Sex-specific quantitative trait loci govern susceptibility to Theiler's murine encephalomyelitis virus-induced demyelination

Susceptibility to Theiler's murine encephalomyelitis virus-induced demyelination (TMEVD), a mouse model for multiple sclerosis (MS), is genetically controlled. Through a mouse-human comparative mapping approach, identification of candidate susceptibility loci for MS based on the location of TMEVD susceptibility loci may be possible. Composite interval mapping (CIM) identified quantitative trait loci (QTL) controlling TMEVD severity in male and female backcross populations derived from susceptible DBA/2J and resistant BALBc/ByJ mice. We report QTL on chromosomes 1, 5, 15, and 16 affecting male mice. In addition, we identified two QTL in female mice located on chromosome 1. Our results support the existence of three linked sex-specific QTL on chromosome 1 with opposing effects on the severity of the clinical signs of TMEV-induced disease in male and female mice.     

Degradation of biphenyl by Mycobacterium sp. strain PYR-1

The metabolism of biphenyl by Mycobacterium sp. PYR-1 was investigated. The Mycobacterium sp. degraded >98% of the biphenyl added within 72 h. Analysis of ethyl acetate extracts of the culture medium by HPLC indicated that benzoic acid was the major metabolite. Other products were 4-hydroxybiphenyl, 4-hydroxybenzoic acid, and 5-oxo-5-phenylpentanoic acid. The metabolites were characterized by mass and 1H NMR spectrometry. Identification of benzoic acid and 5-oxo-5-phenylpentanoic acid indicates that biphenyl degradation by Mycobacterium sp. PYR-1 is generally similar to known pathways. A novel alternative metabolic pathway consisted of monooxygenation at C-4 of biphenyl to give 4-hydroxybiphenyl, with subsequent degradation via ring cleavage to 4-hydroxybenzoic acid.     

Mapping and analysis of quantitative trait loci in experimental populations

Simple statistical methods for the study of quantitative trait loci (QTL), such as analysis of variance, have given way to methods that involve several markers and high-resolution genetic maps. As a result, the mapping community has been provided with statistical and computational tools that have much greater power than ever before for studying and locating multiple and interacting QTL. Apart from their immediate practical applications, the lessons learnt from this evolution of QTL methodology might also be generally relevant to other types of functional genomics approach that are aimed at the dissection of complex phenotypes, such as microarray assessment of gene expression.     

High-throughput quantification of soy isoflavones in human and rodent blood using liquid chromatography with electrospray mass spectrometry and tandem mass spectrometry detection

Soy-containing foods and dietary supplements are widely consumed for putative health benefits (e.g., cancer chemoprevention, beneficial effects on serum lipids associated with cardiovascular health, reduction of osteoporosis, relief of menopausal symptoms). However, studies of soy isoflavones in experimental animals suggest possible adverse effects as well (e.g., enhancement of reproductive organ cancer, modulation of endocrine function, anti-thyroid effects). This paper describes the development and validation of a sensitive high throughput method for quantifying isoflavones in blood from experimental animal and human studies. Serum samples containing genistein, daidzein, and equol were processed using reverse phase solid-phase extraction in the 96-well format for subsequent LC-ES/MS/MS or LC-ES/MS analysis using isotope dilution in conjunction with labeled internal standards. The method was validated by repetitive analysis of spiked blank serum and the intra-day and inter-day accuracy (88-99%) and precision (relative standard deviations from 3 to 13%) of measurement determined. The lower limit of quantification for all isoflavones was approximately 0.005 micro M using MS/MS detection, and 0.03 micro M using MS for genistein and daidzein. The degree of method performance, with respect to throughput, sensitivity and selectivity, makes this approach practical for analysis of large sample sets generated from mechanistic animal studies and human clinical trials of soy isoflavones.     

Inactivation of thyroid peroxidase by soy isoflavones,in vitro and in vivo

Soy-containing foods and dietary supplements are widely consumed for putative health benefits (e.g. cancer chemoprevention, beneficial effects on serum lipids associated with cardiovascular health, reduction of osteoporosis, relief of menopausal symptoms). However, studies of soy isoflavones in experimental animals suggest possible adverse effects as well (e.g. enhancement of reproductive organ cancer, modulation of endocrine function, anti-thyroid effects). This paper reviews the evidence in humans and animals for anti-thyroid effects of soy and its principal isoflavones, genistein and daidzein.     

Equol, a natural estrogenic metabolite from soy isoflavones: convenient preparation and resolution of R- and S-equols and their differing binding and biological activity through estrogen receptors alpha and beta

Equol is a metabolite produced in vivo from the soy phytoestrogen daidzein by the action of gut microflora. It is known to be estrogenic, so human exposure to equol could have significant biological effects. Equol is a chiral molecule that can exist as the enantiomers R-equol and S-equol. To study the biological activity of racemic (+/-)-equol, as well as that of its pure enantiomers, we developed an efficient and convenient method to prepare (+/-)-equol from available isoflavanoid precursors. Furthermore, we optimized a method to separate the enantiomers of equol by chiral HPLC, and we studied for the first time, the activities of the enantiomers on the two estrogen receptors, ERalpha and ERbeta. In binding assays, S-equol has a high binding affinity, preferential for ERbeta (K(i)[ERbeta]=16 nM; beta/alpha=13 fold), that is comparable to that of genistein (K(i)[ERbeta]=6.7 nM; beta/alpha=16), whereas R-equol binds more weakly and with a preference for ERalpha (K(i)[ERalpha]=50 nM; beta/alpha=0.29). All equol isomers have higher affinity for both ERs than does the biosynthetic precursor daidzein. The availability and the in vitro characterization of the equol enantiomers should enable their biological effects to be studied in detail.     

DNA adducts derived from administration of acrylamide and glycidamide to mice and rats

Acrylamide (AA) is an important industrial chemical that is neurotoxic, mutagenic to somatic and germ cells, and carcinogenic in chronic rodent bioassays. Recent findings of AA in many common starchy foods have sparked renewed interest in determining toxic mechanisms and in understanding the cancer, neurotoxicity, and reproductive risks from typical human exposures. Dosing mice and rats with AA (50 mg/kg) led to presence of glycidamide (GA) in serum and tissues. Furthermore, GA-derived DNA adducts of adenine and guanine were formed in all tissues examined, including both target tissues identified in rodent carcinogenicity bioassays and in non-target tissues. Dosing rats and mice with an equimolar amount of GA typically produced higher levels of DNA adducts than observed with AA. Kinetics of DNA adduct formation and accumulation were measured following oral administration of a single dose of AA (50 mg/kg) or from repeat dosing (1 mg/kg/day), respectively. The formation of these DNA adducts is consistent with previously reported mutagenicity of AA and GA in vitro, which involved reaction of GA with adenine and guanine bases. These results provide strong support for a genotoxic mechanism of AA carcinogenicity in rodents. The kinetic/biomarker approaches described here may represent a meaningful way to extrapolate cancer risks to actual human exposures from food, which are much lower.     

A model selection-based interval-mapping method for autopolyploids

While extensive progress has been made in quantitative trait locus (QTL) mapping for diploid species, similar progress in QTL mapping for polyploids has been limited due to the complex genetic architecture of polyploids. To date, QTL mapping in polyploids has focused mainly on tetraploids with dominant and/or codominant markers. Here, we extend this view to include any even ploidy level under a dominant marker system. Our approach first selects the most likely chromosomal marker configurations using a Bayesian selection criterion and then fits an interval-mapping model to each candidate. Profiles of the likelihood-ratio test statistic and the maximum-likelihood estimates (MLEs) of parameters including QTL effects are obtained via the EM algorithm. Putative QTL are then detected using a resampling-based significance threshold, and the corresponding parental configuration is identified to be the underlying parental configuration from which the data are observed. Although presented via pseudo-doubled backcross experiments, this approach can be readily extended to other breeding systems. Our method is applied to single-dose restriction fragment autotetraploid alfalfa data, and the performance is investigated through simulation studies.     

Accounting for variability in the use of permutation testing to detect quantitative trait loci

Locating quantitative trait loci (QTL), or genomic regions associated with known molecular markers, is of increasing interest in a wide variety of applications ranging from human genetics to agricultural genetics. The hope of locating QTL (or genes) affecting a quantitative trait is that it will lead to characterization and possible manipulations of these genes. However, the complexity of both statistical and genetic issues surrounding the location of these regions calls into question the asymptotic statistical results supplying the distribution of the test statistics employed. Coupled with the power of current-day computing, permutation theory was reintroduced for the purpose of estimating the distribution of any test statistic used to test for the location of QTL. Permutation techniques have offered an attractive alternative to significance measures based on asymptotic theory. The ideas of permutation testing are extended in this application to include confidence intervals for the thresholds and p-values estimated in permutation testing procedures. The confidence intervals developed account for the Monte Carlo error associated with practical applications of permutation testing and lead to an effective method of determining an efficient permutation sample size.     

Combining Affymetrix microarray results

Background  

As the use of microarray technology becomes more prevalent it is not unusual to find several laboratories employing the same microarray technology to identify genes related to the same condition in the same species. Although the experimental specifics are similar, typically a different list of statistically significant genes result from each data analysis.