Biomedical subjects

R H Davis

Publications and source records attributed to R H Davis.

At least 19 recordsLinked to original sources

PubMed1992

Ornithine decarboxylase gene of Neurospora crassa: isolation, sequence, and polyamine-mediated regulation of its mRNA.

Ornithine decarboxylase (ODC), which initiates the biosynthesis of the polyamines putrescine, spermidine, and spermine, is encoded by the spe-1 gene of the fungus Neurospora crassa. This gene and its cDNA have been cloned and sequenced. The gene has a single 70-nucleotide intron in the coding sequence. The cDNA, comprising the entire coding region, recognizes a single 2.4-kb mRNA in Northern (RNA) blots. The mRNA transcript, defined by S1 mapping, has an extremely long, 535-base leader without strong secondary-structure features or an upstream reading frame. The translational start of the protein is ambiguous: a Met-Val-Met sequence precedes the Pro known to be the N terminus of the ODC polypeptide. The polypeptide encoded by the N. crassa spe-1 gene (484 amino acids) has 46% amino acid identity with that of Saccharomyces cerevisiae (466 amino acids) and 42% with that of mouse (461 amino acids). Alignment of the longer N. crassa sequence with S. cerevisiae and mouse sequences creates gaps in different sites in the S. cerevisiae and mouse sequences, suggesting that N. crassa ODC is closer to an ancestral form of the enzyme than that of either yeast or mouse ODC. N. crassa ODC, which turns over rapidly in vivo in the presence of polyamines, has two PEST sequences, found in most ODCs and other proteins with rapid turnover. In striking contrast to other eucaryotic organisms, the variation in the rate of ODC synthesis in response to polyamines in N. crassa is largely correlated with proportional changes in the abundance of ODC mRNA. Spermidine is the main effector of repression, while putrescine has a weaker effect. However, putrescine accumulation appears to increase the amount of active ODC that is made from a given amount of ODC mRNA, possibly by improving its translatability. Conversely, prolonged starvation for both putrescine and spermidine leads to the differentially impaired translation of ODC mRNA.

Amino Acid Sequence↗

PubMed1992

Sequestered end products and enzyme regulation: the case of ornithine decarboxylase.

The polyamines (putrescine, spermidine, and spermine) are synthesized by almost all organisms and are universally required for normal growth. Ornithine decarboxylase (ODC), an initial enzyme of polyamine synthesis, is one of the most highly regulated enzymes of eucaryotic organisms. Unusual mechanisms have evolved to control ODC, including rapid, polyamine-mediated turnover of the enzyme and control of the synthetic rate of the protein without change of its mRNA level. The high amplitude of regulation and the rapid variation in the level of the protein led biochemists to infer that polyamines had special cellular roles and that cells maintained polyamine concentrations within narrow limits. This view was sustained in part because of our continuing uncertainty about the actual biochemical roles of polyamines. In this article, we challenge the view that ODC regulation is related to precise adjustment of polyamine levels. In no organism does ODC display allosteric feedback inhibition, and in three types of organism, bacteria, fungi, and mammals, the size of polyamine pools may vary radically without having a profound effect on growth. We suggest that the apparent stability of polyamine pools in unstressed cells is due to their being largely bound to cellular polyanions. We further speculate that allosteric feedback inhibition, if it existed, would be inappropriately responsive to changes in the small, freely diffusible polyamine pool. Instead, mechanisms that control the amount of the ODC protein have appeared in most organisms, and even these are triggered inappropriately by variation of the binding of polyamines to ionic binding sites. In fact, feedback inhibition of ODC might be maladaptive during hypoosmotic stress or at the onset of growth, when organisms appear to require rapid increases in the size of their cellular polyamine pools.

Animals↗

PubMed1992

Aloe vera and the inflamed synovial pouch model.

Administration of air under the skin produced a pouch wall that closely resembled a synovium in that the inner lining was made up of macrophages and fibroblasts. Administration of 1% carrageenan directly into the 7-day-old air pouch produced an inflammation characterized by an increased number of mast cells in pouch fluid as well as an increase in wall vascularity. A punch biopsy weight of the pouch wall did not reveal an increase in 1% carrageenan-treated animals. However, a 10% Aloe vera treatment of carrageenan-inflamed synovial pouches reduced the vascularity 50% and the number of mast cells in synovial fluid 48%. The pouch wall punch biopsy weight was increased by A. vera, which was verified by histologic examination of the inner synovial lining. Aloe vera stimulated the synovial-like membrane, as evidenced by an increased number of fibroblasts, suggesting that A. vera stimulated fibroblasts for growth and repair of the synovial model. The synovial air pouch can be used to study simultaneously the acute anti-inflammatory and fibroblast stimulating activities of A. vera.

Aloe↗

PubMed1991

Calcium modulation of polyamine transport is lost in a putrescine-sensitive mutant of Neurospora crassa.

Putrescine transport in Neurospora is saturable and concentrative in dilute buffers, but in the growth medium putrescine simply equilibrates across the cell membrane. We describe a mutant, puu-1, that can concentrate putrescine from the growth medium because the polyamine transport system has lost its normal sensitivity to Ca2+. The wild type closely resembles the mutant if it is washed with citrate and ethylene glycol bis(beta-aminoethyl ether)N,N'-tetraacetic acid. The mutant phenotype also appears in the wild type after treatment with cycloheximide. The results suggest that putrescine uptake is normally regulated by an unstable Ca(2+)-binding protein that restricts polyamine uptake. This protein is evidently distinct from the polyamine-binding function for uptake, which is normal in mutant and in cycloheximide-treated wild type cells. The puu-1 mutation, stripping of Ca2+, and cycloheximide treatment all cause an impairment of amino acid transport, indicating that other membrane transport functions rely upon the product of the puu-1+ gene. Preliminary evidence suggests that the putrescine carrier is not the Ca(2+)-sensitive, low-affinity K(+)-transport system, but K+ efflux does accompany putrescine uptake.

Biological Transport↗

PubMed1991

Polyamine toxicity in Neurospora crassa: protective role of the vacuole.

We used mutant strains of Neurospora crassa to define the discretionary capacity of this species for excess putrescine. The spe-3 mutant, which accumulates putrescine internally, and the puu-1 mutant, which accumulates toxic levels of putrescine from the medium, both sequestered large excesses of putrescine in vacuoles. Concomitantly in puu-1, inorganic polyphosphate increased modestly and some of the monovalent cation of the vacuole was discharged. These two factors contribute to the increased capacity for polyamines in this fungus. Putrescine, however, can exceed the capacity of vacuoles such that they no longer protect the cytosol from toxic levels of the amine. The puu-1 mutant illustrates the importance of the sequestration of intracellular polyamines, as well as the control of polyamine uptake through the cell membrane.

Amino Acids↗

PubMed1991

Effects of Vibrio cholerae recombinant strains on rabbit ileum in vivo. Enterotoxin production and myoelectric activity.

Previous studies have identified the effects of Vibrio cholerae and its enterotoxin, choleragen (CT A+B+), on the myoelectric activity of rabbit ileal loops in vivo. The response was defined as the migrating action potential complex, the single ring contraction that propels luminal contents aborad. In this study the same rabbit model is used to assess whether migrating action potential complex activity or fluid output is induced by recombinant strains of V. cholerae that produce no subunit of cholera toxin (CT A-B-) or only by the inactive binding subunit (CT A-B+). Three live strains were studied: El Tor wild-type N16961 (CT A+B+) and recombinant strains CVD106 (CT A-B+) and JBK70 (CT A-B-). Controls received sterile culture broth. Migrating action potential complex frequency in animals inoculated with CT A+B+ was significantly increased compared with that in all other experimental groups (P less than 0.01). Fluid output was also increased in animals inoculated with CT A+B+ compared with fluid output in all other groups (P less than 0.05). Migrating action potential complex frequency and fluid output in rabbits given CT A-B+ or CT A-B- did not differ from activity in controls. How these recombinant strains induce diarrhea is unknown, but the mechanism may involve bacterial colonization or production of an unknown toxin.

Animals↗

PubMed1991

Alternative sulphur donors for detoxification of cyanide in the chicken.

1. Urinary excretion of thiocyanate by hens after dosage with cyanide was studied over 3 hr periods during which various sulphur sources were infused. 2. With 20 mumoles cyanide, endogenous sulphur supplies appeared to be almost sufficient. 3. With 45 mumoles cyanide, thiocyanate excretion was doubled with 90 mumoles of sulphur donor. Higher doses of mercaptopyruvate were also effective but not rhodanese substrates (thiosulphate or methanethiosulphonate): they interfered with thiocyanate excretion and may also have suppressed its formation. 4. Mercaptopyruvate and rhodanese substrates also differed in their effects on blood cyanide concentration and on the excretion of isotope from radiolabelled cyanide.

Animals↗

PubMed1991

Intestinal motility changes in rats after enteric serotonergic neuron destruction.

The myenteric plexus consists of several subpopulations of morphologically and chemically distinct neurons known to contain a variety of peptides and amines, one of which is serotonin (5-hydroxytryptamine). These neurons are considered essential for nerve-to-nerve transmission. In the present study, we investigated the effect of 5,6- and 5,7-dihydroxytryptamine (5,6-DHT; 5,7-DHT), indoleamine neurotoxins that selectively and irreversibly injure the serotonergic neurons of the myenteric plexus. Treatment with 5,6-, or 5,7-DHT caused marked disruption of the activity front of the migrating myoelectric complex (MMC), increased its duration, and decreased its propagation velocity. At higher doses, 5,7-DHT also reduced the slow-wave frequency. Immunohistochemical techniques showed that tissue from rats treated with 5,7-DHT was depleted of serotonin-like immunoreactivity within the myenteric plexus neurons. Reserpine also caused motility and immunohistochemical changes similar to those induced by the two neurotoxins. Therefore, destruction of enteric serotonergic neurons disrupts the MMC. These studies support the cellular concepts that serotonergic neurons function as interneurons in the myenteric plexus, modulating and processing the neural stimuli, and that serotonin is an important neurotransmitter in the small intestine.

5,6-Dihydroxytryptamine↗

PubMed1991

Aloe vera as a biologically active vehicle for hydrocortisone acetate.

Aloe vera, as a biological vehicle for hydrocortisone 21-acetate, was tested topically and systemically against acute inflammation. Systemically, the combination of A. vera and hydrocortisone produced a maximum 88.1% inhibition of edema. Polymorphonuclear leukocyte infiltration was reduced 91.1%. The topical inhibition of edema peaked at 97%. The possibility that A. vera has significant potential as a biologically active vehicle for steroids is discussed.

Acute Disease↗

PubMed1991

The isolation of an active inhibitory system from an extract of aloe vera.

An Aloe vera extract was prepared with 50% ethanol. The resultant supernatant and precipitate were tested for anti-inflammatory activity using the croton oil-induced ear-swelling assay. The supernatant fraction decreased inflammation, when applied topically, by 29.2%, and the precipitate decreased inflammation by 12.1%. The authors have shown that the anti-inflammatory activity (inhibitory system) resides in the supernatant of a 50% ethanol extract.

Aloe↗

PubMed1991

Isolation of a stimulatory system in an Aloe extract.

The authors' previous work on a 50% ethanol extract of Aloe vera was done to evaluate anti-inflammatory activity using the croton oil-induced ear swelling assay. The anti-inflammatory activity was found in the supernatant fraction. The supernatant fraction decreased inflammation, when applied topically, by 29.2%, and the precipitate decreased inflammation by 12.1%. However, in the present work, the precipitate fraction decreased the wound diameter by an average of 47.1% (stimulatory system). Little or no wound healing activity was found in the supernatant. Aloe vera appears to act as a modulatory system toward wounds and inflammation and is a potentially valuable tool for managing lower extremity conditions.

Adult↗

PubMed1990

The genetics of polyamine synthesis in Neurospora crassa.

New mutations of the polyamine pathway of Neurospora crassa fell into three categories. The majority affected ornithine decarboxylase and lay at the previously defined spe-1 locus. One mutation, JP100, defining the new spe-2 locus, eliminated S-adenosyl-methionine decarboxylase and led to putrescine accumulation. Revertants of this mutation suggested that the locus encodes the enzyme. Two other mutations, LV105 and JP120, defined a third locus, spe-3. Strains with these mutations also accumulated putrescine and were presumed to lack spermidine synthase activity, which catalyzes the formation of spermidine from putrescine and decarboxylated S-adenosylmethionine. The three spe loci lay within about 20 map units of one another on the right arm of Linkage Group V in the order: centromere-spe-2-spe-1-spe-3. The requirement for spermidine for growth was much less in spe-2 and spe-3 mutants than in spe-1 mutants, which do not accumulate putrescine. This suggested that putrescine fulfills many, but not all, of the functions of spermidine, or that high levels of putrescine render spermidine more effective in its essential roles.

Adenosylmethionine Decarboxylase↗

PubMed1990

Deoxyhypusine/hypusine formation on a 21,000-dalton cellular protein in a Neurospora crassa mutant in vivo and in vitro.

Hypusine formation on an 18,000-dalton cellular protein is a unique spermidine-dependent, post-translational modification that appears to be ubiquitous in mammalian cells. To determine whether this modification also exists in lower eukaryotes, we examined possible labeling in vitro and in vivo of cellular protein(s) by [3H]spermidine in a mutant strain of Neurospora crassa (arge-12 ota aga) in which ornithine and polyamine synthesis could be nutritionally manipulated. Because of poor uptake of polyamines in this organism, [3H]ornithine, the immediate precursor of polyamines, was used for the in vivo labeling experiment. Both in vitro and in vivo labeling resulted in a specific labeling of a 21,000-dalton protein. Radioactive hypusine was recovered from radiolabeled 21,000-dalton protein following acid hydrolysis. The in vitro labeling of the 21,000-dalton protein was dramatically stimulated by NAD+ and NADP+, but not by FMN or FAD, suggesting that an NAD+/NADP(+)-dependent oxidative cleavage of spermidine is involved in deoxyhypusine formation. Isoelectric focusing/sodium dodecyl sulfate two-dimensional gel analysis revealed three isoforms of the in vitro labeled 21,000-dalton protein, with pI values ranging from 5.2 to 6.5. In contrast, the 21,000-dalton protein metabolically labeled in vivo gave only one spot with a pI value of approx. 3.5.

Electrophoresis, Polyacrylamide Gel↗

PubMed1990

Management of polyamine pools and the regulation of ornithine decarboxylase.

The management of polyamine synthesis and polyamine pools differs fundamentally from that of most other small molecular-weight endproducts. The polyamines are vital to growth and important cellular functions, but they are toxic in excess. I argue here that their multivalent cationic character, leading to binding to cell constituents, precludes fluent feedback inhibition of synthesis. This has led to the development of elaborate alternative regulatory mechanisms controlling ornithine decarboxylase, the key initial enzyme of the pathway. Poorly regulated polyamine synthesis and the toxicity of polyamines impose upon cells a need to control uptake and to dispose of excess polyamines. Recent data on polyamine transport suggest unorthodox mechanisms of accomplishing these functions.

Animals↗

PubMed1990

Reduced selenium status in chicks given diets low in sulphur-containing amino acids and containing sodium nitroprusside as a source of cyanide.

1. Two experiments were carried out to study the effect of dietary cyanide, given in the form of sodium nitroprusside (SNP), on the growth and selenium status of chicks fed on diets low in sulphur-containing amino acids. 2. In experiment 1, SNP (0.3 g/kg) depressed growth rate and food intake when included in diets low in total sulphur-containing amino acids. It also reduced plasma and liver glutathione peroxidase activity (GSH-Px) and kidney selenium concentration. These latter variables also showed increases in response to supplements of selenium of 0.1 and 0.2 mg selenium/kg (as sodium selenite) although growth was not affected. 3. In experiment 2, SNP did not affect growth when reduced to 0.1 g/kg in diets low both in methionine and total sulphur-containing amino acids. It did, however, reduce plasma GSH-Px activity, which again increased in response to similar dietary supplements of selenium. 4. It is concluded that dietary cyanide effectively increases the requirement for selenium and could induce deficiency in diets only marginally adequate in selenium, particularly when the supply of sulphur-containing amino acids was marginal or inadequate.

Amino Acids, Sulfur↗

PubMed1990

Vitamin C influence on localized adjuvant arthritis.

This work attempts to determine the influence of vitamin C on locally induced inflammation and arthritis in rat paws, as measured by rat paw swelling, polymorphonuclear leukocyte infiltration, pain, and surface skin temperature. Daily subcutaneous administration of 150 mg/kg of vitamin C over 20 days reduced arthritic swelling, increased pain tolerance, and decreased polymorphonuclear leukocyte infiltration, with no significant change in surface temperature. Vitamin C may provide podiatrists with a supplemental or alternative treatment for patients with rheumatoid arthritis.

Animals↗

PubMed1990

Opportunistic developmental surveillance in general practice.

The importance of the developmental surveillance of children is generally accepted. One method which could be used in general practice is opportunistic surveillance, where the doctor makes use of contacts with children in the surgery to assess their development, and offer advice and health education to parents where appropriate. This study describes an audit of opportunistic developmental surveillance in one general practice. It was found that 95% of a cohort of 78 children had been assessed at least once by the time they were one year old. In addition, at two points in the study, after one and two years, it was found that 81% of children aged two years or less in the practice had had a recent assessment. This level of surveillance compares favourably with reported attendance rates in clinics.

Adolescent↗

PubMed1990

Treatment of peptic ulcer in general practice and in hospital: a comparison of omeprazole and cimetidine.

The aim of this study was to compare duodenal ulcer healing and symptom relief after two and four weeks treatment with omeprazole or cimetidine in groups of patients treated in general practice and as hospital out-patients. It was a randomised, double-blind, parallel group study with stratification for trial centre (hospital or GP). Endoscopy was performed at entry, after two weeks and, if unhealed at two weeks, after four weeks. All endoscopies were carried out in hospitals. In all, 189 patients were randomised (98 omeprazole, 91 cimetidine), 79 (42 per cent) of which by GPs, to either omeprazole 20 mg om (n = 41) or cimetidine 800 mg nocte (n = 38) for two to four weeks. After two weeks, ulcer healing occurred in 56 per cent (omeprazole) and 29 per cent (cimetidine) (p less than 0.05) of patients treated by GPs, and 67 per cent (omeprazole) and 36 per cent (cimetidine) (p less than 0.005) of those treated as hospital out-patients. Similar differences in healing rates were seen after four weeks. Omeprazole produces faster duodenal ulcer healing than cimetidine whether patients are treated as hospital out-patients or by GPs.

Adolescent↗