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Biomedical subjects

R Gill

Publications and source records attributed to R Gill.

At least 73 records · Page 4Linked to original sources
PubMed1995

The neuroprotective effect of the glycine site antagonist 3R-(+)-cis-4-methyl-HA966 (L-687,414) in a rat model of focal ischaemia.

3R-(+)-cis-4-Methyl-HA966 (L-687,414) is a novel and selective, low intrinsic activity, partial agonist at the glycine site of the N-methyl-D-aspartate (NMDA) receptor. Thus, while it acts primarily to block NMDA receptor function in the presence of glycine, it fails to produce a complete block of NMDA receptor activation. In this study, we have investigated its neuroprotective effects in a rat model of focal ischaemia, involving permanent occlusion of the left middle cerebral artery. L-687,414 was administered as a bolus dose of 17.6 mg/kg i.v. straight after the occlusion or as a bolus dose + infusion for 4 h. The doses of L-687,414 used for the infusion studies were 7 mg/kg i.v. + 7 mg/kg/h, 14 mg/kg + 14 mg/kg/h, or 30 mg/kg + 30 mg/kg/h. The 17.6-mg/kg dose gave an estimated peak plasma level of 24 micrograms/ml, which decayed with a t1/2 of 56 min. The three infusion dosing regimens gave mean plasma levels over the 4 h of 11, 25 and 61 micrograms/ml plasma, respectively. The 17.6-mg/kg dose of L-687,414 gave no significant protection against the volume of hemispheric, cortical, or caudate damage when compared with the control group of animals. The lowest infusion dosing regimen of L-687,414 which gave a plasma level of 11 micrograms/ml over the 4 h was also ineffective against the volume of infarction measured in the different brain regions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Activation of natural killer cells by mouse mammary tumor virus C4 in BALB/c and T-cell receptor V beta 2-transgenic mice.

Lymphocytic infiltrates of BALB/c C4 hyperplastic alveolar nodule (HAN) have elevated natural killer (NK) activity, which correlates positively with the progression of C4 HAN to tumor: C4 HAN produces an infectious mouse mammary tumor virus, MMTV(C4), which encodes a superantigen that activates and deletes T-cells with the V beta 2 segment in the T-cell receptor. In this report, NK activation by both MMTV(C4) and MMTV(C4) superantigen was tested. NK activity was measured in naive BALB/c mice, BALB/c mice depleted of V beta 2+ T-cell, or V beta 2-transgenic mice after they received injections of either purified MMTV(C4) or MMTV(C4)-infected splenocytes. Elevated NK activity was observed in BALB/c mice receiving MMTV(C4) or MMTV(C4)-infected splenocytes. Depletion of V beta 2+, but not V beta 8+, T-cells by specific anti-V beta hybridoma before injection of MMTV(C4)-infected cells reduced but did not eliminate NK activation. NK activation in V beta 2-transgenic mice occurred before massive CD4 T-cell deletion took place and was more pronounced than that in the nontransgenic littermates. These results indicate that MMTV activates NK cells through superantigen-dependent and -independent pathways and supports the role of MMTV(C4) in the augmented NK activity observed in C4 HAN infiltrates. The progression of C4 HAN to tumor represents a model system for the analysis of how tumorigenesis may be affected by lesion-associated viruses.

Animals↗
PubMed1994

The neuroprotective effects of the decahydroisoquinoline, LY 215490; a novel AMPA antagonist in focal ischaemia.

LY 215490 (3RS,4aRS,6RS,8aRS)-6-[2-(1(2)H-tetrazole-5- yl)ethyl]decahydroisoquinoline-3-carboxylic acid), a novel, selective, competitive and systemically active AMPA receptor antagonist was tested as a neuroprotective agent against focal ischaemia in a model of permanent MCA occlusion in the rat. LY 215490 was administered at a dose of 10, 30 or 100 mg/kg 30 min prior to and post-MCA occlusion. The animals were allowed to survive for 24 hr, following which time the brains were processed for volumetric analysis of the infarct size. The low dose of LY 215490 was not effective against the infarct volume in the hemisphere, cortex or caudate. The 2 x 30 mg/kg dose of LY 215490 resulted in 25 and 31% protection against the volume of hemispheric and cortical ischaemic damage, respectively. The highest dose of LY 215490 resulted in a reduced neuroprotective effect with 23 and 27% protection against the volume of hemispheric and cortical ischaemic damage, respectively. The slightly reduced neuroprotective effect of the highest dosing regimen may be due to the respiratory problems seen with this dose. Neither of the two neuroprotective doses of LY 215490 produced any reduction in the volume of caudate damage which represents the core of the infarct.

Analysis of Variance↗
PubMed1994

The pharmacology of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA)/kainate antagonists and their role in cerebral ischaemia.

The development of selective, systemically active alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA)/kainate antagonists over the last 4 years has enabled the role of this excitatory amino acid receptor subtype to be scrutinised in the different models of ischaemia. The animal models of cerebral ischaemia can be subdivided into two major categories: focal ischaemia, in which the resulting infarct resembles the clinical condition of stroke; and models of severe forebrain ischaemia, in which there is delayed neuronal degeneration of hippocampal CA1 neurones. The neuropathology in the latter models resembles the clinical condition seen following a cardiac arrest, for example. It is well established that N-methyl-D-aspartate (NMDA) antagonists such as MK-801, 3-(2-carboxypiperazine-4-yl)-propenyl-1-phosphonate (CPPene), DL-(E)-2-amino-4-methyl-5-phosphono-3-pentanoic acid (CGP 37849), and N-(1-naphthyl)-N'-(3-ethylphenyl)-N'-methylguanidine hydrochloride (CNS 1102) are neuroprotective in animal models of focal ischaemia. However, in models of severe forebrain ischaemia NMDA antagonists produced only partial protection. The discovery of 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX) as a systemically active AMPA receptor antagonist enabled the role of this receptor subtype in ischaemia to be investigated. NBQX was shown to be neuroprotective against delayed neuronal degeneration of hippocampal CA1 neurones in animal models of severe forebrain ischaemia. Recent studies have demonstrated that NBQX administration can be delayed by up to 12 h and amelioration of delayed neuronal degeneration of hippocampal CA1 neurones can still be seen. NBQX has also been shown to be neuroprotective in animal models of permanent and temporary middle cerebral artery occlusion. 1-(Aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine (GYKI 52466), a systemically active noncompetitive AMPA/kainate antagonist, was neuroprotective against focal ischaemia but was unable to attenuate hippocampal CA1 neuronal degeneration. Whilst the newer compounds such as (3SR,4aRS,6RS,8aRS)-6-[2-(1H-tetrazol-5-yl )-ethyl]-1,2,3,4,4a,5,6,7,8a-decahydroisoquinoline-3-carboxylic acid (LY 215490) and 6-(1-imidazolyl)-7-nitroquinoxaline-2,3(1H,4H)-dione (YM900) have been demonstrated to be neuroprotective in focal ischaemia models, there is still a lack of information with regard to their efficacy in models of severe forebrain ischaemia. It appears from initial studies that AMPA/kainate antagonists have a better behavioural profile than NMDA antagonists in terms of a lack of phychostimulant and phychotomimetic effects. However, these antagonists have their own problems in that they cause severe depression of glucose utilisation in the central nervous system at neuroprotective doses.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Cloning and expression of a rat neuronal nitric oxide synthase coding sequence in a baculovirus/insect cell system.

A DNA sequence encoding rat neuronal NO synthase (nNOS) was isolated and cloned into the baculovirus expression vector pVL1393 to generate pVLRBNOS. Transfection of Spodoptera frugiperda Sf-21 cells with the construct pVLRBNOS resulted in the synthesis of high levels of neuronal NO synthase. Analysis of the expression pattern revealed soluble, enzymatically active NO synthase in the cytoplasm of cell extracts. Active enzyme could also be purified from culture supernatants using 2'-5' ADP sepharose affinity chromatography. This enzyme was recognised by antibodies to the native nNOS and showed a similar degree of inhibition by arginine analogs as the native nNOS. The majority of the NOS synthesised had accumulated as insoluble "inclusion-body" material. The purification of recombinant nNOS from insect cells should facilitate characterisation of neuronal NO synthase.

Amino Acid Oxidoreductases↗
PubMed1992

The dose-response relationship and therapeutic window for dizocilpine (MK-801) in a rat focal ischaemia model.

The purpose of the present study was to examine the dose-response relationship and the maximum time for which effective therapy could be delayed for the N-methyl-D-aspartate antagonist dizocilpine (MK-801) as a neuroprotective agent in a permanent focal ischaemia model in the rat. The ED50 for dizocilpine in the amelioration of cortical damage in this model was found to be approximately 0.3 mg/kg (single i.p. dose, 30 min post onset of ischaemia) and significant protection was only obtained when therapy (3 mg/kg i.p.) was delayed for one hour or less after the onset of ischaemia. In a further experiment, dizocilpine 3 mg/kg i.p., produced a peak plasma level of 44 ng/ml and had a t1/2 elimination of 1.65 h.

Analysis of Variance↗
PubMed1992

The neuroprotective actions of 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX) in a rat focal ischaemia model.

The neuroprotective effects of NBQX, a selective antagonist for the AMPA/kainate subtype of excitatory amino acid receptors, were investigated in a rat focal ischaemia model, involving permanent occlusion of the left middle cerebral artery (MCA). NBQX (3, 10 or 30 mg/kg) was administered i.v. immediately after MCA occlusion and again 1 h later. The highest dose of NBQX (2 x 30 mg/kg) gave significant protection against hemispheric (24%) and cortical (27%) ischaemic damage. The lower doses of NBQX (2 x 3 or 2 x 10 mg/kg) were ineffective. No protection was seen against caudate damage for any of the doses of NBQX tested. NBQX has a t1/2 of 30 min, therefore, a second experiment was done in which a dose of 30 mg/kg was given as an i.v. bolus followed immediately by an infusion of 10 mg/kg/h for 4 h, dosing was started immediately after MCA occlusion. This dosing regimen resulted in a mean plasma level over the 4 h of 17 micrograms/ml, and significant protection against the volume of hemispheric (29%) and cortical (35%) ischaemic damage, which was slightly better than that achieved with two bolus doses of 30 mg/kg. Once again no protection was seen against caudate damage. We conclude that NBQX, an AMPA/kainate antagonist was neuroprotective in a focal ischaemia model in the rat.

Analysis of Variance↗
PubMed1992

Retention reproducibility of basic drugs in high-performance liquid chromatography on a silica column with a methanol-high-pH buffer eluent. Changes in selectivity with the age of the stationary phase.

The long-term reproducibility of the separation of basic drugs by high-performance liquid chromatography using a silica column with a methanol-high-pH eluent was examined. The relative capacity factors of specific tertiary amines changed systematically compared with the other drugs on storage of the columns for 1 year. The changes were enhanced on storing the column in mobile phase and were accelerated by prolonged washing with water. A comparison of freshly packed columns suggested that similar changes also occurred with the dry silica on storage. These effects may be a contributor to previously observed batch-to-batch differences in the silica.

Caffeine↗
PubMed1992

Mutational activation of the N-ras oncogene assessed in primary clonogenic culture of acute myeloid leukemia (AML): implications for the role of N-ras mutation in AML pathogenesis.

The number of steps involved in the pathogenesis of acute myeloid leukemia (AML) is unclear. The initiating event would be expected to exist in all leukemic cells, but subsequent events may be subclonal. If several genetic events occur, they may cooperate within the same cell or be alternatively acquired by different subclones. These possibilities cannot be adequately analyzed in DNA prepared directly from patient specimens. In this study, N-ras mutations demonstrable in DNA prepared from peripheral blood of 10 patients with AML were examined in primary in vitro colonies (AML-colony-forming units [CFU]) grown from these patients. Both colonies containing the mutant gene and colonies containing normal allele only were obtained from each patient. The proportion of colonies containing no mutant allele varied among patients (5% to 57%). A subset of mutation containing colonies appeared to have lost the normal allele in nine of 10 AML cases analyzed. In the four cases with two N-ras mutations, the two mutations were found to exist in different subclones. In these cases, macroscopic colonies (AML-MCFU) were also obtained using an assay system designed to select for earlier clonogenic cells than in the AML-CFU assay. The N12cys mutation in AML10 was found in the CFU, but not in the MCFU, and the N12asp mutation in AML43 was found in the MCFU, but not in the CFU. These results suggest that N-ras mutation is a postinitiation event in AML that contributes to the outgrowth of more malignant subclones. Where two mutations are found in a case of AML, they appear to have been acquired by separate subclones, which may show different degrees of differentiation.

Base Sequence↗
PubMed1992

The effect of MK-801 on cortical spreading depression in the penumbral zone following focal ischaemia in the rat.

Cortical spreading depression (CSD) is a transient depression of neuronal activity that spreads across the cortical surface. In the present studies, we have investigated CSD activity in the penumbral zone following permanent middle cerebral artery (MCA) occlusion in the rat (n = 16/group), using double-barreled Ca(2+)-sensitive microelectrodes. Measurements of CSD activity were made for 3 h in each animal. During this time, a varying number of spontaneous CSDs were seen in the control group (total was 30, with a range of 0-7/rat). These CSDs were of varying duration: "small" (approximately 1 min) and "big" (5-45 min) CSDs. During a CSD, the extracellular [Ca2+] decreased to 0.11 +/- 0.07 mM (mean +/- SD). After 3 h, the extracellular [Ca2+] in the cortex (penumbral zone) was either normal (10/16 rats) or lowered to 0.5 mM (2/16 rats) or to 0.1 mM (4/16 rats). In the caudate nucleus (ischaemic core area), all rats had an extracellular [Ca2+] of approximately 0.1 mM when measured after the 3 h recording period. Neuropathological evaluation of the brains of the animals, which had been allowed to survive for 24 h after MCA occlusion, revealed ischaemic damage in the dorsolateral cortex and caudate nucleus. Administration of the noncompetitive NMDA antagonist, MK-801 (3 mg/kg i.p.), 30 min after MCA occlusion resulted in 24 and 29% reductions in the volume of hemispheric and cortical damage, respectively, which was highly significant (p less than 0.0001); no protection was seen against caudate damage.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Molecular aspects of the calbindins.

Studies from our laboratory concerning regulation of calbindin include regulation by 1,25-dihydroxycholecalciferol [1,25(OH)2D3], receptor regulation as a possible mechanism for modulating calbindin's response to hormone, tissue specific regulation and regulation by factors other than 1,25(OH)2D3. With regard to receptor regulation, we found that the induction of calbindin mRNA in intestine and kidney by 1,25(OH)2D3 is not accompanied by a corresponding alteration in vitamin D receptor (VDR) mRNA in the vitamin D-deficient, low calcium rat. However, in the vitamin D-replete rat, administration of 1,25(OH)2D3 results in an induction of both calbindin and VDR mRNA in these tissues. These results suggest the presence of an inhibitor of 1,25(OH)2D3-mediated receptor up-regulation in the vitamin D-deficient, low calcium animal. Glucocorticoids can also regulate calbindin gene expression. Dexamethasone treatment (50 micrograms.100 g body weight-1.d-1 for 4 d) results in a 75% decrease in rat intestinal calbindin-D9k mRNA. This decrease may be related to the inhibition of intestinal calcium absorption previously observed after glucocorticoid administration. Kidney calbindin-D28k mRNA is unaffected by glucocorticoid treatment, indicating tissue specificity of the glucocorticoid response. To evaluate more precisely the means whereby 1,25(OH)2D3 and other modulators can influence calbindin gene expression, we isolated the chromosomal gene for calbindin-D28k by screening a mouse genomic library in cosmid. Ros 17/2.8 cells were transfected with recombinant plasmids in which the mouse calbindin promoter is fused to the reporter gene encoding chloramphenicol acetyltransferase. Deletion studies have enabled us to identify sequence elements in the mouse calbindin-D28k gene that confer basal activation and a hormone inducible response.

Animals↗
PubMed1992

Morphoregulatory role of thidiazuron : substitution of auxin and cytokinin requirement for the induction of somatic embryogenesis in geranium hypocotyl cultures.

Somatic embryogenesis was induced in hypocotyl explants of geranium (Pelargonium x hortorum) cultured on media supplemented with various concentrations of N-phenyl-N'-1,2,3-thiadiazol-5-ylurea (thidiazuron). In less than 2 weeks, somatic embryos were observed in treatments containing levels of thidiazuron (TDZ) ranging from 0.2 to 1.0 micromolar. The use of N(6)-benzylaminopurine in combination with indole-3-acetic acid also evoked embryogenesis, but the efficiency of somatic embryo production was significantly lower than that obtained with TDZ. Hypocotyl culture for only 2 days on TDZ-supplemented medium before transfer to a basal medium was sufficient for inducing somatic embryogenesis. This distinction between the induction and expression of embryogenesis may provide an experimental system for studying the developmental biology of somatic embryogenesis. Substitution of the auxin-cytokinin requirement for the induction of somatic embryogenesis by TDZ suggests the possibility of a novel mode of its action by modulation of endogenous growth regulators.

Journal Article↗
PubMed1992

Low frequency of ras oncogene mutations in Philadelphia-positive acute leukemia and report of a novel mutation H61 Leu in a single case.

Activating ras mutations are frequent (25-60%) in chronic myelomonocytic leukemia (CMML) and in acute myeloid leukemia (AML) (30%), in contrast to chronic myeloid leukemia (CML) in which the incidence is very low (0-3%). This might reflect that the leukemic cell in CML is at a level of differentiation in which ras gene activation is not involved or, alternatively, might be due to the presence in CML of the bcrlabl fused gene. We have analyzed the presence of point mutations in codons 12, 13, 59, 61 and 63 of N-, K-, and H-ras genes, in 26 cases of Philadelphia-chromosome-positive, bcrlabl-positive acute leukemia (Ph+ AL), and in eight CMML cases by using the polymerase chain reaction. Aberrant ras genes were detected in a single Ph+ AL case, and in four out of eight CMML patients. The Ph+ AL showing altered ras allele had an unusual point mutation in H-ras gene, substituting leucine for glutamine. This mutation has not been previously found in any hematological disease. Our findings suggest that ras mutations are probably not involved in the pathogenesis of those leukemias in which blast cells contain bcrlabl oncogene activation.

Codon↗
PubMed1991

Multiple K-ras codon 12 mutations in cholangiocarcinomas demonstrated with a sensitive polymerase chain reaction technique.

By using a modified polymerase chain reaction strategy, we have devised an approach to detect a K-ras oncogene mutated at codon 12 in the presence of 1000 normal alleles. This is a considerable improvement in sensitivity on previous assays. Application of this assay to 15 cholangiocarcinomas showed that all contained a K-ras mutation to codon 12 and that nine of the tumors contained two or more mutations. In 11 cases, mutations were present in less than 10% of the cells in the sample. In common with pancreatic adenocarcinomas, in which 75 to 95% of cases contain a mutation in K-ras, cholangiocarcinomas show a very high frequency of ras gene mutation, but within a tumor only a fraction of cells contain a ras mutation. The presence of multiple mutations and the low frequency of mutant alleles in the samples argue against K-ras mutations being the initiating genetic lesion in this tumor, but suggest that ras gene mutation is involved in the stepwise progression of neoplastic cells to full malignancy.

Adenoma, Bile Duct↗
PubMed1991

Striatal A2 receptor regulates apomorphine-induced turning in rats with unilateral dopamine denervation.

The effect of the purine agonist N-ethylcarboxamido-adenosine (NECA) on apomorphine-induced rotation was investigated in rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal pathway. Intrastriatal administration of NECA on the denervated side caused a dose-dependent inhibition of contralateral rotation. This inhibition was prevented by prior intrastriatal injection of theophylline. The adenosine A1 receptor antagonist 8-cyclopentyltheophylline was ineffective at concentrations which block this receptor, but effective in preventing the action of NECA at concentrations which block the adenosine A2 receptor. In the absence of apomorphine, NECA had no effect on behaviour. It is concluded that A2 receptor activation counteracts apomorphine effects in the striatum. Since the A2 receptor may be localized to striatal cholinergic neurones, the possible role of these neurones in purine-induced behaviours is discussed.

Adenosine↗
PubMed1991

Analysis of ras gene mutations in childhood myeloid leukaemia.

Previous studies have shown that approximately 30% of adult acute myeloid leukaemias and 20% of adult acute lymphoid leukaemias contain point mutated ras oncogenes. In order to assess whether ras oncogenes are also involved in childhood leukaemias, we have used polymerase chain reaction (PCR) amplification and synthetic oligonucleotide probes to study the nature and frequency of ras gene mutations in childhood leukaemias, concentrating largely on the acute myeloid leukaemias (AML). Thirty-four childhood presentation AML DNAs were screened for mutations in and around codons 12, 61 and 117 of N-, K- and H-ras. Eight of these samples (24%) contained ras mutations. As in the adult disease, the gene predominantly involved was N-ras (6/8), with occasional activation of K-ras (2/6). The most common base change was a G----A transition at codon 12 or 13 (4/8). Of the patients with mutant ras, 4/8 were diagnosed as AML FAB subtype M5. Five of the 34 childhood AMLs analysed displayed abnormalities of chromosome 7. However, none of these cases contained a mutant ras gene. One AML patient was studied at relapse, 14 months after initial presentation. The presentation mutation (N61p3) was not detectable, although a new mutation (N13Cys) was readily identified. This observation extends our original finding with presentation and relapse samples of adult AML, in which it was uncommon for the relapse sample to contain the same ras mutation as the presentation DNA. In addition, two out of five patients diagnosed as juvenile CML, were found to harbour mutant ras.

Acute Disease↗