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Biomedical subjects

J Reed

Publications and source records attributed to J Reed.

At least 19 recordsLinked to original sources
PubMed1999

Arsenite induces apoptosis via a direct effect on the mitochondrial permeability transition pore.

The molecular mode of action of arsenic, a therapeutic agent employed in the treatment of acute promyelocytic leukemia, has been elusive. Here we provide evidence that arsenic compounds may act on mitochondria to induce apoptosis. Arsenite induces apoptosis accompanied by a loss of the mitochondrial transmembrane potential (Delta Psim). Inhibition of caspases prevents the arsenite-induced nuclear DNA loss, but has no effect on the Delta Psim dissipation and cytolysis induced by arsenite. In contrast, Bcl-2 expression induced by gene transfer prevents all hallmarks of arsenite-induced cell death, including the Delta Psim collapse. PK11195, a ligand of the mitochondrial benzodiazepine receptor, neutralizes this Bcl-2 effect. Mitochondria are required in a cell-free system to mediate arsenite-induced nuclear apoptosis. Arsenite causes the release of an apoptosis-inducing factor (AIF) from the mitochondrial intermembrane space. This effect is prevented by the permeability transition (PT) pore inhibitor cyclosporin A, as well as by Bcl-2, which is known to function as an endogenous PT pore antagonist. Arsenite also opens the purified, reconstituted PT pore in vitro in a cyclosporin A- and Bcl-2-inhibitible fashion. Altogether these data suggest that arsenite can induce apoptosis via a direct effect on the mitochondrial PT pore.

Apoptosis↗
PubMed1999

Molecular dynamics study of the proposed beta-hairpin form of the switch domain from HIV1 gp120 alone and complexed with an inhibitor of CD4 binding.

The strong tendency of beta-hairpin peptides to aggregate can prevent their structural resolution. The polar form of the switch peptide (LAV 15mer) at the CD4-binding domain of HIV1 gp120 is such a peptide, and NMR investigations of its interaction with a class of CD4-binding inhibitors developed in this laboratory have been hindered. Detailed knowledge of the interaction is required for the development of more potent switch inhibitors, that act by disrupting the cooperative folding transition necessary for binding to the CD4 receptor. In carrying out molecular dynamics simulation of the free peptide under polar conditions, we found that the properties of the resulting structure agree closely with those observed by circular dichroism. The same conditions, used to model the peptide/ inhibitor complex, produced a stable bimolecular structure with specific interactions between the inhibitor and side chains on the peptide, (e.g., Trp12 and the LPCR tetrad), known to control the folding transition. These help explain existing data on the relative potency of inhibitor derivatives and provide a basis for improved inhibitor design.

Algorithms↗
PubMed1999

Spontaneous apoptosis and the expression of p53 and Bcl-2 family proteins in locally advanced head and neck cancer.

BACKGROUND: Apoptotic cell death plays a key role in the pathogenesis, aggressiveness, and therapy responsiveness of cancer. The suicidal machinery of apoptosis is genetically controlled. Proteins of the Bcl-2 family as well as p53 are important regulators of apoptosis. OBJECTIVE: To assess the rate of spontaneous apoptosis and the expression of p53 and Bcl-2 family proteins in locally advanced squamous cell carcinomas of the head and neck. DESIGN: Twenty-six patients with locally advanced squamous cell carcinoma of the head and neck were included in the study. The expression of p53, Bcl-2, Mcl-1, Bax, and Bak was assessed by immunohistochemical analysis. The terminal deoxytransferase-mediated deoxyuridine nick end-labeling assay was used to quantify apoptosis by flow cytometry. RESULTS: Tumor cells containing immunostaining for the antiapoptotic proteins Bcl-2 and Mcl-1 were present in 4 (15%) and 24 (92%) of the cases evaluated, respectively, whereas immunopositivity for the proapoptotic proteins Bax and Bak was found in 9 (35%) and 24 (92%) of the samples. Immunoreactivity to p53 was detected in 20 (77%) of the samples. There was a positive correlation between the expression of Bcl-2 and Bax and between Mcl-1 and Bak. A low fraction of apoptotic cells (<2.5%) in the pretreatment tumor samples was significantly correlated with increased 2-year survival in these patients. CONCLUSIONS: Our results establish the frequent expression of the Bcl-2 family proteins Bcl-2, Mcl-1, Bax, and Bak in locally advanced head and neck cancer. In addition, this study suggests that the apoptotic fraction in pretreatment tumor samples might be of prognostic importance for the outcome in these patients.

Adult↗
PubMed1999

Direct effects of phosphorylation on the preferred backbone conformation of peptides: a nuclear magnetic resonance study.

Control of protein activity by phosphorylation appears to work principally by inducing conformational change, but the mechanisms so far reported are dependent on the structural context in which phosphorylation occurs. As the activity of many small peptides is also regulated by phosphorylation, we decided to investigate possible direct consequences of this on the preferred backbone conformation. We have performed 1H nuclear magnetic resonance (NMR) experiments with short model peptides of the pattern Gly-Ser-Xaa-Ser, where Xaa represents Ser, Thr, or Tyr in either phosphorylated or unphosphorylated form and with either free or blocked amino and carboxy termini. The chemical shifts of amide protons and the 3JNH-Halpha coupling constants were estimated from one-dimensional and two-dimensional scalar correlated spectroscopy (COSY) spectra at different pH values. The results clearly indicate a direct structural effect of serine and threonine phosphorylation on the preferred backbone dihedrals independent of the presence of charged groups in the surrounding sequence. Tyrosine phosphorylation does not induce such a charge-independent effect. Additionally, experiments with p-fluoro- and p-nitro-phenylalanine-containing peptides showed that the mere presence of an electronegative group on the aromatic ring of tyrosine does not produce direct structural effects. In the case of serine and threonine phosphorylation a strong dependence of the conformational shift on the protonation level of the phosphoryl group could be observed, showing that phosphorylation induces the strongest effect in its dianionic, i.e., physiological, form. The data reveal a hitherto unknown mechanism that may be added to the repertoire of conformational control of peptides and proteins by phosphorylation.

Amino Acid Sequence↗
PubMed1999

Discharging older people from hospital to care homes: implications for nursing.

This paper summarizes a research study which explored the experiences of older people being discharged from hospital to nursing and residential homes in the North East of England. While there has been considerable research which has looked at the discharge of patients from hospital to their own homes, little literature could be found which addressed discharge to care homes. While this may reflect an assumption that this form of discharge is less problematic, it is arguable that this is only the case for staff - there is a body of literature on re-location which suggests that the move to a care home is a major life event for older people. Taking a qualitative approach, this study interviewed 20 older people and 17 of their family members after discharge from hospital to a care home. We found that few people had been offered opportunities to discuss their move with nurses, and that older people tended to adopt a stoical attitude. In focus groups, interviews and written responses from 23 members of staff in the hospital and in care homes, we found that there was a lack of clarity over whose role it was to initiate such discussions. The paper concludes with some discussion of the implications for nursing practice of changing care interfaces.

Aged↗
PubMed1999

Children's attitudes toward interacting with peers with different craniofacial anomalies.

OBJECTIVE: This study was designed to evaluate children's understanding of different craniofacial anomalies and their willingness to interact with children with such anomalies. DESIGN: This was a between-measures design in which children were randomly allocated to one of three groups. Each group viewed one of three pairs of computer-generated images (nondistinctive, cleft lip, or misshapen nose) of similar-aged children. SETTING: Participants were recruited from two city elementary schools and were interviewed at their schools. PARTICIPANTS: A total of 100 children (aged 7 to 10 years) entered the study, and complete sets of data were obtained for each child. As the majority of the children were white (n = 92), the nonwhite children (n = 8) were excluded from the data analyses. MAIN OUTCOME MEASURES: Participants were asked a number of questions to ascertain their thoughts about the image, and measures were then taken of each child's willingness to interact with the stimulus child. RESULTS: There were no significant differences between the three groups. Boys were significantly more willing to interact with the stimulus images than were girls, and there was a nonsignificant trend for girls to be more likely to spontaneously mention the craniofacial anomaly. Participants gave varied explanations for the condition's causation. CONCLUSIONS: Boys and girls differed in their willingness to interact with unfamiliar peers with and without facial distinctions. Various explanations were given to explain causality of the anomaly. Findings lend some support to the proposal that high "background attractiveness" can overshadow the impact of a craniofacial anomaly.

Analysis of Variance↗
PubMed1998

Salt wars.

Explore the source record for details and available documents.

Health Policy↗
PubMed1998

Automated high resolution optical mapping using arrayed, fluid-fixed DNA molecules.

New mapping approaches construct ordered restriction maps from fluorescence microscope images of individual, endonuclease-digested DNA molecules. In optical mapping, molecules are elongated and fixed onto derivatized glass surfaces, preserving biochemical accessibility and fragment order after enzymatic digestion. Measurements of relative fluorescence intensity and apparent length determine the sizes of restriction fragments, enabling ordered map construction without electrophoretic analysis. The optical mapping system reported here is based on our physical characterization of an effect using fluid flows developed within tiny, evaporating droplets to elongate and fix DNA molecules onto derivatized surfaces. Such evaporation-driven molecular fixation produces well elongated molecules accessible to restriction endonucleases, and notably, DNA polymerase I. We then developed the robotic means to grid DNA spots in well defined arrays that are digested and analyzed in parallel. To effectively harness this effect for high-throughput genome mapping, we developed: (i) machine vision and automatic image acquisition techniques to work with fixed, digested molecules within gridded samples, and (ii) Bayesian inference approaches that are used to analyze machine vision data, automatically producing high-resolution restriction maps from images of individual DNA molecules. The aggregate significance of this work is the development of an integrated system for mapping small insert clones allowing biochemical data obtained from engineered ensembles of individual molecules to be automatically accumulated and analyzed for map construction. These approaches are sufficiently general for varied biochemical analyses of individual molecules using statistically meaningful population sizes.

Animals↗
PubMed1998

A quantitative study of optical mapping surfaces by atomic force microscopy and restriction endonuclease digestion assays.

Many new techniques in biomolecular chemistry and genomic analysis require the immobilization of molecular reagents on specially prepared surfaces. However, the process of molecular fixation often interferes with or precludes the use of standard in vitro biochemical assays. Optical mapping is an emergent technology for genomic analysis which relies on the biochemical activity of DNA fixed to silanized glass surfaces. Optical mapping surfaces have been shown to be compatible with restriction endonucleases and a variety of DNA polymerases. The essential properties of biochemically active surfaces are poorly understood in most of the current technologies which utilize molecular fixation, including optical mapping. The purpose of this study is to use the powerful technique of atomic force microscopy, in combination with informative enzymatic assays, to correlate biochemical activity with microscopic surface structure. The results presented provide meaningful insight into the effect of surface preparation on the biochemical accessibility of surface-bound molecules. Novel analysis which may facilitate the automation of optical mapping is presented.

DNA↗
PubMed1998

The thiol crosslinking agent diamide overcomes the apoptosis-inhibitory effect of Bcl-2 by enforcing mitochondrial permeability transition.

In several different cell lines, Bcl-2 prevents the induction of apoptosis (DNA fragmentation, PARP cleavage, phosphatidylserine exposure) by the pro-oxidant ter-butylhydroperoxide (t-BHP) but has no cytoprotective effect when apoptosis is induced by the thiol crosslinking agent diazenedicarboxylic acid his 5N,N-dimethylamide (diamide). Both t-BHP and diamide cause a disruption of the mitochondrial transmembrane potential delta psi(m) that is not inhibited by the broad spectrum caspase inhibitor z-VAD.fmk, although z-VAD.fmk does prevent nuclear DNA fragmentation and poly(ADP-ribose) polymerase cleavage in these models. Bcl-2 stabilizes the delta psi(m) of t-BHP-treated cells but has no inhibitory effect on the delta psi(m) collapse induced by diamide. As compared to normal controls, isolated mitochondria from Bcl-2 overexpressing cells are relatively resistant to the induction of delta psi(m) disruption by t-BHP in vitro. Such Bcl-2 overexpressing mitochondria also fail to release apoptosis-inducing factor (AIF) and cytochrome c from the intermembrane space, whereas control mitochondria not overexpressing Bcl-2 do liberate AIF and cytochrome c in response to t-BHP. In contrast, Bcl-2 does not confer protection against diamide-triggered delta psi(m) collapse and the release of AIF and cytochrome c. This indicates that Bcl-2 suppresses the permeability transition (PT) and the associated release of intermembrane proteins induced by t-BHP but not by diamide. To further investigate the mode of action of Bcl-2, semi-purified PT pore complexes were reconstituted in liposomes in a cell-free, organelle-free system. Recombinant Bcl-2 or Bcl-X(L) proteins augment the resistance of reconstituted PT pore complexes to pore opening induced by t-BHP. In contrast, mutated Bcl-2 proteins which have lost their cytoprotective potential also lose their PT-modulatory capacity. Again, Bcl-2 fails to confer protection against diamide in this experimental system. The reconstituted PT pore complex itself cannot release cytochrome c encapsulated into liposomes. Altogether these data suggest that pro-oxidants, thiol-reactive agents, and Bcl-2 can regulate the PT pore complex in a direct fashion, independently from their effects on cytochrome c. Furthermore, our results suggest a strategy for inducing apoptosis in cells overexpressing apoptosis-inhibitory Bcl-2 analogs.

Apoptosis↗
PubMed1998

Second trimester abortion using prostaglandin E2 suppositories with or without intracervical Laminaria japonica: a randomized study.

OBJECTIVE: To compare the abortifacient efficacy of vaginal prostaglandin (PG) E2 suppositories with and without pretreatment with intracervical PGE2 gel and Laminaria japonica. METHODS: One hundred seventy-five women between 16 and 20 weeks' gestation requesting abortion were divided randomly into three groups. Forty-one received PGE2 vaginal suppositories alone to induce abortion, 72 had Laminaria placed 24 hours before vaginal PGE2, and 62 were given intracervical 0.5 mg PGE2 gel plus Laminaria 24 hours before vaginal PGE2. Maternal demographic characteristics, induction to delivery time, number of suppositories required, and complications were analyzed. RESULTS: The mean maternal age, gravidity, parity, race, and gestational age were similar among groups. Ninety-five percent of all women delivered within 24 hours. The induction to delivery time was significantly longer in the PGE2-only women (mean +/- standard deviation 689+/-319 minutes) compared with that of those receiving PGE2 plus Laminaria (487+/-321 minutes) and PGE2 plus Laminaria plus gel (547 +/-374 minutes, P = .01). There was a statistically significant difference in the number of suppositories needed to complete the abortion process. The PGE2-only group required more suppositories (median three, range 1-8) compared with PGE2 plus Laminaria (median 3, range 1-3) and PGE2 plus Laminaria plus gel (median 2.5, range 1-9; P = .001). Patients in the PGE2 plus Laminaria plus gel group reported more pain associated with placement (median pain score 4, range 0-10) compared with PGE2 plus Laminaria (median 2, range 0-9; P = .003). There was a lower incidence of febrile episodes in the PGE2-only group (29%) compared with PGE2 plus Laminaria (68%) and PGE2 plus Laminaria plus gel (54%, P = .002). CONCLUSION: Placement of Laminaria japonica 24 hours before PGE2 vaginal suppository-induced abortion resulted in a significantly shorter induction-to-delivery time, and pretreatment with Laminaria japonica decreased the number of suppositories required to complete abortion. Pretreatment with intracervical PGE2 gel increased pain associated with Laminaria placement and did not improve the efficacy of the procedure.

Abortion, Induced↗
PubMed1998

The importance of place for older people moving into care homes.

In an increasingly mobile society, the importance of place can be forgotten in the process of making decisions about where older people should receive care. There is some evidence, however, that issues of location and place are important to older people, both in the way that their sense of self is developed and maintained through identification with places and through their ability to participate in activities. This paper describes a research study which followed a group of older people through the process of moving into care homes, and discusses some of the data produced, which indicated how a sense of place shaped the process of moving.

Aged↗