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Biomedical subjects

M J Bennett

Publications and source records attributed to M J Bennett.

At least 19 recordsLinked to original sources
PubMed1992

Identification of urinary metabolites of (+/-)-2-(p-isobutylphenyl)propionic acid (Ibuprofen) by routine organic acid screening.

Ibuprofen [(+/-)-2-(p-isobutylphenyl)propionic acid] has recently been introduced as a pediatric anti-inflammatory agent. To determine how this agent interferes with urine organic acid analysis, an important pediatric investigation, we have analyzed urine from two subjects pre- and post-Ibuprofen dosage and two subjects on chronic Ibuprofen therapy. A distinctive pattern of drug interference on the organic acid profile was detected. There were three major components, corresponding to unmetabolized Ibuprofen and to the oxidation products hydroxy Ibuprofen and carboxy Ibuprofen. Therefore, the major mechanism of Ibuprofen metabolism appears to be microsomal, although mitochondrial and peroxisomal routes cannot be excluded. Laboratories carrying out routine organic acid analysis should be aware of the nature and magnitude of the organic aciduria caused by Ibuprofen.

Adult↗
PubMed1992

Synthesis and biological activity of CCK heptapeptide analogues. Effects of conformational constraints and standard modifications on receptor subtype selectivity, functional activity in vitro, and appetite suppression in vivo.

A series of modifications of the CCK7 analogue (des-NH2)Tyr(SO3-)-Nle-Gly-Trp-Nle-Asp-Phe-NH2 was prepared and tested for binding to guinea pig CCK-A and CCK-B receptors and in CCK-A-mediated functional assays. Selected analogues also were tested for appetite suppressant activity in rats. Several conformationally restricted residues in the C-terminal tetrapeptide region, including delta Z-Phe33, (N-Me)Phe33, (N-Me)Asp32, (N-Me)Leu31, and 3PP31 (3PP = trans-3-n-propyl-L- proline) were found to be acceptable modifications at one or both receptor subtypes. The (N-Me)Asp32 and (N-Me)Leu31 modifications afforded potent and selective CCK-A and CCK-B ligands, respectively. SAR studies in the N-terminal acyldipeptide region examined structural requirements for the side chain at position 28, where Gly and Pro replacements were found to possess high affinity at both receptor subtypes. Other conformationally restrictive modifications were less active. All of the analogues that showed high affinity (less than 10 nM) for the CCK-A receptor also were full agonists in amylase release and most were full or nearly full agonists in the phosphoinositide (PI) turnover assay, the most notable exception being the delta Z-Phe33 analogue, which showed 69% of the maximal response in the PI assay. Potent activity in suppression of food intake in rats was found for selected analogues.

Amino Acid Sequence↗
PubMed1992

The crystal structure of diphtheria toxin.

The crystal structure of the diphtheria toxin dimer at 2.5 A resolution reveals a Y-shaped molecule of three domains. The catalytic domain, called fragment A, is of the alpha + beta type. Fragment B actually consists of two domains. The transmembrane domain consists of nine alpha-helices, two pairs of which are unusually apolar and may participate in pH-triggered membrane insertion and translocation. The receptor-binding domain is a flattened beta-barrel with a jelly-roll-like topology. Three distinct functions of the toxin, each carried out by a separate structural domain, can be useful in designing chimaeric proteins, such as immunotoxins, in which the receptor-binding domain is substituted with antibodies to target other cell types.

Bacterial Toxins↗
PubMed1992

Selective cleavage of closely-related mRNAs by synthetic ribozymes.

In Phaseolus vulgaris L. (French bean) glutamine synthetase (GS) is encoded by four closely-related genes termed gln-alpha, gln-beta, gln-gamma and gln-delta. We have constructed and characterised in vitro a number of hammerhead ribozymes designed to cleave individual RNAs encoded by these genes. The three ribozymes, termed J1, J2 and J3, were targeted to cleave RNA at the start of the gamma and beta, and the middle of the gamma, GS open reading frames respectively. All three ribozymes successfully discriminated between the four (alpha, beta, gamma and delta) highly homologous sequences, even though the targeted sites of cleavage shared up to 18 out of 22 identical bases with other gene family members. The ribozyme-mediated cleavage reactions were Mg2+ dependent and enhanced at higher temperatures, although the J1 ribozyme retained considerable activity at physiological temperatures. Both J1 and J2 demonstrated a time-dependent cleavage of their targeted GS RNAs, although these two ribozymes differed markedly in their ability to cleave multiple substrate molecules. The rate of cleavage by J1 was found to be reduced in the presence of related GS RNAs and by total leaf poly(A) RNAs. The implications of these results for ribozyme activity in vivo are discussed.

Amino Acid Sequence↗
PubMed1992

Azelaic and pimelic acids: metabolic intermediates or artefacts?

Azelaic and pimelic acids are excreted in elevated amounts in urine in disorders of mitochondrial beta-oxidation and disorders of peroxisomal beta-oxidation, for which they are of significant diagnostic value. We have detected the presence of azelaic, pimelic and even-chain-length dicarboxylic acids (adipic, suberic and sebacic acids) arising artefactually as a result of storage of small sample volumes in plastic containers. Storage of samples for organic acid analysis in glass containers is recommended.

Adipates↗
PubMed1992

3-Methylglutaconic aciduria associated with Pearson syndrome and respiratory chain defects.

3-Methylglutaconic aciduria was detected in four patients with Pearson syndrome, a multitissue disorder with hematologic abnormalities, lactic acidosis resulting from defective oxidative phosphorylation, and deletions in the mitochondrial genome. 3-Methylglutaconic acid may be an additional useful marker for Pearson syndrome and may be a more specific marker than other organic acids identified in this disorder.

Acidosis, Lactic↗
PubMed1992

Determinants of fasting plasma triglyceride levels: metabolism of hepatic and intestinal lipoproteins.

The contribution of the kinetics of exogeneous and endogenous lipoproteins in determining the level of triglyceride in fasting plasma was assessed in a group of 19 normolipidaemic and hypertriglyceridaemic subjects. From data derived during a 9-h infusion of [15N]-glycine, we have assessed very low density lipoprotein apolipoprotein B production, and from data analysed by kinetic modelling obtained following ingestion of retinol and triolein, we have assessed chylomicron and chylomicron remnant clearance in a group of 19 normolipidaemic and hypertriglyceridaemic subjects. A strong positive correlation was observed between the fasting plasma triglyceride level and the reciprocal of the apolipoprotein B fractional synthetic rate (r = 0.83, P less than 0.01). A positive correlation was also found with the rate of clearance of chylomicron remnants (Sf 20-400; r = 0.87, P less than 0.01) and of chylomicrons (Sf greater than 400; r = 0.69, P less than 0.01). No correlation was found between the fasting plasma triglyceride level and either of the plasma post-heparin lipolytic activities. Multivariate analysis revealed that 95% of the variance in triglyceride levels could be explained by the apolipoprotein B fractional synthetic rate and the chylomicron remnant clearance rate. The strong correlation between chylomicron remnant clearance, a measure of exogenous lipid metabolism, and fasting (hence, endogenous) plasma triglyceride levels suggests that remnants of chylomicrons and very low density lipoproteins share some common components of the removal process.

Adult↗
PubMed1992

Comparison of post-mortem urinary and vitreous humour organic acids.

We have analysed organic acid profiles in 74 samples of post-mortem vitreous humour from the sudden infant death syndrome and compared the profiles to those obtained from the corresponding urine or bladder wall swab. There was a high degree of correlation indicating that vitreous humour analysis in high-risk infants is an appropriate analytical strategy when urine is not available. In our patient sample two infants had evidence of abnormal methylmalonic acid metabolism, one had glyceric aciduria (and elevated levels of vitreous humour glyceric acid), one had evidence of pre-existing liver damage as judged by the presence of 4-hydroxyphenyllactic acid, one had a non-ketotic dicarboxylic aciduria indicating inhibited fatty acid oxidation and two patients had significant long-chain 3-hydroxydicarboxylic acids and evidence of paracetamol ingestion.

Carboxylic Acids↗
PubMed1992

Medium chain acyl-coenzyme A dehydrogenase deficiency.

Medium chain acyl-coenzyme A dehydrogenase deficiency (MCADD) was the first metabolic disorder found to be associated with sudden infant death syndrome. This review covers recent advances in the biochemical and molecular understanding of MCADD.

Acyl-CoA Dehydrogenase↗
PubMed1992

Detection and quantitation of acylcarnitines in plasma and blood spots from patients with inborn errors of fatty acid oxidation.

Acylcarnitine profiling in plasma and dried blood spots by radioisotopic exchange/HPLC demonstrates that MCAD deficiency can be reliably detected in the asymptomatic state without L-carnitine therapy. The OC/AcC ratio differentiates MCAD deficiency from normal controls. A longer chain acylcarnitine (r.t. 43 min.) was detected in all 3 patients with a defect in long chain fatty acid oxidation. Detection of C4- and C5-acylcarnitine isomers in plasma helped characterize a metabolic defect affecting branched chain acyl-CoA oxidation in 3 patients. Quantitative data in 2 patients with MCAD deficiency showed that plasma concentrations of OC and AcC are dependent on both the availability of free carnitine and the severity of metabolic decompensation.

Acyl-CoA Dehydrogenase↗
PubMed1992

When do gut flora in the newborn produce 3-phenylpropionic acid? Implications for early diagnosis of medium-chain acyl-CoA dehydrogenase deficiency.

Urinary excretion of 3-phenylpropionylglycine (PPG) is a diagnostic marker for medium-chain acyl-CoA dehydrogenase (MCAD) deficiency. PPG is derived from 3-phenylpropionic acid (PPA), a product of anaerobic bacterial metabolism in the gut. To determine when the infant gut was colonized with PPA-producing bacteria, we cultured stool in prereduced thioglycollate broth from 93 apparently healthy infants. We analyzed the products of bacterial metabolism by gas chromatography/mass spectrometry for the presence of PPA. Trend analysis demonstrated a significant difference (P less than 0.001) in PPA production between early and later infancy. PPA was not detected in 84% of media isolated from stool collected from infants younger than four months. For older infants, 67% of the samples were PPA-positive. Thus, because the normal gut is not sufficiently colonized with PPA-producing bacteria before three to four months of age, PPG analysis alone is not a sensitive marker for the early detection of MCAD deficiency. Using stable isotope dilution mass spectrometry, we measured PPG and n-hexanoylglycine (HG) excretion in two well newborns with MCAD deficiency. HG, believed to be an endogenous metabolite associated with MCAD deficiency, was consistently above normal in all urine samples.

Acyl-CoA Dehydrogenases↗
PubMed1991

Crystallization of diphtheria toxin.

Two new crystal forms (forms III and IV) have been grown of diphtheria toxin (DT), which kills susceptible cells by catalyzing the ADP-ribosylation of elongation factor 2, thereby stopping protein synthesis. Forms III and IV diffract to 2.3 A and 2.7 A resolution, respectively. Both forms belong to space group C2; the unit cell parameters for form III are a = 107.3 A, b = 91.7 A, c = 66.3 A and beta = 94.7 degrees and those for form IV are a = 108.3 A, b = 92.3 A, c = 66.1 A and beta = 90.4 degrees. Both forms have one protein chain per asymmetric unit with the dimeric molecule on a twofold axis of symmetry. Form IV is exceptional among all crystal forms of DT in that it can be grown reproducibly. Thus the form IV crystals should yield a crystallographic structure giving insight into the catalytic, receptor-binding and membrane-insertion properties of DT.

Crystallization↗
PubMed1991

Glutaric aciduria type I: a common cause of episodic encephalopathy and spastic paralysis in the Amish of Lancaster County, Pennsylvania.

We have diagnosed type I glutaric aciduria (GA-I) in 14 children from 7 Old Order Amish families in Lancaster County, Pennsylvania. An otherwise rare disorder, GA-I appears to be a common cause of acute encephalopathy and cerebral palsy among the Amish. The natural history of the disease, which was previously unrecognized in this population, is remarkably variable and ranges from acute infantile encephalopathy and sudden death to static extrapyramidal cerebral palsy to normal adult. Ten patients first manifested the disease between 3 and 18 months at the time of an acute infectious illness. Four of these children died in early childhood, also during acute illnesses. However, there has been little progression of the neurological disease after age 5 years in the surviving children and intellect usually has been preserved, even in children with severe spastic paralysis. When well, patients have plasma glutaric acid concentrations ranging from 4.8 to 14.2 mumol/liter (nl 0-5.6 mumol/liter) and urinary glutaric acid concentrations from 12.5 to 196 mg/g creatinine (nl 0.5-8.4 mg/g creatinine). We have found that GA-I can be diagnosed in the Amish by measurement of urinary glutaric acid concentrations using isotope-dilution gas chromatography/mass spectrometry, whereas the diagnosis can easily be missed by routine urine organic acid gas chromatography.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute Disease↗
PubMed1991

Familial combined hyperlipidaemia: use of stable isotopes to demonstrate overproduction of very low-density lipoprotein apolipoprotein B by the liver.

We have assessed very low-density lipoprotein apolipoprotein B production, using [15N]glycine as an endogenous marker in a 9-hour primed constant infusion protocol, in four adult male subjects with familial combined hyperlipidaemia and in four normolipidaemic adult male controls. The mean very low-density lipoprotein apolipoprotein B absolute synthetic rate was significantly greater in the familial combined hyperlipidaemic subjects than in control subjects (26.31 +/- 8.37 vs 9.36 +/- 4.07 mg/kg per 24 h, p less than 0.05). These results confirm findings using exogenous radioisotope labelling techniques that very low-density lipoprotein apolipoprotein B production is significantly increased in most patients with familial combined hyperlipidaemia. A modified 9-hour protocol can be safely done repeatedly and in children and pregnant women.

Adult↗