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M A Johnson

Publications and source records attributed to M A Johnson.

At least 55 records · Page 3Linked to original sources
PubMed2001

S-nitrosothiols signal the ventilatory response to hypoxia.

Increased ventilation in response to hypoxia has been appreciated for over a century, but the biochemistry underlying this response remains poorly understood. Here we define a pathway in which increased minute ventilation (&Vdot;E ) is signalled by deoxyhaemoglobin-derived S-nitrosothiols (SNOs). Specifically, we demonstrate that S-nitrosocysteinyl glycine (CGSNO) and S-nitroso-l-cysteine (l-CSNO)-but not S-nitroso-d-cysteine (d-CSNO)-reproduce the ventilatory effects of hypoxia at the level of the nucleus tractus solitarius (NTS). We show that plasma from deoxygenated, but not from oxygenated, blood produces the ventilatory effect of both SNOs and hypoxia. Further, this activity is mediated by S-nitrosoglutathione (GSNO), and GSNO activation by gamma-glutamyl transpeptidase (gamma-GT) is required. The normal response to hypoxia is impaired in a knockout mouse lacking gamma-GT. These observations suggest that S-nitrosothiol biochemistry is of central importance to the regulation of breathing.

Animals↗
PubMed2001

Reduced bone mineral density in HIV-positive individuals.

A total of 105 HIV-positive patients underwent dual-energy X-ray absorbtiometry (DEXA) scan to assess bone mineral density (BMD). The prevalence of reduced BMD was found to be 71% and was higher in patients who had ever been treated with protease inhibitors (PI). Our results suggest a possible association between PI and reduced BMD, and further complicate the debate regarding when to commence treatment of HIV and with what agents to start.

Absorptiometry, Photon↗
PubMed2001

Accelerated s-nitrosothiol breakdown by amyotrophic lateral sclerosis mutant copper,zinc-superoxide dismutase.

Mutations in copper,zinc-superoxide dismutase (SOD) have been implicated in familial amyotrophic lateral sclerosis (FALS). We have investigated the breakdown of S-nitrosothiols by wild-type (WT) SOD and two common FALS mutants, alanine-4 valine (A4V) SOD and glycine-37 arginine (G37R) SOD. In the presence of glutathione, A4V SOD and G37R SOD catalyzed S-nitrosoglutathione breakdown three times more efficiently than WT SOD. Indeed, A4V SOD catabolized GSNO more efficiently than WT SOD throughout the physiological range of GSH concentrations. Moreover, a variety of additional S-nitrosothiols were catabolized more readily by A4V SOD than by WT SOD. Initial rate data for fully reduced WT SOD and A4V SOD, and data using ascorbic acid as the reductant, suggest that FALS mutations in SOD may influence the efficiency of reduction of the copper center by glutathione. We have identified a potentially toxic gain of function of two common FALS mutations that may contribute to neurodegeneration in FALS.

Amyotrophic Lateral Sclerosis↗
PubMed2001

Mitochondrial enzyme-deficient hippocampal neurons and choroidal cells in AD.

OBJECTIVE: To determine whether hippocampal neurons and choroidal epithelial cells demonstrate a mitochondrial enzyme deficiency in AD more frequently than in normal aging. BACKGROUND: High levels of mutant mitochondrial DNA (mtDNA) cause a deficiency in cytochrome c oxidase (COX) (complex IV activity) because three of its 13 subunits are encoded for by mtDNA. In contrast, succinate dehydrogenase (SDH) (complex II activity) remains intact because all of its subunits are nuclear encoded. The histologic hallmark of cells containing high levels of mtDNA mutation in both primary mtDNA disorders and normal aging muscle is the presence of COX-deficient SDH-positive cells. METHODS: The authors applied a sequential histochemical method for COX and SDH to hippocampal sections in 17 AD and 17 age-matched control brains. This confers the advantages of both looking at individual cells in situ and measuring the actual mitochondrial complex activity rather than simply the complex quantity. RESULTS: COX-deficient SDH-positive hippocampal neurons and choroidal epithelial cells are more prevalent in patients with AD than in controls. In addition the COX-deficient SDH-positive choroidal cells are associated with an enlargement in size. CONCLUSION: This increase in number of COX-deficient SDH-positive hippocampal pyramidal neurons and choroid epithelial cells provides strong evidence that a substantial mitochondrial enzyme activity defect occurs in individual cells more frequently in AD than in normal aging and that mitochondria may play a significant role in the pathogenesis of AD.

Aged↗
PubMed2001

Vaccination of pigs with a recombinant porcine adenovirus expressing the gD gene from pseudorabies virus.

Five week old, commercially available large white pigs were vaccinated with either a single dose or two doses of a recombinant porcine adenovirus expressing the glycoprotein D gene from pseudorabies virus (PRV). Pigs were monitored for the development of serum neutralizing antibodies to PRV and challenged 3 weeks after final vaccination. Prior to challenge, pigs given 2 doses of the vaccine demonstrated boosted levels of antibody compared with those given a single dose, and all surviving pigs had increased neutralization titres over pre-challenge levels. Following challenge, pigs were monitored for clinical signs of disease, with blood and nasal swabs collected for virus isolation. All control animals became sick with elevated temperatures for 6 days post challenge, whereas; vaccinated animals displayed an increase in body temperature for only 2-3 days. Control pigs and those given a single dose all lost condition, but the group given 2 doses remained healthy. At postmortem, gross lesions of pneumonia only occurred in control animals and those given a single dose of vaccine. Histology carried out on the brains of all animals demonstrated a difference in severity of infection and frequency of immunohistochemical antigen detection between test animals, with control and single dose groups being most severely affected and pigs given 2 doses the least. Virus isolation studies demonstrated that no viraemia could be detected, but virus was found in nasal swabs from some animals in both groups of vaccinates following challenge.

Adenoviridae↗
PubMed2001

A prime-boost vaccination strategy using naked DNA followed by recombinant porcine adenovirus protects pigs from classical swine fever.

Weaned pigs (6-week-old) and 7-day-old pre-weaned piglets were vaccinated with naked plasmid DNA expressing the gp55/E2 gene from classical swine fever virus (CSFV). Both groups of pigs were then given a booster dose of recombinant porcine adenovirus expressing the gp55 gene (rPAV-gp55). Following challenge with CSFV, 100% of weaned pigs and 75% pre-weaned piglets were protected from disease. Weaned pigs given a single dose of rPAV-gp55 were also protected, but showed a slight increase in temperature immediately post-challenge. However, weaned animals given a DNA prime before rPAV-gp55 showed no fluctuation in body temperature following challenge and no pathology in spleen or lymph nodes upon post-mortem. In addition, no CSFV could be re-isolated from the rPAV vaccinated group and from only one pig in the prime-boost group following challenge, suggesting that both vaccination regimes have the potential to reduce or prevent virus shedding following experimental challenge.

Adenoviridae↗
PubMed2001

Paraplegin gene analysis in hereditary spastic paraparesis (HSP) pedigrees in northeast England.

OBJECTIVE: To identify the frequency and characterize the phenotype of paraplegin mutations in the hereditary spastic paraparesis (HSP) population in the northeast of England. BACKGROUND: HSP is a disorder that shows both clinical and genetic heterogeneity. To date, 13 loci have been associated with an HSP phenotype, with the causative gene having been identified in four of these. Two autosomal genes have been identified, paraplegin and spastin, and two X-linked genes have been identified, L1CAM (cell adhesion molecule) and proteolipid protein. METHODS: Thirty HSP pedigrees from the northeast of England were analyzed for mutation in each of the 17 exons of the paraplegin gene. RESULTS: A single family with a paraplegin mutation was identified in which the paraplegin mutation co-segregates with an HSP phenotype in an apparent dominant manner. The authors also describe frequent polymorphism in the paraplegin gene in both the HSP and control populations. CONCLUSION: Mutations in the paraplegin gene are not a common cause of HSP in the northeast of England. The phenotype of the paraplegin-related HSP family described had several striking features including amyotrophy, raised creatine kinase, sensorimotor peripheral neuropathy, and oxidative phosphorylation defect on muscle biopsy.

ATPases Associated with Diverse Cellular Activitie↗
PubMed2001

Reasons for modification and discontinuation of antiretrovirals: results from a single treatment centre.

OBJECTIVE: To describe the reasons for, and factors associated with, modification and discontinuation of highly active antiretroviral therapy (HAART) regimens at a single clinic. SUBJECTS: A total of 556 patients who started HAART at the Royal Free Hospital were included in analyses. Modification was defined as stopping or switching any antiretrovirals in the regimen, whereas discontinuation was defined as the simultaneous stopping of all antiretrovirals included in the initial regimen. Reasons were classified as immunological/virological failure (IVF) and toxicities and patient choice/poor compliance (TPC). RESULTS: The median CD4 count at starting HAART was 171 x 10(6) cells/l and viral load 5.07 log copies/ml. During a median follow-up of 14.2 months, 247 patients (44.4%) modified their HAART regimen, 72 due to IVF (29.1%) and 159 due to TPC (64.4%) and a total of 148 patients (26.6%) discontinued HAART. Older patients were less likely to modify HAART [relative hazard (RH), 0.73 per 10 years; P = 0.0008], as were previously treatment-naive patients (RH, 0.65; P = 0.0050), those in a clinical trial (RH, 0.64; P = 0.027) and those who started nelfinavir (RH, 0.57; P = 0.035). Patients who started with four or more drugs (RH, 2.21, P < 0.0001), who included ritonavir in the initial regimen (RH, 1.41; P = 0.035) or who had higher viral loads during follow-up (RH per log increase, 1.51; P < 0.0001) were more likely to modify HAART. CONCLUSIONS: There was a high rate of modification and discontinuation of HAART regimens in the first 12 months, particularly due to toxicities, patient choice or poor compliance.

Antiretroviral Therapy, Highly Active↗
PubMed2001

New selectable marker/auxotrophic host strain combinations for molecular genetic manipulation of Pichia pastoris.

We describe the isolation and characterization of three new biosynthetic genes-ARG4, ADE1, and URA3-from the methylotrophic yeast Pichia pastoris. The predicted products of the genes share significant sequence similarity to their Saccharomyces cerevisiae counterparts, namely argininosuccinate lyase, PR-aminoimidazolesuccinocarboxamide synthase, and orotidine-5'-phosphate decarboxylase, respectively. Along with the previously described HIS4 gene, each gene was incorporated as the yeast selectable marker into a set of shuttle vectors designed to express foreign genes in P. pastoris. In addition, we have constructed a series of host strains containing all possible combinations of ade1, arg4, his4, and ura3 auxotrophies to be used with these new vectors.

Amino Acid Sequence↗
PubMed2001

Mitochondrial DNA mutations in disease and ageing.

The chronological accumulation of mitochondrial DNA mutations has been proposed as a potential mechanism in the physiological processes of ageing and age-related disease. We discuss the evidence behind this theory and relate some of the ageing mitochondrial changes to mitochondrial DNA disorders. In particular, we describe the aggregation of cytochrome c oxidase-deficient cells in both skeletal muscle and the CNS in normal ageing as seen in the mitochondrial DNA disorders. These mitochondrial enzyme-deficient cells have been shown to occur in significant quantities in both muscle and CNS in patients with mitochondrial DNA disorders. In both ageing and mtDNA disorder muscle these cytochrome c-deficient fibres contain high levels of a single mutant strain of mitochondrial DNA. Whether these mutations are a primary or secondary event in the physiology of ageing remains to be determined.

Aging↗
PubMed2001

Replicative fitness in vivo of HIV-1 variants with multiple drug resistance-associated mutations.

The relative fitness of HIV-1 viral variants containing a broad range of drug resistance-associated mutations has been little studied in vivo. Understanding the relative fitness associated with viruses containing mutations may aid future therapeutic management. The aim of this study was to investigate the relative fitness of mutant viruses by assessing a cohort of patients who had developed resistance to many drugs and subsequently stopped all therapy. Eleven patients were assessed for drug resistance associated mutations in the protease (PR) and reverse transcriptase (RT) genes before and, at multiple time points, after stopping therapy. Relative fitness was calculated as a function of the rate of disappearance of mutant viruses when therapy was stopped. The least fit viruses were associated with the RT mutation M184I/V (11.6% less fit) and the PR mutations D30N (12.4% less fit) and M46I/L (21% less fit). Mutations at these codons were associated with significant reductions in fitness levels compared to wild-type viruses. Mutations at codons 10, 20, 36, and 63 in the PR gene remained fairly constant when therapy was stopped and may not significantly reduce viral fitness. The rapid re-population of wild-type viruses may allow the recycling of antiretroviral drugs prescribed previously.

Adult↗
PubMed2001

Pichia pastoris Pex14p, a phosphorylated peroxisomal membrane protein, is part of a PTS-receptor docking complex and interacts with many peroxins.

The peroxisomal protein import machinery plays a central role in the assembly of this organelle in all eukaryotes. Genes encoding components of this machinery, termed peroxins or Pex proteins, have been isolated and characterized in several yeast species and in mammals, including humans. Here we report on one of these components, Pex14p, from the methylotrophic yeast Pichia pastoris. Work in other organisms has shown that Pex14p is located on the cytoplasmic surface of the peroxisomal membrane and binds peroxisomal targeting signal (PTS) receptors carrying proteins bound for the peroxisomal matrix, results that have led to the hypothesis that Pex14p is a receptor-docking protein. P. pastoris Pex14p (PpPex14p) behaves like an integral membrane protein, with its C-terminus exposed on the cytosolic side of the peroxisomal membrane. PpPex14p complexes with many peroxins, including Pex3p (Snyder et al., 1999b), Pex5p, Pex7p, Pex13p, Pex17p, itself, and a previously unreported peroxin, Pex8p. A portion of Pex14p is phosphorylated, but both phosphorylated and unphosphorylated forms of Pex14p interact with several peroxins. The interactions between Pex14p and other peroxins provide clues regarding the function of Pex14p in peroxisomal protein import.

Amino Acid Sequence↗
PubMed2001

Oral and sub-cutaneous vaccination of commercial pigs with a recombinant porcine adenovirus expressing the classical swine fever virus gp55 gene.

A recombinant porcine adenovirus expressing the classical swine fever virus (CSFV) gp55/E2 gene was administered to commercially available pigs via oral or subcutaneous routes and their susceptibility to oral and subcutaneous challenge with CSFV was determined. 100% of animals vaccinated and challenged subcutaneously were protected. In the groups of pigs vaccinated either orally or subcutaneously and then challenged orally, 60% of animals were protected. Before challenge, neutralising antibodies to CSFV were detected in 60% of pigs vaccinated subcutaneously, but in none of those given the vaccine orally. CSFV antigen was found in the spleens of surviving pigs that had been vaccinated orally. In contrast, subcutaneous vaccination was shown to preclude the presence of CSFV in the spleen of animals that survived challenge.

Adenoviruses, Porcine↗
PubMed2001

Washout periods for brimonidine 0.2% and latanoprost 0.005%.

PURPOSE: To evaluate the intraocular pressure washout time after discontinuing brimonidine 0.2% twice daily and latanoprost 0.005% once every evening. METHODS: We discontinued brimonidine or latanoprost in a masked fashion from primary open-angle glaucoma or ocular hypertensive patients. The intraocular pressure was measured twice weekly until patients returned to untreated baseline. RESULTS: In 32 patients, the mean longest eye washout time for brimonidine (n = 15) was 3.3 +/- 3.0 weeks and for latanoprost (n = 17) was 4.4 +/- 3.2 weeks (P =.24). In all but one patient, brimonidine returned to baseline by 5 weeks and latanoprost returned by 8 weeks. CONCLUSION: After discontinuing latanoprost or brimonidine, a wide variation exists in washout times among individuals, with latanoprost demonstrating a trend to a longer washout period.

Adrenergic alpha-Agonists↗
PubMed2001

Strategy for mutation analysis in the autosomal recessive limb-girdle muscular dystrophies.

We describe a strategy for molecular diagnosis in the autosomal recessive limb-girdle muscular dystrophies, a highly heterogeneous group of inherited muscle-wasting diseases. Genetic mutation analysis is directed by immunoanalysis of muscle biopsies using antibodies against a panel of muscular dystrophy-associated proteins. Performing the molecular analysis in this way greatly increases the chance that mutations will be found in the first gene examined. The use of this strategy can significantly decrease the time involved in determining the genetic fault in a patient with a clinical diagnosis of recessive limb-girdle muscular dystrophy, as well as having a feedback effect, which is useful in helping clinicians to identify subtle clinical differences between the subtypes of the disease. The use of this approach has so far helped us to identify mutations in ten sarcoglycanopathy (limb-girdle muscular dystrophy 2C-2F) patients, and seven calpainopathy (limb-girdle muscular dystrophy 2A) patients.

Calpain↗
PubMed2001

Radiotherapeutic efficacy of (153)Sm-CMDTPA-Tyr(3)-octreotate in tumor-bearing rats.

A number of radiolabeled somatostatin analogs have been evaluated in animal tumor models for radiotherapeutic efficacy. The majority of the agents tested have used either high-energy beta-emitters, such as Y-90 or Re-188, or the Auger electron-emitting radionuclide, In-111. Because a medium-energy beta-emitter might have equivalent efficacy compared to high-energy emitters, and lower toxicity to non-target tissues, we have evaluated the therapeutic potential of the beta-emitting nuclide, Sm-153, chelated to the somatostatin analog, CMDTPA-Tyr(3)-octreotate. Using an in vitro binding assay, this octreotate derivative was shown to have high affinity for the somatostatin subtype-2 receptor (IC(50) = 2.7 nM). Biodistribution studies in CA20948 tumor-bearing Lewis rats demonstrate that the Sm-153 labeled compound has high uptake and retention in tumor tissue (1.7% injected dose/g tissue, 4 hrs post injection) and has rapid overall clearance properties from non-target tissue. Radiotherapy studies were carried out using (153)Sm-CMDTPA-Tyr(3)-octreotate and CA20948 tumor bearing Lewis rats at 7 days post implant. Dose regimens consisting of single and multiple i.v. injections of 5.0 mCi/rat (185 MBq) were employed over a time span of 7 days. Suppression of tumor growth rate was observed in all treated animals compared to untreated controls. Greater inhibition of tumor growth was observed in animals that received multiple doses. These studies indicate that medium-energy beta-emitting isotopes have considerable potential for the treatment of somatostatin receptor-positive tumors.

Animals↗
PubMed2001

A novel pantothenate kinase gene (PANK2) is defective in Hallervorden-Spatz syndrome.

Hallervorden-Spatz syndrome (HSS) is an autosomal recessive neurodegenerative disorder associated with iron accumulation in the brain. Clinical features include extrapyramidal dysfunction, onset in childhood, and a relentlessly progressive course. Histologic study reveals iron deposits in the basal ganglia. In this respect, HSS may serve as a model for complex neurodegenerative diseases, such as Parkinson disease, Alzheimer disease, Huntington disease and human immunodeficiency virus (HIV) encephalopathy, in which pathologic accumulation of iron in the brain is also observed. Thus, understanding the biochemical defect in HSS may provide key insights into the regulation of iron metabolism and its perturbation in this and other neurodegenerative diseases. Here we show that HSS is caused by a defect in a novel pantothenate kinase gene and propose a mechanism for oxidative stress in the pathophysiology of the disease.

Amino Acid Sequence↗