ABSTRACT
The most common cause of autosomal recessive familial Parkinson's disease (PD) are mutations in the PRKN/PARK2 gene encoding an E3 ubiquitin protein-ligase PARKIN. We report the generation of an iPSC cell line from the fibroblasts of a male PD patient carrying a common missense variant in exon 7 (p.Arg275Trp), and a 133 kb deletion encompassing exon 8, using transiently-present Sendai virus. The established line displays typical human primed iPSC morphology and expression of pluripotency-associated markers, normal karyotype without SNP array-detectable copy number variations and can give rise to derivatives of all three embryonic germ layers. We envisage the usefulness of this iPSC line, carrying a common and well-studied missense mutation in the RING1 domain of the PARKIN protein, for the elucidation of PARKIN-dependent mechanisms of PD using in vitro and in vivo models.
Subject(s)
Induced Pluripotent Stem Cells , Parkinson Disease , Humans , Male , Induced Pluripotent Stem Cells/metabolism , Parkinson Disease/genetics , Parkinson Disease/metabolism , DNA Copy Number Variations , Mutation/genetics , Ubiquitin-Protein Ligases/genetics , Ubiquitin-Protein Ligases/metabolismABSTRACT
Forensic science has yet to take full advantage of single cell analysis. Its greatest benefit is the ability to alleviate the challenges associated with DNA mixture analysis, which remains a significant hurdle in forensic science. Many of the factors that cause complexity in mixture interpretation are absent in single cell analyses-multiple contributors, varied levels of contribution, and allele masking. This study revisits single cell analyses in the context of forensic identification, introducing previously unseen depth to the characterization of data generated from single cells using a novel pipeline that includes recovery of single cells using the DEPArray NxT and amplification using the PowerPlex Fusion 6c kit with varied PCR cycles (29, 30, and 31). The resulting allelic signal was assessed using analytical thresholds of 10, 100, and 150RFU. The mean peak heights across the sample sets generally increased as cycle number increased, 75.0 ± 85.3, 147.1 ± 172.6, and 226.1 ± 298.2 RFU, for 29, 30, and 31 cycles, respectively. The average proportion of allele/locus dropout was most significantly impacted by changes in the detection threshold, whereas increases in PCR cycle number had less impact. Overall data quality improved notably when increasing PCR from 29 to 30 cycles, less improvement and more volatility was introduced at 31 cycles. The average random match probabilities for the 29, 30, and 31 cycle sets at 150RFU are 1 in 2.4 × 1018 ± 1.46 × 1019, 1 in 1.49 × 1025 ± 5.8 × 1025, and 1 in 1.83 × 1024 ± 8.09 × 1024, respectively. This demonstrates the current power of single cell analysis in removing the need for complex mixture analysis.
Subject(s)
Forensic Sciences , Single-Cell Analysis/methods , Alleles , DNA Fingerprinting/methods , Humans , Polymerase Chain Reaction/methods , ProbabilityABSTRACT
Bipolar disorder (BD) is a highly heritable and heterogeneous mental illness whose manifestations often include impulsive and risk-taking behavior. This particular phenotype suggests that abnormal striatal function could be involved in BD etiology, yet most transcriptomic studies of this disorder have concentrated on cortical brain regions. We believe we report the first transcriptome sequencing of the postmortem human dorsal striatum comparing bipolar (18) and control (17) subjects. Fourteen genes were detected as differentially expressed at a 5% false discovery rate, including a few immune response genes such as NLRC5, S100A12, LILRA4 and FCGBP, as well as an assortment of non-protein coding genes. Functional pathway analysis found an enrichment of upregulated genes across many immune/inflammation pathways and an enrichment of downregulated genes among oxidative phosphorylation pathways. Co-expression network analysis revealed 20 modules of highly interconnected genes; two of the modules were significantly enriched for BD susceptibility single-nucleotide polymorphisms deriving from a large genome-wide association study data set. Remarkably, the module with the highest genetic association signal for BD, which contained many genes from signaling pathways, was also enriched in markers characteristic of gene expression in dorsal striatum medium spiny neurons-unlike most other modules, which showed no such regional and neuronal specificity. These findings draw a link between BD etiology at the gene level and a specific brain region, and highlight striatal signaling pathways as potential targets for the development of novel treatments to manage BD.
Subject(s)
Bipolar Disorder/genetics , Corpus Striatum/metabolism , Transcriptome , Adult , Aged , Aged, 80 and over , Autopsy , Bipolar Disorder/metabolism , Brain/metabolism , Central Nervous System/metabolism , Energy Metabolism/genetics , Female , Gene Expression Profiling , Gene Regulatory Networks/genetics , Genetic Predisposition to Disease/genetics , Genome-Wide Association Study , Humans , Immunity, Active/genetics , Male , Middle Aged , Polymorphism, Single Nucleotide/genetics , Transcriptome/geneticsABSTRACT
One-third of type-2 diabetic patients respond poorly to metformin. Despite extensive research, the impact of genetic and nongenetic factors on long-term outcome is unknown. In this study we combine nonlinear mixed effect modeling with computational genetic methodologies to identify predictors of long-term response. In all, 1,056 patients contributed their genetic, demographic, and long-term HbA1c data. The top nine variants (of 12,000 variants in 267 candidate genes) accounted for approximately one-third of the variability in the disease progression parameter. Average serum creatinine level, age, and weight were determinants of symptomatic response; however, explaining negligible variability. Two single nucleotide polymorphisms (SNPs) in CSMD1 gene (rs2617102, rs2954625) and one SNP in a pharmacologically relevant SLC22A2 gene (rs316009) influenced disease progression, with minor alleles leading to less and more favorable outcomes, respectively. Overall, our study highlights the influence of genetic factors on long-term HbA1c response and provides a computational model, which when validated, may be used to individualize treatment.
Subject(s)
Diabetes Mellitus, Type 2/metabolism , Disease Progression , Glycated Hemoglobin/metabolism , Membrane Proteins/genetics , Metformin/therapeutic use , Organic Cation Transport Proteins/genetics , Pharmacogenomic Variants/genetics , Adult , Aged , Aged, 80 and over , Diabetes Mellitus, Type 2/drug therapy , Female , Humans , Hypoglycemic Agents/therapeutic use , Longitudinal Studies , Male , Middle Aged , Nonlinear Dynamics , Organic Cation Transporter 2 , Polymorphism, Single Nucleotide/genetics , Tumor Suppressor Proteins , Young AdultABSTRACT
Bipolar disorder is a highly heritable neuropsychiatric disorder affecting nearly 2.5% of the population. Prior genetic studies identified a panel of common and rare single-nucleotide polymorphisms associated with the disease that map to the first intron of the PDE10A gene. RNA sequencing of striatal brain tissue from bipolar and healthy control subjects identified a novel transcript of PDE10A, named PDE10A19, that codes for a PDE10A isoform with a unique N terminus. Genomic sequences that can encode the novel N terminus were conserved in other primates but not rodents. The RNA transcript was expressed at equal or greater levels in the human striatum compared with the two annotated transcripts, PDE10A1 and PDE10A2. The PDE10A19 transcript was detected in polysomal fractions; western blotting experiments confirmed that the RNA transcript is translated into protein. Immunocytochemistry studies using transfected mouse striatal and cortical neurons demonstrated that the PDE10A19 protein distributes to the cytosol, like PDE10A1, and unlike PDE10A2, which is associated with plasma membranes. Immunoprecipitation and immunocytochemical experiments revealed that the PDE10A19 isoform interacts physically with PDE10A2 and, when expressed at elevated levels, interferes with the plasma membrane localization of PDE10A2. These studies illustrate the complexity of PDE10A gene expression in the human brain and highlight the need to unravel the gene's complex and complete coding capabilities along with its transcriptional and translational regulation to guide the development of therapeutic agents that target the protein for the treatment of neuropsychiatric illness.
Subject(s)
Bipolar Disorder/genetics , Bipolar Disorder/metabolism , Corpus Striatum/metabolism , Gene Expression/genetics , Phosphoric Diester Hydrolases/genetics , Phosphoric Diester Hydrolases/metabolism , Bipolar Disorder/pathology , Blotting, Western , Humans , Pathology, Molecular , Protein Isoforms/geneticsABSTRACT
Pathogenic mutations in the OPA1 gene can be associated with Autosomal Dominant Optic Atrophy (ADOA). In approximately 20 % of patients with OPA1 mutations, a more complex neurodegenerative disorder with extraocular manifestations, known as ADOA Plus, can arise. 12 members of a multigenerational family were assessed clinically and screened for a genetic mutation in OPA1. Eight family members displayed manifestations consistent with ADOA Plus and four did not. Affected members of the oldest available generation displayed the most severe phenotype, which included severe optic atrophy, deafness, ptosis, ophthalmoplegia, proximal myopathy, neuropathy and ataxia. The next generation was less severely affected but several members displayed manifestations only after the fifth decade. Genetic analysis revealed a heterozygous variant in the OPA1 gene (c.1053T>A, p.Asp351Glu) that segregated with disease. The affected family members described here exhibited visual loss later than is typical for OPA1-related disease, as well as later onset of other neurological abnormalities in the fifth or sixth decades of life that progressed to severe neurological disability by the seventh decade. These findings expand the clinical spectrum of OPA1-related disease associated with a novel OPA1 mutation.
Subject(s)
GTP Phosphohydrolases/genetics , Optic Atrophy, Autosomal Dominant/genetics , Optic Atrophy, Autosomal Dominant/physiopathology , Adult , Aged , Australia , Female , Humans , Male , Mutation , Pedigree , PhenotypeABSTRACT
Primary auditory neurons (ANs) in the mammalian cochlea play a critical role in hearing as they transmit auditory information in the form of electrical signals from mechanosensory cochlear hair cells in the inner ear to the brainstem. Their progressive degeneration is associated with disease conditions, excessive noise exposure and aging. Replacement of ANs, which lack the ability to regenerate spontaneously, would have a significant impact on research and advancement in cochlear implants in addition to the amelioration of hearing impairment. The aim of this study was to induce a neuronal phenotype in endogenous non-neural cells in the cochlea, which is the essential organ of hearing. Overexpression of a neurogenic basic helix-loop-helix transcription factor, Ascl1, in the cochlear non-sensory epithelial cells induced neurons at high efficiency at embryonic, postnatal and juvenile stages. Moreover, induced neurons showed typical properties of neuron morphology, gene expression and electrophysiology. Our data indicate that Ascl1 alone or Ascl1 and NeuroD1 is sufficient to reprogram cochlear non-sensory epithelial cells into functional neurons. Generation of neurons from non-neural cells in the cochlea is an important step for the regeneration of ANs in the mature mammalian cochlea.
Subject(s)
Auditory Pathways/cytology , Cell Differentiation/physiology , Cochlea/cytology , Neurons/cytology , Regeneration/physiology , Animals , Auditory Pathways/physiology , Cell Count , Cochlea/physiology , Hearing/physiology , Mice , Neurons/physiologyABSTRACT
One-third of type 2 diabetes patients do not respond to metformin. Genetic variants in metformin transporters have been extensively studied as a likely contributor to this high failure rate. Here, we investigate, for the first time, the effect of genetic variants in transcription factors on metformin pharmacokinetics (PK) and response. Overall, 546 patients and healthy volunteers contributed their genome-wide, pharmacokinetic (235 subjects), and HbA1c data (440 patients) for this analysis. Five variants in specificity protein 1 (SP1), a transcription factor that modulates the expression of metformin transporters, were associated with changes in treatment HbA1c (P < 0.01) and metformin secretory clearance (P < 0.05). Population pharmacokinetic modeling further confirmed a 24% reduction in apparent clearance in homozygous carriers of one such variant, rs784888. Genetic variants in other transcription factors, peroxisome proliferator-activated receptor-α and hepatocyte nuclear factor 4-α, were significantly associated with HbA1c change only. Overall, our study highlights the importance of genetic variants in transcription factors as modulators of metformin PK and response.
Subject(s)
Diabetes Mellitus, Type 2/drug therapy , Diabetes Mellitus, Type 2/genetics , Hypoglycemic Agents/pharmacokinetics , Metformin/pharmacokinetics , Pharmacogenetics , Polymorphism, Single Nucleotide , Transcription Factors/genetics , Adult , Aged , Aged, 80 and over , Biomarkers/metabolism , Diabetes Mellitus, Type 2/blood , Female , Genome-Wide Association Study , Glycated Hemoglobin/metabolism , Hepatocyte Nuclear Factor 4/genetics , Homozygote , Humans , Hypoglycemic Agents/administration & dosage , Male , Metformin/administration & dosage , Middle Aged , Models, Biological , Multivariate Analysis , PPAR alpha/genetics , Phenotype , Retrospective Studies , Sp1 Transcription Factor/genetics , Treatment Outcome , United States , Young AdultABSTRACT
The spiral ganglion conveys afferent auditory information predominantly through a single class of type I neurons that receive signals from inner hair cell sensory receptors. These auditory primary afferents, like in other systems (Puopolo and Belluzzi, 1998; Gascon and Moqrich, 2010; Leao et al., 2012) possess a marked diversity in their electrophysiological features (Taberner and Liberman, 2005). Consistent with these observations, when the auditory primary afferents were assessed in neuronal explants separated from their peripheral and central targets it was found that individual neurons were markedly heterogeneous in their endogenous electrophysiological features. One aspect of this heterogeneity, obvious throughout the ganglion, was their wide range of excitability as assessed by voltage threshold measurements (Liu and Davis, 2007). Thus, while neurons in the base differed significantly from apical and middle neurons in their voltage thresholds, each region showed distinctly wide ranges of values. To determine whether the resting membrane potentials (RMPs) of these neurons correlate with the threshold distribution and to identify the ion channel regulatory elements underlying heterogeneous neuronal excitability in the ganglion, patch-clamp recordings were made from postnatal day (P5-8) murine spiral ganglion neurons in vitro. We found that RMP mirrored the tonotopic threshold distribution, and contributed an additional level of heterogeneity in each cochlear location. Pharmacological experiments further indicated that threshold and RMP was coupled through the Kv1 current, which had a dual impact on both electrophysiological parameters. Whereas, hyperpolarization-activated cationic channels decoupled these two processes by primarily affecting RMP without altering threshold level. Thus, beyond mechanical and synaptic specializations, ion channel regulation of intrinsic membrane properties imbues spiral ganglion neurons with different excitability levels, a feature that contributes to primary auditory afferent diversity.
Subject(s)
Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels/metabolism , Neurons/metabolism , Shaker Superfamily of Potassium Channels/metabolism , Spiral Ganglion/cytology , 4-Aminopyridine/pharmacology , Animals , Animals, Newborn , Biophysical Phenomena/drug effects , Biophysics , Cadmium Chloride/pharmacology , Elapid Venoms/pharmacology , Electric Stimulation , Membrane Potentials/drug effects , Membrane Potentials/genetics , Mice , Mice, Inbred C57BL , Neurotoxins/pharmacology , Patch-Clamp Techniques , Potassium Channel Blockers/pharmacology , Tetrodotoxin/pharmacologyABSTRACT
Efforts to define the genetic architecture underlying variable statin response have met with limited success, possibly because previous studies were limited to effect based on a single dose. We leveraged electronic medical records (EMRs) to extract potency (ED50) and efficacy (Emax) of statin dose-response curves and tested them for association with 144 preselected variants. Two large biobanks were used to construct dose-response curves for 2,026 and 2,252 subjects on simvastatin and atorvastatin, respectively. Atorvastatin was more efficacious, was more potent, and demonstrated less interindividual variability than simvastatin. A pharmacodynamic variant emerging from randomized trials (PRDM16) was associated with Emax for both. For atorvastatin, Emax was 51.7 mg/dl in subjects homozygous for the minor allele vs. 75.0 mg/dl for those homozygous for the major allele. We also identified several loci associated with ED50. The extraction of rigorously defined traits from EMRs for pharmacogenetic studies represents a promising approach to further understand the genetic factors contributing to drug response.
Subject(s)
Electronic Health Records , Hydroxymethylglutaryl-CoA Reductase Inhibitors/administration & dosage , Hydroxymethylglutaryl-CoA Reductase Inhibitors/therapeutic use , Hyperlipidemias/drug therapy , Hyperlipidemias/genetics , Algorithms , Alleles , Atorvastatin , Cholesterol, LDL/blood , Cohort Studies , Databases, Factual , Dose-Response Relationship, Drug , Genotype , Heptanoic Acids/administration & dosage , Heptanoic Acids/therapeutic use , Humans , Lipid Metabolism/drug effects , Lipid Metabolism/genetics , Lipids/blood , Phenotype , Polymorphism, Single Nucleotide , Pyrroles/administration & dosage , Pyrroles/therapeutic use , Randomized Controlled Trials as Topic , Simvastatin/administration & dosage , Simvastatin/therapeutic useABSTRACT
BACKGROUND AND PURPOSE: Previous work in our laboratory showed opioid agents inhibit cytokine expression in astrocytes. Recently, Watkins and colleagues hypothesized that opioid agonists activate toll-like receptor 4 (TLR4) signalling, which leads to neuroinflammation. To test this hypothesis, we characterized LPS and opioid effects on TLR4 signalling in reporter cells. EXPERIMENTAL APPROACH: NF-κB reporter cells expressing high levels of TLR4 were used to compare LPS and opioid effects on NF-κB activation, a pathway activated by TLR4 stimulation. KEY RESULTS: LPS increased TLR4 signalling in a concentration-dependent manner and was antagonized by LPS antagonist (LPS-RS, from Rhodobacter sphaeroides). A concentration ratio analysis showed that LPS-RS was a competitive antagonist. The opioid agonists, morphine and fentanyl, produced minor activation of TLR4 signalling when given alone. When tested following LPS stimulation, opioid agonists inhibited NF-κB activation but this inhibition was not blocked by the general opioid antagonist, naloxone, nor by the selective µ opioid receptor antagonist, ß-FNA. Indeed, both naloxone and ß-FNA also inhibited NF-κB activation in reporter cells. Further examination of fentanyl and ß-FNA effects revealed that both opioid agents inhibited LPS signalling in a non-competitive fashion. CONCLUSIONS AND IMPLICATIONS: These results show that LPS-RS is a competitive antagonist at the TLR4 complex, and that both opioid agonists and antagonists inhibit LPS signalling in a non-competitive fashion through a non-GPCR, opioid site(s) in the TLR4 signalling pathway. If confirmed, existing opioid agents or other drug molecules more selective at this novel site may provide a new therapeutic approach to the treatment of neuroinflammation.
Subject(s)
Analgesics, Opioid/pharmacology , Lipopolysaccharides/pharmacology , Nerve Tissue Proteins/metabolism , Signal Transduction/drug effects , Toll-Like Receptor 4/metabolism , Analgesics, Opioid/agonists , Analgesics, Opioid/antagonists & inhibitors , Binding, Competitive , Drug Antagonism , Escherichia coli K12/metabolism , Genes, Reporter/drug effects , HEK293 Cells , Humans , Kinetics , Ligands , Lipopolysaccharide Receptors/genetics , Lipopolysaccharide Receptors/metabolism , Lipopolysaccharides/antagonists & inhibitors , Lymphocyte Antigen 96/genetics , Lymphocyte Antigen 96/metabolism , NF-kappa B p50 Subunit/agonists , NF-kappa B p50 Subunit/genetics , NF-kappa B p50 Subunit/metabolism , Narcotic Antagonists/chemistry , Narcotic Antagonists/pharmacology , Nerve Tissue Proteins/agonists , Nerve Tissue Proteins/antagonists & inhibitors , Nerve Tissue Proteins/genetics , Receptors, Opioid, mu/agonists , Receptors, Opioid, mu/antagonists & inhibitors , Receptors, Opioid, mu/metabolism , Recombinant Proteins/agonists , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/metabolism , Rhodobacter sphaeroides/metabolism , Toll-Like Receptor 4/agonists , Toll-Like Receptor 4/antagonists & inhibitors , Toll-Like Receptor 4/geneticsABSTRACT
Interindividual variation in response to metformin, first-line therapy for type 2 diabetes, is substantial. Given that transporters are determinants of metformin pharmacokinetics, we examined the effects of promoter variants in both multidrug and toxin extrusion protein 1 (MATE1) (g.-66T â C, rs2252281) and MATE2 (g.-130G â A, rs12943590) on variation in metformin disposition and response. The pharmacokinetics and glucose-lowering effects of metformin were assessed in healthy volunteers (n = 57) receiving metformin. The renal and secretory clearances of metformin were higher (22% and 26%, respectively) in carriers of variant MATE2 who were also MATE1 reference (P < 0.05). Both MATE genotypes were associated with altered post-metformin glucose tolerance, with variant carriers of MATE1 and MATE2 having an enhanced (P < 0.01) and reduced (P < 0.05) response, respectively. Consistent with these results, patients with diabetes (n = 145) carrying the MATE1 variant showed enhanced metformin response. These findings suggest that promoter variants of MATE1 and MATE2 are important determinants of metformin disposition and response in healthy volunteers and diabetic patients.
Subject(s)
Diabetes Mellitus, Type 2/metabolism , Hypoglycemic Agents/pharmacokinetics , Metformin/pharmacokinetics , Organic Cation Transport Proteins/genetics , Diabetes Mellitus, Type 2/genetics , Female , Genotype , Humans , Hypoglycemic Agents/pharmacology , Kidney/drug effects , Kidney/metabolism , Male , Metformin/pharmacology , Organic Cation Transport Proteins/metabolismABSTRACT
BACKGROUND/AIM: The nuclear POLG gene encodes the catalytic subunit of DNA polymerase gamma (polγ), the only polymerase involved in the replication and proofreading of mitochondrial DNA. As a consequence, POLG mutations can cause disease through impaired replication of mitochondrial DNA. To date, over 150 different mutations have been identified, with a growing number of associated phenotypes described. The aim of this study was to determine the prevalence of POLG mutations in an adult population of Australian patients with mitochondrial disease, displaying symptoms commonly associated with POLG-related diseases. METHODS: The clinical presentations of 322 patients from a specialist adult mitochondrial disease clinic were reviewed. Nineteen exhibited a cluster of three or more predefined clinical manifestations suggestive of POLG-related disease: progressive external ophthalmoplegia, seizures and/or an abnormal electroencephalogram, neuropathy, ataxia, liver function abnormalities, migraine or dysphagia/dysarthria. Patients were screened for mutations by direct nucleotide sequencing of the coding and exon-flanking intronic regions of POLG. RESULTS: Five of the 19 patients (26%) displaying a phenotype suggestive of POLG-related disease were found to have informative POLG coding mutations (p.T851A, p.N468D, p.Y831C, p.G517V and novel p.P163S variant). Literature and analysis of these mutations revealed that two of these patients had pathogenic mutations known to cause POLG-related disease (patient #1: p.T851A and p.P163S; patient #2: p.T851A and p.N468D). CONCLUSIONS: We conclude that the prevalence of pathogenic POLG mutations in our selected adult Australian cohort with suggestive clinical manifestations was 10%. A further 16% of patients had POLG variants but are unlikely to be responsible for causing their disease.
Subject(s)
DNA-Directed DNA Polymerase/genetics , Mitochondrial Diseases/diagnosis , Mitochondrial Diseases/genetics , Mutation/genetics , Adult , Amino Acid Sequence , Australia/epidemiology , Cohort Studies , DNA Polymerase gamma , Female , Humans , Middle Aged , Mitochondrial Diseases/epidemiology , Molecular Sequence Data , Prevalence , Young AdultABSTRACT
The genetic basis for bipolar disorder (BPD) is complex with the involvement of multiple genes. As it is well established that cyclic adenosine monophosphate (cAMP) signaling regulates behavior, we tested variants in 29 genes that encode components of this signaling pathway for associations with BPD type I (BPD I) and BPD type II (BPD II). A total of 1172 individuals with BPD I, 516 individuals with BPD II and 1728 controls were analyzed. Single SNP (single-nucleotide polymorphism), haplotype and SNP × SNP interactions were examined for association with BPD. Several statistically significant single-SNP associations were observed between BPD I and variants in the PDE10A gene and between BPD II and variants in the DISC1 and GNAS genes. Haplotype analysis supported the conclusion that variation in these genes is associated with BPD. We followed-up PDE10A's association with BPD I by sequencing a 23-kb region in 30 subjects homozygous for seven minor allele risk SNPs and discovered eight additional rare variants (minor allele frequency < 1%). These single-nucleotide variants were genotyped in 999 BPD cases and 801 controls. We obtained a significant association for these variants in the combined sample using multiple methods for rare variant analysis. After using newly developed methods to account for potential bias from sequencing BPD cases only, the results remained significant. In addition, SNP × SNP interaction studies suggested that variants in several cAMP signaling pathway genes interact to increase the risk of BPD. This report is among the first to use multiple rare variant analysis methods following common tagSNPs associations with BPD.
Subject(s)
Bipolar Disorder/genetics , Cyclic AMP/genetics , Genetic Predisposition to Disease/genetics , Adult , Chromosome Mapping/methods , Female , Gene Frequency/genetics , Genotype , Haplotypes/genetics , Humans , Linkage Disequilibrium/genetics , Male , Polymorphism, Single Nucleotide , Signal Transduction/geneticsABSTRACT
Environmental exposure to mercury can cause a number of adverse effects in humans including the disruption of endocrine function that may result in sex-specific effects. The present study was designed to characterize sex-specific effects of chronic inorganic mercury exposure on toll-like receptor (TLR) 2 and TLR4 and inflammatory signaling in the liver of prairie voles (Microtus ochrogaster). Following 10 weeks of exposure to mercury via drinking water, effects on protein expression levels of TLR2 and TLR4 and the downstream p38 mitogen-activated protein kinase (p38 MAPK) and nuclear factor-kappa (NF-κB) signaling pathways were assessed. Using immunoblot analysis, we found that mercury exposure significantly enhanced the expression of TLR4 and activated p38 MAPK and NF-κB pathways in vole livers. This is the first report indicating that TLR4 may serve as a sensor for chronic mercury exposure in the liver. Further, compared to females, mercury-treated male voles exhibited significant increases in activated p38 MAPK and a greater extent of liver damage. Together, these findings establish sex-specific liver immunomodulation and cellular signaling following chronic inorganic mercury exposure. Furthermore, this study also supports the use of voles as biomarkers of environmental mercury contamination and offers a promising in vivo tool to test various therapeutic strategies for mercury detoxification.
Subject(s)
Environmental Pollutants/toxicity , Liver/drug effects , Mercuric Chloride/toxicity , NF-kappa B/metabolism , Signal Transduction/drug effects , Toll-Like Receptor 4/biosynthesis , Animals , Arvicolinae , Blotting, Western , Liver/enzymology , Liver/immunology , Liver/metabolism , Liver/pathology , Male , Sex Factors , Toll-Like Receptor 2/biosynthesis , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
Multidrug and toxin extrusion 2 (MATE2-K (SLC47A2)), a polyspecific organic cation exporter, facilitates the renal elimination of the antidiabetes drug metformin. In this study, we characterized genetic variants of MATE2-K, determined their association with metformin response, and elucidated their impact by means of a comparative protein structure model. Four nonsynonymous variants and four variants in the MATE2-K basal promoter region were identified from ethnically diverse populations. Two nonsynonymous variants-c.485C>T and c.1177G>A-were shown to be associated with significantly lower metformin uptake and reduction in protein expression levels. MATE2-K basal promoter haplotypes containing the most common variant, g.-130G>A (>26% allele frequency), were associated with a significant increase in luciferase activities and reduced binding to the transcriptional repressor myeloid zinc finger 1 (MZF-1). Patients with diabetes who were homozygous for g.-130A had a significantly poorer response to metformin treatment, assessed as relative change in glycated hemoglobin (HbA1c) (-0.027 (-0.076, 0.033)), as compared with carriers of the reference allele, g.-130G (-0.15 (-0.17, -0.13)) (P=0.002). Our study showed that MATE2-K plays a role in the antidiabetes response to metformin.
Subject(s)
Diabetes Mellitus, Type 2/drug therapy , Hypoglycemic Agents/pharmacokinetics , Metformin/pharmacokinetics , Organic Cation Transport Proteins/genetics , Adult , Aged , Alleles , Animals , Female , Genetic Variation , Glycated Hemoglobin/metabolism , HCT116 Cells , HEK293 Cells , Haplotypes , Humans , Hypoglycemic Agents/pharmacology , LLC-PK1 Cells , Luciferases/metabolism , Male , Metformin/pharmacology , Middle Aged , Polymorphism, Genetic , Promoter Regions, Genetic , Racial Groups/genetics , Retrospective Studies , Swine , Treatment OutcomeABSTRACT
Health-care information technology and genotyping technology are both advancing rapidly, creating new opportunities for medical and scientific discovery. The convergence of these two technologies is now facilitating genetic association studies of unprecedented size within the context of routine clinical care. As a result, the medical community will soon be presented with a number of novel opportunities to bring functional genomics to the bedside in the area of pharmacotherapy. By linking biological material to comprehensive medical records, large multi-institutional biobanks are now poised to advance the field of pharmacogenomics through three distinct mechanisms: (i) retrospective assessment of previously known findings in a clinical practice-based setting, (ii) discovery of new associations in huge observational cohorts, and (iii) prospective application in a setting capable of providing real-time decision support. This review explores each of these translational mechanisms within a historical framework.
Subject(s)
Electronic Health Records/trends , Pharmaceutical Preparations/administration & dosage , Pharmacogenetics/trends , Decision Support Techniques , Genetic Association Studies/methods , Genomics , Genotype , Humans , Research DesignABSTRACT
Common genetic variants of the liver-specific human organic cation transporter 1 (OCT1; SLC22A1) have reduced transport capacity for substrates such as the antidiabetic drug metformin. The effect of the reduced OCT1 function on drug interactions associated with OCT1 has not been investigated and was, therefore, the focus of the study presented here. HEK293 cells expressing human OCT1-reference or the variants R61C, V408M, M420del and G465R were first used to study the kinetics and inhibition pattern of the OCT1 substrate 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP(+)). In the second part OCT1-mediated (14)C-metformin uptake was studied in the presence of drugs administered concomitantly with metformin. Transport studies using ASP(+) showed that the function of the variants decreased in the following order: OCT1-reference=V408M=M420del >R61C >>G465R. Variants M420del and R61C were more sensitive to drug inhibition, with IC(50) values up to 23 times lower than those of the OCT1-reference. Uptake studies using (14)C-metformin were in qualitative agreement with those using ASP(+), with the exception that a larger reduction in transport capacity was observed for M420del. Concomitantly administered drugs, such as verapamil and amitriptyline, revealed potential drug-drug interactions at clinical plasma concentrations of metformin for OCT1-M420del.
Subject(s)
Genotype , Hypoglycemic Agents/pharmacology , Metformin/pharmacology , Organic Cation Transporter 1/antagonists & inhibitors , Cell Line , Drug Interactions , Green Fluorescent Proteins/metabolism , Humans , Hypoglycemic Agents/chemistry , Inhibitory Concentration 50 , Kinetics , Metformin/chemistry , Methylamines/chemistry , Organic Cation Transporter 1/metabolism , Pyridinium Compounds/chemistryABSTRACT
Integrins are heterodimeric transmembrane cell adhesion receptors that are essential for a wide range of biological functions via cell-matrix and cell-cell interactions. Recent studies have provided evidence that some of the subunits in the integrin family are involved in synaptic and behavioral plasticity. To further understand the role of integrins in the mammalian central nervous system, we generated a postnatal forebrain and excitatory neuron-specific knockout of alpha8-integrin in the mouse. Behavioral studies showed that the mutant mice are normal in multiple hippocampal-dependent learning tasks, including a T-maze, non-match-to-place working memory task for which other integrin subunits like alpha3- and beta1-integrin are required. In contrast, mice mutant for alpha8-integrin exhibited a specific impairment of long-term potentiation (LTP) at Schaffer collateral-CA1 synapses, whereas basal synaptic transmission, paired-pulse facilitation and long-term depression (LTD) remained unaffected. Because LTP is also impaired in the absence of alpha3-integrin, our results indicate that multiple integrin molecules are required for the normal expression of LTP, and different integrins display distinct roles in behavioral and neurophysiological processes like synaptic plasticity.