Identification of prescription opioid misuse and abuse behaviors and risk factors in chronic pain patients using the Prescription Opioid Misuse and Abuse Questionnaire (POMAQ).

Objectives: To identify patient risk factors associated with prescription opioid misuse and abuse as well as groupings of misuse and abuse behaviors as measured by the Prescription Opioid Misuse and Abuse Questionnaire (POMAQ).

Methods: Adults with chronic pain requiring long-term treatment with opioids completed the POMAQ and other study questionnaires. Latent class analysis (LCA) was used to examine underlying subgroups exhibiting particular risk profiles.

Inhaled vitamin A is more effective than intramuscular dosing in mitigating hyperoxia-induced lung injury in a neonatal rat model of bronchopulmonary dysplasia.

Prevention of bronchopulmonary dysplasia (BPD) in premature-birth babies continues to be an unmet medical need. Intramuscular vitamin A is currently employed in preterm neonates to prevent BPD but requires intramuscular injections in fragile neonates. We hypothesized that noninvasive inhaled delivery of vitamin A, targeted to lung, would be a more effective and tolerable strategy.

Preschool Deployment of Evidence-Based Social Communication Intervention: JASPER in the Classroom.

Few research-developed early intervention models have been deployed to and tested in real world preschool programs. In this study, teaching staff implemented a social communication modularized intervention, JASPER, in their daily program. Sixty-six preschool children with autism in twelve classrooms (12 teachers) were randomized to receive immediate JASPER training (IT) or were waitlisted (WL) for 3 months with a 1-month follow up.

Investigation of peptide biomarker stability in plasma samples using time-course MS analysis.

Peptide biomarkers in plasma or serum are subject to proteolytic degradation caused by intrinsic peptidase activities, resulting in a potential barrier in translating a discovered biomarker into clinical application. This chapter describes a method using time-course MALDI-TOF MS analysis to investigate the stability of a plasma peptide biomarker under a variety of preanalytical situations. A synthesized peptide with the same primary sequence as a potential endogenous biomarker is spiked into a blood sample, and the sample is incubated over time at r.

Minimizing preanalytical variation of plasma samples by proper blood collection and handling.

Blood samples collected for proteome studies are subject to a variety of preanalytical instability, among which intrinsic proteolysis activities cause a broad spectrum of protein and peptide degradation. This chapter describes two MALDI MS-based methods for plasma peptidomic analyses; a direct MALDI-TOF MS and an LC MALDI-TOF MS. Using these methods, we compared peptides and their time-dependent changes in traditional serum, four plasma samples with different anticoagulants and additives: EDTA-based, citrate-based, or heparin-based, and EDTA-based with protease inhibitors.

Application of free flow electrophoresis to the analysis of the urine proteome.

Urine is a complex fluid, which is thought to contain valuable diagnostic information regarding general health. In particular, there is great diagnostic potential in the peptide and/or protein content of urine, but the information is present in low abundance. Most traditional proteomic techniques lack sufficient sensitivity/dynamic range, especially for dilute and/or complex samples.

Resolution of adiponectin oligomers in human plasma using free flow electrophoresis.

Adiponectin is an important adipocytokine hormone which circulates in blood as homo-oligomers (trimer, hexamer and high molecular weight (HMW) forms) as well as a truncated form corresponding to the globular domain. Free flow electrophoresis (FFE) used in zone electrophoresis mode revealed the presence of isoforms within these oligomeric forms in plasma. HMW adiponectin oligomer showed two isoforms which carry different charge density at pH 4.

Intrinsic peptidase activity causes a sequential multi-step reaction (SMSR) in digestion of human plasma peptides.

Human plasma and serum samples, including protein and peptide biomarkers, are subjected to preanalytical variations and instability caused by intrinsic proteases. In this study, we directly investigated the stability of peptide biomarkers by spiking an isotopically labeled peptide into human plasma and serum samples and then monitoring its time-dependent change. Fibrinogen peptide A (FPA) was used as a model substrate, and its degradation in a conventional serum and plasma either with citrate, heparin, or EDTA as the anticoagulant, or EDTA plus protease inhibitors (inhibited plasma), was measured using time-course MALDI-TOF MS analysis.

Top-down proteomics on a high-field Fourier transform ion cyclotron resonance mass spectrometer.

Mass spectrometry is the tool of choice for sequencing peptides and determining the sites of posttranslational modifications; however, this bottom-up approach lacks in providing global information about the modification states of proteins including the number and types of isoforms and their stoichiometry. Recently, various techniques and mass spectrometers, such as high-field Fourier Transform Ion Cyclotron Resonance (FTICR) mass spectrometers, have been developed to study intact proteins (top-down proteomics). While the protein molecular mass and the qualitative and quantitative information about protein isoforms can be revealed by FTICR-MS analysis, their primary structure (including the identification of modifications and their exact locations in the amino acid sequence) can directly be determined using the MS/MS capability offered by the FTICR mass spectrometer.

Thrombin induces broad spectrum proteolysis in human serum samples.

Background: During clotting, a thrombin cleaves fibrinogen releasing fibrinopeptide A (FPA). FPA is easily identified in serum using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). Using MALDI-TOF MS, we observed multiple, progressively shorter fragments of serum FPA.

Free-flow electrophoresis for top-down proteomics by Fourier transform ion cyclotron resonance mass spectrometry.

High-efficiency prefractionation of complex protein mixtures is critical for top-down proteomics, i.e., the analysis of intact proteins by MS.

Inhibition of intrinsic proteolytic activities moderates preanalytical variability and instability of human plasma.

Human plasma and serum proteins are subject to intrinsic proteolytic degradation both during and after blood collection. By monitoring peptides, we investigated the stability of plasma and serum samples and the effects of anticoagulants and protease inhibitors on the plasma samples. Serum and plasma were subjected to time-course incubation, and the peptides (750-3200 Da) were extracted and analyzed with MALDI-TOF MS.

Region-specific detection of neuroblastoma loss of heterozygosity at multiple loci simultaneously using a SNP-based tag-array platform.

Many cancers are characterized by chromosomal aberrations that may be predictive of disease outcome. Human neuroblastomas are characterized by somatically acquired copy number changes, including loss of heterozygosity (LOH) at multiple chromosomal loci, and these aberrations are strongly associated with clinical phenotype including patient outcome. We developed a method to assess region-specific LOH by genotyping multiple SNPs simultaneously in DNA from tumor tissues.

HUPO Plasma Proteome Project specimen collection and handling: towards the standardization of parameters for plasma proteome samples.

There is a substantial list of pre-analytical variables that can alter the analysis of blood-derived samples. We have undertaken studies on some of these issues including choice of sample type, stability during storage, use of protease inhibitors, and clinical standardization. As there is a wide range of sample variables and a broad spectrum of analytical techniques in the HUPO PPP effort, it is not possible to define a single list of pre-analytical standards for samples or their processing.

High-density single-nucleotide polymorphism maps of the human genome.

Here we report a large, extensively characterized set of single-nucleotide polymorphisms (SNPs) covering the human genome. We determined the allele frequencies of 55,018 SNPs in African Americans, Asians (Japanese-Chinese), and European Americans as part of The SNP Consortium's Allele Frequency Project. A subset of 8333 SNPs was also characterized in Koreans.

beta-Adrenergic receptor polymorphisms and responses during titration of metoprolol controlled release/extended release in heart failure.

Objective: beta-Blockers require careful initiation and titration when used in patients with heart failure. Some patients tolerate beta-blocker therapy initiation without difficulty, whereas in other patients this period presents clinical challenges. We tested the hypothesis that polymorphisms at codons 389 (Arg389Gly) and 49 (Ser49Gly) of the beta(1)-adrenergic receptor would be associated with differences in initial tolerability of beta-blocker therapy in patients with heart failure.

Pharmacokinetics and CYP2D6 genotypes do not predict metoprolol adverse events or efficacy in hypertension.

Objective: Beta-Blocker use can be associated with adverse effects that may have an impact on adherence or harm patients. The commonly prescribed beta-blocker metoprolol is metabolized by the polymorphic cytochrome P450 (CYP) 2D6 enzyme, resulting in widely variable drug exposure. We investigated whether metoprolol plasma concentrations, CYP2D6 polymorphisms, or genotype-derived phenotype was associated with adverse effects or efficacy in patients with hypertension.

Auto-validation of fluorescent primer extension genotyping assay using signal clustering and neural networks.

Background: SNP genotyping typically incorporates a review step to ensure that the genotype calls for a particular SNP are correct. For high-throughput genotyping, such as that provided by the GenomeLab SNPstream instrument from Beckman Coulter, Inc., the manual review used for low-volume genotyping becomes a major bottleneck.

The thermodynamics of template-directed DNA synthesis: base insertion and extension enthalpies.

We used stopped-flow calorimetry to measure the overall enthalpy change associated with template-directed nucleotide insertion and DNA extension. Specifically, we used families of hairpin self-priming templates in conjunction with an exonuclease-free DNA polymerase to study primer extension by one or more dA or dT residues. Our results reveal exothermic heats between -9.

Advanced computational techniques for re-sequencing DNA with polymerase signaling assay arrays.

Re-sequencing, the identification of the specific variants in a sequence of interest compared with a known genomic sequence, is a ubiquitous task in today's biology. Universal arrays, which interrogate all possible oligonucleotides of a certain length in a target sequence, have been suggested for computationally determining a polynucleotide sequence from its oligonucleotide content. We present here new methods that use such arrays for re-sequencing.

Chromosome-wide distribution of haplotype blocks and the role of recombination hot spots.

Recent studies of human populations suggest that the genome consists of chromosome segments that are ancestrally conserved ('haplotype blocks'; refs. 1-3) and have discrete boundaries defined by recombination hot spots. Using publicly available genetic markers, we have constructed a first-generation haplotype map of chromosome 19.

SNPstream UHT: ultra-high throughput SNP genotyping for pharmacogenomics and drug discovery.

Single nucleotide polymorphism (SNP) genotyping is playing an increasing role in genome mapping, pharmacogenetic studies, and drug discovery. To date, genome-wide scans and studies involving thousands of SNPs and samples have been hampered by the lack of a system that can perform genotyping with cost-effective throughput, accuracy, and reliability. To address this need, Orrhid has developed an automated, ultra-high throughput system, SNPstream UHT, which uses multiplexed PCR in conjunction with our next generation SNP-IT tag array single base extension genotyping technology The system employs oligonucleotide microarrays manufactured in a 384-well format on a novel glass-bottomed plate.

Effects of 3,N4-ethenodeoxycytidine on duplex stability and energetics.

The exocyclic cytosine adduct 3,N4-ethenocytosine is highly mutagenic in mammalian cells. We describe the impact of this adduct on DNA duplex stability. The adduct does not disrupt the overall B-form DNA structure; however, structural accommodation of the adduct is necessary at the lesion site.

A quantitative method for evaluating the stabilities of nucleic acids.

We report a general method for screening, in solution, the impact of deviations from canonical Watson-Crick composition on the thermodynamic stability of nucleic acid duplexes. We demonstrate how fluorescence resonance energy transfer (FRET) can be used to detect directly free energy differences between an initially formed "reference" duplex (usually a Watson-Crick duplex) and a related "test" duplex containing a lesion/alteration of interest (e.g.

Mammographic changes after hormone replacement therapy in patients who have undergone breast irradiation.

Objective: We examined three patients who began hormone replacement therapy after lumpectomy and breast irradiation. In these women increased tissue density appeared on mammography only or disproportionately in the nonirradiated breast. To our knowledge, this observation has not been reported.