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Biomedical subjects

C M Jones

Publications and source records attributed to C M Jones.

At least 55 records · Page 3Linked to original sources
PubMed1997

Use of reduced sorbent bed and disk membrane solid-phase extraction for the analysis of pharmaceutical compounds in biological fluids, with applications in the 96-well format.

Significant improvements in the isolation of pharmaceutical compounds from plasma, serum and urine, have been achieved using ultra low mass sorbent bed and thin disk solid-phase extraction (SPE) material. The use of low sorbent masses or disk SPE material has allowed a significant reduction in solvent usage and extraction times. The reduction in solvent volumes required has allowed elution volumes to be reduced to as low as 30 microl with high and consistent analyte recovery. Several SPE RP-HPLC methods have been developed using these materials, including LC-MS methods. When the chromatographic conditions allow the eluent to be injected directly or injected after dilution with distilled water Empore disks are the extraction media of choice due to the materials low elution volume requirements. When operated in the 96-well microtitre format this micro-extraction provides a very efficient throughput and requires little sample manipulation.

Androstadienes↗
PubMed1997

Can a two-state MWC allosteric model explain hemoglobin kinetics?

We have analyzed the nanosecond-millisecond kinetics of ligand binding and conformational changes in hemoglobin. The kinetics were determined from measurements of precise time-resolved optical spectra following nanosecond photodissociation of the heme-carbon monoxide complex. To fit the data, it was necessary to extend the two-state allosteric model of Monod, Wyman, and Changeux (MWC) to include geminate ligand rebinding and nonexponential tertiary relaxation within the R quaternary structure. Considerable simplification of the model is obtained by using a linear free energy relation for the rates of quaternary transitions, and by incorporating concepts from recent studies on the physics of geminate rebinding and conformational changes in myoglobin. The model, described by 85 coupled differential equations, quantitatively explains a demanding set of complex kinetic data. Moreover, with the same set of kinetic parameters it simultaneously fits the equilibrium data on ligand binding and the distribution of ligation states. The present results, together with those from single-crystal oxygen binding studies, indicate that the two-state MWC allosteric model has survived its most critical tests.

Allosteric Site↗
PubMed1997

Antigen-specific CD8+ T cell subset distribution in lymph nodes draining the site of herpes simplex virus infection.

Inoculation with replicating virus leads to an increase in T cell numbers within lymph nodes that drain the site of infection. This increase has been associated with a nonspecific proliferation of bystander cells, with only a minority thought to be directed to the infectious agent. Such an assumption is largely based on precursor cytotoxic T lymphocyte (CTL) estimations using limiting dilution analysis. Recently, studies using more advanced molecular approaches have suggested that such functionally derived precursor frequencies considerably underestimate the proportion of T cells specific for the antigen under investigation. We have defined T cell receptor sequences characteristic of CTL populations directed to a dominant determinant of the herpes simplex virus (HSV) glycoprotein B (gB). In this investigation, we used this receptor signature as a probe to directly monitor changes occurring within lymph nodes draining the sites of active infection with HSV. We found that although lymph node CD8+ T cell numbers increase as a consequence of HSV infection, the majority of these cells are small resting cells that are not enriched for gB-specific receptors. In contrast, a significant proportion of activated T cells are highly enriched for CTL bearing gB-specific receptors. Our results are therefore consistent with a nonspecific migration of CTL precursors into the lymph nodes draining the site of infection, followed by the activation and proliferation of the antigen-specific subset that normally makes up a small proportion of the naive T cell repertoire.

Amino Acid Sequence↗
PubMed1997

Evidence for cooperation between TCR V region and junctional sequences in determining a dominant cytotoxic T lymphocyte response to herpes simplex virus glycoprotein B.

TCR repertoire availability has the potential to influence the immune response to foreign antigens. Here we have analysed how changes in V region availability influence the H-2b-restricted cytotoxic T lymphocyte (CTL) response to a dominant peptide determinant derived from the herpes simplex virus glycoprotein B (gB). We have previously shown that C57BL/6 mice mount a gB-specific, Kb-restricted CTL response which is dominated by a TCRBV10+ population and a TCRBV8S1+ subpopulation, both containing highly conserved CDR3 elements. We find that this dominant gB-specific CTL pool is lost in C57/L mice which have a different TCRBV haplotype. A population of CTL with diverse TCRBV and junctional sequence usage, which otherwise represents a minor subset in the gB-specific response, appears to emerge as a consequence of this TCRBV gene variation. The loss of preferential V region-encoded complementarity determining regions (CDR) 1- and/or CDR2-ligand interactions in this emerging population also results in a change in CDR3 sequence usage and a corresponding focusing of an otherwise promiscuous pattern of cross-reactivity with a panel of gB498-505 substitution analogues. This suggests that the difference between the two distinct TCR populations is the relative contributions of the CDR towards ligand recognition. Therefore, preferential V region-ligand interaction, at the expense of CDR3 peptide recognition, appears to control the dominant TCR selection in the C57BL/6 response to this peptide determinant.

Amino Acid Sequence↗
PubMed1997

Social deprivation and tooth decay in Scottish schoolchildren.

OBJECTIVES: To quantify the association between dental decay in Scottish Schoolchildren & social deprivation as measured by the Carstairs Index from the 1991 census. DESIGN: An ecological study using data from the Scottish Health Boards Dental Epidemiology Programme and The Public Health Common Data set. SETTING: Scottish Health Boards from 1992-3 to 1994-5. SUBJECTS: Random samples of 5,920 five-year-olds, 5,344 12-year-olds and 6,007 14-year-olds across the 15 Health Boards in Scotland. RESULTS: Positive correlations were demonstrated in all age groups; 12-year-olds (r = 0.72) in 1992-3, 5-year-olds (r = 0.8) in 1993-4 and 14-year-olds (r = 0.55) in 1994-5. Dental decay in all age groups was positively and significantly associated with deprivation as measured by the Carstairs index. CONCLUSION: Tooth decay was confirmed as a disease associated with social deprivation in Scotland. The increasing polarisation of decay to socio-economically deprived groups of the population, suggests a 'whole population' approach such as water fluoridation would prevent tooth decay in these deprived groups most effectively. Nevertheless all evidence-based interventions should be used to try to narrow the "Dental Health Divide' by improving the dental health of deprived individuals.

Adolescent↗
PubMed1996

Signalling by TGF-beta family members: short-range effects of Xnr-2 and BMP-4 contrast with the long-range effects of activin.

BACKGROUND: One way of establishing a morphogen gradient in a developing embryo involves the localized synthesis of an inducing molecule followed by its diffusion into surrounding tissues. The morphogen-like effects of the mesoderm-inducing factor activin provide support for this idea in amphibian development. The questions remain, however, of how activin exerts its long-range effects, and whether long-range signalling is a property of all transforming growth factor beta (TGF-beta) family members. RESULTS: We compare the signalling ranges of activin and two other TGF-beta family members, Xnr-2 and BMP-4. Unlike activin, Xnr-2 and BMP-4 act over short distances. Furthermore, the effects of constitutively active activin receptors are strictly cell-autonomous. These observations suggest that the long-range effects of activin occur through protein diffusion and that "relay' mechanisms are not initiated by any of these TGF-beta family members. Mechanisms limiting the signalling range of Xnr-2 were addressed by studying Xnr-2 processing and secretion. An activin-Xnr-2 fusion protein signals over many cell diameters, suggesting that regulated processing or secretion is one limiting factor. Disaggregation and reaggregation of Xnr-2-producing tissues also extends the range of Xnr-2, suggesting that components of intact tissue restrict spread of the protein. CONCLUSIONS: The long-range effects of activin are likely to occur through the diffusion of activin protein. The short-range effects of Xnr-2 and BMP-4 emphasize that long-range diffusion is not a general property of TGF-beta-related molecules. Finally, signalling ranges may be regulated by constraints on processing or secretion and by interactions with extracellular components of embryonic tissues.

Activin Receptors, Type I↗
PubMed1996

Photoaffinity labelling of a receptor structure for macrophage cytotoxicity inducing factor (MCF).

Monocyte cytotoxicity inducing factor (MCF) is a cytokine derived from CD4+ lymphocytes which was isolated using its ability to cause human blood monocytes to become cytotoxic for tumour targets. An N-terminal derived biologically active peptide (GAAVLEDSQ) of the intact molecule was previously used to demonstrate a single class of high affinity binding sites on human blood monocytes and U937 cells. Photoaffinity labelling was carried out to identify the receptor. The N-terminal nonapeptide fragment (P2) could be cross-linked to a single molecular species on the surface of U937 cells when cross-linking was performed on intact cells. Under fully reducing conditions, this molecule had an average molecular weight of 58 kD. In contrast, the apparent molecular weight when determined under nonreducing conditions was 158-163 kD. Octylglucoside solubilized U937 membranes were then applied to a P2 ligand affinity column. The major protein peak when eluted had an apparent molecular weight of 73 kD when determined by a 12% SDS PAGE gel. Photoaffinity labelling of the ligand affinity product was carried out in solution. Gel electrophoresis under nonreducing conditions demonstrated cross-linking to a 163 kD and a 29 kD protein, while under reducing and denaturing conditions additional species were seen at 97 kD, 67 kD and 50 kD. Photoaffinity cross-linking performed both on whole cells and partially purified receptor in solution, and could be specifically abolished by the addition of unlabelled peptide. These data present evidence that the single high affinity binding site corresponds to a U937 membrane protein having an average molecular weight of 58 kD which exists as an apparent homodimer under native conditions, but may have additional low molecular weight components which were not available for binding and derivatization using intact cells.

Affinity Labels↗
PubMed1996

A T cell clone with three potential TCR alpha chain rearrangements expresses only one receptor combination at the cell surface.

In determining the T cell receptor (TcR) usage of various T cell clones that recognize peptide antigens derived from allergens, a particular clone (AC20) was found, that apparently expressed three different species of mRNA encoding alpha chains. The logical conclusion that the cells were not clonal was refuted by the finding of only a single beta chain rearrangement. One of the alpha chains (V alpha20), was not in frame, but two V alpha8 transcripts of different lengths were both potentially translatable. Sequence analysis suggested that the shorter transcript was generated by a secondary splice event from the longer, through the use of a splice donor sequence encoded by the J alpha38 gene segment. The efficiency of excision of the intervening sequence is such that approximately equal amounts of the long and short transcripts occur in the steady state pool of mRNA. This phenomenon has been reported previously in TcR alpha rearrangements, but it has never been made clear whether these truncated chains can form a functional TcR. Reconstitution of a TcR negative cell line with these transcripts showed that only the full length alpha chain was able to pair efficiently with the beta chain to generate a functional receptor at the cell surface.

Amino Acid Sequence↗
PubMed1996

Dental electro-anaesthesia in children: a pilot study.

Sixty-six children aged 10-17 years, with previous experience of local anaesthesia, had one or more restorations placed while using the Cedeta Mark 2 electronic dental anaesthesia system. A control group of 121 children were treated routinely using injected local anaesthetic. Following all treatment the patients used a 10 cm visual analogue scale to subjectively report the severity of any pain they had experienced. Eighteen (27%) of the study group required injection of local anaesthetic to complete the episode of treatment. The study group had significantly higher pain scores than the control group. It was concluded that further research is warranted to assess the application of this technique to special groups, but that it is not at present a substitute for local anaesthesia by injection in routine paediatric dental practice.

Adolescent↗
PubMed1996

Oligoclonal V gene usage by T lymphocytes in bronchoalveolar lavage fluid from sarcoidosis patients.

The T-cell antigen receptor (TCR) repertoire was examined in lymphocytes isolated from the lungs and blood of 12 sarcoidosis patients and nine control patients. This analysis, by polymerase chain reaction (PCR), examined the variable (V)-domain genes of both the alpha and beta chains of the TCR. This is the first study to examine the usage of all known V alpha gene segments in sarcoidosis. A similar degree of diversity was observed in the TCR repertoire in the lungs and blood of the sarcoidosis patients. However, 11 of the 12 sarcoidosis patients showed an increased use of particular TCR V alpha and V beta genes in lung T cells as compared with blood. The pattern of TCR V gene bias in the lung T cells was specific for each patient. The clonality of selected V genes was examined by determining the third complementarity-determining region (CDR3) length polymorphism of particular PCR products. The majority of lung T cells with biased TCR V gene segments were oligoclonal. Altogether, these results suggest oligoclonal expansion of lung T cells in response to a local antigenic stimulus, with additional nonspecific T-cell accumulation. The variability in the V gene segments used by the expanded T-cell subsets in different sarcoidosis patients may reflect different epitopes or antigens being recognized in the lung, as well as variations in major histocompatibility complex (MHC) haplotype between the patients.

Adult↗
PubMed1996

Bone morphogenetic protein-4 (BMP-4) acts during gastrula stages to cause ventralization of Xenopus embryos.

Injection of RNA encoding BMP-4 into the early Xenopus embryo suppresses formation of dorsal and anterior cell types. To understand this phenomenon, it is necessary to know the stage at which BMP-4 acts. In this paper, we present three lines of evidence showing that BMP-4 misexpression has no effect on the initial steps of mesoderm induction, either dorsal or ventral, but instead causes ventralization during gastrulation. Firstly, activation of organizer-specific genes such as goosecoid, Xnot, pintallavis and noggin occurs normally in embryos injected with BMP-4 RNA, but transcript levels are then rapidly down-regulated as gastrulation proceeds. Similarly, BMP-4 does not affect the initial activation of goosecoid by activin in animal caps, but expression then declines precipitously. Secondly, embryos made ventral by injection with BMP-4 RNA cannot be rescued by grafts of Spemann's organizer at gastrula stages. Such embryos therefore differ from those made ventral by UV-irradiation, where the defect occurs early and rescue can be effected by the organizer. Finally, the dorsalizing effects of the organizer, and of the candidate dorsalizing signal noggin, both of which exert their effects during gastrulation, can be counteracted by BMP-4. Together, these experiments demonstrate that BMP-4 can act during gastrulation both to promote ventral mesoderm differentiation and to attenuate dorsalizing signals derived from the organizer.

Animals↗
PubMed1996

Index of Orthodontic Treatment Need, its use in a dental epidemiology survey calibration exercise.

The Index of Orthodontic Treatment Need (IOTN) was included in a pre-survey calibration exercise of 18 dental epidemiologists. Nine months later 16 of the epidemiologists were re-calibrated in the IOTN. The Index was readily accepted by the experienced dental epidemiological examiners and each examination was estimated to be extended by, on average, less than two minutes. The inter-examiner agreement of the dental health component using the weighted kappa statistic improved from a mean of 0.53 (moderate agreement) for the first calibration to 0.66 (good agreement) at the second calibration. The mean inter-examiner agreement on the aesthetic component using the weighted kappa statistic reduced from 0.52 for the first calibration to 0.49 at the second. Using sensitivity and specificity to measure agreement on the dichotomous decision of those defined as having a definite treatment need, mean sensitivity rose from 0.72 to 0.79 and specificity from 0.90 to 0.97. It is concluded that dental examiners for epidemiological surveys can be trained to use the IOTN by using a pre-calibration exercise and that agreement on the dental health component improves after the index has been used for some months.

Adolescent↗
PubMed1995

Inductive signals. Revolving vertebrates.

An old idea about the relationship between arthropod and vertebrate body plans has been given new life by studies of the signalling genes controlling dorsal and ventral development in Drosophila and Xenopus.

Amino Acid Sequence↗
PubMed1995

Children's ratings of dental injection and treatment pain, and the influence of the time taken to administer the injection.

Three hundred and eight children aged 3-16 years (mean age 10.2), who were undergoing routine dental treatment, recorded on visual analogue scales their ratings of pain associated with injection and treatment. The injection pain scores were examined for their relationship to age, gender, time taken to administer the injection and injection type. The treatment pain scores were compared between groups who had teeth either extracted or restored, and between groups assessed by the operator as having total or partial anaesthesia. A significant inverse correlation was found between subjective injection pain and injection duration. The difference in treatment pain scores was significant between groups assessed by the operator as having total or partial anaesthesia. Inferior dental nerve blocks were rated significantly more painful than buccal infiltrations. Age, gender, and the operative procedure performed had no statistically significant relationship to the injection pain scores or treatment pain scores. The visual analogue pain scale was found to be unsuitable for use by children under 7 years of age.

Administration, Buccal↗
PubMed1995

Nodal-related signals induce axial mesoderm and dorsalize mesoderm during gastrulation.

Mouse embryos homozygous for a null mutation in nodal arrest development at early gastrulation and contain little or no embryonic mesoderm. Here, two Xenopus nodal-related genes (Xnr-1 and Xnr-2) are identified and shown to be expressed transiently during embryogenesis, first within the vegetal region of late blastulae and later in the marginal zone during gastrulation, with enrichment in the dorsal lip. Xnrs and mouse nodal function as dose-dependent dorsoanterior and ventral mesoderm inducers in whole embryos and explanted animal caps. Using a plasmid vector to produce Xnr proteins during gastrulation, we show that, in contrast to activin and other TGF beta-like molecules, Xnr-1 and Xnr-2 can dorsalize ventral marginal zone explants and induce muscle differentiation. Xnr signalling also rescues a complete embryonic axis in UV-ventralized embryos. The patterns of Xnr expression, the activities of the proteins and the phenotype of mouse nodal mutants, all argue strongly that a signaling pathway involving nodal, or nodal-related peptides, is an essential conserved element in mesoderm differentiation associated with vertebrate gastrulation and axial patterning.

Amino Acid Sequence↗