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Biomedical subjects

Mark D Johnson

Publications and source records attributed to Mark D Johnson.

At least 19 recordsLinked to original sources
PubMed2007

Aberrant splicing of cyclin-dependent kinase-associated protein phosphatase KAP increases proliferation and migration in glioblastoma.

The cyclin-dependent kinase (Cdk)-associated protein phosphatase KAP is a dual-specificity phosphatase of which the only known function is to dephosphorylate Cdk2 and inhibit cell cycle progression. Paradoxically, we find increased KAP mRNA expression in malignant astrocytomas, which correlates with increasing histologic grade and decreased patient survival. We have resolved this apparent paradox with the discovery of aberrant KAP splicing in malignant astrocytomas that leads to increased expression of KAP-related transcripts but decreased KAP protein expression. In addition, the aberrant splicing generates a dominant negative KAP variant that increases proliferation. We provide the first evidence that KAP not only regulates proliferation but also inhibits migration by decreasing cdc2 mRNA and protein expression. The effect of KAP on cdc2 expression requires its phosphatase activity but does not involve direct dephosphorylation of cdc2. Thus, KAP regulates both cdc2-dependent migration and Cdk2-dependent proliferation, and its loss due to aberrant splicing increases malignancy in human gliomas.

Alternative Splicing↗
PubMed2006

A genome-wide screen reveals functional gene clusters in the cancer genome and identifies EphA2 as a mitogen in glioblastoma.

A novel genome-wide screen that combines patient outcome analysis with array comparative genomic hybridization and mRNA expression profiling was developed to identify genes with copy number alterations, aberrant mRNA expression, and relevance to survival in glioblastoma. The method led to the discovery of physical gene clusters within the cancer genome with boundaries defined by physical proximity, correlated mRNA expression patterns, and survival relatedness. These boundaries delineate a novel genomic interval called the functional common region (FCR). Many FCRs contained genes of high biological relevance to cancer and were used to pinpoint functionally significant DNA alterations that were too small or infrequent to be reliably identified using standard algorithms. One such FCR contained the EphA2 receptor tyrosine kinase. Validation experiments showed that EphA2 mRNA overexpression correlated inversely with patient survival in a panel of 21 glioblastomas, and ligand-mediated EphA2 receptor activation increased glioblastoma proliferation and tumor growth via a mitogen-activated protein kinase-dependent pathway. This novel genome-wide approach greatly expanded the list of target genes in glioblastoma and represents a powerful new strategy to identify the upstream determinants of tumor phenotype in a range of human cancers.

Brain Neoplasms↗
PubMed2005

Drug-induced oxidative stress in rat liver from a toxicogenomics perspective.

Macrophage activators (MA), peroxisome proliferators (PP), and oxidative stressors/reactive metabolites (OS/RM) all produce oxidative stress and hepatotoxicity in rats. However, these three classes of hepatotoxicants give three distinct gene transcriptional profiles on cDNA microarrays, an indication that rat hepatocytes respond/adapt quite differently to these three classes of oxidative stressors. The differential gene responses largely reflect differential activation of transcription factors: MA activate Stat-3 and NFkB, PP activate PPARa, and OS/RM activate Nrf2. We have used gene signature profiles for each of these three classes of hepatotoxicants to categorize over 100 paradigm (and 50+ in-house proprietary) compounds as to their oxidative stress potential in rat liver. In addition to a role for microarrays in predictive toxicology, analyses of small subsets of these signature profiles, genes within a specific pathway, or even single genes often provide important insights into possible mechanisms involved in the toxicities of these compounds.

Animals↗
PubMed2005

Comparative analysis of algorithms for identifying amplifications and deletions in array CGH data.

MOTIVATION: Array Comparative Genomic Hybridization (CGH) can reveal chromosomal aberrations in the genomic DNA. These amplifications and deletions at the DNA level are important in the pathogenesis of cancer and other diseases. While a large number of approaches have been proposed for analyzing the large array CGH datasets, the relative merits of these methods in practice are not clear. RESULTS: We compare 11 different algorithms for analyzing array CGH data. These include both segment detection methods and smoothing methods, based on diverse techniques such as mixture models, Hidden Markov Models, maximum likelihood, regression, wavelets and genetic algorithms. We compute the Receiver Operating Characteristic (ROC) curves using simulated data to quantify sensitivity and specificity for various levels of signal-to-noise ratio and different sizes of abnormalities. We also characterize their performance on chromosomal regions of interest in a real dataset obtained from patients with Glioblastoma Multiforme. While comparisons of this type are difficult due to possibly sub-optimal choice of parameters in the methods, they nevertheless reveal general characteristics that are helpful to the biological investigator.

Algorithms↗
PubMed2005

Chemical reactions between arsenic and zero-valent iron in water.

Batch experiments and X-ray photoelectron spectroscopic (XPS) analyses were performed to study the reactions between arsenate [As(V)], arsenite [As(III)] and zero-valent iron [Fe(0)]. The As(III) removal rate was higher than that for As(V) when iron filings (80-120 mesh) were mixed with arsenic solutions purged with nitrogen gas in the pH range of 4-7. XPS spectra of the reacted iron coupons showed the reduction of As(III) to As(0). Soluble As(III) was formed when As(V) reacted with Fe(0) under anoxic conditions. However, no As(0) was detected on the iron coupons after 5 days of reaction in the As(V)-Fe(0) system. The removal of the arsenic species by Fe(0) was attributed to electrochemical reduction of As(III) to sparsely soluble As(0) and adsorption of As(III) and As(V) to iron hydroxides formed on the Fe(0) surface under anoxic conditions. When the solutions were open to atmospheric air, the removal rates of As(V) and As(III) were much higher than under the anoxic conditions, and As(V) removal was faster than As(III). The rapid removal of As(III) and As(V) was caused by adsorption on ferric hydroxides formed readily through oxidation of Fe(0) by dissolved oxygen.

Adsorption↗
PubMed2005

The proteomics of neurodegeneration.

The continuing improvement and refinement of proteomic and bioinformatic tools has made it possible to obtain increasing amounts of structural and functional information about proteins on a global scale. The emerging field of neuroproteomics promises to provide powerful strategies for further characterizing neuronal dysfunction and cell loss associated with neurodegenerative diseases. Neuroproteomic studies have thus far revealed relatively comprehensive quantitative changes and post-translational modifications (mostly oxidative damage) of high abundance proteins, confirming deficits in energy production, protein degradation, antioxidant protein function, and cytoskeletal regulation associated with neurodegenerative diseases such as Alzheimer and Parkinson disease. The identification of changes in low-abundance proteins and characterization of their functions based on protein-protein interactions still await further development of proteomic methodologies and more dedicated application of these technologies by neuroscientists. Once accomplished, however, the resulting information will certainly provide a truly comprehensive view of neurodegeneration-associated changes in protein expression, facilitating the identification of novel biomarkers for the early detection of neurodegenerative diseases and new targets for therapeutic intervention.

Alzheimer Disease↗
PubMed2004

A gene expression signature for oxidant stress/reactive metabolites in rat liver.

Formation of free radicals and other reactive molecules is responsible for the adverse effects produced by a number of hepatotoxic compounds. cDNA microarray technology was used to compare transcriptional profiles elicited by training and testing sets of 15 oxidant stressors/reactive metabolite treatments to those produced by approximately 85 other paradigm compounds (mostly hepatotoxicants) to determine a shared signature profile for oxidant stress-associated hepatotoxicity. Initially, 100 genes were chosen that responded significantly different to oxidant stressors/reactive metabolites (OS/RM) compared to other samples in the database, then a 25-gene subset was selected by multivariate analysis. Many of the selected genes (e.g., aflatoxin aldehyde reductase, diaphorase, epoxide hydrolase, heme oxgenase and several glutathione transferases) are well-characterized oxidant stress/Nrf-2-responsive genes. Less than 10 other compounds co-cluster with our training and testing set compounds and these are known to generate OS/RMs as part of their mechanisms of toxicity. Using OS/RM signature gene sets, compounds previously associated with macrophage activation formed a distinct cluster separate from OS/RM and other compounds. A 69-gene set was chosen to maximally separate compounds in control, macrophage activator, peroxisome proliferator and OS/RM classes. The ease with which these 'oxidative stressor' classes can be separated indicates a role for microarray technology in early prediction and classification of hepatotoxicants. The ability to rapidly screen the oxidant stress potential of compounds may aid in avoidance of some idiosyncratic drug reactions as well as overtly toxic compounds.

Animals↗
PubMed2004

Inverse gene expression patterns for macrophage activating hepatotoxicants and peroxisome proliferators in rat liver.

Macrophage activation contributes to adverse effects produced by a number of hepatotoxic compounds. Transcriptional profiles elicited by two macrophage activators, LPS and zymosan A, were compared to those produced by 100 paradigm compounds (mostly hepatotoxicants) using cDNA microarrays. Several hepatotoxicants previously reported to activate liver macrophages produced transcriptional responses similar to LPS and zymosan, and these were used to construct a gene signature profile for macrophage activators in the liver. Measurement of cytokine mRNAs in the same liver samples by RT-PCR independently confirmed that these compounds are associated with macrophage activation. In addition to expected effects on acute phase proteins and metabolic pathways that are regulated by LPS and inflammation, a strong induction was observed for many endoplasmic reticulum-associated stress/chaperone proteins. Additionally, many genes in our macrophage activator signature profile were well-characterized PPARalpha-induced genes which were repressed by macrophage activators. A shared gene signature profile for peroxisome proliferators was determined using a training set of clofibrate, WY 14643, diethylhexylphthalate, diisononylphthalate, perfluorodecanoic acid, perfluoroheptanoic acid, and perfluorooctanoic acid. The signature profile included macrophage activator-induced genes that were repressed by peroxisome proliferators. NSAIDs comprised an interesting pharmacological class in that some compounds, notably diflunisal, co-clustered with peroxisome proliferators whereas several others co-clustered with macrophage activators, possibly due to endotoxin exposure secondary to their adverse effects on the gastrointestinal system. While much of these data confirmed findings from the literature, the transcriptional patterns detected using this toxicogenomics approach showed relationships between genes and biological pathways requiring complex analysis to be discerned.

Animals↗
PubMed2004

Proteome analysis of DNA damage-induced neuronal death using high throughput mass spectrometry.

Isotope-coded affinity tag reagents and high throughput mass spectrometry were used to quantitate changes in the expression of 150 proteins in mouse wild-type (p53(+/+)) cortical neurons undergoing DNA damage-induced death. Immunological techniques confirmed several of the changes in protein expression, but microarray analysis indicated that many of these changes were not accompanied by altered mRNA expression. Proteome analysis revealed perturbations in mitochondrial function, free radical production, and neuritogenesis that were not observed in p53-deficient neurons. Changes in Tau, cofilin, and other proteins recapitulated abnormalities observed in neurodegenerative states in vivo. Additionally, DNA damage caused a p53-dependent decrease in expression of members of the protein kinase A (PKA) signaling pathway. PKA inhibition promoted death in the absence of DNA damage, revealing a novel mechanism by which endogenous down-regulation of PKA signaling may contribute to p53-dependent neuronal death. These data demonstrate the power of high throughput mass spectrometry for quantitative analysis of the neuronal proteome.

Animals↗
PubMed2004

Determination of cytochrome P450 1A2 and cytochrome P450 3A4 induction in cryopreserved human hepatocytes.

Freshly prepared human hepatocytes are considered as the 'gold standard' for in vitro testing of drug candidates. However, several disadvantages are associated with the use of this model system. The availability of hepatocytes is often low and consequently the planning of the experiments rendered difficult. In addition, the quality of the available cells is in some cases poor. As an alternative, cryopreserved human hepatocytes were validated as a model to study cytochrome P450 1A2 (CYP1A2) and cytochrome P450 3A4 (CYP3A4) induction. In a single blinded experiment, hepatocytes from three separate lots were incubated with three concentrations of different compounds, and compared to non-treated cells and cells incubated with omeprazole or rifampicin. CYP1A2 and CYP3A4 induction was determined by measuring 7-ethoxyresorufin-O-deethylation activity and 6beta-hydroxytestosterone formation, respectively. CYP1A2 and CYP3A4 mRNA and protein expression were analyzed by TaqMan QRT-PCR and immunodetection. Cells responded well to the prototypical inducers with a mean 38.8- and 6.2-fold induction of CYP1A2 and CYP3A4 activity, respectively. Similar as with fresh human hepatocytes, high batch-to-batch variation of CYP1A2 and CYP3A4 induction was observed. Except for 1 and 10 microM rosiglitazone, the glitazones did not significantly affect CYP1A2. A similar result was observed for CYP3A4 activity although CYP3A4 mRNA and protein expression were dose-dependently upregulated. In conclusion, cryopreserved human hepatocytes may be a good alternative to fresh hepatocytes to study CYP1A and 3A induction.

Cells, Cultured↗
PubMed2004

Peripheral stimulation for treatment of trigeminal postherpetic neuralgia and trigeminal posttraumatic neuropathic pain: a pilot study.

OBJECTIVE: Trigeminal neuropathic pain (TNP) after facial trauma or herpes zoster infection is often refractory to treatment. Peripheral nerve stimulation has been used to treat occipital neuralgia; however, efficacy in controlling facial TNP or postherpetic neuralgia is unknown. A retrospective case series of patients who underwent subcutaneous placement of stimulating electrodes for treatment of V(1) or V(2) TNP secondary to herpetic infection or facial trauma is presented. METHODS: Ten patients received implanted subcutaneous pulse generators and quadripolar electrodes for peripheral stimulation of the trigeminal nerve supraorbital or infraorbital branches. Long-term treatment results were determined by retrospective review of medical records (1998-2003) and by independent observers interviewing patients using a standard questionnaire. Surgical complication rate, preoperative symptom duration, degree of pain relief, preoperative and postoperative work status, postoperative changes in medication usage, and overall degree of therapy satisfaction were assessed. Mean follow-up was 26.6 +/- 4.7 months. RESULTS: Peripheral nerve stimulation provided at least 50% pain relief in 70% of patients with TNP or postherpetic neuralgia. Medication use declined in 70% of patients, and 80% indicated that they were mostly or completely satisfied with treatment overall. There were no treatment failures (<50% pain relief and a lack of decrease in medication use) in the posttraumatic group, and two failures (50%) occurred in the postherpetic group. The complication rate requiring reoperation was 30%. CONCLUSION: Peripheral nerve stimulation of the supraorbital or infraorbital branches of the trigeminal nerve is an effective method for relief of TNP after facial trauma or herpetic infection. A prospective trial using this novel approach to treat these disorders is thus warranted.

Adult↗
PubMed2003

Proteomics in the postgenomic age.

Technical developments in the field of proteomics are poised to generate advances in our understanding of protein structure, function, and organization in complex signaling and regulatory networks. Improvements in mass spectrometry instrumentation, the implementation of protein arrays, and the development of robust informatics software are providing sensitive, high-throughput technologies for large-scale identification and quantitation of protein expression, protein modifications, subcellular localization, protein function, and protein-protein interactions. These advances have significant implications for understanding how cellular proteomes are regulated in health and disease.

Animals↗
PubMed2003

p53-dependent cell death signaling in neurons.

The p53 tumor suppressor gene is a sequence-specific transcription factor that activates the expression of genes engaged in promoting growth arrest or cell death in response to multiple forms of cellular stress. p53 expression is elevated in damaged neurons in acute models of injury such as ischemia and epilepsy and in brain tissue samples derived from animal models and patients with chronic neurodegenerative diseases. p53 deficiency or p53 inhibition protects neurons from a wide variety of acute toxic insults. Signal transduction pathways associated with p53-induced neuronal cell death are being characterized, suggesting that intervention may prove effective in maintaining neuronal viability and restoring function following neural injury and disease.

Animals↗
PubMed2003

Acute congestive heart failure after laparoscopic cholecystectomy: a case report.

Compared with open procedures, laparoscopic surgery is safe with a low incidence of complications. In rare circumstances, however, intraoperative complications such as acute pulmonary edema have been reported. The patient described herein is a 59-year-old woman with obesity, gastroesophageal reflux disease, and chronic obstructive pulmonary disease who developed acute congestive heart failure (CHF) and cardiomegaly immediately following laparoscopic cholecystectomy. She required emergent reintubation, diuresis, and admission to the intensive care unit for postoperative mechanical ventilation. Potential causes of pulmonary edema associated with laparoscopic surgery (extreme Trendelenburg position, venous carbon dioxide embolism, absorption of crystalloid irrigation fluid, cardiopulmonary disease, adverse drug reactions, negative pressure [postobstructive pulmonary edema]) were considered. A process of exclusion revealed that the hemodynamic changes induced by insufflation with an intra-abdominal pressure of 20 mm Hg were the most likely causes of the CHF. Suggestions to prevent occurrence of CHF are tight control of hemodynamics with use of invasive monitoring in high-risk patients and gentle, slow insufflation of the abdomen to an intra-abdominal pressure of 15 mm Hg or less. Intraoperative and/or postoperative CHF should be treated with diuretics, intravenous nitroglycerin, arterial vasodilators, and/or inotropic agents as needed.

Acute Disease↗
PubMed2002

Peg3/Pw1 is a mediator between p53 and Bax in DNA damage-induced neuronal death.

Neuronal cell death after DNA damage requires p53 and Bax, but the mechanism by which p53 activation leads to Bax translocation and cell death in neurons is not known. We report here that Peg3/Pw1 is up-regulated after DNA damage in cortical neurons in a p53-dependent manner. Overexpression of Peg3/Pw1 leads to decreased neuronal viability. The deleterious effect of Peg3/Pw1 on neuronal survival is abrogated by deletion of either p53 or Bax, indicating an essential role for both in Peg3/Pw1-mediated neuronal death. Moreover, overexpression of a Peg3/Pw1 dominant negative protein inhibits Bax translocation and neuronal cell death after DNA damage. These findings implicate Peg3/Pw1 as a mediator between p53 and Bax in a neuronal cell death pathway activated by DNA damage.

Adenoviridae↗
PubMed2002

Proteome analysis of camptothecin-treated cortical neurons using isotope-coded affinity tags.

Isotope-coded affinity tags (ICATs) were employed to identify and quantitate changes in protein expression between control and camptothecin-treated mouse cortical neurons. Proteins extracted from control cortical neurons and those treated with camptothecin were labeled with the light and heavy isotopic versions of the ICAT reagents, respectively. ICAT-labeled samples were combined, proteolytically digested, and the derivatized peptides isolated using immobilized avidin chromatography. The peptides thus isolated were analyzed by reversed-phase liquid chromatography coupled directly to either a conventional ion-trap mass spectrometer (IT-MS) or a Fourier transform ion cyclotron resonance mass spectrometer (FTICR). While a majority of the peptide identifications were accomplished using IT-MS, FTICR was used to quantitate the relative abundances of the ICAT-labeled peptides taking advantage of its high resolution, sensitivity, and duty cycle. By using this combination of MS technologies we have thus far identified and quantified the expression of greater than 125 proteins from control and camptothecin-treated mouse cortical neurons. While proteins from most functional classes of proteins were identified, a particularly large percentage of the enzymes involved in glycolysis and the tricarboxylic acid cycle were observed.

Animals↗
PubMed2002

Neuronal survival and cell death signaling pathways.

Neuronal viability is maintained through a complex interacting network of signaling pathways that can be perturbed in response to a multitude of cellular stresses. A shift in the balance of signaling pathways after stress or in response to pathology can have drastic consequences for the function or the fate of a neuron. There is significant evidence that acutely injured and degenerating neurons may die by an active mechanism of cell death. This process involves the activation of discrete signaling pathways that ultimately compromise mitochondrial structure, energy metabolism and nuclear integrity. In this review we examine recent evidence pertaining to the presence and activation of anti- and pro-cell death regulatory pathways in nervous system injury and degeneration.

Calpain↗