Age-related hearing impairment (ARHI), or presbycusis, is the most prevalent sensory impairment in the elderly. ARHI is a complex disease caused by an interaction between environmental and genetic factors. Here we describe the results of the first whole genome association study for ARHI. The study was performed using 846 cases and 846 controls selected from 3434 individuals collected by eight centers in six European countries. DNA pools for cases and controls were allelotyped on the Affymetrix 500K GeneChip for each center separately. The 252 top-ranked single nucleotide polymorphisms (SNPs) identified in a non-Finnish European sample group (1332 samples) and the 177 top-ranked SNPs from a Finnish sample group (360 samples) were confirmed using individual genotyping. Subsequently, the 23 most interesting SNPs were individually genotyped in an independent European replication group (138 samples). This resulted in the identification of a highly significant and replicated SNP located in GRM7, the gene encoding metabotropic glutamate receptor type 7. Also in the Finnish sample group, two GRM7 SNPs were significant, albeit in a different region of the gene. As the Finnish are genetically distinct from the rest of the European population, this may be due to allelic heterogeneity. We performed histochemical studies in human and mouse and showed that mGluR7 is expressed in hair cells and in spiral ganglion cells of the inner ear. Together these data indicate that common alleles of GRM7 contribute to an individual's risk of developing ARHI, possibly through a mechanism of altered susceptibility to glutamate excitotoxicity.

Familial combined hyperlipidemia (FCHL, MIM-144250) is a common, multifactorial and heterogeneous dyslipidemia predisposing to premature coronary artery disease and characterized by elevated plasma triglycerides, cholesterol, or both. We identified a mutant mouse strain, HcB-19/Dem (HcB-19), that shares features with FCHL, including hypertriglyceridemia, hypercholesterolemia, elevated plasma apolipoprotein B and increased secretion of triglyceride-rich lipoproteins. The hyperlipidemia results from spontaneous mutation at a locus, Hyplip1, on distal mouse chromosome 3 in a region syntenic with a 1q21-q23 FCHL locus identified in Finnish, German, Chinese and US families. We fine-mapped Hyplip1 to roughly 160 kb, constructed a BAC contig and sequenced overlapping BACs to identify 13 candidate genes. We found substantially decreased mRNA expression for thioredoxin interacting protein (Txnip). Sequencing of the critical region revealed a Txnip nonsense mutation in HcB-19 that is absent in its normolipidemic parental strains. Txnip encodes a cytoplasmic protein that binds and inhibits thioredoxin, a major regulator of cellular redox state. The mutant mice have decreased CO2 production but increased ketone body synthesis, suggesting that altered redox status down-regulates the citric-acid cycle, sparing fatty acids for triglyceride and ketone body production. These results reveal a new pathway of potential clinical significance that contributes to plasma lipid metabolism.

Thioredoxin-interacting protein (Txnip) is a ubiquitous protein that binds with high affinity to thioredoxin and inhibits its ability to reduce sulfhydryl groups via NADPH oxidation. HcB-19 mice contain a nonsense mutation in Txnip that eliminates its expression. Unlike normal animals, HcB-19 mice have ϳ3-fold increase in insulin levels when fasted. The C-peptide/insulin ratio is normal, suggesting that the hyperinsulinemia is due to increased insulin secretion. Fasted HcB-19 mice are hypoglycemic, hypertriglyceridemic, and have higher than normal levels of ketone bodies. Ablation of pancreatic ␤-cells with streptozotocin completely blocks the fasting-induced hypoglycemia/hypertriglyceridemia, suggesting that these abnormalities are due to excess insulin secretion. This is supported by increased hepatic mRNA levels of the insulin-inducible, lipogenic transcription factor sterol-responsive element-binding protein-1c and two of its targets, acetyl-CoA carboxylase and fatty acid synthase. During a prolonged fast, the hyperinsulinemia up-regulates lipogenesis but fails to down-regulate hepatic phosphoenolpyruvate carboxykinase mRNA expression. Hepatic ratios of reduced:oxidized glutathione, established regulators of gluconeogenic/glycolytic/lipogenic enzymes, were elevated 30% in HcB-19 mice, suggesting a loss of Txnip-enhanced sulfhydryl reduction. The altered hepatic enzymatic profiles of HcB-19 mice divert phosphoenolpyruvate to glyceroneogenesis and lipogenesis rather than gluconeogenesis. Our findings implicate Txnip-modulated sulfhydryl redox as a central regulator of insulin secretion in ␤-cells and regulation of many of the branch-points of gluconeogenesis/glycolysis/lipogenesis.

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We now show that Txnip-deficient mice in the fed state exhibit a metabolic profile similar to fasted mice, including increased levels of plasma ketone bodies and free fatty acids, decreased glucose, and increased hepatic expression of peroxisome proliferator-activated receptor-␥ coactivator-1 ␣ , phosphoenolpyruvate carboxykinase, glucose-6-phosphatase, and acylCoA oxidase. Dramatic differences in the expression of key metabolic enzymes were also observed in other tissues, and the fat-to-muscle ratio of Txnip-deficient mice was increased by ‫ف‬ 40%. We demonstrate an effect of Txnip on the redox status, as the Txnip-deficient mice in the fed state had a significant increase in the ratio of NADH to NAD ؉ . Surprisingly, we observed that Txnip-deficient mice and wild-type mice had similar levels of thioredoxin activity, suggesting that the effects of Txnip deficiency may be mediated in part by other interactions. These results indicate a role for Txnip in the metabolic response to feeding and the maintenance of the redox status.

The effect of decreased fatty acid oxidation on liver lipid metabolism in HcB-19 mice, a mouse model of hyperlipidemia (Txnip(-/-)), was investigated using metabolic labeling. De novo cholesterol synthesis and de novo lipogenesis were quantified using 1-(13)C(1) acetic acid, and liver triacylglycerol (TAG) derived from dietary fatty acids was quantified using dietary glyceryl tri(hexandecanoate-d(31)). Tissue samples were analyzed for TAG, free cholesterol (FC), and cholesterol ester (CE) content. Txnip(-/-) mice had significantly elevated (P < 0.05) serum nonesterified fatty acids compared with wild-type (WT) littermates; their livers weighed more and contained more TAG and total cholesterol. Txnip(-/-) liver also contained measurable CE; CE was not detectable in WT mice. Liver CE content was elevated despite lower cholesterol fractional synthesis rates (16 vs. 31%/d in Txnip(-/-) and WT mice, respectively). FC absolute synthesis rate (ASR) in WT mice (0.28 +/- 0.0 micromol/d) was similar to the combined synthesis rates of FC (0.13 +/- 0.10 micromol/d) and CE (0.10 +/- 0.00 micromol/d) in Txnip(-/-) mice. Lipogenesis, as assessed by TAG-palmitate ASR, was significantly greater in Txnip(-/-) mice (1.47 +/- 0.08 vs. 0.49 +/- 0.06 micro mol/d) and liver fatty acid synthase activity was also higher (7.96 +/- 2.53 vs. 4.83 +/- 1.44 U/mg protein). Both elevated lipogenesis and increased fatty acid reesterification to glycerol and cholesterol contributed to fat in the livers of Txnip(-/-) mice. These data support elevated fatty acid synthesis as the primary contributor to liver TAG in Txnip(-/-) mice, although increased esterification of fatty acids also contributed to excess liver TAG. The absolute total cholesterol synthesis rate was not altered, but esterification of fatty acids to cholesterol provided an additional means to buffer physiologically the negative results of excess fatty acid availability.

(http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. metabolic pathway gene expression in mice

In order to evaluate metabolic pathways associated with obesity, global gene-expression data were integrated with phenotypic and genetic segregation analyses, identifying 13 metabolic pathways the genes of which are coordinately regulated in association with obesity. Four genomic regions were found to control the coordinated expression of these pathways and novel genes potentially associated with the identified pathways were identified.

Noise-induced hearing loss (NIHL) is one of the more common sources of environmentally induced hearing loss in adults. In a mouse model, Castaneous (CAST/Ei) is an inbred strain that is resistant to NIHL, while the C57BL/6J strain is susceptible. We have used the genome-tagged mice (GTM) library of congenic strains, carrying defined segments of the CAST/Ei genome introgressed onto the C57BL/6J background, to search for loci modifying the noise-induced damage seen in the C57BL/6J strain. NIHL was induced by exposing 6-8-week old mice to 108 dB SPL intensity noise. We tested the hearing of each mouse strain up to 23 days after noise exposure using auditory brainstem response (ABR). This study identifies a number of genetic loci that modify the initial response to damaging noise, as well as long-term recovery. The data suggest that multiple alleles within the CAST/Ei genome modify the pathogenesis of NIHL and that screening congenic libraries for loci that underlie traits of interest can be easily carried out in a high-throughput fashion.