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H A Simmonds

Publications and source records attributed to H A Simmonds.

At least 163 records · Page 9Linked to original sources
PubMed1986

Prenatal exclusion of purine nucleoside phosphorylase deficiency.

We report on the prenatal exclusion of purine nucleoside phosphorylase (PNP) deficiency in a fetus whose parents were known to be heterozygotes for the enzyme defect. Prenatal investigation was performed in the 16th week of gestation on amniotic fluid and cultured amnion cells using sensitive techniques. The results suggested that the fetus was either normal or a heterozygote. PNP activity in cord red cells confirmed the heterozygous status of the baby.

Adult↗
PubMed1985

Use of a fundamental elution protocol for the development of reversed-phase high-performance liquid chromatography enabling rapid simultaneous determination of purines, pyrimidines and allied compounds commonly found in human biological fluids.

Many high-performance liquid chromatographic methods for the analysis of nucleosides and bases have been developed for a specific requirement. Most are not directly applicable to the separation of all the purines and pyrimidines found in biological fluids. A method has been developed to separate a wide range of compounds found in human urine and plasma in a single run without unduly long run times. Special attention has been paid to sample preparation. Isocratic systems derived from this method for the analysis of specific compounds and enzyme assay mixtures over even shorter periods are described.

Body Fluids↗
PubMed1985

Human B lymphocytes and thymocytes but not peripheral blood mononuclear cells accumulate high dATP levels in conditions simulating ADA deficiency.

Inherited adenosine deaminase (ADA) deficiency is associated with a lymphospecific cytotoxicity affecting both dividing and non-dividing cells. The metabolic basis for this was investigated using different cell types and the potentially toxic metabolite 2'-deoxyadenosine (dAR) in short-term experiments under physiological conditions simulating ADA deficiency (1 mM Pi 8.7 microM dAR). In the uncultured cells, [8-14C] dAR alone was metabolized almost completely only by thymocytes and tonsil-derived B-lymphocytes. The greater percentage of counts (greater than 75%) were in the medium (deoxyinosine, hypoxanthine). Cellular counts were predominantly in adenine nucleotides, and to a lesser extent guanine nucleotides. Interestingly, both thymocytes and tonsil-derived B-lymphocytes, and a partially ADA deficient B lymphoblast line, accumulated detectable amounts of dATP even in the absence of ADA inhibition. Peripheral blood lymphocytes (PBMs) did not, and showed little dAR metabolism. In experiments simulating ADA deficiency varying amounts of 2'-deoxycoformycin (2'dCF) were needed to completely inhibit ADA (20-60 microM), with thymocytes requiring the highest amount. ADA inhibited thymocytes and tonsillar B-lymphocytes accumulated very high dATP levels, which were sustained to an equal extent by both over a 60-min period; PBMs accumulated the lowest values. Results in cultured cells reflected findings in previous studies. Some counts were also found in ATP by a route excluding ADA or PNP. These results again question the hypothesis that B-cells are more resistant than T-cells to the toxic effects of dAR because of an inability to accumulate and sustain elevated dATP levels and underline the lack of comparability between enzyme activity in intact as distinct from lysed cells. They cast doubt on the validity of cultured cells as a model for ADA deficiency and suggest the observed toxicity in some instances might result from altered ATP or GTP pools through inadequate ADA inhibition. They indicate that combined immunodeficiency in ADA deficiency could relate to an equal sensitivity of B-cells and T-cell precursors to the toxic effects of dATP accumulation.

Adenosine Deaminase↗
PubMed1985

Superoxide radicals, immunodeficiency and xanthine oxidase activity: man is not a mouse!

The importance of intact adenosine deaminase (ADA) activity in the generation of superoxide anion by xanthine oxidase has been disputed in studies using human neutrophils or mouse macrophages. The latter demonstrated a positive correlation between ADA activity and superoxide production during phagocytosis. The immunodeficiency in inherited ADA deficiency was related to a defect in this process. Since there is considerable interspecies variation in the tissue distribution of xanthine oxidase, the metabolism of [8-14C]deoxyadenosine (dAR), the toxic metabolite which accumulates in inherited ADA deficiency, was investigated in human peritoneal macrophages. Evaluation of the distribution of radiolabel in both cell and medium demonstrated that human macrophages with intact ADA metabolize dAR under physiological conditions to deoxyinosine and hypoxanthine exclusively. The hypoxanthine is further metabolized within the cell to ATP and GTP, via IMP. No xanthine or uric acid could be detected, confirming that in human macrophages xanthine oxidase activity is insignificant, as it is in most other human cells and tissues, except liver and intestinal mucosa. Thus production of superoxide radicals in such cells via this route would be impossible, and consequently unaffected either by ADA deficiency or the xanthine oxidase inhibitor allopurinol.

Deoxyadenosines↗
PubMed1985

An inborn error of purine metabolism, deafness and neurodevelopmental abnormality.

A syndrome of hyperuricemia, sensorineural deafness, mild mental handicap and congenital disequilibrium in a four-year-old boy is probably inherited as a sex-linked condition since his mother has sensorineural deafness and similar biochemical abnormalities. There is evidence of a superactive PP-ribose-P synthetase, normal purine salvage enzymes, and severe depletion of nicotinamide adenine dinucleotide and guanine triphosphate in red cells.

Deafness↗
PubMed1985

EHNA is a poor inhibitor of deoxyadenosine catabolism in cultured human lymphocytes.

Erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA) has been used by many workers as enzyme inhibitor in vitro to simulate the in vivo situation in inherited adenosine deaminase (ADA) deficiency. In this study the metabolism of 8-14C deoxyadenosine (dAR) has been followed in cultured lymphocytes from patients deficient in enzymes associated with the catabolism and salvage of dAR, in the absence and presence of 10 microM EHNA. The results show that EHNA, at these concentrations, does not prevent the catabolism of dAR and thus does not provide a valid model for investigating the toxicity to the immune system in inherited ADA deficiency.

Adenine↗
PubMed1984

Metabolism of deoxynucleosides by lymphocytes in long-term culture deficient in different purine enzymes.

The metabolism of 8-14C-labelled 2'-deoxyadenosine (dAR) and 2'-deoxyguanosine (dGR) has been investigated using lymphocytes in long-term culture transformed by Epstein-Barr (EB) virus (B-cells) from eight patients with different inherited purine enzyme defects. The use of such lines enabled accurate mapping of the route of metabolism by acting as a 'trap' for the radiolabel at specific points. With either substrate (25 microM) most of the label was recovered in the medium. Using dAR, less than 30% of the radiolabel was incorporated into cellular nucleotides. For dGR, values were less than 18%. Studies with dAR alone confirmed the principal route of metabolism was to hypoxanthine, with further metabolism (by lines with intact salvage pathways) to ATP and GTP in the ratio of approximately 4:1. Lack of accumulation of deoxyinosine in the purine nucleoside phosphorylase (PNP) deficient line, or hypoxanthine in the hypoxanthine guanine phosphoribosyltransferase (HGPRT) deficient line, using dAR together with the adenosine deaminase (ADA) inhibitor 2'-deoxycoformycin (dCF) at 10 microM, confirmed the effectiveness of ADA inhibition. Nevertheless, some ATP was still formed by all lines in the presence of dCF by a route as yet unknown. Only the ADA deficient lines formed dATP with dAR alone. However, some dATP was formed by all lines in the presence of dCF. A partially HGPRT deficient line formed extremely high dATP levels, well in excess of those formed by the T-cell line CEM. Studies with dGR revealed some interesting differences, a large proportion of the substrate being metabolized predominantly to xanthine by most enzyme deficient lines. In the PNP deficient line most of the substrate remained unmetabolized, but some dGTP was formed. No other enzyme deficient line formed any dGTP--with or without the PNP inhibitor 8-aminoguanosine (8-NH2GR)--with one exception. Again this was the partially HGPRT deficient line, which with the inhibitor again formed more dGTP than the T-cell line. Within the cells most of the substrate was metabolized to GTP, except in the PNP, and totally HGPRT deficient lines. Levels of GTP formed were not altered by the inhibitor, reflecting the lack of effective PNP inhibition by 8-NH2GR. Some counts were also found in ATP and IMP, confirming the existence of this route in mammalian cells of lymphoid origin. The results also support previous studies by us using cell lines with intact purine pathways, which demonstrated that, contrary to current beliefs, some B-cell lines are capable of accumulating high levels of deoxynucleotides.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Deaminase Inhibitors↗