these variants present. Finally, our in silico multiomics analysis revealed several hub genes, like DRD2, PTPRD, FOXP1, COMT, CTNNAP2, to be synergistic regulated each other in the etiology of smoking.

Single nucleotide polymorphisms rs148582811 regulates its host gene ARVCF expression to affect nicotine-associated hippocampus-dependent memory.

Although numerous susceptibility loci are nominated for nicotine dependence (ND), no report showed any association of ARVCF with ND. Through genome-wide sequencing analysis, we first identified genetic variants associated nominally with ND and then replicated them in an independent sample. Of the six replicated variants, rs148582811 in ARVCF located in the enhancer-associated marker peak is attractive. The effective-median-based Mendelian randomization analysis indicated that ARVCF is a causal gene for ND. RNA-seq analysis detected decreased ARVCF expression in smokers compared to nonsmokers. Luciferase reporter assays indicated that rs148582811 and its located DNA fragment allele-specifically regulated ARVCF expression. Immunoprecipitation analysis revealed that transcription factor X-ray repair cross-complementing protein 5 (XRCC5) bound to the DNA fragment containing rs148582811 and allele-specifically regulated ARVCF expression at the mRNA and protein levels. With the Arvcf knockout mouse model, we showed that Arvcf deletion not only impairs hippocampus-dependent learning and memory, but also alleviated nicotine-induced memory deficits.

Heme metabolism mediates the effects of smoking on gut microbiome.

INTRODUCTION: The number of smokers worldwide increased greatly during the past decades and reached 1.14 billion in 2019, becoming a leading risk factor for human health. Tobacco smoking has wide effects on human genetics, epigenetics, transcriptome, and gut microbiome. Although many studies have revealed effects of smoking on host transcriptome, research on the relationship among smoking, host gene expression, and the gut microbiome is limited. METHODS: We first explored transcriptome and metagenome profile differences between smokers and non-smokers. To evaluate the relationship between host gene expression and gut microbiome, we then applied bi-directional mediation analysis to infer causal relationships between smoking, gene expression, and gut microbes. RESULTS: Metagenome and transcriptome analyses revealed 71 differential species and 324 differential expressed genes between smokers and non-smokers. With smoking as an exposure variable, we identified 272 significant causal relationships between gene expression and gut microbes, among which there were 247 genes that mediate the effect of smoking on gut microbes. Pathway-based enrichment analysis showed that these genes were significantly enriched in heme metabolic pathway, which mainly mediated the changes of Bacteroides finegoldii and Lachnospiraceae bacterium 9_1_43BFAA. Additionally, by performing metabolome data analysis in the Integrated Human Microbiome project (iHMP) database, we verified the correlation between the intermediate products of the heme metabolism pathway (porphobilinogen, bilirubin, and biliverdin) and gut microbiome. CONCLUSIONS: By investigating the bi-directional interaction between smoking-related host gene expression and gut microbes, this study provided evidence for the mediation of smoking on gut microbes through co-involvement or interaction of heme metabolism. IMPLICATIONS: By comparing the metagenome and transcriptome sequencing profiles between 34 smokers and 33 age- and gender-matched non-smokers, we are the first to reveal causal relationships among tobacco smoking, host gene expression and gut microbes. These findings offer insight into how smoking affects gut microbes through host gene expression and metabolism, which highlights the importance of heme metabolism in modulating the effects of smoking on gut microbiome.

An HBV susceptibility variant of KNG1 modulates the therapeutic effects of interferons α and λ1 in HBV infection by promoting MAVS lysosomal degradation.

BACKGROUND: Hepatitis B virus (HBV) infection is one of the main causes of hepatocellular carcinoma (HCC). The relationship between HBV infection and the host genome as well as their underlying mechanisms remain largely unknown. METHODS: In this study, we performed a whole-genome exon sequencing analysis of 300 sib-pairs of Chinese HBV-infected families with the goal of identifying variants and genes involved in HBV infection. A site-direct mutant plasmid was used to investigate the function of SNP rs76438938 in KNG1. The functional and mechanical studies of KNG1 were conducted with in vitro liver cell lines and a hydrodynamic injection model in vivo. The impact of KNG1 on HBV infection therapy was determined in hepatocytes treated with IFN-α/λ1. FINDINGS: Our whole-exon association study of 300 families with hepatitis B infection found that SNP rs76438938 in KNG1 significantly increased the risk for HBV infection, and the rs76438938-T allele was found to promote HBV replication by increasing the stability of KNG1 mRNA. By competitively binding HSP90A with MAVS, KNG1 can inhibit the expression of types I and III IFNs by promoting MAVS lysosomal degradation. Such suppression of IFN expression and promotion of HBV replication by Kng1 were further demonstrated with an animal model in vivo. Lastly, we showed that the rs76438938-C allele can improve the therapeutic effect of IFN-α and -λ1 in HBV infection. INTERPRETATION: This study identified a SNP, rs76438938, in a newly discovered host gene, KNG1, for its involvement in HBV infection and treatment effect through modulating the cellular antiviral process. FUNDING: This study was supported in part by the Independent Task of State Key Laboratory for Diagnosis and Treatment of Infectious Diseases of the First Affiliated Hospital of Zhejiang University, the China Precision Medicine Initiative (2016YFC0906300), and the Research Center for Air Pollution and Health of Zhejiang University.

Education plays a crucial role in the pathway from poverty to smoking: a Mendelian randomization study.

BACKGROUND AND AIMS: Disproportionately high rates of smoking have been found in low-income communities, but the causal direction and role of education in this relationship remains less well understood. Here, we used bidirectional Mendelian randomization (MR) to measure the causal relationships between smoking, income and education. DESIGN: Two-sample univariable and multivariable MR analyses were conducted to evaluate the total and direct effect of income and education on tobacco smoking. The effects of smoking on education and income were explored with reverse MR analysis. SETTING: European ancestry. PARTICIPANTS: The most recent large-scale genome-wide association study (GWAS) summary data on educational attainment, household income and smoking (n = 143 210-766 345). MEASUREMENTS: Genetic variants for exposures including income, education and smoking. FINDINGS: Both income and education had protective effects against smoking, especially for smoking initiation (education: ß = -0.447, 95% CI = -0.508 to -0.387, P < 0.001; income: ß = -0.290, 95% CI = -0.43 to -0.149, P < 0.001) and cessation (education: ß = -0.364, 95% CI = -0.429 to -0.298, P < 0.001; income: ß = -0.323, 95% CI = -0.448 to -0.197, P < 0.001). Here, higher scores in cessation indicated a lower likelihood of quitting according to the coding scheme. There was little evidence that income influenced smoking once education was controlled for, whereas education could significantly affect smoking behaviours independently of income (P = 3.40 × 10-10 -0.0272). The reverse MR results suggested that smoking may result in a loss of years of schooling (ß = -0.190, 95% CI = -0.297 to -0.083, P < 0.001) and reduced earnings (ß = -0.204, 95% CI = -0.347 to -0.060, P = 0.006). CONCLUSIONS: Education appears to play an important role in the relationship between income and smoking. There is a bidirectional association of smoking with socioeconomic status.

Landscape analysis of lncRNAs shows that DDX11-AS1 promotes cell-cycle progression in liver cancer through the PARP1/p53 axis.

Although long non-coding RNAs (lncRNAs) play important roles in tumorigenesis, the underlying mechanisms are unclear. Transcriptomic analysis of 33 hepatocellular carcinoma (HCC) samples revealed that the most enriched pathway for differentially expressed genes was related to the cell cycle process, where DDX11-AS1 is the most significant lncRNA. Upregulation of DDX11-AS1 expression through demethylation was significantly associated with a poor prognosis. Further mechanistic studies revealed that DDX11-AS1 promoted the growth of HCC by interacting with PARP1 through attenuating its binding to p53, leading to downregulated expression of p53 for inhibiting the transcription of downstream genes such as p21. Knockdown of DDX11-AS1 expression in xenograft mice using anti-DDX11-AS1 oligonucleotide suppressed liver tumor proliferation. These findings indicate that DDX11-AS1 plays a role in the development of liver cancer by affecting the cell cycle.

Genome-wide DNA methylation analysis reveals significant impact of long-term ambient air pollution exposure on biological functions related to mitochondria and immune response.

Exposure to long-term ambient air pollution is believed to have adverse effects on human health. However, the mechanisms underlying these impacts are poorly understood. DNA methylation, a crucial epigenetic modification, is susceptible to environmental factors and likely involved in these processes. We conducted a whole-genome bisulfite sequencing study on 120 participants from a highly polluted region (HPR) and a less polluted region (LPR) in China, where the HPR had much higher concentrations of five air pollutants (PM2.5, PM10, SO2, NO2, and CO) (fold difference 1.6 to 6.6 times; P value 1.80E-07 to 3.19E-23). Genome-wide methylation analysis revealed 371 DMRs in subjects from the two areas and these DMRs were located primarily in gene regulatory elements such as promoters and enhancers. Gene enrichment analysis showed that DMR-related genes were significantly enriched in diseases related to pulmonary disorders and cancers and in biological processes related to mitochondrial assembly and cytokine production. Further, HPR participants showed a higher mtDNA copy number. Of those identified DMRs, 15 were significantly correlated with mtDNA copy number. Finally, cytokine assay indicated that an increased plasma interleukin-5 level was associated with greater air pollution. Taken together, our findings suggest that exposure to long-term ambient air pollution can lead to alterations in DNA methylation whose functions relate to mitochondria and immune responses.

Identification of 34 genes conferring genetic and pharmacological risk for the comorbidity of schizophrenia and smoking behaviors.

The prevalence of smoking is significantly higher in persons with schizophrenia (SCZ) than in the general population. However, the biological mechanisms of the comorbidity of smoking and SCZ are largely unknown. This study aimed to reveal shared biological pathways for the two diseases by analyzing data from two genome-wide association studies with a total sample size of 153,898. With pathway-based analysis, we first discovered 18 significantly enriched pathways shared by SCZ and smoking, which were classified into five groups: postsynaptic density, cadherin binding, dendritic spine, long-term depression, and axon guidance. Then, by using an integrative analysis of genetic, epigenetic, and expression data, we found not only 34 critical genes (e.g., PRKCZ, ARHGEF3, and CDKN1A) but also various risk-associated SNPs in these genes, which convey susceptibility to the comorbidity of the two disorders. Finally, using both in vivo and in vitro data, we demonstrated that the expression profiles of the 34 genes were significantly altered by multiple psychotropic drugs. Together, this multi-omics study not only reveals target genes for new drugs to treat SCZ but also reveals new insights into the shared genetic vulnerabilities of SCZ and smoking behaviors.

Genetic and Epigenetic Analysis Revealing Variants in the NCAM1-TTC12-ANKK1-DRD2 Cluster Associated Significantly With Nicotine Dependence in Chinese Han Smokers.

BACKGROUNDS: Although studies have demonstrated that the NCAM1-TTC12-ANKK1-DRD2 gene cluster plays essential roles in addictions in subjects of European and African origin, study of Chinese Han subjects is limited. Further, the underlying biological mechanisms of detected associations are largely unknown. METHODS: Sixty-four single-nucleotide polymorphisms (SNPs) in this cluster were analyzed for association with Fagerstrom Test for Nicotine Dependence score (FTND) and cigarettes per day (CPD) in male Chinese Han smokers (N = 2616). Next-generation bisulfite sequencing was used to discover smoking-associated differentially methylated regions (DMRs). Both cis-eQTL and cis-mQTL analyses were applied to assess the cis-regulatory effects of these risk SNPs. RESULTS: Association analysis revealed that rs4648317 was significantly associated with FTND and CPD (p = .00018; p = .00072). Moreover, 14 additional SNPs were marginally significantly associated with FTND or CPD (p = .05-.01). Haplotype-based association analysis showed that one haplotype in DRD2, C-T-A-G, formed by rs4245148, rs4581480, rs4648317, and rs11214613, was significantly associated with CPD (p = .0005) and marginally associated with FTND (p = .003). Further, we identified four significant smoking-associated DMRs, three of which are located in the DRD2/ANKK1 region (p = .0012-.00005). Finally, we found five significant CpG-SNP pairs (p = 7.9 × 10-9-6.6 × 10-6) formed by risk SNPs rs4648317, rs11604671, and rs2734849 and three methylation loci. CONCLUSIONS: We found two missense variants (rs11604671; rs2734849) and an intronic variant (rs4648317) with significant effects on ND and further explored their mechanisms of action through expression and methylation analysis. We found the majority of smoking-related DMRs are located in the ANKK1/DRD2 region, indicating a likely causative relation between non-synonymous SNPs and DMRs. IMPLICATIONS: This study shows that there exist significant association of variants and haplotypes in ANKK1/DRD2 region with ND in Chinese male smokers. Further, this study also shows that DNA methylation plays an important role in mediating such associations.

Transcriptomics Curation of SARS-CoV-2 Related Host Genes in Mice With COVID-19 Comorbidity: A Pilot Study.

The pandemic of coronavirus disease 2019 (COVID-19), a respiratory disease caused by a novel severe acute respiratory syndrome coronavirus-2, is causing substantial morbidity and mortality. Along with the respiratory symptoms, underlying diseases in senior patients, such as diabetes, hypertension, and coronary heart disease, are the most common comorbidities, which cause more severe outcomes and even death. During cellular attachment and entry of severe acute respiratory syndrome coronavirus-2, the key protein involved is the angiotensin I converting enzyme 2 (ACE2), which is located on the membrane of host cells. Here, we aim to curate an expression profile of Ace2 and other COVID-19 related genes across the available diabetes murine strains. Based on strictly manual curation and bioinformatics analysis of the publicly deposited expression datasets, Ace2 and other potentially involved genes such as Furin, Tmprss2, Ang, and Ang2 were examined. We found that Ace2 expression is rather ubiquitous in three selected diabetes prone strains (db/db, ob/ob and diet-induced obese). With the most abundant datasets present, the liver shows a medium Ace2 expression level compared with the lungs, pancreatic islets, brain and even T cells. Age is a more critical factor for Ace2 expression in db/db compared with the other two strains. Besides Ace2, the other four host genes showed varied levels of correlation to each other. To accelerate research on the interaction between COVID-19 and underlying diseases, the Murine4Covid transcriptomics database (www.geneureka.org/Murine4Covid) will facilitate the design of research on COVID-19 and comorbidities.

Establishment of a diabetes mellitus type 1 model in the common marmoset.

Common marmosets have attracted considerable attention as a small standard primate model in biomedical research. However, no marmoset diabetes model is available. Here, we established a marmoset diabetes model via the combination of partial pancreatectomy and intravenous streptozotocin (STZ). A partial pancreatectomy was performed in 11 common marmosets and multiple STZ doses were intravenously administered. Diabetes was diagnosed upon sustained hyperglycaemia (nonfasting blood glucose level >200 mg/dl). Blood glucose and biochemistry were periodically assessed, in addition to glucose tolerance testing, continual blood glucose determination using a continuous glucose monitoring system, urine testing and histological evaluation. In 8 of the 11 animals (73%), diabetes mellitus was induced. The diabetic marmosets also showed abnormal intravenous and oral glucose tolerance test results. Blood glucose levels decreased in response to human insulin administration. The hyperglycaemic state was irreversible and persisted for more than 3 months, and the animals' condition was manageable via daily insulin administration. Thus, diabetes can be successfully induced and maintained in the common marmoset via partial pancreatectomy and STZ administration. This protocol effectively generates a valuable animal model for studying disease pathogenesis, risk factors and therapeutic interventions, including islet transplantation.

Identification of LIFR, PIK3R1, and MMP12 as Novel Prognostic Signatures in Gallbladder Cancer Using Network-Based Module Analysis.

Background: Gallbladder cancer (GBC) is a rare and aggressive malignancy of the biliary tract with a dismal survival rate. Effective biomarkers and therapeutic targets are urgently needed. Methods: We analyzed gene expression profiles of GBC to identify differentially expressed genes (DEGs) and then used these DEGs to identify functional module biomarkers based on protein functional interaction (FI) networks. We further evaluated the module-gene protein expression and clinical significance with immunohistochemistry staining (IHC) in a tissue microarray (TMA) from 80 GBC samples. Results: Five functional modules were identified. Module 0 included classical cancer signaling pathways, such as Ras and PI3K-Akt; and modules 1-4 included genes associated with muscle cells, fibrinogen, extracellular matrix, and integrins, respectively. We validated the expression of LIFR, PIK3R1, and MMP12, which were hubs or functional nodes in modules. Compared with paired peritumoural tissues, we found that the expression of LIFR (P = 0.002) and PIK3R1 (P = 0.046) proteins were significantly downregulated, and MMP12 (P = 0.006) was significantly upregulated. Further prognostic analysis showed that patients with low expression of LIFR had shorter overall survival than those with high expression (log-rank test P = 0.028), the same trend as for PIK3R1 (P = 0.053) and MMP12 (P = 0.006). Multivariate analysis indicated that expression of MMP12 protein (hazard ratio [HR] = 0.429; 95% confidence interval [CI] 0.198, 0.930; P = 0.032) was one of the significant independent prognostic factors for overall survival. Conclusions: We found a highly reliable FI network, which revealed LIFR, PIK3R1, and MMP12 as novel prognostic biomarker candidates for GBC. These findings could accelerate biomarker discovery and therapeutic development in this cancer.

Association and cis-mQTL analysis of variants in serotonergic genes associated with nicotine dependence in Chinese Han smokers.

Variants in serotonergic genes are implicated in nicotine dependence (ND) in subjects of European and African origin, but their involvement with smoking in Asians is largely unknown. Moreover, mechanisms underlying the ND risk-associated single-nucleotide polymorphisms (SNPs) in these genes are rarely investigated. The Fagerström Test for Nicotine Dependence (FTND) score was used to assess ND in 2616 male Chinese Han smokers. Both association and interaction analysis were used to examine the association of variants in the serotonergic genes with FTND. Further, expression and methylation quantitative trait loci (cis-mQTL) analysis was employed to determine the association of individual SNPs with the extent of methylation of each CpG locus. Individual SNP-based association analysis revealed that rs1176744 in HTR3B was marginally associated with FTND (p = 0.042). Haplotype-based association analysis found that one major haplotype, T-T-A-G, formed by SNPs rs3758987-rs4938056-rs1176744-rs2276305, located in the 5' region of HTR3B, showed a significant association with FTND (p = 0.00025). Further, a significant genetic interactive effect affecting ND was detected among SNPs rs10160548 in HTR3A, and rs3758987, rs2276305, and rs1672717 in HTR3B (p = 0.0074). Finally, we found four CpG sites (CpG_4543549, CpG_4543464, CpG_4543682, and CpG_4546888) to be significantly associated with three cis-mQTL SNPs (i.e., rs3758987, rs4938056, and rs1176744) located in our detected haplotype within HTR3B. In sum, we showed SNP rs1176744 (Tyr129Ser) to be associated with ND. Together with the SNPs rs3758987 and rs4938056 in HTR3B, they formed a major haplotype, which had significant association with ND. We further showed these SNPs contribute to ND through four methylated sites in HTR3B. All these findings suggest that variants in the serotonergic system play an important role in ND in the Chinese Han population. More importantly, these findings demonstrated that the involvement of this system in ND is through gene-by-gene interaction and methylation.

Detection of Significant Association Between Variants in Cannabinoid Receptor 1 Gene (CNR1) and Personality in African-American Population.

Background: Several studies have revealed significant associations between single nucleotide polymorphisms (SNPs) in the cannabinoid receptor 1 (CNR1) gene and a broad spectrum of psychiatric disorders such as major depressive disorder (MDD), attention deficit hyperactivity disorder (ADHD), and schizophrenia. Personality traits that are highly related to susceptibility to these conditions have been associated with the CNR1 variants in subjects of Caucasian origin. However, there are no reported studies regarding the effects of CNR1 polymorphisms on personality traits in the African-American (AA) population. Methods: We performed an imputation-based association analysis for 26 CNR1 variants with five dimensions of personality in 3,046 AAs. Results: SNPs rs806372 and rs2180619 showed a significant association with extraversion after Bonferroni correction for multiple testing (p < 0.0019). Further, several extraversion-associated SNPs were significantly associated with conscientiousness, agreeableness, and openness. SNP priority score analysis indicated that SNPs rs806368, rs806371, and rs2180619 play a role in the modulation of personality and psychiatric conditions. Conclusion:CNR1 is important in determining personality traits in the AA population.

Association and cis-mQTL analysis of variants in CHRNA3-A5, CHRNA7, CHRNB2, and CHRNB4 in relation to nicotine dependence in a Chinese Han population.

Nicotine dependence (ND) is a worldwide health problem. Numerous genetic studies have demonstrated a significant association of variants in nicotinic acetylcholine receptors (nAChRs) with smoking behaviors. However, most of these studies enrolled only subjects of European or African ancestry. In addition, although an increasing body of evidence implies a causal connection of single-nucleotide polymorphisms (SNPs) and epigenetic regulation of gene expression, few studies of this issue have been reported. In this study, we performed both association and interaction analysis for 67 SNPs in CHRNA3-A5, CHRNA7, CHRNB2, and CHRNB4 with ND in a Chinese Han population (N = 5055). We further analyzed cis-mQTL for the three most significant SNPs and 5580 potential methylation loci within these target gene regions. Our results indicated that the SNPs rs1948 and rs7178270 in CHRNB4 and rs3743075 in CHRNA3 were significantly associated with the Fagerström Test for Nicotine Dependence (FTND) score (p = 6.6 × 10-5; p = 2.0 × 10-4, and p = 7.0 × 10-4, respectively). Haplotype-based association analysis revealed that two major haplotypes, T-G and C-A, formed by rs3743075-rs3743074 in CHRNA3, and other two major haplotypes, A-G-C and G-C-C, formed by rs1948-rs7178270-rs17487223 in CHRNB4, were significantly associated with the FTND score (p ≤ 8.0 × 10-4). Further, we found evidence for the presence of significant interaction among variants within CHRNA3/B4/A5, CHRNA4/B2/A5, and CHRNA7 in affecting ND, with corresponding p values of 5.8 × 10-6, 8.0 × 10-5, and 0.012, respectively. Finally, we identified two CpG sites (CpG_2975 and CpG_3007) in CHRNA3 that are significantly associated with three cis-mQTL SNPs (rs1948, rs7178270, rs3743075) in the CHRNA5/A3/B4 cluster (p ≤ 1.9 × 10-6), which formed four significant CpG-SNP pairs in our sample. Together, we revealed at least three novel SNPs in CHRNA3 and CHRNB4 to be significantly associated with the FTND score. Further, we showed that these significant variants contribute to ND via two methylated sites, and we demonstrated significant interaction affecting ND among variants in CHRNA5/A3/B4, CHRNA7, and CHRNA4/B2/A5. In sum, these findings provide robust evidence that SNPs in nAChR genes convey a risk of ND in the Chinese Han population.

Prevalence of Cigarette Smoking and Nicotine Dependence in Men and Women Residing in Two Provinces in China.

AIMS: Although it is known that there is a high smoking prevalence among Chinese, key issues such as social and environmental factors impacting smoking initiation and persistence, the percentage of smokers considered nicotine dependence (ND), and the availability and use of ND treatments have rarely been investigated. METHODS: To address these issues, from 2012 to 2014, we conducted a large-scale study in the Zhejiang and Shanxi provinces of China using the Fagerström Test for Nicotine Dependence and other validated questionnaires. RESULTS: Of the 17,057 subjects, consisting of 13,476 males and 3,581 females aged 15 years or older, the prevalence of male smoking was 66.1% [95% confidence interval (CI) 65.5%, 66.9%] and that of female smoking was 3.2% (95% CI 3.0%, 3.8%). Among males, 25.8% (95% CI 25.0%, 26.5%) were low-to-moderate ND, and 11.8% (95% CI 11.2%, 12.3%) were high ND (H-ND), persons who have significant difficulty quitting without treatment. The degrees of ND were related to age, extent of education, and annual family income. The social-environmental factors examined conveyed a higher risk for smoking initiation, which is particularly true for the influence of smoking by friends. Furthermore, current smokers had a significantly higher risk of suffering respiratory and digestive symptoms. CONCLUSION: These data not only show a high smoking prevalence in Chinese men but also reveal that a relatively large number of smokers are H-ND. Considering that few Chinese smokers seek ND treatment, a comprehensive smoking prevention and treatment program designed specifically for Chinese is greatly needed.

Significant association of the CHRNB3-CHRNA6 gene cluster with nicotine dependence in the Chinese Han population.

Although numerous studies have revealed significant associations between variants in the nicotinic acetylcholine receptors (nAChR) subunits and nicotine dependence (ND), only few studies were performed in Chinese subjects. Here, we performed association and interaction analysis for 20 single nucleotide polymorphisms (SNPs) in the CHRNB3-CHRNA6 gene cluster with ND in a Chinese Han population (N = 5,055). We found nominally significant associations for all tested SNPs with ND measured by the Fagerström Test for Nicotine Dependence score; of these, 11 SNPs remained significant after Bonferroni correction for multiple tests (p = 9 × 10-4~2 × 10-3). Further conditional analysis indicated that no other SNP was significantly associated with ND independent of the most-highly significant SNP, rs6474414. Also, our haplotype-based association analysis indicated that each haplotype block was significantly associated with ND (p < 0.01). Further, we provide the first evidence of the genetic interaction of these two genes in affecting ND in this sample with an empirical p-value of 0.0015. Finally, our meta-analysis of samples with Asian and European origins for five SNPs in CHRNB3 showed significant associations with ND, with p-values ranging from 6.86 × 10-14 for rs13280604 to 6.50 × 10-8 for rs4950. This represents the first study showing that CHRNB3/A6 are highly associated with ND in a large Chinese Han sample.

Fetal Gene Therapy for Ornithine Transcarbamylase Deficiency by Intrahepatic Plasmid DNA-Micro-Bubble Injection Combined with Hepatic Ultrasound Insonation.

We evaluated the therapeutic efficacy of hepatic transfection of plasmid DNA using micro-bubbles and ultrasound insonation for fetal correction of ornithine transcarbamylase (OTC) deficiency in mice. Twenty-three sparse-fur heterozygous pregnant mice (day 16 of gestation) were divided into three groups: injection of plasmid-DNA micro-bubble mixture into fetal liver with ultrasound insonation (Tr, n = 11); control group 1 (C1), injection of plasmid-DNA micro-bubble mixture into fetal liver with no insonation (n = 5); and control group 2 (C2), injection of saline-micro-bubble mixture into fetal liver with ultrasound insonation (n = 7). Levels of blood ammonia and urinary orotic acid were significantly lower in the Tr group than in the C1 and C2 groups (p < 0.05, p < 0.01, respectively), whereas OTC activity was not different between groups. Therefore, ultrasound insonation with micro-bubbles enhanced plasmid DNA transfection into fetal mouse liver, leading to one of the therapeutic methods in ammonia metabolism. This might provide more time for OTC-deficient infants until liver transplantation.

Meta-Analysis of the COMT Val158Met Polymorphism in Major Depressive Disorder: Effect of Ethnicity.

The COMT (catechol-O-methyltransferase) Val158Met polymorphism (rs4680) is a potential susceptibility variant for major depressive disorder (MDD). Although many genetic studies have examined the association between MDD and this polymorphism, the results were inconclusive. In the present study, we conducted a series of meta-analyses of samples consisting of 2905 MDD cases and 2403 controls with the goal of determining whether this variant indeed has any effect on MDD. We revealed a significant association in the comparison of Val/Val + Val/Met vs. Met/Met (OR =1.180; 95 % CI = 1.019, 1.367; P = 0.027), Val/Met vs. Val/Val (OR =1.18; 95 % CI = 1.038, 1.361; P = 0.013), and Val/Met vs. Met/Met (OR =1.229; 95 % CI = 1.053, 1.435; P = 0.009). Further meta-analyses of samples with European ancestry demonstrated a significant association of this SNP with MDD susceptibility in Val/Val + Val/Met vs. Met/Met (OR =1.231, 95 % CI = 1.046, 1.449; P = 0.013) and Val/Met vs. Met/Met (OR =1.284, 95 % CI = 1.050, 1.484; P = 0.012). For the samples with East Asian ancestry, we found a significant association in both allelic (Val vs. Met: OR =0.835; 95 % CI = 0.714, 0.975; P = 0.023) and genotypic (Met/Met + Val/Met vs. Val/Val: OR =1.431, 95 % CI = 1.143, 1.791; P = 0.002; Val/Met vs. Val/Val: OR =1.482, 95 % CI = 1.171, 1.871; P = 0.001) analyses. No evidence of heterogeneity among studies or publication bias was observed. Together, our results indicate that the COMT Val158Met polymorphism is a vulnerability factor for MDD with distinct effects in different ethnic populations.

Contribution of Variants in CHRNA5/A3/B4 Gene Cluster on Chromosome 15 to Tobacco Smoking: From Genetic Association to Mechanism.

Cigarette smoking is the major cause of preventable death and morbidity throughout the world. Many compounds are present in tobacco, but nicotine is the primary addictive one. Nicotine exerts its physiological and pharmacological roles in the brain through neuronal nicotinic acetylcholine receptors (nAChRs), which are ligand-gated ion channels consisting of five membrane-spanning subunits that can modulate the release of neurotransmitters, such as dopamine, glutamate, and GABA and mediate fast signal transmission at synapses. Considering that there are 12 nAChR subunits, it is highly likely that subunits other than α4 and ß2, which have been intensively investigated, also are involved in nicotine addiction. Consistent with this hypothesis, a number of genome-wide association studies (GWAS) and subsequent candidate gene-based associated studies investigating the genetic variants associated with nicotine dependence (ND) and smoking-related phenotypes have shed light on the CHRNA5/A3/B4 gene cluster on chromosome 15, which encodes the α5, α3, and ß4 nAChR subunits, respectively. These studies demonstrate two groups of risk variants in this region. The first one is marked by single nucleotide polymorphism (SNP) rs16969968 in exon 5 of CHRNA5, which changes an aspartic acid residue into asparagine at position 398 (D398N) of the α5 subunit protein sequence, and it is tightly linked SNP rs1051730 in CHRNA3. The second one is SNP rs578776 in the 3'-untranslated region (UTR) of CHRNA3, which has a low correlation with rs16969968. Although the detailed molecular mechanisms underlying these associations remain to be further elucidated, recent findings have shown that α5* (where "*" indicates the presence of additional subunits) nAChRs located in the medial habenulo-interpeduncular nucleus (mHb-IPN) are involved in the control of nicotine self-administration in rodents. Disruption of α5* nAChR signaling diminishes the aversive effects of nicotine on the mHb-IPN pathway and thereby permits more nicotine consumption. To gain a better understanding of the function of the highly significant genetic variants identified in this region in controlling smoking-related behaviors, in this communication, we provide an up-to-date review of the progress of studies focusing on the CHRNA5/A3/B4 gene cluster and its role in ND.