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Biomedical subjects

M A Johnson

Publications and source records attributed to M A Johnson.

At least 181 records · Page 10Linked to original sources
PubMed1997

Development of a point mutation assay for the detection of human cytomegalovirus UL97 mutations associated with ganciclovir resistance.

A point mutation assay was developed to detect the quantitative prevalence of mutations at codons 460 (M to I; M to V), 520 (H to Q), 594 (A to V) and 595 (L to F; L to S) within the UL97 gene of human cytomegalovirus which segregate with ganciclovir resistance. Synthetic mixtures of wild-type and mutant plasmids containing the UL97 gene were amplified by nested polymerase chain reaction and the 700 base pair amplicon subsequently subjected to the point mutation assay. In plasmid reconstruction experiments, there was a high correlation between experimentally derived percentage mutant with the theoretical values. The assay was then used to assess the changes in the genetic composition of the UL97 gene in three patients on prolonged ganciclovir therapy. All three patients developed genotypic resistance against ganciclovir involving mutation at codon L595S, L595F and double mutation at codons L595F and M460I. In one patient, alteration of therapy to foscarnet did not affect the composition of UL97 and virus remained genotypically resistant to ganciclovir. In contrast, in two patients whose therapy was altered to cidofovir (HPMPC), repopulation with cytomegalovirus strains carrying the wild-type (ganciclovir-sensitive) codon at positions 595 and 460 occurred. The potential use of this assay for the rapid detection of cytomegalovirus resistance in patients on long-term ganciclovir therapy is discussed.

Acquired Immunodeficiency Syndrome↗
PubMed1997

The relationship between beta-2-microglobulin, CD4 lymphocyte count, AIDS and death in HIV-positive individuals.

The relationship, in 539 individuals infected with the human immunodeficiency virus (HIV), between two prognostic markers, the CD4 count and beta-2-microglobulin (B2M), and the development of the acquired immunodeficiency syndrome (AIDS) and death was investigated. Cox proportional hazards models were used to determine the risk of AIDS or death. In a multivariate model which adjusted for demographic factors and treatment, the most recent measurements of B2M (relative hazard (RH) 1.37 per g/l higher) and CD4 count (RH 2.17 per log-unit lower) were both significantly associated with the development of AIDS. Similarly, in a multivariate model which additionally adjusted for the development of AIDS as a time dependent covariate, there was a strong relationship with risk of death for the most recent measurements of B2M (RH 1.34 per g/l higher), and CD4 lymphocyte count (RH 1.91 per log-unit lower). A difference in the level of B2M could be used among patients with similar CD4 counts as an indicator of increased risk of progression to AIDS or death. Using the most recent values of these markers provides a better estimate of the risk of AIDS or death, compared to the more common method of analysis, where baseline values of the markers are used.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Progression of HIV disease is associated with increased expression of Fc gammaRI and CR1 on alveolar macrophages.

The expression of receptors for complement and the Fc region of immunoglobulin by alveolar macrophages (AM) constitutes a valuable aid to effector function of these cells. However, during HIV infection such expression may also act to increase binding of immune complexes, thus facilitating viral infection of these cells. This study was designed to determine whether changes in the expression of these receptors occurs in situ during HIV infection. Lung macrophages were isolated by bronchoalveolar lavage in groups of HIV+ subjects segregated on the basis of peripheral CD4 count. A group of normal subjects was also investigated. Expression of CR1 and Fc gammaRI was quantified by measuring the optical density of reaction product following controlled immunoperoxidase staining with MoAbs CD35 and CD64. Both CR1 and Fc gammaRI were increased over normal in all HIV+ subjects. This increase was progressive with advancing disease as determined by correlation with declining peripheral CD4 count. Comparison of asymptomatic and symptomatic subjects with HIV infection showed no difference in CR1 expression but a rise in Fc gammaRI expression in the latter group. An overall inverse correlation was also found between peripheral CD4 count and Fc gammaRI expression, but not CR1 expression. These data demonstrate a significant increase in the expression of these receptors on AM from HIV+ subjects, and show that this increase may occur before any symptoms in these patients.

Adult↗
PubMed1997

Cytochrome P4501A (CYP1A) induction in rat and man by the benzodioxino derivative, fluparoxan.

1. Fluparoxan is an alpha 2-adrenoceptor antagonist that has a relatively planar, tricyclic structure and was considered a potential substrate and inducer of cytochrome P4501A (CYP1A) enzymes. 2. Structure-activity analysis indicated some potential for CYP1A interaction, although its greater log P and molecular depth, compared with many CYP1A inducers, suggested fluparoxan would be a weak ligand for the aryl hydrocarbon (Ah) receptor and only a weak inducer. 3. In vitro, fluparoxan showed little affinity for the CYP1A enzymes. The compound was not metabolized by human CYP1A1 or 1A2 heterologously expressed in yeast and its rate of metabolism in rat and human microsomes was unaffected by the addition of the 1A inhibitor alpha-naphthoflavone. Furthermore, Ki's for fluparoxan against EROD activity were > 4000-fold higher than those of alpha-naphthoflavone. 4. In vivo, however, fluparoxan did show some capacity for CYP1A induction. In rat, hepatic EROD activity increased approximately 40-fold with seven once-daily oral doses of fluparoxan (50 mg/kg, solution), and immunoblotting studies confirmed induction of CYP1A2, though not of 1A1. In man, administration of 11 twice-daily oral doses of fluparoxan (8 mg tablet) produced some reduction in plasma levels of orally administered phenacetin and in the ratio of phenacetin AUC/urinary paracetamol, consistent with increased O-deethylation.

Adolescent↗
PubMed1997

Promoting HIV prevention: a problem identification approach to interventions in post-HIV test counselling.

HIV antibody testing now occurs in a wide variety of clinical settings. Counselling at the time of HIV testing involves more than obtaining informed consent and there can be an active role for HIV prevention strategies. Whilst research has not clearly identified the effectiveness of either HIV testing or counselling for HIV prevention, HIV testing does allow for focused discussions with individuals about risk behaviour. Therefore all efforts to refine interventions to enhance HIV prevention must be encouraged. Interventions which encourage clients to examine beliefs about behaviours and relationships can bring about an increase in perceived choices, and thereby lead to changes in behaviour. A model is presented for conducting post-HIV test counselling for use by health professionals involved in HIV testing. Examples of questions and question styles are provided to illustrate this framework which addresses prevailing beliefs and encourages contributions from clients.

AIDS Serodiagnosis↗
PubMed1997

Intracellular mitochondrial triplasmy in a patient with two heteroplasmic base changes.

We report the clinical, biochemical, and genetic investigation of a patient with a severe mitochondrial encephalomyopathy. Genetic studies identified a novel, heteroplasmic tRNA mutation at nt 10010. This T-->C transition is located in the DHU loop of mitochondrial tRNA(Gly). In skeletal muscle, it was present at lower levels in cytochrome c oxidase (COX)-normal (87.2% +/- 11%) compared with COX-deficient fibers (97.3% +/- 2.6%); it was found in skin fibroblasts and blood cells, but at lower levels of heteroplasmy (15% +/- 6% and 17% +/- 10%, respectively). A second, heteroplasmic transition (A-->G), at nt 5656, showed a different distribution than the tRNA(Gly) mutation, with very low levels in skeletal muscle (< 3%) but higher levels in blood (22.7% +/- 3%) and skin fibroblasts (21% +/- 2%). These transitions were followed both in vivo, by repeat biopsy and blood sampling, and in vitro, by establishing primary cultures of myoblasts and skin fibroblasts. Repeat muscle biopsy showed a dramatic increase in COX-deficient fibers, but not of the tRNAGly mutation. Indeed, no significant change in heteroplasmy was measured for either substitution in muscle or blood. In vitro analysis gave very different results. The T10010C was not found in cultured myoblasts, even at early passage. In uncloned fibroblasts, the T10010C was stable (approximately 10%) for several passages but then gradually was lost. In contrast, the A5656G rose progressively from 27% to 91%. In cloned fibroblasts, different combinations of both base-pair changes and wild type could be identified, confirming the presence of clonal, intracellular triplasmy.

Adult↗
PubMed1997

Cytotoxic mechanisms in inflammatory myopathies. Co-expression of Fas and protective Bcl-2 in muscle fibres and inflammatory cells.

Expression of the Fas 'death receptor', Fas (CD95/APO-1) renders cells susceptible to programmed cell death ('apoptosis'), whereas Bcl-2 protects cells from apoptosis. Using fluorescence immunohistochemistry, we analysed Fas and Bcl-2 expression in muscle from five patients with polymyositis (PM), four patients with inclusion body myositis (IBM), three patients with dermatomyositis (DM), three patients with Duchenne muscular dystrophy (DMD) and three nonmyopathic controls. Fas (CD95) and Bcl-2 were not detected in control muscle, but expressed in muscle fibres and inflammatory cells in PM, IBM, DM and DMD. The proportion of Fas+ muscle fibres ranged from < 1 to 50%, and was higher in PM and IBM than in DM and DMD. On average, the Fas+ muscle fibres were smaller (median diameter, 10 microns; range, 7-32 microns) than the Fas- fibres (median, 36 microns; range, 10-60 microns). Less than 10% of the Fas+ muscle fibres co-expressed the regeneration marker CD56 (neural cell adhesion molecule N-CAM). In PM and IBM, the proportion of Fas+ muscle fibres was higher among fibres invaded or contacted by T cells than among fibres not contacted by T cells (P < 0.01). The proportion of Fas+ fibres co-expressing Bcl-2 was 76 +/- 16% in PM, 100% in IBM and 63 +/- 23% in DM. Fas and Bcl-2 expression was also noted in inflammatory cells in PM, IBM, DM and DMD. Using the terminal deoxytransferase-catalysed DNA nick end labelling technique for detection of nuclear DNA fragmentation, none of myonuclei, and < 0.1% of inflammatory cell nuclei, showed signs of apoptosis. Our results suggest that, although Fas expression confers susceptibility to Fas-mediated apoptosis, Fas-expressing muscle fibres and inflammatory cells are protected by the anti-apoptotic protein Bcl-2.

Adolescent↗
PubMed1997

Functionally relevant changes occur in HIV-infected individuals' alveolar macrophages prior to the onset of respiratory disease.

OBJECTIVE: We have compared the phenotypic and functional changes found in alveolar macrophages recovered from the lungs of 39 HIV-positive individuals with no respiratory disease with those from 33 HIV-positive individuals with pneumonitis and 31 healthy controls. METHODS: Bronchoalveolar lavage (BAL) cell cytospin preparations were stained using monoclonal antibody immunoperoxidase and double immunofluorescence techniques. Cytokine levels within supernatant BAL were determined using enzyme immunoassay. RESULTS: There were marked differences in alveolar macrophage phenotype between the three groups. In particular, the relative proportion of cells staining RFD1+RFD7- (inducer cells) was reduced in the HIV-positive individuals without respiratory disease. This was correlated with measures of declining systemic immunity. Patients with pneumonitis had the highest levels of measured cytokines [interleukin-1 beta, tumour necrosis factor-alpha and transforming growth factor (TGF)-beta 2], followed by the HIV-positive individuals without respiratory disease. In this latter population a negative correlation was found between active (non acid dissociated) TGF-beta 2 and blood CD4 cell count. CONCLUSIONS: The differences between the three groups suggest that alterations of potential relevance to the pulmonary immune response are occurring in alveolar macrophages prior to the onset of respiratory disease. This study confirms the importance of investigating asymptomatic HIV-positive individuals.

AIDS-Related Opportunistic Infections↗
PubMed1997

Cytomegalovirus (CMV) viraemia detected by polymerase chain reaction identifies a group of HIV-positive patients at high risk of CMV disease.

BACKGROUND: Cytomegalovirus (CMV) disease is a major cause of morbidity in patients with HIV infection. Despite treatment, CMV retinitis causes substantial visual loss, especially in patients with CD4 cell counts below 50 x 10(6)/l. Although routine ophthalmological screening of these patients has been recommended, no controlled trials have evaluated how frequently it should be performed. The aim of this study was to assess whether CMV polymerase chain reaction (PCR) results could direct ophthalmological screening to patients at high risk of CMV retinitis. METHODS: In a prospective study of HIV-positive patients with CD4 cell counts below 50 x 10(6)/l, CMV viraemia was detected by qualitative PCR of whole blood. Patients who were CMV PCR-viraemic were allocated to monthly virological and ophthalmological follow-up; patients who were PCR-negative received 3-monthly virological and ophthalmological follow-up. CMV viral load was determined in all CMV-positive samples using a quantitative competitive PCR. RESULTS: Nineteen out of 97 patients developed CMV disease over the first 12 months of the study. Sixteen (59%) out of 27 patients who were CMV-positive developed disease compared with three (4%) out of 70 of patients who were PCR-negative (P = 0.0001). A positive CMV PCR result was significantly associated with the development of disease (P = 0.0001), with a relative hazard of 20.15 [95% confidence interval (CI), 5.80-69.98]. Median CMV viral load was significantly higher in those individuals who went on to develop CMV disease (P = 0.02). In PCR-positive patients, each 0.25 log10 increase in viral load increased the risk of disease (relative hazard, 1.37; 95% CI, 1.15-1.63; P = 0.0004). CONCLUSIONS: CMV PCR predicts the development of CMV disease and can be used to target ophthalmological resources to those patients at highest risk of retinitis. Asymptomatic patients who are PCR-positive represent a high-risk group in whom controlled trials of pre-emptive therapy could be conducted.

AIDS-Related Opportunistic Infections↗
PubMed1997

Micronutrient supplementation and infection in institutionalized elders.

A randomized, double-blind, placebo-controlled clinical trial was conducted to determine the effects of a low-dose (< 2 times the Recommended Dietary Allowance) micronutrient supplement containing trace minerals (zinc and selenium) and/or vitamins/provitamins (vitamin C, alpha-tocopherol, and beta-carotene) on the incidence of respiratory and urogenital infections in institutionalized elderly. After 2 years, there was a significant decrease in the mean number of infections in elders given trace elements (p < 0.01) but not vitamins.

Aged↗
PubMed1997

Prospective study of human herpesvirus 6, human herpesvirus 7, and cytomegalovirus infections in human immunodeficiency virus-positive patients.

Blood samples from human immunodeficiency virus (HIV)-positive patients were monitored for cytomegalovirus (CMV), human herpesvirus 6 (HHV-6), and HHV-7 by PCR. We detected CMV in 17% of the patients, HHV-6 in 6%, and HHV-7 in 3%. The viral loads of CMV were significantly higher than those of HHV-6 (P = 0.007) or HHV-7 (P = 0.01). Detection of CMV and HHV-6 was associated with low and high CD4 counts, respectively.

CD4 Lymphocyte Count↗
PubMed1997

Factors of nutritional health-seeking behaviors. Findings from the Georgia Centenarian Study.

Health-seeking behaviors are described by Harris and Guten (1979) as any behavior of an individual that promotes, protects, or maintains one's health, regardless of actual or perceived health status. The purpose of this study was to determine if nutritional health-seeking behavior (Bausell, 1986) comprised one or more factors for older adults (N = 256). Participants were in their 60s (n = 90), 80s (n = 91), and 100+ (n = 75). Exploratory factor analysis indicated that nutritional health-seeking behavior items formed two factors: avoid (i.e., avoiding unhealthy nutritional behavior) and seek (i.e., attempting or seeking healthy nutritional behavior). Multiple regression analysis revealed that risk factors for poor nutritional health-seeking behaviors in older adults include advanced age, low economic resources, and male gender. Protective factors included in the personality factors of self-discipline, enthusiasm, sensitivity, and warmth.

Age Factors↗
PubMed1997

A novel mitochondrial tRNA isoleucine gene mutation causing chronic progressive external ophthalmoplegia.

We report a sporadic case of chronic progressive external ophthalmoplegia that developed during childhood and was associated with ragged-red and cytochrome c oxidase (COX)-negative fibers in skeletal muscle. Sequencing of all the mitochondrial transfer RNA (tRNA) genes identified a single potentially pathogenic mutation--a T to C transition at position 4274 in the tRNA(Ile) gene. This mutation was not present in skeletal muscle from 79 controls, and the level of the mutation in COX-negative fibers was significantly greater than the level in COX-positive fibers.

Adult↗
PubMed1997

Nucleotide and amino acid sequence analysis of the 100K protein of a serotype 3 porcine adenovirus.

The genomic region between map units 69 and 78 of a type 3 porcine adenovirus (PAV3) was sequenced and analysed. An open reading frame (ORF) of 2514 nucleotides encoding a polypeptide of 838 amino acids and approximately 94.1 kDa was found. The size and location of the ORF suggested it was the PAV3 homologue of the 100K gene and this was confirmed by nucleotide sequence comparison with the 100K of human adenovirus type 2. Amino acid sequence alignment of the predicted polypeptide with the sequences of the 100K proteins of four human adenoviruses and type 10 fowl adenovirus revealed sequence identities of between 31% and 52%. Although amino acid conservation was present throughout the entire sequences compared, lower identity was noted in both the amino- and carboxy-termini.

Adenoviruses, Human↗
PubMed1997

Nucleotide sequence of canine herpesvirus homologues of herpes simplex virus type 1 US2, US3, glycoproteins I and E, US8.5 and US9 genes.

The partial nucleotide sequence of two BamHI fragments that span the unique short region (US), terminal repeat region (TR) and internal repeat region (IR) of canine herpesvirus (CHV) has been determined. Data obtained revealed several open reading frames (ORF's) identified as the US2, US3, gI, gE and US9 homologues of herpes simplex virus type 1 (HSV1). The CHV homologues also show significant identity in amino acid sequence with those encoded by feline herpesvirus type 1 (FHV1), bovine herpesvirus (BHV1) and equine herpesvirus (EHV1). Translation of another ORF showed little amino acid identity with the gene products of other alpha-herpesviruses. Its genomic position relative to the other CHV homologues would suggest it is the US8.5 gene of CHV.

Alphaherpesvirinae↗