Biomedical subjects

C R Jones

Publications and source records attributed to C R Jones.

At least 73 records · Page 4Linked to original sources

PubMed1992

Induction of various cytochromes CYP2B, CYP2C and CYP3A by phenobarbitone in non-human primates.

Male and female patas (Erythrocebus patas) and cynomolgus (Macaca fascicularis) monkeys were treated with phenobarbitone (PB) and examined for the induction of various cytochrome P450 (P450)-mediated drug metabolizing enzymes. Hydroxylation of testosterone at the 6 beta, 2 beta, and 15 beta positions, metabolites normally associated with CYP3A P450s increased 2- to 5-fold in PB-treated animals. Induction of this P450 family was confirmed by the use of polyclonal antisera directed against the human CYP3A enzymes which inhibited both induced and constitutive 6 beta- and 15 beta-hydroxylase activities in both species of monkeys. The enzymatic activities testosterone 16 beta-hydroxylation, pentoxyresorufin O-dealkylation [PROD], and benzyloxyresorufin O-dealkylation [BZROD] typically associated with the rodent CYP2B subfamily in rodents were also examined. Testosterone 16 beta-hydroxylation activity was highly induced up to 15-fold in both species of monkeys to maximal levels similar to those induced in PB-treated male rats. BZROD was similarly induced up to 10-fold in both species of monkeys, but the maximal levels of BZROD achieved were substantially lower than those obtained in PB-treated rats. Finally, PROD yielded an idiosyncratic response showing substantial induction (> 30-fold) in certain patas monkeys (4 out of 8) but minimal (< 5-fold) induction in the other patas monkeys (4 out of 8) or any of the cynomolgus monkeys (0 out of 8). Immunodetection of various cytochromes using polyclonal antisera directed against rat cytochromes confirmed the induction of CYP3A proteins as well as the induction of protein(s) immunologically cross-reactive with rat CYP2B P450s. BZROD, PROD, or testosterone 16 beta-hydroxylase activities, were not inhibited by antibody to CYP3A P450s. However, high concentrations of polyclonal antiserum directed against rat CYP2B inhibited all three activities in PB-induced patas monkeys. In contrast, this antiserum failed to inhibit the hydroxylation of testosterone at the 6 beta or 2 beta positions. Induction of the CYP2C subfamily was observed by immunochemical detection. Interestingly, induction of this subfamily appeared to be more pronounced in cynomolgus than in patas monkeys. Finally, we failed to observe significant sex-dependent differences in P450-mediated enzymatic activities in either control or induced monkeys. These results confirm the induction of a similar spectrum of P450 proteins by PB in non-human primates to that which has previously been observed in rodents.

Animals↗

PubMed1992

Effect of Elicitation and Changes in Extracellular pH on the Cytoplasmic and Vacuolar pH of Suspension-Cultured Soybean Cells.

We have employed both (31)P nuclear magnetic resonance spectroscopy and two intracellular fluorescent pH indicator dyes to monitor the pH of the vacuole and cytoplasm of suspension-cultured soybean cells (Glycine max Merr cv Kent). For the (31)P nuclear magnetic resonance studies, a flow cell was constructed that allowed perfusion of the cells in oxygenated growth medium throughout the experiment. When the perfusion medium was transiently adjusted to a pH higher than that of the ambient growth medium, a rapid elevation of vacuolar pH was observed followed by a slow (approximately 30 minute) return to near resting pH. In contrast, the concurrent pH changes in the cytoplasm were usually fourfold smaller. These data indicate that extracellular pH changes are rapidly communicated to the vacuole in soybean cells without significantly perturbing cytoplasmic pH. When elicitors were dissolved in a medium of altered pH and introduced into the cell suspension, the pH of the vacuole, as above, quickly reflected the pH of the added elicitor solution. In contrast, when the pH of either a polygalacturonic acid or Verticillium dahliae elicitor preparation was adjusted to the same pH as the ambient medium, no significant change in either vacuolar or cytoplasmic pH was observed during the 35 minute experiment. These results were confirmed in experiments with pH-sensitive fluorescent dyes. We conclude that suspension-cultured soybean cells do not respond to elicitation by significantly changing the pH of their vacuolar or cytoplasmic compartments.

Journal Article↗

PubMed1991

Endothelins: functional and autoradiographic studies in guinea pig trachea.

The presence of binding sites for [125I]endothelin-1 and the contractile activities of endothelins (ETs) and sarafotoxin S6b and the endothelin fragment ET(16-21) were investigated in guinea pig trachea. ETs and sarafotoxin S6b (0.1-100 nM) induced potent contractile responses in guinea pig trachea with EC50 values ranging from 1.57 to 12.97 nM. Epithelium removal increased the potencies of ET-1, ET-2 and S6b, but not that of ET-3, and maximal responses to ET-1 and ET-2 were also increased. Effects of epithelium removal were partially mimicked by phosphoramidon (10 microM), an enkephalinase inhibitor, suggesting that enkephalinase (EC.3.4.24.11.) is able to degrade ET-1 and ET-2. ET-3-induced contractions were not affected by phosphoramidon. Autoradiographic studies suggested the presence of at least two specific binding sites for [125]ET-1 in guinea pig airway smooth muscle. The correlation between Kd and EC50 values suggests that the binding sites identified in the airway smooth muscle represent functional receptors for ETs. ET(16-21) and ET(16-21)-NH2 were less potent agonists than the ETs in guinea pig trachea and 10 microM ET(16-21) was unable to inhibit [125I]ET-1 binding in guinea pig airway smooth muscle. Therefore, these results suggest that the C-terminal hexapeptide of ET-1 cannot be used to classify ET receptors in guinea pig trachea.

Animals↗

PubMed1991

Two-dimensional NMR and structure determination of salmon calcitonin in methanol.

The structure of the 32-residue peptide salmon calcitonin (sCT) in 90% MeOH-10% H2O has been investigated by two-dimensional NMR techniques and molecular modeling. Sequential assignments for nearly all of the 32 spin systems have been obtained, and results indicate that the heptaresidue loop formed by the disulfide bond between Cys-1 and Cys-7 is followed by an alpha-helical segment from Val-8 through Tyr-22. A region of conformational heterogeneity is observed for residues 20-25, resulting from the slow isomerism of the cis and trans forms of Pro-23. The C-terminal segment is found to exist in an extended conformation.

Amides↗

PubMed1991

Induction of cytochrome P450 and other drug metabolizing enzymes in rat liver following dietary exposure to Aroclor 1254.

Selected drug metabolizing activities were measured in female F344/NCr rats exposed to graded dietary concentrations of Aroclor 1254 (1 to 1000 ppm) for 7 days or to lower concentrations of Aroclor (1 to 10 ppm) for up to 28 days. Following the 7-day exposure, the hepatic O-dealkylation of ethoxyresorufin (ETR), mediated primarily by cytochrome P450IA, was increased 60-, 10-, and 4-fold by 33, 10, and 3 ppm Aroclor, respectively. In rats exposed to 10 and 3 ppm Aroclor for 28 days, this activity was increased approximately 30- and 10-fold, respectively. Hepatic ETR O-dealkylase activities correlated with Aroclor concentrations in the livers of exposed rats (r = 0.99, p less than 0.01). Although the O-dealkylation of benzyloxyresorufin was highly increased by 7-days dietary exposure to 1000 ppm Aroclor, the levels of Aroclor necessary for detection of induction were substantially higher than those required for detection of ETR O-dealkylase induction. Examination of the non-P450-mediated drug metabolizing activities, epoxide hydrolase and DT-diaphorase, similarly showed limited (approximately 10-fold) increases. In contrast, aldehyde dehydrogenase (benzaldehyde, NADP+) activity was highly increased (greater than 40-fold) at 1000 ppm, however this activity was increased to only a limited extent at lower Aroclor concentrations (e.g. approximately 3-fold at 33 ppm). These results support the potential use of cytochrome P450 activities as potential biomarkers for environmental exposure to PCBs and related compounds.

Aldehyde Dehydrogenase↗

PubMed1991

Autoradiography of adrenoceptors in rat and human brain: alpha-adrenoceptor and idazoxan binding sites.

This chapter reviews the current classification of adrenoceptors, and notes the difficulties of combining the molecular biological and pharmacological classifications of adrenoceptors. Possibilities for mapping the distribution of the proposed subtypes of adrenoceptors using currently available ligands are discussed, and the autoradiographic visualisation of the broad subtypes of alpha 1-, alpha 2-, beta 1-, and beta 2-adrenoceptors in the rat, monkey and human brain described and illustrated. The non-selectivity of ligands currently being used to label alpha-adrenoceptors is shown; we compare the distribution of [3H]idazoxan binding sites with the distribution of alpha 2-adrenoceptors visualised using other ligands. Resolution limitations of current autoradiographic approaches are considered and we shown how in situ hybridisation can complement data from receptor labelling studies used to localise receptors to pre- or postsynaptic sites.

Animals↗

PubMed1991

Proliferation in human bladder carcinoma measured by Ki-67 antibody labelling: its potential clinical importance.

Ki-67 is a monoclonal antibody which recognises a human nuclear antigen expressed in proliferating cells. The antibody was used to assess proliferation in primary human bladder tumours from 64 patients. Ki-67 index (the number of Ki-67 positive tumour cells divided by the total number of tumour cells %) was derived from 59 tumours. A wide range of Ki-67 indices were recorded, range 3.0-65.8%, mean 20.2%. The Ki-67 index correlated with known prognostic factors: T stage (P = 0.002) and histological grade (P less than 0.001), early stage disease and more differentiated tumours having lower Ki-67 indices. Patients with invasive disease (21 patients) had significantly higher Ki-67 indices than those with non-invasive disease (P = 0.01). Patients with metastatic disease at presentation (four cases) all had a Ki-67 index of greater than or equal to 29%. Ki-67 antibody staining is a simple technique for assessing the proliferation fraction than can be performed on a small amount of tissue taken at routine biopsy without prior injection of thymidine analogues.

Aged↗

PubMed1991

Endothelin receptor heterogeneity; structure activity, autoradiographic and functional studies.

Radioligand binding studies were used to define the binding constants for inhibition of [125I]endothelin-1 binding by endothelin-1 (ET-1), endothelin-3 (ET-3) and a synthetic analogue of ET-1, [Ala1,3,11,15]ET-1 in rat aorta and cerebellum. The inhibition curves for the three substances were similar in the cerebellum (Ki, ET-1 = ET-3 = [Ala1,3,11,15]ET-1) but differed in the aorta (Ki, ET-1 much less than [Ala1,3, 11,15]ET-1) with the ET-3 inhibition curves being biphasic, indicating binding to two sites. In functional studies in the aorta, in contrast to findings in the cerebellum, [Ala1,3,11,15]ET-1 was inactive and ET-3 was a partial agonist. These findings support the idea of heterogeneity of endothelin receptors. Autoradiographic studies were performed to identify the cellular localisation of endothelin-1 receptors in the two tissues. In the aorta binding was to all elements of the vasculature including the endothelium, smooth muscle, periadventitial connective tissue and nerves. In the cerebellum binding sites were located predominantly over the neuronal elements of the granular cell layer. The cellular localisation of the high and low affinity sites for ET-3, and the functional significance of the high affinity binding site for ET-3, remain to be determined.

Animals↗

PubMed1990

Effect of distortions in the deoxyribose phosphate backbone conformation of duplex oligodeoxyribonucleotide dodecamers containing GT, GG, GA, AC, and GU base-pair mismatches on 31P NMR spectra.

We have previously suggested that variations in the 31P chemical shifts of individual phosphates in duplex oligonucleotides are attributable to torsional angle changes in the deoxyribose phosphate backbone. This hypothesis is not directly supported by analysis of the 1H/31P two-dimensional J-resolved spectra of a number of mismatch dodecamer oligonucleotide duplexes including the following sequences: d-(CGTGAATTCGCG), d(CGUGAATTCGCG), d(CGGGAATTCGCG), d(CGAGAATTCGCG), and d(CGCGAATTCACG). The 31P NMR signals of the dodecamer mismatch duplexes were assigned by 2D 1H/31P pure absorption phase constant time (PAC) heteronuclear correlation spectra. From the assigned H3' and H4' signals, the 31P signals of the base-pair mismatch dodecamers were identified. JH3'-P coupling constants for each of the phosphates of the dodecamers were obtained from 1H/31P J-resolved selective proton flip 2D spectra. By use of a modified Karplus relationship, the C4'-C3'-O3'-P torsional angles (epsilon) were obtained. JH3'-P coupling constants were measured for many of the oligonucleotides as a function of temperature. There exists a good linear correlation between 31P chemical shifts and the epsilon torsional angle. This correlation can be further extended to the C3'-O3'-P-O5' torsional angle (zeta) by using a linear relationship between epsilon and zeta obtained from crystal structure studies. The 31P chemical shifts follow the general observation that the more internally the phosphate is located within the oligonucleotide sequence, the more upfield the 31P resonance occurs. In addition, 31P chemical shifts show sequence- and site-specific variations. Analysis of the backbone torsional angle variations from the coupling constant analysis has provided additional information regarding the origin of these variations in 31P chemical shifts.

Base Sequence↗

PubMed1990

Subtypes of alpha 1-adrenoceptors in hippocampus of pigs, guinea-pigs, calves and humans: regional differences.

Radioligand binding studies were performed with membranes of guinea-pig, pig, calf and human hippocampus using [125I]BE 2254 (also known as [125I]HEAT) as the radioligand. [125I]BE 2254 bound with similar high affinity to saturable populations of recognition sites in all four membrane preparations. Competition curves obtained with a variety of ligands (e.g., WB 4101, benoxathian, 5-methyl-urapidil) were biphasic and the profiles of the high- and low-affinity components of [125I]BE 2254 binding were similar in all four membrane preparations. The data suggest that [125I]BE 2254 labels two subtypes of alpha 1-adrenoceptors in the hippocampus of these species. [3H]WB 4101 was used to label alpha 1A recognition sites in pig hippocampus membranes. [3H]WB 4101 recognized with high affinity an apparently homogeneous class of sites, as suggested by monophasic saturation and competition experiments. The rank order of affinity of the compounds for the high-affinity component of [125I]BE 2254 binding was similar to the rank order of affinity of these drugs for [3H]WB 4101 sites. The pharmacological profile of the low-affinity component of [125I]BE 2254 binding was similar to that described recently for the alpha 1B-adrenoceptor cloned from DDT1 cells. In autoradiographic studies with human hippocampal slices, CEC (chloroethylclonidine), an alkylating agent described to show selectivity for alpha 1B-adrenoceptors, displaced preferentially [125I]BE 2254 binding from the molecular layer of the dentate gyrus. In contrast, WB 4101 an alpha 1A-adrenoceptor-selective ligand, displaced preferentially [125I]BE 2254 binding in the hilus and the CA3 region. The data show that 2 subtypes of alpha 1-adrenergic recognition sites can be identified in the hippocampus.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Antagonists↗

PubMed1990

Binding characteristics of [125I]endothelin-1 in guinea-pig trachea and its displacement by endothelin-1, endothelin-3 and endothelin (16-21).

The presence of binding sites for endothelin-1 (ET-1) and endothelin-3 (ET-3) in airway epithelium, submucosa and smooth muscle of guinea-pig trachea was investigated using in vitro autoradiography. We also examined the ability of the C-terminal hexapeptide of endothelin, ET(16-21) to inhibit specific [125I]ET-1, binding. ET-1 appeared to bind to a single class of binding sites (nH not different from unity). In contrast, nH values for ET-3 displacement of [125I]ET-1 specific binding were different from unity, suggesting that these peptides bound to more than one site. ET (16-21) did not affect [125I]ET-1 binding. The present results suggest that the binding sites identified in guinea pig airway smooth muscle may be related to receptors mediating contractile responses to the endothelins.

Animals↗

PubMed1990

Urothelial grafts in mice.

Mouse bladder epithelium has been successfully transplanted to the bladders of syngeneic mice and has survived for at least twenty weeks. The fate of the transplanted tissue was followed using a fluorescein label. The recipient bladders were prepared by stripping the urothelium either by a surgical or a chemical method. The possibility of adopting a comparable technique for the treatment of early bladder cancer in man is discussed.

Animals↗

PubMed1990

Task demand and cardiovascular response magnitude: further evidence of the mediating role of success importance.

The interactive effects of success importance and task demand upon cardiovascular reactivity were investigated in 2 experiments. In Experiment 1, Ss learned that success on an easy or difficult memory task would allow them to avoid a mild or severe noise. As expected, pretask and task elevations in heart rate and systolic blood pressure increased with difficulty only when the noise was to be severe. In Experiment 2, Ss listened to a victim, attending either to how she must feel or to technical features of the presentation. Then they were given the chance to earn a donation by succeeding on an easy or difficult memory task. Pretask cardiovascular data accorded with predictions predicated on the assumption that the need to help would be greater among victim perspective Ss. For them, systolic elevations increased with difficulty, whereas for technical perspective Ss systolic elevations were low regardless of task demand.

Achievement↗

PubMed1990

Two-dimensional 1H and 31P NMR spectra and restrained molecular dynamics structure of an oligodeoxyribonucleotide duplex refined via a hybrid relaxation matrix procedure.

Assignment of the 1H and 31P resonances of a decamer DNA duplex, d(CGCTTAAGCG)2 was determined by two-dimensional COSY, NOESY and 1H-31P Pure Absorption phase Constant time (PAC) heteronuclear correlation spectroscopy. The solution structure of the decamer was calculated by an iterative hybrid relaxation matrix method combined with NOESY-distance restrained molecular dynamics. The distances from the 2D NOESY spectra were calculated from the relaxation rate matrix which were evaluated from a hybrid NOESY volume matrix comprising elements from the experiment and those calculated from an initial structure. The hybrid matrix-derived distances were then used in a restrained molecular dynamics procedure to obtain a new structure that better approximates the NOESY spectra. The resulting partially refined structure was then used to calculate an improved theoretical NOESY volume matrix which is once again merged with the experimental matrix until refinement is complete. JH3'-P coupling constants for each of the phosphates of the decamer were obtained from 1H-31P J-resolved selective proton flip 2D spectra. By using a modified Karplus relationship the C4'-C3'-O3'-P torsional angles (epsilon) were obtained. Comparison of the 31P chemical shifts and JH3'-P coupling constants of this sequence has allowed a greater insight into the various factors responsible for 31P chemical shift variations in oligonucleotides. It also provides an important probe of the sequence-dependent structural variation of the deoxyribose phosphate backbone of DNA in solution. These correlations are consistent with the hypothesis that changes in local helical structure perturb the deoxyribose phosphate backbone. The variation of the 31P chemical shift, and the degree of this variation from one base step to the next is proposed as a potential probe of local helical conformation within the DNA double helix. The pattern of calculated epsilon and zeta torsional angles from the restrained molecular dynamics refinement agrees quite well with the measured JH3'-P coupling constants. Thus, the local helical parameters determine the length of the phosphodiester backbone which in turn constrains the phosphate in various allowed conformations.

Absorption↗

PubMed1990

Effect of calcium and beta-adrenoceptor antagonists on beta-adrenoceptor density and Gs alpha expression in human atria.

In 21 patients undergoing open heart surgery the effects of the beta-adrenoceptor antagonists propranolol and sotalol and the calcium antagonist nifedipine (alone or in combination) on right atrial beta-adrenoceptor density and G protein expression were studied. The non-selective beta-adrenoceptor antagonists increased both right atrial beta 1- and beta 2-adrenoceptor density; this increase was not affected by nifedipine. There was no correlation between beta-adrenoceptor density and expression of the G protein subunit Gs alpha suggesting independent regulation of the gene expression for G proteins and their coupled receptors.

Adrenergic beta-Antagonists↗

PubMed1990

Receptor modification in the brains of spontaneously hypertensive and Wistar-Kyoto rats: regionally specific and selective increase in cerebellar beta 2-adrenoceptors.

Quantitative in vitro autoradiography was used to study beta-adrenergic and benzodiazepine receptor density in discrete regions from sagittal brain sections of 20 week old hypertensive (SHR) and normotensive (WKY) rats. The density of beta-adrenoceptors was increased by 68% in the granule cell layer of the cerebellum of the SHR without a change in receptor affinity; this increase was specific for receptors of the beta 2-subtype. On the other hand, benzodiazepine receptor density was unchanged in the cerebella of SHR. These results indicate that brain beta-adrenoceptors are differentially modulated by the hypertensive state which may be either a cause or a consequence of alterations in adrenergic nervous system activity found in the SHR.

Animals↗

PubMed1990

Autoradiographic visualization of A1 adenosine receptors in rat brain with [3H]8-cyclopentyl-1,3-dipropylxanthine.

A1 adenosine receptors were labeled in rat brain sections with the antagonist [3H]8-cyclopentyl-1,3-dipropylxanthine ([3H]DPCPX) and visualized at the light microscopic level using autoradiography. The specific binding of [3H]DPCPX to the sections showed the pharmacological characteristics of A1 adenosine receptors and was accompanied by very low levels of nonspecific binding. Whereas GTP had no significant effect on [3H]DPCPX binding to rat brain membranes, the addition of 100 microM GTP increased the apparent affinity of [3H]DPCPX to tissue sections fivefold (from 1.83 to 0.35 nM), enhancing it to the affinity measured in membranes. However, GTP altered neither the binding capacity nor the distribution of binding sites in tissue sections. It is suggested that a competitive antagonism with endogenous adenosine explains the lower affinity of [3H]DPCPX in the absence of GTP. The autoradiographic pattern of [3H]DPCPX binding was characteristic for A1 adenosine receptors. Distinct labeling of the different layers of the cerebellar cortex was shown by photomicrographs generated with the coverslip technique. In addition, several fiber tracts were found to be labeled. The high selectivity for A1 adenosine receptors and low nonspecific binding of [3H]DPCPX, the ability to produce high-resolution autoradiograms, together with the fact that the effects of endogenous adenosine can be eliminated by the addition of GTP make [3H]DPCPX a very useful tool in the autoradiographic study of A1 adenosine receptors.

Animals↗

PubMed1990

Binding of [125I]-endothelin-1 to rat cerebellar homogenates and its interactions with some analogues.

1. [125I]-endothelin-1, over the concentration range 6 pM-10 nM, bound to a single site in homogenates of rat cerebellum with high affinity (Kd = 2.8 +/- 0.6 x 10(-10) M). The site was present in a concentration of 321 +/- 58 fmol mg-1 protein. 2. The rates of association and dissociation of [125I]-endothelin-1 with the binding site were slow (at 25 degrees C, k+1 = 8.0 +/- 1.3 x 10(5) M-1s-1; k-1 = 2.6 x 10(-4)s-1) and, on addition of a maximally displacing concentration of endothelin-1 (100 nM), 94.0 +/- 8.4% of the [125I]-endothelin-1 was still bound after 14 h. 3. [125I]-endothelin-1 binding was inhibited by a number of naturally occurring or genetically encoded members of the endothelin/sarafotoxin family of peptides. The order of potency was endothelin-3 = sarafotoxin S6b greater than endothelin-2 = endothelin-1 much greater than porcine proendothelin1-39. 4. Binding was also inhibited by analogues in which either one or both of the cystine disulphide bridges had been replaced by substitution with 2 or 4 alanine residues. The tetra-alanyl substituted analogue, [Ala1,3,11,15]endothelin-1, was equipotent with endothelin-1 at inhibiting the binding of [125I]-endothelin-1. [Ala3,11]endothelin-1 and [Ala1,15]endothelin-1, analogues which each contained one of the disulphide bridges from the parent peptide, were respectively 3 and 14 times less potent than the parent peptide. An analogue in which the Glu10 residue had been anisylated was 25 fold less potent than endothelin-1. 5. It is concluded that the structural requirements for binding to the cerebellar sites for [1251]_ endothelin-1 do not require the presence of the disulphide bridges characteristic of the endothelin/ sarafotoxin family. Rather, the binding may be more sensitive to the presence of bulky side chain substituents, at least in the smaller intramolecular loop.

Animals↗