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Biomedical subjects

M D Brown

Publications and source records attributed to M D Brown.

At least 145 records · Page 8Linked to original sources
PubMed1994

Epidemic neuropathy in Cuba not associated with mitochondrial DNA mutations found in Leber's hereditary optic neuropathy patients. Cuba Neuropathy Field Investigation Team.

An epidemic neuropathy in Cuba has caused bilateral optic neuropathies in more than 26,000 people during the past three years. Various pathogenetic factors have been proposed, including toxins, nutritional deficiencies, and an underlying genetic predisposition involving mitochondrial DNA. As part of a case-control collaborative investigation, 135 Cuban blood samples were analyzed for the most common mitochondrial DNA mutations associated with Leber's hereditary optic neuropathy. None of the participants tested were found to have the mitochondrial DNA mutations at nucleotide positions 11778, 3460, 14484, 7444, or 9804. Of 57 definite case subjects and 69 normal control subjects, three case and three control subjects had the mutation at nucleotide position 9438, three different case and three different control subjects had the mutation at position 13708, and one case and one control subject had the mutation at position 15257 in association with the mutation at position 13708. The most common mitochondrial DNA mutations associated with Leber's hereditary optic neuropathy do not appear to be contributing factors in the epidemic neuropathy in Cuba. We also identified a large Cuban family with maternally related members who experienced visual loss consistent with the diagnosis of Leber's hereditary optic neuropathy. Maternal family members harbored the highly pathogenetic mutation at nucleotide position 11778.

Adolescent↗
PubMed1994

A mitochondrial DNA mutation at nucleotide pair 14459 of the NADH dehydrogenase subunit 6 gene associated with maternally inherited Leber hereditary optic neuropathy and dystonia.

A five-generation Hispanic family expressing maternally transmitted Leber hereditary optic neuropathy and/or early-onset dystonia associated with bilateral basal ganglia lesions was studied. Buffy coat mitochondrial DNA (mtDNA) from a severely affected child was amplified by the polymerase chain reaction and greater than 90% sequenced. The mtDNA proved to be a Native American haplogroup D genotype and differed from the standard "Cambridge" sequence at 40 nucleotide positions. One of these variants, a G-to-A transition at nucleotide pair (np) 14459, changed a moderately conserved alanine to a valine at NADH dehydrogenase subunit 6 (ND6) residue 72. The np 14459 variant was not found in any of 38 Native American haplogroup D mtDNAs, nor was it detected in 108 Asian, 103 Caucasian, or 99 African mtDNAs. Six maternal relatives in three generations were tested and were found to harbor the mutation, with one female affected with Leber hereditary optic neuropathy being heteroplasmic. Thus, the np 14459 G-to-A missense mutation is specific to this family, alters a moderately conserved amino acid in a complex I gene, is a unique mtDNA variant in Native American haplogroup D, and is heteroplasmic, suggesting that it is the disease-causing mutation.

Adenine↗
PubMed1994

Molecular basis of mitochondrial DNA disease.

Mitochondrial ATP production via oxidative phosphorylation (OXPHOS) is essential for normal function and maintenance of human organ systems. Since OXPHOS biogenesis depends on both nuclear- and mitochondrial-encoded gene products, mutations in both genomes can result in impaired electron transport and ATP synthesis, thus causing tissue dysfunction and, ultimately, human disease. Over 30 mitochondrial DNA (mtDNA) point mutations and over 100 mtDNA rearrangements have now been identified as etiological factors in human disease. Because of the unique characteristics of mtDNA genetics, genotype/phenotype associations are often complex and disease expression can be influenced by a number of factors, including the presence of nuclear modifying or susceptibility alleles. Accordingly, these mutations result in an extraordinarily broad spectrum of clinical phenotypes ranging from systemic, lethal pediatric disease to late-onset, tissue-specific neurodegenerative disorders. In spite of its complexity, an understanding of the molecular basis of mitochondrial DNA disease will be essential as the first step toward rationale and permanent curative therapy.

DNA, Mitochondrial↗
PubMed1994

Long term bradycardia by electrical pacing: a new method for studying heart rate reduction.

OBJECTIVE: The aim was to develop a method of inducing bradycardia on a chronic basis by electrical pacing and to study its effect upon myocardial capillarity in pigs. METHODS: Farm pigs were instrumented with Medtronic dual chamber telemetric pacemakers and two leads, either both atrial or one atrial and one ventricular. Bradycardia was achieved by linking pacing stimuli to endogenous atrial or ventricular events for the different electrode positions respectively, and monitored postoperatively for 4-5 weeks by implanted telemetric ECG devices. Myocardial capillary supply (lectin staining) and myocyte cross sectional areas were then estimated in sections of the left ventricle. RESULTS: Heart rates were reduced by 40-50 beats.min-1 at operation from a resting rate of 110(SEM 3) beats.min-1 in eight animals and bradycardia maintained upon recovery from anaesthesia for up to five weeks, with no obvious limitations for the animals. After this time, heart rates measured under anaesthesia were found to remain low for up to 2 h after pacing had been switched off. There was no evidence of myocyte hypertrophy, yet mean capillary density.mm-2 was significantly increased by pacing, from 1470(50) to 1734(82), p < 0.02. CONCLUSIONS: It is possible to produce primary heart rate reduction in the pig by a novel means of pacing on a chronic basis, leading to increased myocardial capillary supply without myocyte hypertrophy. This method can provide a basis for future investigations of the beneficial effects of heart rate reduction in the vascularly compromised heart.

Animals↗
PubMed1994

VP2 sequences of recent European 'very virulent' isolates of infectious bursal disease virus are closely related to each other but are distinct from those of 'classical' strains.

Part of genomic segment A of five recent European 'very virulent' (VV) isolates of infectious bursal disease virus, encoding the major, conformational, protective epitope, have been reverse-transcribed, amplified and cloned. Sequence analysis of 582 bases of VP2 showed that the four U.K. isolates are closely related to each other, differing by no more than two bases, but are distinct from the 'classical' type 1 strains (differing by at least 29 bases, or four amino acids, from any other strain and by 15 bases, or three amino acids, from the consensus sequence of all other strains). The U.K. isolates differ from an earlier VV isolate from the Netherlands by no more than five bases or two amino acids. The U.K. isolates differ by only one or two bases, with no amino acid changes, from the recently published sequence of a Japanese VV isolate.

Amino Acid Sequence↗
PubMed1994

A non-directed, hydroxylamine-generated suppressor mutation in the P3 pairing region of the bacteriophage T4 td intron partially restores self-splicing capability.

Hydroxylamine (HA) mutagenesis of an HA-induced splicing-defective bacteriophage T4 td intron mutant with a mutation in the intron P3 RNA pairing region was used to generate pseudorevertants. Because HA can only cause GC to AT transitions, the original mutant (H104A) could not undergo true reversion, yet the compensatory mutation on the opposite side of the P3 helix, which was complementary to the original H104A mutation, could occur. A pseudorevertant was isolated that contained both the original H104A mutation and the compensatory mutation HS9. By phenotypic and molecular genetic criteria, this double mutant (H104A-HS9) was shown to be able to undergo significant RNA splicing, thus confirming the existence and functional importance of the long-range P3 pairing region in this phage intron. The second-site suppressor mutation (HS9) was isolated by phage cross and also exhibited some self-splicing ability. A correlation exists between the strength of P3 helix Watson-Crick base pairing and the apparent level of splicing when wild-type, H104A, HS9, and H104A-HS9 are compared. This suggests that the primary role of the P3 RNA pairing region in the T4 td intron is structural in contributing to the critical RNA secondary structure.

Aged↗
PubMed1994

Effect of torbafylline on muscle blood flow, performance, and capillary supply in ischemic muscles subjected to varying levels of activity.

Torbafylline, a novel xanthine derivative, was given to rats by gavage (2 x 25 mg.kg-1.day-1, morning and evening) to study its effect upon fast muscles (tibialis anterior and extensor digitorum longus) made ischemic by unilateral ligation of the common iliac artery and subjected to two types of activity imposed by chronic electrical stimulation at 10 Hz: (i) strenuous, 6 h.day-1 as 3 x 2 h with 90- to 120-min intervals or (ii) mild, 105 min.day-1 as 7 x 10-15 min with 90- or 85-min intervals for 12-14 days. Some of the deleterious effects on ischemic muscles of strenuous activity (reduced blood flow during contractions, less resistance to fatigue) were remedied by torbafylline treatment, and values normalized. Most notably, torbafylline significantly reduced the degree of capillary endothelial cell swelling. In addition, the degree of atrophy was reduced and fatigue resistance improved in muscles contralateral to ischemic, which had been impaired with the strenuous regime. Torbafylline treatment had little further effect on ischemic muscles subjected to mild stimulation, which on its own improved functional hyperemia, resistance to fatigue, and the capillaries per fibre ratio in tibialis anterior, although it did significantly increase the capillary per fibre ratio in extensor digitorum longus. These data indicate a possible role for torbafylline as an adjunct to exercise therapy for chronic muscle ischemia.

Animals↗
PubMed1994

Effect of long-term electrical stimulation on vascular supply and fatigue in chronically ischemic muscles.

Fast skeletal muscles of Sprague-Dawley rats [tibialis anterior (TA) and extensor digitorum longus (EDL)] were subjected to ischemia by unilateral ligation of the common iliac artery. In some animals, ischemia was combined with indirect electrical stimulation at 10 Hz either for 3 x 2 h (strenuous activity) or for 7 x 10-min bouts/day (mild activity). After 2 wk, muscle blood flow and fatigue were measured during 5-min isometric supramaximal twitch contractions at 4 Hz. Terminal arteriole diameters were assessed in TA by intravital microscopy at rest and during contractions. Vascular perfusion pressure in the muscles was estimated from measurements in the carotid and saphenous arteries below the site of ligation. Capillary supply was expressed in TA and EDL as capillary-to-fiber ratio on the basis of histochemical staining for capillaries. Strenuous stimulation of ischemic muscles increased their atrophy, failed to restore blood flow, and actually worsened fatigue. In contrast, mild stimulation improved perfusion pressure, increased capillary-to-fiber ratio in the glycolytic part of TA, restored dilatation of terminal arterioles during muscle contractions, and improved blood flow and muscle fatigue so that they were no longer significantly different from control muscles. Thus, an attenuated intermittent protocol may be indicated in the treatment of muscle ischemia.

Animals↗
PubMed1994

The effect of long-term bradycardia on heart microvascular supply and performance.

Bradycardia has been shown to be beneficial for the normal and ischaemic heart because it improves diastolic perfusion and oxygen supply demand balance. Experimentally, a chronically induced decrease in heart rate, either by electrical pacing or pharmacological means, was found previously to increase myocardial capillary supply in normal rabbit and rat hearts. These studies have been extended to a larger mammal, the pig, in which a direct bradycardia (approximately 30% decrease in heart rate) was induced by electrical pacing for 4-5 weeks. There was no evidence of heart hypertrophy and capillary density was found to be significantly increased in the left, but not right, ventricle. Cardiac function during dobutamine inotropic challenge was better in pig hearts which had been paced bradycardially. They performed greater stroke work-higher stroke flow output at lower heart rate--for similar coronary blood flow, thus demonstrating an improved economy of flow utilisation. Heart rate reduction may facilitate capillary growth in the absence of cardiac hypertrophy by prolonging diastolic perfusion, and/or mechanical stretch of vessels due to increased stroke volume capacity. In either case, capillaries would be exposed to increased wall tension which could trigger angiogenesis.

Animals↗
PubMed1994

Alterations of annexin expression in pathological neuronal and glial reactions. Immunohistochemical localization of annexins I, II (p36 and p11 subunits), IV, and VI in the human hippocampus.

Annexins are Ca(2+)-dependent membrane-binding proteins that are potentially important in Ca(2+)-induced neurotoxicity or neuroprotection. To address the possible involvement of annexins in cellular reactions to brain injury and neurodegenerative disease, we studied the immunohistochemical localization of annexins I, II (p36 and p11), IV, and VI in the adult human hippocampus. Formalin-fixed, paraffin-embedded tissue from autopsy cases representing hypoxic-ischemic injury, seizure disorders, Alzheimer's disease, and age-related controls were examined. Neurons showed cytoplasmic immunoreactivity for annexin I, whereas annexin VI was distributed in patterns suggesting plasma membrane and perisynaptic locations. The cytoarchitectural distribution of annexin VI within neurons was altered in pathological states and annexin VI was strongly associated with neuronal granulovacuolar bodies in Alzheimer's disease. Reactive astrocytes expressed annexins I, II (p36 and p11), and IV, whereas quiescent astrocytes were minimally immunoreactive. Significant annexin immunoreactivity was also detected in oligodendrocytes (annexin IV), ependymocytes (I, II, and IV), choroid plexus (I, IV, and VI), meningothelium (I, II, IV, and VI), and vascular endothelium (II and IV) and smooth muscle (I, IV, and VI). This is the first comparative study of immunoreactivities for multiple annexins in human brain. Neurons and glia display selective and different profiles of annexin protein expression and show immunohistochemical changes in pathological conditions, which suggest involvement of annexins in neuronal and glial reactions to injury.

Adult↗
PubMed1993

Mitochondrial DNA variants observed in Alzheimer disease and Parkinson disease patients.

Mitochondrial DNA (mtDNA) variants associated with Alzheimer disease (AD) and Parkinson disease (PD) were sought by restriction endonuclease analysis in a cohort of 71 late-onset Caucasian patients. A tRNA(Gln) gene variant at nucleotide pair (np) 4336 that altered a moderately conserved nucleotide was present in 9/173 (5.2%) of the patients surveyed but in only 0.7% of the general Caucasian controls. One of these patients harbored an additional novel 12S rRNA 5-nucleotide insertion at np 956-965, while a second had a missense variant at np 3397 that converted a highly conserved methionine to a valine. This latter mutation was also found in an independent AD + PD patient, as was a heteroplasmic 16S rRNA variant at np 3196. Additional studies will be required to determine the significance, if any, of these mutations.

Aged↗
PubMed1993

Distribution and characterization of mutations induced by nitrous acid or hydroxylamine in the intron-containing thymidylate synthase gene of bacteriophage T4.

The detailed distribution and characterization of 51 hydroxylamine (HA)-induced and 59 nitrous acid (NA)-induced mutations in the intron-containing bacteriophage T4 thymidylate synthase (td) gene is reported here. Mutations were mapped in 10 regions of the td gene by recombinational marker rescue using plasmid or M13 subclones of the td gene. Phage crosses using deletion mutants with known breakpoints in the 3' end of the td intron subdivided HA and NA mutations which mapped in this region. At least 31 of the mutations map within the 1-kb group I self-splicing intron. Intron mutations mapped only in the 5' and 3' ends of the intron sequence, in accordance with the hypothesis that the 5' and 3' domains of the T4 td intron are essential for correct RNA splicing. RNA sequence analysis of a number of mapped td mutations has identified two intron nucleotides and one exon nucleotide where both HA- and NA-induced mutations commonly occur. These three loci are characterized by a GC dinucleotide, with the mutations occurring at the cytosine residue. Thus, these data indicate at least three potential sites of both HA- and NA-induced mutagenic hotspot activity within the td gene.

Bacteriophage T4↗
PubMed1993

Proinsulin and insulin concentrations following intravenous glucose challenges in normal, obese, and non-insulin-dependent diabetic subjects.

We investigated the effects of different intravenous (IV) glucose challenges on insulin and proinsulin secretion. On separate occasions, seven normal controls and five obese and five non-insulin-dependent diabetic (NIDDM) subjects each received an IV glucose tolerance test (IVGTT), a hyperglycemic clamp (HY), and a 60-minute, standardized, low-dose, continuous infusion of glucose (CIG) in a randomized fashion. Basal proinsulin concentrations in NIDDM subjects (8.4 +/- 5.0 pmol/L) were significantly higher compared with those of normal (1.1 +/- 0.2) and obese subjects (1.5 +/- 0.4; both P < .05). Basal molar proinsulin:insulin ratio (P:I) was also significantly higher in NIDDM subjects (22% +/- 12%) compared with normal (1.0%) and obese subjects (1.6% +/- 0.8%; both P < .01). Proinsulin concentrations did not increase significantly in any group during the first 10 minutes of the IV glucose challenges. However, during HY, significant increases in proinsulin concentration occurred after 60 minutes in each group. In normal and obese subjects, IV glucose administration resulted in significant acute increases in insulin concentrations compared with the characteristic blunted response in NIDDM subjects. The P:I ratio in normal and obese subjects did not change in the first 10 minutes after IV glucose administration. However, by the end of HY, the P:I ratio had increased significantly in normal subjects by 1% to 5% +/- 2% (P < .05), and in obese subjects by 1% to 5% +/- 1% (P < .02). In NIDDM subjects, both HY (19% +/- 10% to 27% +/- 12%) and IVGTT (18% +/- 9% to 43% +/- 16%) resulted in a transient increase in the basal P:I ratio by 5 minutes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

A proflavin-induced frameshift hotspot in the thymidylate synthase gene of bacteriophage T4.

Twenty-one independent thymidylate synthase deficient (td) mutants were isolated after proflavin mutagenesis of T4D0 phage. A strikingly high proportion of these mutations (17 of 21; 80%) mapped in a small 122 nucleotide (nt) region which spans the 5' splice site of this intron-containing gene. This region comprises only 14% of the total td exon sequence. RNA sequence analysis of these mutants identified a series of frameshift insertion/deletion mutations and indicated a hotspot for proflavin-induced mutations in the 3' end of exon I of the td gene. The mutant sequences at the hotspot site fully support a previously proposed mutagenic mechanism for proflavin-induced mutations in which frameshifts are produced as a consequence of exonuclease or DNA polymerase activity at the 3' ends of nicks in the DNA produced by perturbation of the T4-encoded type II topoisomerase activity by the acridine. Sixteen of the seventeen DNA mutations in the hotspot region can be explained by the model as a consequence of enzymatic processing of nicks at two phosphodiester bonds staggered by 4 base pairs (bp) and located on opposite strands of the DNA. Thus, these mutants exhibit precisely the symmetry expected of topoisomerase-mediated mutagenesis. The DNA sequences of the td hotspot mutants, when considered with the sequences of proflavin-induced mutants in the T4 rIIB and lysozyme genes, confirm the view that proflavin-induced mutations in diverse bacteriophage T4 DNA sequences are all produced by the topoisomerase-dependent mechanisms and do not support the view that classical misalignments in DNA repeats are hotspots for proflavin-induced mutagenesis in T4.

Bacteriophage T4↗
PubMed1993

Merkel cell carcinoma.

Merkel cell carcinoma is a malignant neuroendocrine tumor with features of epithelial differentiation. Biologically aggressive, it may be difficult to diagnose and, particularly in its late stages, even more difficult to treat effectively. This article addresses what is known and what is still controversial about the histogenesis, diagnosis, and management of Merkel cell carcinoma and the structure and function of the Merkel cell from which it is believed to be derived. The incidence, clinical presentation and diagnosis, ultrastructure, immunocytochemistry, treatment, and prognosis of this tumor will be discussed.

Carcinoma, Merkel Cell↗