Mark W Duncan | University of Colorado Denver (original) (raw)

Papers by Mark W Duncan

Research paper thumbnail of Discovery and Validation of Protein Abundance Differences between Follicular Thyroid Neoplasms

Cancer Research, 2008

Distinguishing between benign follicular thyroid adenoma (FTA) and malignant follicular thyroid c... more Distinguishing between benign follicular thyroid adenoma (FTA) and malignant follicular thyroid carcinoma (FTC) by cytologic features alone is not possible. Molecular markers may aid distinguishing FTA from FTC in patients with indeterminate cytology. The aim of this study is to define protein abundance differences between FTC from FTA through a discovery (proteomics) and validation (immunohistochemistry) approach. Difference gel electrophoresis (DIGE) and peptide mass fingerprinting were performed on protein extracts from five patients with FTC and compared with six patients with FTA. Individual gel comparisons (i.e., each FTC extract versus FTA pool) were also performed for the five FTC patients. Immunohistochemical validation studies were performed on three of the identified proteins. Based on DIGE images, 680 protein spots were matched on individual gels. Of these, 102 spots showed statistically significant differences in abundance between FTC and FTA in the individual gel analyses and were therefore studied further. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to identify 54 of these protein spots. Three candidates involved in protein folding (heat shock protein gp96, protein disulfide isomerase A3, and calreticulin) were studied by immunohistochemistry. Moderate calreticulin immunohistochemical staining was the best single marker with a high negative predictive value (88%); combining all three markers (any marker less than moderate staining) had the best positive predictive value (75%) while still retaining a good negative predictive value (68%). With DIGE, we identified 54 proteins differentially abundant between FTC and FTA. Three of these were validated by immunohistochemistry. These findings provide further insights into the diagnosis, prognosis, and pathophysiology of follicular-derived thyroid neoplasms.

Research paper thumbnail of Proteomic Analysis of Mouse Lung Neoplasia *

CHEST Journal, 2002

can be found online on the World Wide Web at:

Research paper thumbnail of Identification of contaminants in proteomics mass spectrometry data

Mass spectrometry (MS) is a widely used method for protein identification. Peptide mass fingerpri... more Mass spectrometry (MS) is a widely used method for protein identification. Peptide mass fingerprinting is the protein identification technique in which MS is employed to determine the masses of peptide fragments generated following enzymatic digestion of proteins. The masses of peptides are then submitted to a recognition program, e.g., MASCOT or MSFIT, for identification of a protein. The strategy is hampered, however, because not only are the peptide masses determined, but also the masses of multiple contaminants that are also present in the sample. Although the masses of some common and known contaminants are removed (e.g., peptides generated by trypsin autolysis), many others are inadvertently incorporated into the analysis. In this paper we present an approach for automa tic identification of contaminant masses so that they can be removed prior to the identification process. For this purpose we have developed an algorithm that clusters mass values. We calculate the frequencies of all masses and then identify contaminants. We propose that masses with frequency higher than a given value are contaminants. In our analysis of 3,029 digested proteins, yielding 78,384 masses, we identified 16 possible contaminants. Of these 16, four are known trypsin autolysis peptides. Removing these contaminant masses from the database search will lead to more accurate and reliable protein identification.

Research paper thumbnail of Mass Spectrometry as an Approach for the Characterization and Identification of Components in Human Tear Fluid

Research paper thumbnail of Mass spectrometric techniques applied to the analysis of human tears: a focus on the peptide and protein constituents

Advances in experimental medicine and biology, 2002

Research paper thumbnail of The Proteinaceous Composition of Human Tear Fluid as Determined by Mass Spectrometry

Research paper thumbnail of Practical quantitative biomedical applications of MALDI-TOF mass spectrometry

Journal of the American Society for Mass Spectrometry, 2002

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS) is use... more Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS) is used to obtain fast and accurate determinations of molecular mass, but quantitative determinations are generally made by other techniques. In this study we illustrate the practical utility of automated MALDI-TOFMS as a tool for quantifying a diverse array of biomolecules covering an extensive molecular weight range, and present in biological extracts and fluids. Growth hormone was measured in rat pituitary tissue; insulin in human pancreatic tissue; homovanillic acid in human urine; and LVV-hemorphin-7, epinephrine and norepinephrine in human adrenal and pheochromocytoma tissues. Internal standards including compounds of similar molecular weight, structural analogs or isotopomers were incorporated into each analysis. We report on the current practical limitations of quantitative MALDI-TOFMS and highlight some of the potential benefits of this technique as a quantitative tool. (J Am Soc

Research paper thumbnail of Alpha-Linolenic Acid: An Omega-3 Fatty Acid with Neuroprotective Properties-Ready for Use in the Stroke Clinic?

BioMed research international, 2015

Alpha-linolenic acid (ALA) is plant-based essential omega-3 polyunsaturated fatty acids that must... more Alpha-linolenic acid (ALA) is plant-based essential omega-3 polyunsaturated fatty acids that must be obtained through the diet. This could explain in part why the severe deficiency in omega-3 intake pointed by numerous epidemiologic studies may increase the brain's vulnerability representing an important risk factor in the development and/or deterioration of certain cardio- and neuropathologies. The roles of ALA in neurological disorders remain unclear, especially in stroke that is a leading cause of death. We and others have identified ALA as a potential nutraceutical to protect the brain from stroke, characterized by its pleiotropic effects in neuroprotection, vasodilation of brain arteries, and neuroplasticity. This review highlights how chronic administration of ALA protects against rodent models of hypoxic-ischemic injury and exerts an anti-depressant-like activity, effects that likely involve multiple mechanisms in brain, and may be applied in stroke prevention. One major ...

Research paper thumbnail of Mature adipocyte proteome reveals differentially altered protein abundances between lean, overweight and morbidly obese human subjects

Molecular and Cellular Endocrinology, 2015

Overweight (OW) and obese individuals are considered to be graded parts of the scale having incre... more Overweight (OW) and obese individuals are considered to be graded parts of the scale having increasing weight as a common feature. They may not, however, be part of the same continuum and may differ metabolically. In this study we applied an untargeted proteomic approach to compare protein abundances in mature adipocytes derived from the subcutaneous adipose tissue of overweight and morbidly obese female subjects to those of lean age matched controls. Mature adipocytes were isolated from liposuction samples of abdominal subcutaneous adipose tissue collected from both lean (L; n = 7, 23.3 ± 0.4 kg/m(2); mean BMI ± SD), overweight (OW; n = 8, 27.9 ± 0.6 kg/m(2); mean BMI ± SD) and morbidly obese (MOB; n = 7, 44.8 ± 3.8 kg/m(2); mean BMI ± SD) individuals. Total protein extracts were then compared by two-dimensional difference in gel electrophoresis (2D DIGE). One hundred and ten differentially expressed protein spots (i.e., fitting the statistical criteria ANOVA test, p < 0.05; fold-change ≥1.5) were detected, and of these, 89 were identified by MALDI-TOF mass spectrometry. Of these, 66 protein spots were common to both groups whereas 23 were unique to the MOB group. Significant differences were evident in the abundances of key proteins involved in glucose and lipid metabolism, energy regulation, cytoskeletal structure and redox control signaling pathways. Differences in the abundance of some chaperones were also evident. The differentially abundant proteins were investigated using Ingenuity Pathway Analysis (IPA) to establish their associations with known biological functions. The network identified in the OW group with the highest score relates to-: cell-to-cell signaling and interaction; in contrast, in the MOB group the major interacting pathways are associated with lipid metabolism, small molecule biochemistry and cancer. The differences in abundance of the differentially regulated proteins were validated by immunoblotting. These findings provide insights into metabolic differences in OW and MOB individuals.

Research paper thumbnail of Purification and Structural Characterization of Siderophore (Corynebactin) from Corynebacterium diphtheriae

PLOS One, 2012

During infection, Corynebacterium diphtheriae must compete with host iron-sequestering mechanisms... more During infection, Corynebacterium diphtheriae must compete with host iron-sequestering mechanisms for iron. C. diphtheriae can acquire iron by a siderophore-dependent iron-uptake pathway, by uptake and degradation of heme, or both. Previous studies showed that production of siderophore (corynebactin) by C. diphtheriae is repressed under high-iron growth conditions by the iron-activated diphtheria toxin repressor (DtxR) and that partially purified corynebactin fails

Research paper thumbnail of Utility of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the analysis of low molecular weight compounds

Rapid Communications in Mass Spectrometry, 1995

ABSTRACT

Research paper thumbnail of Circulating Cytokines and Growth Factors in Pediatric Pulmonary Hypertension

Mediators of Inflammation, 2012

Background. Management of pediatric pulmonary hypertension (PH) remains challenging. We have asse... more Background. Management of pediatric pulmonary hypertension (PH) remains challenging. We have assessed a panel of circulating proteins in children with PH to investigate their value as predictive and/or prognostic biomarkers. From these determinations, we aim to develop a practical, noninvasive tool to aid in the management of pediatric PH. Methods. Twelve cytokines and growth factors putatively associated with lung or vascular disease were examined in plasma specimens from 70 children with PH using multiplex protein array technology. Associations between hemodynamics, adverse events, and protein markers were evaluated. Results. Epidermal growth factor (EGF) and IL-6 were associated with important hemodynamics. Of the twelve proteins, VEGF and IL-6 were significantly, univariately associated with the occurrence of an adverse event, with odds ratios (95% confidence intervals) of 0.56 (0.33-0.98) and 1.69 (1.03-2.77), respectively. When hemodynamic predictors were combined with protein markers, the ability to predict adverse outcomes within the following year significantly increased. Conclusions. Specific circulating proteins are associated with hemodynamic variables in pediatric PH. If confirmed in additional cohorts, measurement of these proteins could aid patient care and design of clinical trials by identifying patients at risk for adverse events. These findings also further support a role for inflammation in pediatric PH.

Research paper thumbnail of Phenolic compounds in the nearshore waters of Sydney, Australia

Marine and Freshwater Research, 1996

ABSTRACT

Research paper thumbnail of Prediction of benefit from EGFR TKIs by proteomic analysis of pretreatment serum

Journal of Thoracic Oncology, 2007

Research paper thumbnail of Vaccine Impedes the Development of Reflux-induced Esophageal Cancer in a Surgical Rat Model: Efficacy of the Vaccine in a Post-Barrett’s Esophagus Setting

Digestive Diseases and Sciences, 2008

Background & Aims Wedevelopedagranulocyte-macrophage-colony-stimulatingfactor(GM-CSF)tumorvaccine... more Background & Aims Wedevelopedagranulocyte-macrophage-colony-stimulatingfactor(GM-CSF)tumorvaccineforesophageal cancer.Weevaluatedtheeffectivenessofthevaccineasapreventionoptioninasurgicalrefluxratmodelofesophagealcancer. Methods A surgical model involving a jejuno-esophagostomy was used to create Barrett's esophagus and esophageal cancer in rats. No carcinogen exposure was utilized. Cell lines derived from these tumors were stably passaged in vitro. GM-CSF-secreting tumor cells were generated using stable transfection. All rats underwent a total gastrectomy, followed by a jejuno-esophagostomy. The surgery promoted the reflux of duodenal contents into the esophagus. All animals were administered either a GM-CSF secreting whole cell vaccine or a phosphate-buffered saline (PBS) placebo injection 4, 6, 14, and 16 weeks post-surgery. Results While 15 of 16 animals in the non-vaccinated placebo group developed esophageal cancer, 94% (15 of 16), animals in the vaccine group had an incidence of cancer of 25% (4 of 16) (p<0.05). Barrett's esophagus was seen in 100% (16 of 16) of the controls and 83% (13 of 16) of the vaccinated animals. Conclusions A GM-CSF-secreting whole cell tumor vaccine impeded esophageal tumor growth, but not the development of Barrett's esophagus, in a clinically relevant surgical reflux model.

Research paper thumbnail of Serum E-cadherin does not associate with response to gefitinib in patients with non-small cell lung cancer (NSCLC)

Journal of Thoracic Oncology, 2007

Research paper thumbnail of Assessment of post-mortem-induced changes to the mouse brain proteome

Journal of Neurochemistry, 2008

This study was designed to assess the influence of high-energy head-focused microwave irradiation... more This study was designed to assess the influence of high-energy head-focused microwave irradiation and the post-mortem interval on measurements of the mouse brain proteome. Difference gel electrophoresis was used to compare mouse brain protein levels in animals killed by decapitation, where the tissue was held at 25 degrees C for selected time intervals post-mortem, and by high-energy head-focused microwave irradiation followed by immediate resection. Microwave-mediated killing was used because it comprehensively snap-inactivates enzymes while largely retaining brain cytoarchitecture. Of the 912 protein spots common to at least eight of 10 gels analyzed, 35 (3.8%) showed significant differences in levels (t-test; p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05) depending on whether animals were killed by microwave irradiation or decapitation. When animals were killed by decapitation, 43 protein spots (4.7%) showed changes in levels over the post-mortem interval (anova; p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05). The vast majority of the near 1000 proteins evident on a 2D gel were stable for up to 4 h. These data have important implications for studies of proteins in the brain, whether based on analysis of tissue derived from animal models or from humans.

Research paper thumbnail of Nicotine and Cotinine Adducts of a Melanin Intermediate Demonstrated by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

Chemical Research in Toxicology, 2001

Pigmentation is a major factor in the incorporation of many drugs into hair. In an attempt to elu... more Pigmentation is a major factor in the incorporation of many drugs into hair. In an attempt to elucidate potential mechanisms of drug-melanin interaction, melanin was synthesized in vitro in the presence of nicotine, which we have shown to have a substantial interaction with melanin, and cotinine, a primary nicotine metabolite. L-DOPA, a precursor of eumelanin, was oxidized and oligomerized with tyrosinase. Nicotine, cotinine, and/or their deuterated analogues were added to the oligomerization reaction mixture in a 10:1 L-DOPA:drug ratio. A black precipitate formed within 60 min. Aliquots were removed from the incubation mixture at 60, 120, and 360 min. MALDI-TOF MS determinations were carried out on each sample to provide a mean and standard error for the masses of interest. Internal calibration allowed accurate mass measurement of the products. A careful comparison of the spectra of samples prepared both with and without drug indicated the presence of masses corresponding to the protonated drug, melanin oligomers, and nicotine or continine adducts of the monomeric melanin intermediate dopaquinone (DOPAQ). Additional support for the presence of drug-melanin adducts was provided by employing deuterated analogues of nicotine and L-DOPA in the reaction and observing that the masses shifted accordingly. Structures of the adducts were further confirmed by select ion gating and postsource decay analysis.

Research paper thumbnail of Relation between normal rectal methylation, smoking status, and the presence or absence of colorectal adenomas

Cancer, 2010

BACKGROUND-Colorectal cancer (CRC) is 1 of the leading causes of death in the Western world. CRC ... more BACKGROUND-Colorectal cancer (CRC) is 1 of the leading causes of death in the Western world. CRC develops from premalignant lesions, chiefly colorectal adenomas. Currently, the most accurate and recommended screening method for finding colorectal adenomas is colonoscopy performed on all individuals aged >50 years. However, the costs and risks associated with this procedure are relatively high. The objectives of the current study were to correlate epigenetic alterations that occur in normal rectal mucosa, smoking status, and age with the presence or absence of concomitant colorectal adenomas and to assess the potential clinical value of methylation in normal rectal biopsies as a screening assay for the presence of polyps and, hence, the need for a full colonoscopy.

Research paper thumbnail of Analysis of docetaxel pharmacokinetics in humans with the inclusion of later sampling time-points afforded by the use of a sensitive tandem LCMS assay

Cancer Chemotherapy and Pharmacology, 2003

Docetaxel is a semisynthetic taxane derived from the needles of the European yew ( Taxus baccata)... more Docetaxel is a semisynthetic taxane derived from the needles of the European yew ( Taxus baccata) and it is an important chemotherapeutic agent in the treatment of recurrent ovarian, breast and non-small-cell lung cancers. Traditional dosing regimens with docetaxel involve doses of 60-100 mg/m(2) by infusion every 3 weeks. Now weekly low-dose (30-36 mg/m(2)) regimens are being evaluated in phase I trials. Such low-dose studies require a more sensitive, specific and rapid assay of docetaxel in biological fluids for the determination of pharmacokinetic parameters. Because docetaxel is primarily metabolized by CYP3A4 and is highly protein-bound in the plasma, there is potential for drug-drug interactions and high interpatient variability in pharmacokinetics. Therefore, pharmacokinetic studies are an important component to understanding the therapeutic variability of docetaxel-containing chemotherapeutic regimens. To this end, we developed an analytical assay for docetaxel based upon tandem LCMS and paclitaxel as an internal standard. The sensitivity of the new assay allowed us to monitor plasma levels of docetaxel out to 48 h after the end of the infusion in patients enrolled in a phase I trial of exisulind (orally, twice daily) receiving weekly docetaxel doses of 30 or 36 mg/m(2) where plasma docetaxel levels are below the lower limit of quantitation for traditional HPLC/UV-based assays at later time-points. The inclusion of the 48-h time-point had significant effects on the calculated pharmacokinetic parameters when using either a three-compartment or non-compartmental analysis. The terminal half-life was significantly increased when the 48-h time-point was included in the pharmacokinetic analysis, and the use of model parameters derived with the inclusion of the 48-h time-point were able to more accurately predict plasma levels at later times. The results reflect the importance of accurate and sensitive analytical methods for the determination of pharmacokinetic parameters and the effect of this later time-point on docetaxel pharmacokinetic modeling. Further, with the increased use of weekly docetaxel in combination with other agents, the inclusion of these later sampling time-points and sensitive methods for drug level determinations are important components in the description of pharmacokinetic drug interactions.

Research paper thumbnail of Discovery and Validation of Protein Abundance Differences between Follicular Thyroid Neoplasms

Cancer Research, 2008

Distinguishing between benign follicular thyroid adenoma (FTA) and malignant follicular thyroid c... more Distinguishing between benign follicular thyroid adenoma (FTA) and malignant follicular thyroid carcinoma (FTC) by cytologic features alone is not possible. Molecular markers may aid distinguishing FTA from FTC in patients with indeterminate cytology. The aim of this study is to define protein abundance differences between FTC from FTA through a discovery (proteomics) and validation (immunohistochemistry) approach. Difference gel electrophoresis (DIGE) and peptide mass fingerprinting were performed on protein extracts from five patients with FTC and compared with six patients with FTA. Individual gel comparisons (i.e., each FTC extract versus FTA pool) were also performed for the five FTC patients. Immunohistochemical validation studies were performed on three of the identified proteins. Based on DIGE images, 680 protein spots were matched on individual gels. Of these, 102 spots showed statistically significant differences in abundance between FTC and FTA in the individual gel analyses and were therefore studied further. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to identify 54 of these protein spots. Three candidates involved in protein folding (heat shock protein gp96, protein disulfide isomerase A3, and calreticulin) were studied by immunohistochemistry. Moderate calreticulin immunohistochemical staining was the best single marker with a high negative predictive value (88%); combining all three markers (any marker less than moderate staining) had the best positive predictive value (75%) while still retaining a good negative predictive value (68%). With DIGE, we identified 54 proteins differentially abundant between FTC and FTA. Three of these were validated by immunohistochemistry. These findings provide further insights into the diagnosis, prognosis, and pathophysiology of follicular-derived thyroid neoplasms.

Research paper thumbnail of Proteomic Analysis of Mouse Lung Neoplasia *

CHEST Journal, 2002

can be found online on the World Wide Web at:

Research paper thumbnail of Identification of contaminants in proteomics mass spectrometry data

Mass spectrometry (MS) is a widely used method for protein identification. Peptide mass fingerpri... more Mass spectrometry (MS) is a widely used method for protein identification. Peptide mass fingerprinting is the protein identification technique in which MS is employed to determine the masses of peptide fragments generated following enzymatic digestion of proteins. The masses of peptides are then submitted to a recognition program, e.g., MASCOT or MSFIT, for identification of a protein. The strategy is hampered, however, because not only are the peptide masses determined, but also the masses of multiple contaminants that are also present in the sample. Although the masses of some common and known contaminants are removed (e.g., peptides generated by trypsin autolysis), many others are inadvertently incorporated into the analysis. In this paper we present an approach for automa tic identification of contaminant masses so that they can be removed prior to the identification process. For this purpose we have developed an algorithm that clusters mass values. We calculate the frequencies of all masses and then identify contaminants. We propose that masses with frequency higher than a given value are contaminants. In our analysis of 3,029 digested proteins, yielding 78,384 masses, we identified 16 possible contaminants. Of these 16, four are known trypsin autolysis peptides. Removing these contaminant masses from the database search will lead to more accurate and reliable protein identification.

Research paper thumbnail of Mass Spectrometry as an Approach for the Characterization and Identification of Components in Human Tear Fluid

Research paper thumbnail of Mass spectrometric techniques applied to the analysis of human tears: a focus on the peptide and protein constituents

Advances in experimental medicine and biology, 2002

Research paper thumbnail of The Proteinaceous Composition of Human Tear Fluid as Determined by Mass Spectrometry

Research paper thumbnail of Practical quantitative biomedical applications of MALDI-TOF mass spectrometry

Journal of the American Society for Mass Spectrometry, 2002

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS) is use... more Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS) is used to obtain fast and accurate determinations of molecular mass, but quantitative determinations are generally made by other techniques. In this study we illustrate the practical utility of automated MALDI-TOFMS as a tool for quantifying a diverse array of biomolecules covering an extensive molecular weight range, and present in biological extracts and fluids. Growth hormone was measured in rat pituitary tissue; insulin in human pancreatic tissue; homovanillic acid in human urine; and LVV-hemorphin-7, epinephrine and norepinephrine in human adrenal and pheochromocytoma tissues. Internal standards including compounds of similar molecular weight, structural analogs or isotopomers were incorporated into each analysis. We report on the current practical limitations of quantitative MALDI-TOFMS and highlight some of the potential benefits of this technique as a quantitative tool. (J Am Soc

Research paper thumbnail of Alpha-Linolenic Acid: An Omega-3 Fatty Acid with Neuroprotective Properties-Ready for Use in the Stroke Clinic?

BioMed research international, 2015

Alpha-linolenic acid (ALA) is plant-based essential omega-3 polyunsaturated fatty acids that must... more Alpha-linolenic acid (ALA) is plant-based essential omega-3 polyunsaturated fatty acids that must be obtained through the diet. This could explain in part why the severe deficiency in omega-3 intake pointed by numerous epidemiologic studies may increase the brain's vulnerability representing an important risk factor in the development and/or deterioration of certain cardio- and neuropathologies. The roles of ALA in neurological disorders remain unclear, especially in stroke that is a leading cause of death. We and others have identified ALA as a potential nutraceutical to protect the brain from stroke, characterized by its pleiotropic effects in neuroprotection, vasodilation of brain arteries, and neuroplasticity. This review highlights how chronic administration of ALA protects against rodent models of hypoxic-ischemic injury and exerts an anti-depressant-like activity, effects that likely involve multiple mechanisms in brain, and may be applied in stroke prevention. One major ...

Research paper thumbnail of Mature adipocyte proteome reveals differentially altered protein abundances between lean, overweight and morbidly obese human subjects

Molecular and Cellular Endocrinology, 2015

Overweight (OW) and obese individuals are considered to be graded parts of the scale having incre... more Overweight (OW) and obese individuals are considered to be graded parts of the scale having increasing weight as a common feature. They may not, however, be part of the same continuum and may differ metabolically. In this study we applied an untargeted proteomic approach to compare protein abundances in mature adipocytes derived from the subcutaneous adipose tissue of overweight and morbidly obese female subjects to those of lean age matched controls. Mature adipocytes were isolated from liposuction samples of abdominal subcutaneous adipose tissue collected from both lean (L; n = 7, 23.3 ± 0.4 kg/m(2); mean BMI ± SD), overweight (OW; n = 8, 27.9 ± 0.6 kg/m(2); mean BMI ± SD) and morbidly obese (MOB; n = 7, 44.8 ± 3.8 kg/m(2); mean BMI ± SD) individuals. Total protein extracts were then compared by two-dimensional difference in gel electrophoresis (2D DIGE). One hundred and ten differentially expressed protein spots (i.e., fitting the statistical criteria ANOVA test, p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05; fold-change ≥1.5) were detected, and of these, 89 were identified by MALDI-TOF mass spectrometry. Of these, 66 protein spots were common to both groups whereas 23 were unique to the MOB group. Significant differences were evident in the abundances of key proteins involved in glucose and lipid metabolism, energy regulation, cytoskeletal structure and redox control signaling pathways. Differences in the abundance of some chaperones were also evident. The differentially abundant proteins were investigated using Ingenuity Pathway Analysis (IPA) to establish their associations with known biological functions. The network identified in the OW group with the highest score relates to-: cell-to-cell signaling and interaction; in contrast, in the MOB group the major interacting pathways are associated with lipid metabolism, small molecule biochemistry and cancer. The differences in abundance of the differentially regulated proteins were validated by immunoblotting. These findings provide insights into metabolic differences in OW and MOB individuals.

Research paper thumbnail of Purification and Structural Characterization of Siderophore (Corynebactin) from Corynebacterium diphtheriae

PLOS One, 2012

During infection, Corynebacterium diphtheriae must compete with host iron-sequestering mechanisms... more During infection, Corynebacterium diphtheriae must compete with host iron-sequestering mechanisms for iron. C. diphtheriae can acquire iron by a siderophore-dependent iron-uptake pathway, by uptake and degradation of heme, or both. Previous studies showed that production of siderophore (corynebactin) by C. diphtheriae is repressed under high-iron growth conditions by the iron-activated diphtheria toxin repressor (DtxR) and that partially purified corynebactin fails

Research paper thumbnail of Utility of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the analysis of low molecular weight compounds

Rapid Communications in Mass Spectrometry, 1995

ABSTRACT

Research paper thumbnail of Circulating Cytokines and Growth Factors in Pediatric Pulmonary Hypertension

Mediators of Inflammation, 2012

Background. Management of pediatric pulmonary hypertension (PH) remains challenging. We have asse... more Background. Management of pediatric pulmonary hypertension (PH) remains challenging. We have assessed a panel of circulating proteins in children with PH to investigate their value as predictive and/or prognostic biomarkers. From these determinations, we aim to develop a practical, noninvasive tool to aid in the management of pediatric PH. Methods. Twelve cytokines and growth factors putatively associated with lung or vascular disease were examined in plasma specimens from 70 children with PH using multiplex protein array technology. Associations between hemodynamics, adverse events, and protein markers were evaluated. Results. Epidermal growth factor (EGF) and IL-6 were associated with important hemodynamics. Of the twelve proteins, VEGF and IL-6 were significantly, univariately associated with the occurrence of an adverse event, with odds ratios (95% confidence intervals) of 0.56 (0.33-0.98) and 1.69 (1.03-2.77), respectively. When hemodynamic predictors were combined with protein markers, the ability to predict adverse outcomes within the following year significantly increased. Conclusions. Specific circulating proteins are associated with hemodynamic variables in pediatric PH. If confirmed in additional cohorts, measurement of these proteins could aid patient care and design of clinical trials by identifying patients at risk for adverse events. These findings also further support a role for inflammation in pediatric PH.

Research paper thumbnail of Phenolic compounds in the nearshore waters of Sydney, Australia

Marine and Freshwater Research, 1996

ABSTRACT

Research paper thumbnail of Prediction of benefit from EGFR TKIs by proteomic analysis of pretreatment serum

Journal of Thoracic Oncology, 2007

Research paper thumbnail of Vaccine Impedes the Development of Reflux-induced Esophageal Cancer in a Surgical Rat Model: Efficacy of the Vaccine in a Post-Barrett’s Esophagus Setting

Digestive Diseases and Sciences, 2008

Background & Aims Wedevelopedagranulocyte-macrophage-colony-stimulatingfactor(GM-CSF)tumorvaccine... more Background & Aims Wedevelopedagranulocyte-macrophage-colony-stimulatingfactor(GM-CSF)tumorvaccineforesophageal cancer.Weevaluatedtheeffectivenessofthevaccineasapreventionoptioninasurgicalrefluxratmodelofesophagealcancer. Methods A surgical model involving a jejuno-esophagostomy was used to create Barrett's esophagus and esophageal cancer in rats. No carcinogen exposure was utilized. Cell lines derived from these tumors were stably passaged in vitro. GM-CSF-secreting tumor cells were generated using stable transfection. All rats underwent a total gastrectomy, followed by a jejuno-esophagostomy. The surgery promoted the reflux of duodenal contents into the esophagus. All animals were administered either a GM-CSF secreting whole cell vaccine or a phosphate-buffered saline (PBS) placebo injection 4, 6, 14, and 16 weeks post-surgery. Results While 15 of 16 animals in the non-vaccinated placebo group developed esophageal cancer, 94% (15 of 16), animals in the vaccine group had an incidence of cancer of 25% (4 of 16) (p<0.05). Barrett's esophagus was seen in 100% (16 of 16) of the controls and 83% (13 of 16) of the vaccinated animals. Conclusions A GM-CSF-secreting whole cell tumor vaccine impeded esophageal tumor growth, but not the development of Barrett's esophagus, in a clinically relevant surgical reflux model.

Research paper thumbnail of Serum E-cadherin does not associate with response to gefitinib in patients with non-small cell lung cancer (NSCLC)

Journal of Thoracic Oncology, 2007

Research paper thumbnail of Assessment of post-mortem-induced changes to the mouse brain proteome

Journal of Neurochemistry, 2008

This study was designed to assess the influence of high-energy head-focused microwave irradiation... more This study was designed to assess the influence of high-energy head-focused microwave irradiation and the post-mortem interval on measurements of the mouse brain proteome. Difference gel electrophoresis was used to compare mouse brain protein levels in animals killed by decapitation, where the tissue was held at 25 degrees C for selected time intervals post-mortem, and by high-energy head-focused microwave irradiation followed by immediate resection. Microwave-mediated killing was used because it comprehensively snap-inactivates enzymes while largely retaining brain cytoarchitecture. Of the 912 protein spots common to at least eight of 10 gels analyzed, 35 (3.8%) showed significant differences in levels (t-test; p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05) depending on whether animals were killed by microwave irradiation or decapitation. When animals were killed by decapitation, 43 protein spots (4.7%) showed changes in levels over the post-mortem interval (anova; p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05). The vast majority of the near 1000 proteins evident on a 2D gel were stable for up to 4 h. These data have important implications for studies of proteins in the brain, whether based on analysis of tissue derived from animal models or from humans.

Research paper thumbnail of Nicotine and Cotinine Adducts of a Melanin Intermediate Demonstrated by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

Chemical Research in Toxicology, 2001

Pigmentation is a major factor in the incorporation of many drugs into hair. In an attempt to elu... more Pigmentation is a major factor in the incorporation of many drugs into hair. In an attempt to elucidate potential mechanisms of drug-melanin interaction, melanin was synthesized in vitro in the presence of nicotine, which we have shown to have a substantial interaction with melanin, and cotinine, a primary nicotine metabolite. L-DOPA, a precursor of eumelanin, was oxidized and oligomerized with tyrosinase. Nicotine, cotinine, and/or their deuterated analogues were added to the oligomerization reaction mixture in a 10:1 L-DOPA:drug ratio. A black precipitate formed within 60 min. Aliquots were removed from the incubation mixture at 60, 120, and 360 min. MALDI-TOF MS determinations were carried out on each sample to provide a mean and standard error for the masses of interest. Internal calibration allowed accurate mass measurement of the products. A careful comparison of the spectra of samples prepared both with and without drug indicated the presence of masses corresponding to the protonated drug, melanin oligomers, and nicotine or continine adducts of the monomeric melanin intermediate dopaquinone (DOPAQ). Additional support for the presence of drug-melanin adducts was provided by employing deuterated analogues of nicotine and L-DOPA in the reaction and observing that the masses shifted accordingly. Structures of the adducts were further confirmed by select ion gating and postsource decay analysis.

Research paper thumbnail of Relation between normal rectal methylation, smoking status, and the presence or absence of colorectal adenomas

Cancer, 2010

BACKGROUND-Colorectal cancer (CRC) is 1 of the leading causes of death in the Western world. CRC ... more BACKGROUND-Colorectal cancer (CRC) is 1 of the leading causes of death in the Western world. CRC develops from premalignant lesions, chiefly colorectal adenomas. Currently, the most accurate and recommended screening method for finding colorectal adenomas is colonoscopy performed on all individuals aged >50 years. However, the costs and risks associated with this procedure are relatively high. The objectives of the current study were to correlate epigenetic alterations that occur in normal rectal mucosa, smoking status, and age with the presence or absence of concomitant colorectal adenomas and to assess the potential clinical value of methylation in normal rectal biopsies as a screening assay for the presence of polyps and, hence, the need for a full colonoscopy.

Research paper thumbnail of Analysis of docetaxel pharmacokinetics in humans with the inclusion of later sampling time-points afforded by the use of a sensitive tandem LCMS assay

Cancer Chemotherapy and Pharmacology, 2003

Docetaxel is a semisynthetic taxane derived from the needles of the European yew ( Taxus baccata)... more Docetaxel is a semisynthetic taxane derived from the needles of the European yew ( Taxus baccata) and it is an important chemotherapeutic agent in the treatment of recurrent ovarian, breast and non-small-cell lung cancers. Traditional dosing regimens with docetaxel involve doses of 60-100 mg/m(2) by infusion every 3 weeks. Now weekly low-dose (30-36 mg/m(2)) regimens are being evaluated in phase I trials. Such low-dose studies require a more sensitive, specific and rapid assay of docetaxel in biological fluids for the determination of pharmacokinetic parameters. Because docetaxel is primarily metabolized by CYP3A4 and is highly protein-bound in the plasma, there is potential for drug-drug interactions and high interpatient variability in pharmacokinetics. Therefore, pharmacokinetic studies are an important component to understanding the therapeutic variability of docetaxel-containing chemotherapeutic regimens. To this end, we developed an analytical assay for docetaxel based upon tandem LCMS and paclitaxel as an internal standard. The sensitivity of the new assay allowed us to monitor plasma levels of docetaxel out to 48 h after the end of the infusion in patients enrolled in a phase I trial of exisulind (orally, twice daily) receiving weekly docetaxel doses of 30 or 36 mg/m(2) where plasma docetaxel levels are below the lower limit of quantitation for traditional HPLC/UV-based assays at later time-points. The inclusion of the 48-h time-point had significant effects on the calculated pharmacokinetic parameters when using either a three-compartment or non-compartmental analysis. The terminal half-life was significantly increased when the 48-h time-point was included in the pharmacokinetic analysis, and the use of model parameters derived with the inclusion of the 48-h time-point were able to more accurately predict plasma levels at later times. The results reflect the importance of accurate and sensitive analytical methods for the determination of pharmacokinetic parameters and the effect of this later time-point on docetaxel pharmacokinetic modeling. Further, with the increased use of weekly docetaxel in combination with other agents, the inclusion of these later sampling time-points and sensitive methods for drug level determinations are important components in the description of pharmacokinetic drug interactions.