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Biomedical subjects

P Sellers

Publications and source records attributed to P Sellers.

At least 19 recordsLinked to original sources
PubMed1999

A comparative study of the Philips HP 735, Braun/Oral B D7 and the Oral B 35 Advantage toothbrushes.

The aim of this 3-group, 3-treatment, single-blind, parallel group study was to evaluate and compare the efficacies of the Philips/Jordan HP 735 powered toothbrush, the Braun/Oral B D7 powered toothbrush and the Oral B Advantage B35 manual toothbrush in a cohort of 75 young adults (18-25 years). Following an appointment for screening, full mouth mean (+/-sd) modified Turesky plaque index (PI) and Löe & Silness gingival index (GI) were recorded at baseline. After 24 h abstinence from all oral hygiene measures, PI was recorded and each subject was given one of the test brushes with detailed instructions for use. The subject then brushed under supervision for 90 s, during which time mean (+/-sd) toothbrushing forces (TBF) were recorded. PI were recorded immediately after supervised brushing and the subjects were then discharged for 6 weeks to use the allocated toothbrush at home. After 6 weeks, PI, GI and TBF were again recorded. Comparisons between the brushing groups for all parameters, at baseline, 24 h and 6 weeks were tested using ANCOVA. There were no significant differences for PI and GI between groups at baseline, or for PI following supervised brushing at 24 h. After 6 weeks subjects using the powered brushes had lower mean PI (+/-sd) scores than those using manual brushes but the differences were significant only at interproximal sites; HP 735 1.44 (0.52), D7 1.44 (0.53), B35 1.75 (0.51) (p=0.05). At 6 weeks, mean (+/-sd) GI were; HP 735 1.49 (0.21), D7 1.61 (0.21), B35 1.64 (0.22) (p=0.033). Mean GI scores for the HP 735 group were similar at baseline and at 6 weeks although for the other brushes, the GI scores actually increased over this period. Mean (+/-sd) TBF (grammes/force) at baseline and 6 weeks respectively for the brushes were; HP 735 233 (205), D7 159 (58), B35 279 (122) (p=0.026): HP 735 194 (86), D7 141 (57), B35 297 (113) (p=0.0001). The within-group variability for the HP 735 TBF reduced considerably over 6 weeks, a likely consequence of the click-force threshold feature of this brush.

Adolescent↗
PubMed1997

Enumeration of flux routes through complex biochemical reactions.

In the present work, a general algorithm for enumeration of the flux routes via which a chemical species "flows" through a complex biochemical reaction is outlined and presented by way of a biological example, a kinetic model for potassium ion permeation through a voltage-gated ion channel. The algorithm is readily amenable to computer based implementation and when used in conjunction with an existing algorithm provides a convenient means for simulation of the equilibrium and steady-state isotope exchange kinetics of complex biochemical reactions.

Algorithms↗
PubMed1996

Adsorption during Heat Treatment Related to the Thermal Unfolding/Aggregation of beta-Lactoglobulins A and B

Adsorption onto chromium surfaces during heat treatment (65-68°C) of beta-lactoglobulin A and B in phosphate buffer, pH 6.88, was investigated by in situ ellipsometry. Thermal unfolding and in situ heat-induced aggregation under the same conditions were studied by differential scanning calorimetry and dynamic light scattering, respectively. A lag phase was observed for buildup of the thickness of the adsorbed layer of beta-lactoglobulin A and B at 68°C. The lag period was found to increase with decreasing temperature. The difference in adsorption rate between the variants was found to be in agreement with the aggregation difference in solution, that is, the A variant aggregated more rapidly after a somewhat slower beginning. These results could be related to a difference in the thermal unfolding behavior. By applying a non-two state model the second of two transitions was suggested to involve the activation of the free sulfydryl group. The results indicated that fouling proceeds via attachment of aggregates with an activated free sulfydryl group through an interchange reaction with a disulfide bond in the adsorbed protein layer.

Journal Article↗
PubMed1995

Thermal denaturation of beta-lactoglobulin: effect of protein concentration at pH 6.75 and 8.05.

Previous work on the thermal denaturation of beta-lactoglobulin at about neutral pH and concentrations generally above 50 mg/ml has shown that the temperature of the maximum in the thermogram increases only slightly with concentration. Likewise, there is little if any concentration dependence at acid pH over a wide concentration range. However, so far as we are aware, no work has been described on the thermal denaturation of beta-lactoglobulin in the physiological range of protein concentration and pH appropriate to milk. We report measurements at pH 6.75 and 8.05 in the concentration range 2-120 mg/ml and show that below about 50 mg/ml the position of the maximum becomes strongly dependent on concentration, passing through a minimum near 25 mg/ml and increasing towards the lowest concentrations where measurements were practicable. Moreover, the narrow, well defined and nearly symmetrical thermal transition observed at high protein concentrations contrasts with a broader and more asymmetric curve at lower concentrations. An explanation for the behaviour seen at the lower protein concentrations is suggested, based on the temperature- and concentration-dependent dissociation of the beta-lactoglobulin dimer and an associated conformational transition. The position of the maximum in the thermogram has a marked dependence on the rate of heating down to the lowest rate investigated of 10 degrees C per hour, showing the importance of slow kinetic effects in the denaturation of this protein.

Animals↗
PubMed1994

A calbindin D9k mutant with reduced calcium affinity and enhanced cooperativity. Metal ion binding, stability, and structural studies.

In the native calcium-binding protein calbindin D9k (M(r) 8.700; 75aa; 2 EF-hands), the backbone carbonyl oxygen of Glu60 coordinates the Ca2+ ion in the C-terminal site (site II). The carboxylate group of the same residue forms a hydrogen bond to a water molecule that constitutes a Ca2+ ligand in the N-terminal site (site I). The mutant E60D, with the charge-conserving substitution Glu60-->Asp, has been prepared to study the role of Glu60 in subjoining the two Ca(2+)-binding sites and its role in the cooperative Ca2+ binding. Ca(2+)-binding studies of the mutant show that the overall affinity for calcium has decreased by a factor of 38 in comparison with wild-type calbindin D9k. The largest reduction is seen in the first macroscopic binding step. The Ca2+ affinities for both sites in the protein are reduced to a similar extent. In contrast, the mutation leads to a large increase in the cooperativity of calcium binding. Differential scanning calorimetry has been used to determine the thermal stability which is almost as high as in the wild-type protein. Cadmium binding has been assessed with 1H and 113Cd NMR. X-ray crystallographic studies of the E60D mutant in its calcium-bound form show very small structural changes relative to the wild-type protein. Almost all differences are within the error limits of the method. The largest crystallographic effects are seen in the crystal packing. Two E60D molecules with slightly different structure are found in the asymmetric unit in contrast to the single molecule in the wild-type crystal.(ABSTRACT TRUNCATED AT 250 WORDS)

Aspartic Acid↗
PubMed1994

The effect of tecogalan sodium on the development of the collateral circulation after acute arterial occlusion.

The survival of tissues distal to major arterial occlusions is critically dependent on the development of collateral blood vessels. The biochemical events fundamental to their evolution remain undefined. To determine the role of endogenous basic fibroblast growth factor in the development of the collateral circulation we evaluated the dose response to Tecogalan Sodium (DS-4152) in a rat model of hind limb ischaemia. Treatment with DS-4152 inhibited the recovery following acute arterial occlusion and impaired limb viability. The inhibitory effects were more pronounced in the calf than in the adductor muscles and tended to be more marked in the groups treated with the higher dose regimens. The results provide further evidence that collateral development and tissue recovery following acute arterial occlusion are dependent on endogenous basic fibroblast growth factor and that this dependence is greater in the more ischaemic, distal tissues.

Acute Disease↗
PubMed1994

The role of PDGF-BB on the development of the collateral circulation after acute arterial occlusion.

The survival of tissues in the presence of arterial occlusion is critically dependent on the development of collateral blood vessels. Identification of the biochemical mediators and their mechanism of action is fundamental to an understanding of the evolution of the collateral circulation. The ability of PDGF-BB to promote this was evaluated in an animal model of hind limb ischaemia. In order to obtain significant quantities of this mitogen for use in our animal model, human recombinant PDGF-BB was expressed in a Chinese Hamster Ovary (CHO) cell line. The transfected CHO cells produced 544 micrograms/l of PDGF-BB in serum free medium (SFM). The 30 kDa form of PDGF-BB was purified to homogeneity as judged by silver staining and amino-acid sequencing. Purified PDGF-BB was shown to be bioactive by a cell proliferation assay. The exogenous administration of PDGF-BB enhanced the recovery of blood flow after acute arterial occlusion. The results suggest that PDGF-BB may have therapeutic value in promoting collateral development following arterial occlusion.

Acute Disease↗
PubMed1993

Disulfide bonds in homo- and heterodimers of EF-hand subdomains of calbindin D9k: stability, calcium binding, and NMR studies.

The effect of decreased protein flexibility on the stability and calcium binding properties of calbindin D9k has been addressed in studies of a disulfide bridged calbindin D9k mutant, denoted (L39C + P43M + I73C), with substitutions Leu 39-->Cys, Ile 73-->Cys, and Pro 43-->Met. Backbone 1H NMR assignments show that the disulfide bond, which forms spontaneously under air oxidation, is well accommodated. The disulfide is inserted on the opposite end of the protein molecule with respect to the calcium sites, to avoid direct interference with these sites, as confirmed by 113Cd NMR. The effect of the disulfide bond on calcium binding was assessed by titrations in the presence of a chromophoric chelator. A small but significant effect on the cooperativity was found, as well as a very modest reduction in calcium affinity. The disulfide bond increases Tm, the transition midpoint of thermal denaturation, of calcium free calbindin D9k from 85 to 95 degrees C and Cm, the urea concentration of half denaturation, from 5.3 to 8.0 M. Calbindins with one covalent bond linking the two EF-hand subdomains are equally stable regardless if the covalent link is the 43-44 peptide bond or the disulfide bond. Kinetic remixing experiments show that separated CNBr fragments of (L39C + P43M + I73C), each comprising one EF-hand, form disulfide linked homodimers. Each homodimer binds two calcium ions with positive co-operativity, and an average affinity of 10(6) M-1. Disulfide linkage dramatically increases the stability of each homodimer. For the homodimer of the C-terminal fragment Tm increases from 59 +/- 2 without covalent linkage to 91 +/- 2 degrees C with disulfide, and Cm from approximately 1.5 to 7.5 M. The overall topology of this homodimer is derived from 1H NMR assignments and a few key NOEs.

Animals↗
PubMed1993

Plasma amyloid precursor protein is decreased in Alzheimer's disease.

Alzheimer's disease is characterized by amyloid deposits whose major protein component is beta A4. beta A4 is a product of the amyloid precursor protein (APP). APP was assayed in partially purified plasma samples from 16 sporadic Alzheimer's disease patients, 12 age-matched controls, 15 Down's syndrome individuals aged 19-36 years and 8 young to middle-aged controls (22-51 years). 14 of the 16 Alzheimer's disease patients had decreased plasma APP when compared with age-matched controls. 14 of the 15 Down's syndrome individuals had similar levels of APP when compared with age-matched and elderly non-demented controls by immunoblotting, whereas one had levels of APP less than controls. Taken together with results from a previous report (Lancet 1992; 340: 453-454), the decreased plasma APP levels mirror the changes observed with cerebrospinal fluid APP levels in Alzheimer's disease.

Adult↗
PubMed1992

Dissection of calbindin D9k into two Ca(2+)-binding subdomains by a combination of mutagenesis and chemical cleavage.

Calbindin D9k is a 75-residue globular protein made up of two Ca2+ binding subdomains of the EF-hand type. In order to examine the subdomains independently, a method was devised to selectively cleave the loop between them. Using site-directed mutagenesis, a unique methionine was substituted for Pro43 in the loop, thus allowing cleavage using cyanogen bromide. Agarose gel electrophoresis shows that the fragments have a high affinity for one another, although less so in the absence of calcium. 1H-NMR spectra of the fragments indicate that the structures of the heterodimers are changed little from that of the intact protein. However, the Ca2+ binding constants of the individual subdomains are several orders of magnitude lower than for the corresponding sites in the uncleaved protein.

Calbindins↗
PubMed1991

Ca2+ binding in proteins of the calmodulin superfamily: cooperativity, electrostatic contributions and molecular mechanisms.

In a large number of intracellular regulatory proteins of the calmodulin superfamily a pair of closely interacting helix-loop-helix Ca2+ binding sites ('EF hands') constitute the functional unit--an arrangement that enables cooperative binding. We have recently made detailed experimental studies of the binding of Ca2+ ions to calmodulin, its tryptic fragments TR1C and TR2C (which each constitute a globular domain of a pair of EF hands) and calbindin D9k. Macroscopic Ca2+ binding constants have been obtained over a range of ionic strengths (0 to 0.15 M KCl). For calmodulin the measurements indicate that the two separate globular domains TR1C and TR2C retain the Ca2+ binding properties they have in the intact molecule, with positive cooperativity within each domain. The absolute value of the free energy of interaction between the two sites in each domain, a measure of the cooperativity, increases with ionic strength and is greater than or equal to 10 kJ mol-1 at 0.15 M KCl. Two-dimensional 1H NMR studies show that the addition of KCl does not alter the conformation of the protein. In the case of calbindin D9k several categories of mutants have been studied. One group encompasses the effect of protein surface charges 5 to 15 A from the Ca2+ binding sites. Two-dimensional 1H NMR shows that neither the addition of KCl, nor mutations that neutralize the surface charges, change the protein conformation. Although the global structure of calbindin D9k is largely unchanged upon binding of calcium, the structure with only one cation bound is more similar to the (Ca2+)2 form. Interestingly, the dynamical properties of the Ca(2+)-free and the (Ca2+)2-forms of calbindin differ greatly. For example, the rate of NH/ND exchange of the Ca(2+)-free form is on average 200 times faster than that of the (Ca2+)2-form. The results obtained so far point to a non-negligible entropic contribution to the observed cooperativity of Ca2+ binding.

Amides↗
PubMed1991

Thermodynamics of Ca2+ binding to calmodulin and its tryptic fragments.

The binding of Ca2+ to calmodulin and its two tryptic fragments has been studied using microcalorimetry. The binding process is accompanied by the uptake or release of protons, depending on the ionic strength. With no added salt, the total enthalpy change for the binding of four calcium ions to calmodulin is -41 kJ mol-1 but in the presence of 0.15 mM KCl delta Htot is +17 kJ mol-1. The mode of binding of Ca2+ is also completely different with and without added salt. It is also shown that for the C-terminal fragment of calmodulin, TR2C, the drastic reduction in delta Gtot for the binding process on increasing the ionic strength is largely an enthalpic effect. Domain interactions in calmodulin are indicated by the fact that the sum of the enthalpies of calcium binding to the two tryptic fragments is not the same as the total binding enthalpy to calmodulin itself. The binding of Ca2+ to calmodulin has also been studied calorimetrically at different temperatures in the range 21-37 degrees C. delta Cp is large and negative in this interval.

Animals↗
PubMed1987

Structure-function relationships in EF-hand Ca2+-binding proteins. Protein engineering and biophysical studies of calbindin D9k.

Genes encoding the minor A component of bovine calbindins D9k--the smallest protein known with a pair of EF-hand calcium-binding sites--with amino acid substitutions and/or deletions have been synthesized and expressed in Escherichia coli and characterized with different biophysical techniques. The mutations are confined to the N-terminal Ca2+-binding site and constitute Pro-20----Gly (M1), Pro-20----Gly and Asn-21 deleted (M2), Pro-20 deleted (M3), and Tyr-13----Phe (M4). 1H, 43Ca, and 113Cd NMR studies show that the structural changes induced are primarily localized in the modified region, with hardly any effects on the C-terminal Ca2+-binding site. The Ca2+ exchange rate for the N-terminal site changes from 3 s-1 in the wild-type protein (M0) and M4 to 5000 s-1 in M2 and M3, whereas there is no detectable variation in the Ca2+ exchange from the C-terminal site. The macroscopic Ca2+-binding constants have been obtained from equilibration in the presence of the fluorescent chelator 2-[[2-[bis(carboxymethyl)-amino]- 5-methylphenoxy]methyl]-6-methoxy-8-[bis(carboxymethyl)amino]quinoline or by using a Ca2+-selective electrode. The Ca2+ affinity of M4 was similar to that of M0, whereas the largest differences were found for the second stoichiometric step in M2 and M3. Microcalorimetric data show that the enthalpy of Ca2+ binding is negative (-8 to -13 kJ.mol-1) for all sites except the N-terminal site in M2 and M3 (+5 kJ.mol-1). The binding entropy is strongly positive in all cases. Cooperative Ca2+ binding in M0 and M4 was established through the values of the macroscopic Ca2+-binding constants. Through the observed changes in the 1H NMR spectra during Ca2+ titrations we could obtain ratios between site binding constants in M0 and M4. These ratios in combination with the macroscopic binding constants yielded the interaction free energy between the sites delta delta G as -5.1 +/- 0.4 kJ.mol-1 (M0) and less than -3.9 kJ.mol-1 (M4). There is evidence (from 113Cd NMR) for site-site interactions also in M1, M2, and M3, but the magnitude of delta delta G could not be determined because of sequential Ca2+ binding.

Aminoquinolines↗
PubMed1983

Effect of retinoids on oral leukoplakia.

This two phase study was designed to observe the toxicity and effectiveness of retinoids on oral leukoplakia. The study design included patients who had visible and measurable oral leukoplakia without history of synchronous oral cancer or oral cancer within the previous 2 years. Documentation of the lesion by direct measurement and photography, as well as a biopsy at the beginning and at the end of the study (for verification of the histologic appearance of the lesion), were performed in each patient. Sixteen patients with oral leukoplakia were treated with 13-cis-retinoic acid formulated in a troche in the strength of 1 mg. Three patients received 3 mg/day, 8 patients received 5 mg/day, and 5 patients received 10 mg/day. The initial visible responding event appeared to be a thinning of the leukoplakia with reduction in the whitish surface leaving a reddish, velvety epithelium. If resolution occurred, the velvety area became pink, assuming the color and texture of the normal adjacent mucosa. Toxicity of the drug appeared to be acceptable among the evaluable patients. Of the 11 patients, 3 demonstrated complete response and 6 demonstrated partial response after 6 months of treatment with the drug under study. Recurrences developed in two of the three patients with complete response, and neither of the two showed complete histologic and cytologic regression. One of the patients with partial response went on to complete response after cessation of treatment. Under the condition of our study, a treatment effect was observed with small amounts of topical 13-cis-retinoic acid and that the level of toxicity was acceptable.

Administration, Topical↗