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Biomedical subjects

R J Evans

Publications and source records attributed to R J Evans.

At least 19 recordsLinked to original sources
PubMed1995

Mapping receptor binding sites in interleukin (IL)-1 receptor antagonist and IL-1 beta by site-directed mutagenesis. Identification of a single site in IL-1ra and two sites in IL-1 beta.

Interleukin-1 receptor antagonist (IL-1ra), an IL-1 family member, binds with high affinity to the type I IL-1 receptor (IL-1RI), blocking IL-1 binding but not inducing an IL-1-like response. Extensive site-directed mutagenesis has been used to identify residues in IL-1ra and IL-1 beta involved in binding to IL-1RI. These analyses have revealed the presence of two discrete receptor binding sites on IL-1 beta. Only one of these sites is present on IL-1ra, consisting of residues Trp-16, Gln-20, Tyr-34, Gln-36, and Tyr-147. Interestingly, the absent second site is at the location of the major structural difference between IL-1ra and IL-1 beta, which are otherwise structurally similar. The two receptor binding sites on IL-1 beta are also present on IL-1 alpha. Thus, it appears that the two IL-1 agonist molecules have two sites for IL-1RI binding, and the homologous antagonist molecule, IL-1ra, has only one. Based on these observations, a hypothesis is presented to account for the difference in activity between the agonist and antagonist proteins. It is proposed that the presence of the two receptor binding sites may be necessary for agonist activity.

Amino Acid Sequence↗
PubMed1995

Fatal acute pancreatitis caused by valproic acid.

We describe a case of a 31-year-old woman with cerebral palsy who developed fatal acute hemorrhagic pancreatitis while being treated with valproic acid to control her seizure activity. Acute pancreatitis is usually due to alcohol ingestion or biliary tract disease, and unusual causes include trauma, metabolic diseases, or drugs. Valproic acid is considered a safe drug, although rare cases of severe toxicity such as hepatitis and acute pancreatitis, including two fatalities, have been reported. Our review of the literature revealed that most patients who developed acute pancreatitis had serum levels of the drug within the therapeutic range, and most of the cases occurred either secondary to a recent increase in the dose or to initiation of treatment. It also appeared that the fatalities occurred due to a delayed diagnosis of acute pancreatitis, either resulting from an unsuspected diagnosis or to the deteriorated mental status of the patients receiving the drug, which precluded their ability to elaborate symptomatology. We believe that early diagnosis and withdrawal of the drug are significant factors determining the course of valproic-acid-associated pancreatitis.

Acute Disease↗
PubMed1995

Long-term inhibition of murine experimental autoimmune encephalomyelitis using CTLA-4-Fc supports a key role for CD28 costimulation.

T cell activation involves not only recognition of antigen presented by the MHC, but also nonspecific interactions termed "costimulation." The costimulatory molecules B7-1 and B7-2 are ligands on antigen-presenting cells for the CD28 and CTLA-4 receptors on T cells. Previously, a fusion protein consisting of human CTLA-4 linked to human Fc was shown to bind B7-1 and B7-2 with high avidity and to prevent specific T cell activation. Here we investigated the effects of a recombinant fusion protein consisting of the extracellular domain of human CTLA-4 bound to mouse IgG2a Fc (CTLA-4-Fc) upon experimental autoimmune encephalomyelitis, a T cell-mediated disease that serves as a model for multiple sclerosis. CTLA-4-Fc prevented experimental autoimmune encephalomyelitis in 26 of 28 CTLA-4-Fc-treated mice (median maximum score 0), whereas 28 of 30 mice treated with control mouse IgG2a developed disease (median maximum score 2.75). Less inflammation and virtually no demyelination or axonal loss occurred in CTLA-4-Fc-treated compared with control-treated mice. Activated splenocytes from CTLA-4-Fc-treated mice were able to transfer disease adoptively to naive recipients. These results indicate a key role for the B7/CD28 system in the development of actively induced murine experimental autoimmune encephalomyelitis, suggesting an area of investigation with therapeutic potential for multiple sclerosis.

Abatacept↗
PubMed1995

Pharmacological characterization of heterologously expressed ATP-gated cation channels (P2x purinoceptors).

cDNAs encoding P2x purinoceptors from human bladder smooth muscle and from rat PC-12 cells were expressed in oocytes and human embryonic kidney 293 cells. Agonist potencies of 2-methylthio-ATP = 2-chloro-ATP = ATP > = 2'- and 3'-O-(4-benzoylbenzoyl)-ATP > or = adenosine-5'-O-(3-thio)-triphosphate > or = P1,P5-di(adenosine-5') pentaphosphate >> ADP prevailed for both P2x purinoceptors. There were two main differences in agonist sensitivity between the two receptors. First, ATP was 10 times more potent at the receptor from bladder (EC50, 0.8 microM) than at the receptor from PC-12 cells (EC50, 8.2 microM). Second, alpha,beta-methylene-ATP and L- and D-beta,gamma-methylene-ATP were agonists in cells expressing the bladder smooth muscle receptor (EC50, 1-3 microM) but were ineffective in cells expressing the PC-12 receptor. The P2 purinoceptor antagonists suramin, pyridoxal phosphate 6-azophenyl-2',4'-disulfonic acid, and pyridoxal-5-phosphate acted similarly at both receptor forms, producing noncompetitive inhibition, with IC50 values of 1-5 microM for suramin and pyridoxal phosphate 6-azophenyl-2',4'-disulfonic acid and 10-20 microM for pyridoxal-5-phosphate. 4,4'-Diisothiocyanatostilbene-2,2'-disulfonic acid distinguished receptor subtypes, producing potent inhibition of the bladder smooth muscle P2x-mediated response, with an IC50 value of 3 microM; it inhibited the PC-12 form by < 40% at 100 or 300 microM. This study thus defines the pharmacological properties of homo-oligomeric forms of these two types of cloned P2x receptor channels.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1994

A new class of ligand-gated ion channel defined by P2x receptor for extracellular ATP.

Extracellular ATP exerts its effects through P2 purinoceptors: these are ligand-gated ion channels (P2x) or G-protein-coupled receptors (P2Y, P2U). ATP at P2x receptors mediates synaptic transmission between neurons and from neurons to smooth muscle, being responsible, for example, for sympathetic vasoconstriction in small arteries and arterioles. We have now cloned a complementary DNA encoding the P2x receptor from rat vas deferens and expressed it in Xenopus oocytes and mammalian cells. ATP activates a cation-selective ion channel with relatively high calcium permeability. Structural predictions suggest that the protein (399 amino acids long) is mostly extracellular and contains only two transmembrane domains plus a pore-forming motif which resembles that of potassium channels. The P2x receptor thus defines a new family of ligand-gated ion channels.

Adenosine Triphosphate↗
PubMed1994

Alexithymia in somatoform disorder patients with chronic pain.

The prevalence of alexithymia in fifty-five motor vehicle accident survivors who had chronic pain complaints and met DSM-III-R criteria for somatoform pain disorder was 53%. Alexithymic and non-alexithymic patients did not differ on self-reports of current pain severity or in the number of pain locations. Alexithymic patients were found to use significantly more words to describe their pain, suggesting they may have a more diffuse style in communicating their pain experience.

Accidents, Traffic↗
PubMed1994

Endotoxin-induced platelet aggregation in heparinised equine whole blood in vitro.

Endotoxaemia is a leading cause of death among horses. Thrombocytopenia is a common finding in clinical and experimentally-induced cases of endotoxaemia and can lead to coagulopathies, including disseminated intravascular coagulopathy which is usually fatal. In this study it was shown that endotoxin (3 ng ml-1 to 25 micrograms ml-1) can aggregate equine platelets in heparinised whole blood in vitro. The endotoxin-induced aggregation (EIA) was shown to be dependent on the presence of leucocytes in the blood and did not occur when detoxified endotoxin was used, suggesting that lipid A was necessary for the response. Aspirin (1 mmol litre-1) had no effect on EIA whereas apyrase (40 micrograms ml-1) completely abolished it and CV3988 (3 to 30 mumol litre-1) (a competitive antagonist of platelet-activating factor) inhibited the response in a concentration-dependent manner. It is concluded that endotoxin activates equine platelets at low concentrations through an indirect mechanism that involves calcium, leucocytes, adenine nucleotides and platelet-activating factor.

Animals↗
PubMed1994

Morphological properties and projections of electrophysiologically characterized neurons in the guinea-pig submucosal plexus.

Intracellular recordings were made from 73 guinea-pig submucosal neurons using neurobiotin-filled microelectrodes; subsequently, neuropeptide immunoreactivity, morphology and nerve fibre projections were determined. Five distinct groups of cells could be distinguished: S cells with inhibitory input (61%), S cells without inhibitory input (19%), AH cells (8%), S-AH cells (5%), and glial networks. S cells with inhibitory input were immunoreactive for vasoactive intestinal polypeptide and showed Dogiel Type III morphology with the axon branching and coursing through two to 12 ganglia; varicosities and tufts of varicosities were observed surrounding other cell bodies as well as over blood vessels. S cells without inhibitory input primarily were immunoreactive for neuropeptide Y; they also showed Dogiel Type III morphology and similar, though shorter, axonal projections and varicose features surrounding other neurons. AH cells, which most likely contained substance P, lacked synaptic input and exhibited Dogiel Type II morphology; they branched more extensively than S cells and also formed varicose tufts within other ganglia. S-AH cells combined electrophysiological properties of S cells with inhibitory input and AH cells and did not show consistent morphological or histochemical characteristics. Typical glial networks were observed; in addition, on two occasions unusual networks of dye and electrical coupling between S cells without inhibitory input and a glial complex were observed. These results suggest that vasoactive intestinal polypeptide-containing S cells may act as interneurons which mediate a slow excitatory synaptic potential; that neuropeptide Y-containing S cells, which are known to be cholinergic, may play a role as cholinergic interneurons mediating the nicotinic fast excitatory synaptic potential; and that AH neurons also may provide cholinergic innervation to other submucosal neurons in addition to their previously described dual projections into mucosa and myenteric plexus.

Action Potentials↗
PubMed1994

The effects of aspirin and paracetamol on the aggregation of equine blood platelets.

The responses of equine blood platelets in citrated platelet-rich plasma to arachidonic acid, U44069 (prostaglandin endoperoxide analogue), adenosine 5'-diphosphate, platelet-activating factor or collagen were investigated by turbidimetric aggregometry. Pre-treatment of the platelets with aspirin (1 mmol/l) or paracetamol (1.3 mmol/l) abolished shape change and aggregation in response to arachidonic acid; decreased the rate of aggregation in response to collagen, with no separate effect on shape change; had no marked effect on aggregation caused by the other agonists; but in no case transformed irreversible aggregation to reversible aggregation. We conclude that thromboxane A2 generation is of minor importance in the aggregation of equine platelets, and in particular that thromboxane A2 is not a significant mediator of irreversible aggregation.

Acetaminophen↗
PubMed1994

Characterization of P2-purinoceptors in the smooth muscle of the rat tail artery: a comparison between contractile and electrophysiological responses.

1. The electrophysiological actions of the P2-purinoceptor agonists, adenosine 5'-triphosphate (ATP), 2-methylthioATP (2-meSATP), alpha, beta-methyleneATP (alpha, beta-meATP) and uridine 5'-triphosphate (UTP) were studied under concentration and voltage-clamp conditions in acutely dissociated rat tail artery smooth muscle cells. For comparison, their actions as vasoconstrictors were studied in intact ring preparations. 2. Rapid application of ATP (100 nM-1 microM) via a U-tube superfusion system activated concentration-dependent inward currents with a latency to onset of less than 3 ms. The inward current decayed by more than 95% during a 2 s application of 300 nM and 1 microM ATP. 3. 2-meSATP (100 mM-1 microM) and alpha, beta-meATP (100 nM-1 microM) also evoked transient inward currents. The agonist order of potency was ATP = 2-meSATP > or = alpha, beta-meATP. UTP (300 nM-1 microM) did not produce a change in the holding current. 4. A second application of ATP (300 nM and 1 microM) 10 min after the first, evoked currents which were one third of the initial amplitude. This decline was dependent upon activation of the P2-purinoceptor. Similar results were seen with 2-meSATP and alpha, beta-meATP (both 300 nM and 1 microM). Cross-desensitization was seen between ATP and 2-meSATP or alpha, beta-meATP. 5. Inward currents evoked by ATP, 2-meSATP and alpha, beta-meATP (all 1 microM) were abolished by the P2-purinoceptor antagonist suramin (100 microM). 6. Alpha, beta-meATP (100 nM-30 micro M), 2-meSATP (3 micro M- 100 micro M), ATP (3 micro M-I mM) and UTP (3 ELM-I mM)produced concentration-dependent contractions of rat tail artery rings. When measured at a level equal to 50% of the maximum response to noradrenaline, the rank order of agonist potency was alpha,beta-meATP>>2-meSATP >UTP >ATP.7. This study shows that the rank order of agonist potency at the P2X-purinoceptor which mediates contractions of the rat isolated tail artery is very different from the potency order for evoking the inward current which initiates the contractions. It is concluded that this difference may be due to the relative absence of breakdown of some of the agonists in the single cell system compared with artery rings.

Adenosine Triphosphate↗
PubMed1993

Effects of phospholipase A2 inhibitors on coupling of alpha 2-adrenoceptors to inwardly rectifying potassium currents in guinea-pig submucosal neurones.

1. Noradrenaline hyperpolarizes guinea-pig submucosal neurones by opening inwardly rectifying potassium channels. Intracellular recordings were made from submucosal neurones and the possible involvement of the phospholipase A2 pathway in this response was examined. 2. The non-specific phospholipase A2 inhibitors, quinacrine (10 microM) and 4-bromophenacyl bromide (4-BPB, 10 microM) inhibited nerve-evoked inhibitory synaptic potentials (i.p.s.ps) and hyperpolarizations to somatostatin and UK 14304. Quinacrine and 4-BPB also blocked the inward rectification present in current-voltage curves in the absence of somatostatin or UK 14304. 3. The more selective phospholipase A2 inhibitor, cyclosporin A (10 microM) and the lipoxygenase and cyclo-oxygenase inhibitor, eicosatetraynoic acid (ETYA, 20 microM) and nordihydroguairetic acid (NDGA, 20 microM) did not alter i.p.s.ps or hyperpolarizations to UK 14304. 4. Exogenously applied arachidonic acid (1-300 microM) did not mimic the i.p.s.p. or the hyperpolarization to UK 14304. 5. We conclude that arachidonic acid or its eicosanoid metabolites produced by phospholipase A2 stimulation are unlikely to be involved in the receptor G-protein coupled activation of potassium currents in submucosal neurones. The inhibition of the noradrenaline-induced hyperpolarization by quinacrine and 4-BPB is most likely due primarily to blockade of the basal inwardly rectifying potassium conductance present in these neurones.

Acetylcholine↗
PubMed1993

Intracranial haemorrhage associated with ingestion of 'ecstasy'.

A case of a patient with intracranial haemorrhage thought to have been associated with ingestion of 'Ecstasy' [3-4 methylenedioxymethamphetamine (MDMA)] is presented. The case illustrates the importance of drug analysis in cases involving illicit drug use.

3,4-Methylenedioxyamphetamine↗
PubMed1993

Potassium currents and their modulation by muscarine and substance P in neuronal cultures from adult guinea pig celiac ganglia.

1. Intracellular microelectrode and whole-cell patch-clamp recordings were obtained from adult guinea pig celiac ganglion neurons grown in tissue culture for 7-14 days. Over 90% of neurons showed phasic-type action-potential discharge with the use of either type of recording electrode; they stained immunohistochemically for catecholamines, tyrosine hydroxylase, and neuropeptide Y. Input resistance (140 M omega) and action-potential amplitude (103 mV) were significantly greater with whole-cell than with microelectrode recordings, but other passive electrical properties were similar. 2. Five potassium currents were characterized: an apamin-sensitive after hyperpolarizing current (IAHP), an apamin and tetraethylammonium-insensitive slow IAHP, an M-like current, a transient outward IA current, and a delayed rectifier IK current. A hyperpolarization-activated cationic Ih current was also present. The first three currents were not observed with whole-cell recordings. 3. Cadmium (200 microM), cobalt (1 mM), lanthanum (30 microM), or a low calcium/high magnesium solution blocked both IAHPS and the M-like current; barium (1 mM) also blocked these currents. 4. Kinetics of the M-like current were best described by a double exponential fit to deactivating tail currents with time constants of 50 and 390 ms at -50 mV. The apamin-sensitive and slow IAHP decayed exponentially with time constants of 145 ms and 3.5 s, respectively. There was no correlation between occurrence of M-like current (95% of neurons) and slow IAHP (40% of neurons), nor any correlation between magnitude of M-like current and IAHP in those cells exhibiting both currents. 5. Muscarine and substance P (SP) caused depolarizations or inward currents (under voltage clamp) at the resting potential (-55 mV) associated with a decreased membrane conductance. The slow IAHP and the M-like current, but not the apamin-sensitive IAHP nor the IA, were blocked by muscarine and SP (IC50 3 microM and 100 nM, respectively). Muscarine and SP also decreased a "leak" potassium current. 6. We conclude that celiac neurons express two calcium-dependent IAHP currents and a calcium-dependent M-current; these are seen by fine-tipped intracellular microelectrodes but not by whole-cell patch electrodes. These currents are not required for spike frequency accommodation. Muscarine and SP reduce these currents, as well as voltage-independent leakage potassium current.

Animals↗
PubMed1992

ATP mediates fast synaptic transmission in mammalian neurons.

In addition to its diverse functions inside cells, ATP can act at several types of cell-surface receptor. One of these (P2X-purinoceptor) is believed to be a ligand-gated cation channel. The presence of P2X receptors on autonomic, sensory and central neurons suggests that ATP might be released to act as a fast excitatory synaptic transmitter. Here we record excitatory synaptic potentials and currents from cultured coeliac ganglion neurons which are mimicked by ATP, blocked by the P2-purinoceptor antagonist suramin, desensitized by alpha,beta-methylene-ATP and unaffected by antagonists acting at nicotine, 5-hydroxytryptamine, N-methyl-D-aspartate (NMDA), non-NMDA glutamate, gamma-aminobutyric acid (GABA), noradrenaline or adenosine receptors. We conclude that ATP is the neurotransmitter at this neuroneuronal synapse.

Action Potentials↗