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Biomedical subjects

F McDonald

Publications and source records attributed to F McDonald.

At least 19 recordsLinked to original sources
PubMed1998

An analysis of Ca2+ release by DGEA: mobilization of two functionally distinct internal stores in Saos-2 cells.

Osteoblasts can be activated by their collagen matrix and in particular the DGEA peptide motif. We have reported that DGEA is able to activate Ca2+ signaling pathways in the human osteoblast-like cell line, Saos-2, by a tyrosine kinase-dependent pathway (T. J. McCann, W. T. Mason, M. C. Meikle, and F. McDonald. Matrix Biol. 16: 271-280, 1997). In the present study, we show that this activity is due to coupling of the signal to intracellular Ca2+ stores, since the DGEA action is not blocked by La3+ but is lost when Ca2+ stores are depleted with 2 microM and blocked by 10 microM ryanodine. The activated stores also differ functionally from those activated by thrombin, as blockade with U-73122 obstructs only thrombin-activated Ca2+ release. We have shown that the DGEA activity was not due to its high-charge density, since the two acidic residues can be substituted with their uncharged homologues (asparagine and glutamine) without significant loss of activity. This was in turn measured by an adhesion assay that also demonstrated this level of specificity. Furthermore, by constructing DGEA bound to FITC, we have shown that DGEA binding was dependent on divalent cations. We have also demonstrated that an intact actin cytoskeleton is not required for Ca2+ activation by inhibiting actin polymerization with the addition of cytochalasin B. These data strengthen the argument that collagen has a significant role in regulating osteoblast function via this peptide motif.

Amino Acid Sequence↗
PubMed1997

Mineral trioxide aggregate stimulates a biological response in human osteoblasts.

We report a novel material that appears to stimulate cytokine production in human osteoblasts and allow good adherence of the cells to the material. We have examined cultured osteoblasts (MG-63) in the presence of mineral trioxide aggregate (MTA) as set in moist conditions; secondly, we examined the behavior of these MG-63 cells with respect to cytokine and osteocalcin production and alkaline phosphatase activity. Standard ELISA assays were used for assessment of interleukin (IL)-1 alpha, IL-1 beta, IL-6, macrophage colony stimulating factor (M-CSF), and osteocalcin. Furthermore the levels of alkaline phosphatase were measured to establish the level of differentiation of the cells. Cells without MTA served as controls. Cells also were grown in the presence of polymethylmethacrylate (PMA), the commonly used orthopedic cement. In all dishes cells were seen adhering to the base and MTA at 6 h and had increased to confluence at 144 h. IL-1 alpha (175.1 +/- 32.6 pg/mL), IL-1 beta (154.0 +/- 26.7 pg/mL), and IL-6 (214.7 +/- 21.8 pg/mL) were raised when the cells were grown in the presence of MTA at 144 h, with raised values at all time intervals. M-CSF appeared to be unaffected although the overall value was high (7,045.0 +/- 89.5 pg/mL). In contrast, cells grown in the absence of MTA produced negligible amounts of these cytokines (< pg/mL) as did those cells grown in the presence of PMA. Osteocalcin production increased when cells were grown on MTA from 3.8 +/- 0.87 ng/mL to 19.7 +/- 2.8 ng/mL. No osteocalcin could be detected with PMA. Cells in contact with MTA also appeared to have levels of alkaline phosphatase similar to those reported elsewhere (4.3 +/- 0.21 mumol/mg protein/min). No cells could be found attached to PMA and so no alkaline phosphatase activity could be measured.

Alkaline Phosphatase↗
PubMed1997

A collagen peptide motif activates tyrosine kinase-dependent calcium signalling pathways in human osteoblast-like cells.

A collagen peptide motif (DGEA) which is a putative alpha 2 beta 1 integrin binding site was examined for its ability to activate Ca2+ signalling pathways in the human osteoblast-like cell line SaOS-2. We show that these cells express both alpha 2 beta 1 integrin subunits (by immunocytochemistry) and that an anti-beta 1 monoclonal antibody (DF5) mobilizes Ca2+ in these cells. DGEA elevated intracellular Ca2+ in fura-2-loaded cells, in a concentration- and sequence-dependent fashion, with an EC50 of 250 microM. The tyrosine kinase inhibitor herbimycin A reduced the number of cells responding to DGEA and to transforming growth factor alpha. Thrombin also stimulated a rise in intracellular Ca2+, but the number of cells responding was not reduced by herbimycin A. The DGEA response was dependent on extracellular Ca2+, but was not due to Ca2+ influx, since it was blocked by thapsigargin and not by lanthanum. Using three different anti-alpha 2 monoclonal antibodies, we were unable to show that the DGEA-induced Ca2+ signal was mediated by the alpha 2 beta 1 integrin. In summary, the DGEA collagen motif does appear to activate receptor-mediated Ca2+ signalling events in SaOS-2 cells, in a divalent cation-dependent manner, but we were unable to demonstrate a role for alpha 2 beta 1 integrin in this response.

Benzoquinones↗
PubMed1996

Calcium waves in fluid flow stimulated osteoblasts are G protein mediated.

Calcium (Ca2+) entry upon cell perturbation has been examined in transformed human osteoblast cells (U-2/OS). The cells were deformed by fluid flow from a patch pipette held in proximity to the cell by applying a positive pressure (+50 mm Hg) for the passage of saline over the membrane. Intracellular calcium [Ca2+]i was examined following loading with 5 microM Fura-2 AM. The changes in ratio were determined at 330-ms intervals. Waves of [Ca2+]i were seen spreading along the length of the individual cell following stimulation (n = 30). The initial change in Ca2+ at the site of stimulation occurred within 660 ms after applying the stimulus. Following 1.3 (+/- 0.33) s of raised [Ca2+]i, the values returned to those of predeformation. The Ca2+ response following fluid flow stimulation was blocked by 300 microM Cd2+, a specific blocker of Ca2+ channels, demonstrating an extracellular source of Ca2+. Preincubation with cholera toxin (250 ng/ml for 6 h) prolonged the elevation of Ca2+ induced by fluid flow stimulation (n = 20). In contrast, pertussis toxin (250 ng/ml for 6 h) completely eliminated the Ca2+ response to fluid flow stimulation (n = 20). Cells maintained in solutions free of Ca2+ demonstrated no change in [Ca2+]i. Tetraethylammonium (6 mM) had no effect on the response (n = 10). In addition pretreatment with ryanodine (2 and 10 microM; each group n = 10) in media showed a reduced wave of Ca2+ in response to mechanical deformation. The response to a phospholipase C inhibitor also eliminated the response to the mechanical deformation (n = 10). In addition cells that demonstrated changes in Ca(2+)-containing media lost the ability to respond when EGTA was added to the media. Following this, 2 microM ryanodine was added to the cells, demonstrating a response too small to replicate the fluid flow stimulated wave, but supporting the view that the cells were vital following preincubation.

Biological Transport↗
PubMed1996

Effect of probe design and bandwidth on laser Doppler readings from vital and root-filled teeth.

Laser Doppler flowmetry has been shown to be useful in assessing blood flow in teeth. This study investigated the effect of probe design and bandwidth on laser Doppler readings from vital and root-filled teeth using an 810 nm light source, and established the sensitivity and specificity of each probe/bandwidth combination. Readings were taken from 20 human subjects with a root-filled tooth and a vital contralateral tooth using each of the probes with 0.125 mm, 0.375 mm and 0.5 mm fibre separations and three bandwidths (3.1 kHz, 14.9 kHz and 20 kHz). Ten pairs of traces from each group were examined by 10 trained observers who indicated whether the traces had come from a vital or root-filled tooth. The sensitivity and specificity of each combination were calculated from the accuracy of their replies. This was repeated for five observers but with additional information from Fourier analysis. Median readings from vital teeth were higher than those from root-filled teeth for all combinations. This difference was only significant at the 95% confidence level for the 0.5 mm probe with the two lower bandwidths. The 0.125 mm fibre separation probe showed good specificity. The other two probes had better sensitivity but poor specificity. The best specificity and sensitivity was shown by the 0.5 mm probe/3.1 kHz bandwidth combination. All sensitivities and specificities increased when additional information from Fourier analysis was available, but the 0.5 mm probe/3.1 kHz combination still had the best sensitivity and specificity.

Blood Flow Velocity↗
PubMed1996

Pattern of transmission of laser light in teeth.

This study examined the transmission of helium neon laser light in 20 dog and human teeth. The effect of probe position and angulation was observed both macroscopically and at a microscopic level using confocal microscopy. In all teeth in both species, laser light was transmitted through teeth to the pulpal surface with the light following the path of the enamel prisms and dentinal tubules. Probe angulation did not affect the pattern of light transmission, nor did probe position; however, the position of the probe on the tooth surface determined which section of the pulp was illuminated. Enamel and dentine together are able to collect and distribute light within the tooth, with both enamel prisms and dentinal tubules acting as optical fibres.

Absorption↗
PubMed1996

Effect of wavelength and bandwidth on the clinical reliability of laser Doppler recordings.

The aim of this study was to investigate the effect of wavelength and bandwidth on laser Doppler flowmeter signals from vital and root-filled teeth, and to establish their sensitivity and specificity. Twenty human subjects were recruited, each with a vital tooth and the contralateral tooth root filled but not restored apart from the palatal access cavity. Readings were taken from these teeth for 3 min at 20 Hz for each of 3.1 kHz, 14.9 kHz and 22.1 kHz bandwidths using a modified laser Doppler blood flow monitor which permitted simultaneous recording using 810 nm and 633 nm light sources with a probe of four optical fibres with 0.25 mm separation. Ten traces from each combination of variables was examined by 10 trained observers who indicated if the traces came from vital or root-filled teeth judged by the amplitude and regularity of pulsatility of the trace. From the accuracy of their replies, sensitivity and specificity were calculated. Median flux values were higher for vital teeth than for root-filled teeth and for the 810 nm wavelength than for the 633 nm wavelength. Flux values increased with wider bandwidth using the 810 nm light source. With the 633 nm light source, the highest flux values were recorded using the 3.1 kHz bandwidth. Using the Mann-Whitney U test, there was a highly significant difference between readings from vital and root-filled teeth for the 3.1 kHz/810 nm wave length combination (p<0.003) and a significant difference for the 3.1 kHz/633 nm wavelength group (p<0.02). Comparison of other groups showed no significant difference (p>0.05). The 810 nm wavelength showed good sensitivity but poor specificity at 14.9 and 22.1 kHz bandwidths. The 633 nm wavelength showed good specificity, but poor sensitivity, at 14.9 and 22.1 kHz bandwidths. The 3.1 kHz bandwidth showed the best sensitivity and specificity for both wavelengths. Sensitivity and specificity were increased if the results of fast Fourier analysis were considered in addition to observations of flux values and pulsatility of traces. The 810 nm/3.1 kHz combination offered the greatest sensitivity and specificity as a test to distinguish between root-filled and vital teeth. This combination was best when the results of Fourier analysis were considered in addition to visual observations.

Artifacts↗
PubMed1996

Effects of prilocaine local anaesthetic solutions on pulpal blood flow in maxillary canines.

The effects of prilocaine local anaesthetic solutions on pulpal blood flow (PBF) in maxillary canines were investigated in nine adult subjects. Buccal infiltration of 2 ml of the following solutions were carried out: 3% prilocaine; 3% prilocaine with 0.03 IU/ml felypressin; and 3% prilocaine with 1:100,000 adrenaline. Blood flow in the anaesthetized tooth was monitored by a laser Doppler flowmeter and data stored in a computer. An electric pulp tester was used to assess pulpal anaesthesia. The duration of anaesthesia was recorded. After administration of plain prilocaine, PBF changed little throughout the experiment in all nine subjects. When prilocaine with felypressin was injected. PBF fluctuated greatly, but there was no sustained increase or decrease. In comparison, prilocaine with adrenaline caused a significant decline in PBF in every subject (p < 0.05), but then gradually started to return to the pre-injection level: there was no such change in PBF of the contralateral tooth. Injection of 3% plain prilocaine achieved a short duration of pulpal anaesthesia (median 7 min) in only three subjects. When prilocaine with felypressin was injected, eight out of nine subjects experienced pulpal anaesthesia (median duration 10.5 min). Injection of prilocaine with adrenaline caused pulpal anaesthesia in six of nine subjects (median duration 10.5 min). The use of vasoconstrictors with prilocaine anaesthetics had less pronounced effects on blood flow and shorter periods of anaesthesia than those reported previously for lignocaine with adrenaline.

Adult↗
PubMed1996

Muscle spindles in the jaw-closer muscles of the domestic cat.

The objectives of this study were to identify the exact location of spindles in jaw-closer muscles of the cat, to count the total number of spindles and to compare their distribution with the distribution of slow extrafusal fibres. The jaw-closer muscle group with all the skeletal attachments intact was fixed in a modified Carnoy solution, decalcified and processed through to wax. Complete series of sections were cut transverse, sagittal and perpendicular to the anterior temporalis muscle. At regular intervals, serial sections were stained by the Weigert-van Gieson method or immunostained for myosin isoforms. Spindle counts were made only from muscle areas where fibres were cut in transverse section, and the spindles were followed individually. The extrafusal fibres were identified by indirect immunoperoxidase staining with antibodies specific for the slow (type I) and fast (type IIM) isoforms of myosin found in jaw-closer muscles. The mean numbers and locations of spindles found were 13 in medial pterygoid (close to inferior border), 123 (one count only) in the deep anterior portion of the temporalis muscle (between the coronoid process of the mandible and the cranium), 50.5 in a small deep zone of superficial masseter anterior to the temporomandibular joint, and 50 in zygomaticomandibularis (the deepest portion of masseter). Most of the spindles were simple spindles. Spindle complexes (4 or more spindle units fused in parallel) were rare and were found only in zygomaticomandibularis and in masseter. Most parts of the jaw-closer muscles had no spindles and contained only fast fibres. Cosegregation of muscle spindles with slow fibres was found in most parts of this muscle group, but the distribution of spindles was more restricted than that of slow fibres.

Animals↗
PubMed1995

In vitro cytotoxicity testing of neodymium-iron-boron magnets.

Neodymium-iron-boron magnets are being increasingly used in orthodontic and orthopedic appliances. The purpose of this study was to investigate the in vitro cytotoxicity of these new "super" magnets. A modified agar overlay technique was performed using both L929 mouse fibroblasts and adult human oral mucosal fibroblasts. The neodyium-iron-boron magnets were tested in the following forms: uncoated magnetized, uncoated demagnetized, parylene coated demagnetized, and parylene coated magnetized. Results revealed that uncoated magnetized, uncoated demagnetized, and parylene coated magnetized were cytotoxic to both L929 mouse fibroblasts and human oral mucosal fibroblasts. However the parylene coated demagnetized neodymium magnets were only cytotoxic to the human mucosal fibroblasts. This study supports the hypothesis that these are cytotoxic effects of possibly corrosion products and of magnetism. The specific toxic effects of parylene were not examined in this present study. Statistical analysis (Mann-Whitney U-test, p = 0.014) suggested that, in this experiment, the human oral mucosal fibroblasts were more sensitive to the effects of these rare earth magnets. With the evidence of a biological effect, the use of implanted magnets must at present be limited until the safety can be assured.

Animals↗
PubMed1995

Effect of corrosion products (neodymium iron boron) on oral fibroblast proliferation.

The biological effects of the corrosion products of neodymium iron boron (Nd2Fe14B) magnets are largely unknown. The aim of this study was to identify the types of corrosion product and to evaluate the effect of the corrosion products (CP) of Nd2Fe14B magnets on the proliferation of human oral mucosal fibroblasts. Uncoated Nd2Fe14B magnets were stored in saline at 37 degrees C for 6 months and the corrosion products collected. 100 microL of a cell suspension (human oral mucosal fibroblasts [14 x 10(4) cells/mL]) was aliquoted into 72 wells of a 96-well plate, the remaining plates receiving culture medium only. After 12 h incubation at 37 degrees C, each well then received 100 microL of either (A) culture medium, (B) 100% CP, (C) 50% CP, or (D) 0% CP. The plates were reincubated at 37 degrees C for a further 48, 96, or 144 h. Fibroblast proliferation was assessed using the methylene blue uptake/elution technique. The compounds in the corrosion product were examined using quantitative X-ray analysis. Statistical analysis (ANOVA, Bonferroni's test 0.05, SAS v 6.04), showed that at each time point, the cell numbers in groups B, C, and D were significantly lower than group A. Within groups B, C, and D no significant differences were found, despite the suggestion of a dose response effect. Fibroblast proliferation in the presence of corrosion products was significantly lower than with culture medium. Fibroblast proliferation did occur in the presence of 0, 50, and 100% CP. The actual corrosion products appeared to be salts of iron but 3.2% (+/- 0.6) of neodymium chloride (NdCl3) was found.

Alloys↗
PubMed1995

Loading of cells and a possible upper limit of load response with respect to strain energy density.

A model has been established to examine the effects of mechanically loading bone on the ionic distribution in osteocytes. The ionic concentrations are determined from the fluorescence of specific ionic indicators SPFI, PBFI, and SPQ. A deformation was induced in the calvariae of neonatal rats which were used as the experimental subjects. The ionic distribution was not uniform across the calvarial specimen, and with the aid of finite element stress analysis we examined the distribution of strain energy density within the specimen. A similarity between the pattern of distribution of strain energy density and the potassium and sodium ions was observed, whereas this was not the case with chloride ions.

Animals↗
PubMed1995

Physical and chemical properties of a new root-end filling material.

This study determined the chemical composition, pH, and radiopacity of mineral trioxide aggregate (MTA), and also compared the setting time, compressive strength, and solubility of this material with those of amalgam, Super-EBA, and Intermediate Restorative Material (IRM). X-ray energy dispersive spectrometer in conjunction with the scanning electron microscope were used to determine the composition of MTA, and the pH value of MTA was assessed with a pH meter using a temperature-compensated electrode. The radiopacity of MTA was determined according to the method described by the International Organization for Standardization. The setting time and compressive strength of these materials were determined according to methods recommended by the British Standards Institution. The degree of solubility of the materials was assessed according to modified American Dental Association specifications. The results showed that the main molecules present in MTA are calcium and phosphorous ions. In addition, MTA has a pH of 10.2 initially, which rises to 12.5 three hours after mixing. MTA is more radiopaque than Super-EBA and IRM. Amalgam had the shortest setting time (4 min) and MTA the longest (2 h 45 min). At 24 h MTA had the lowest compressive strength (40 MPa) among the materials, but it increased after 21 days to 67 MPa. Finally, except for IRM, none of the materials tested showed any solubility under the conditions of this study.

Aluminum Compounds↗
PubMed1995

The effect of mechanical deformation on the distribution of ions in fibroblasts.

The extracellular and intracellular sodium, potassium and chloride concentrations were determined in fibroblast cells located in the rat calvarium. The ionic values were determined by fluorescence microscopy after incubation with the fluorescent probes, sodium-binding benzofuran isophthalate (SBFI), potassium-binding benzofuran isophthalate (PBFI) and 6-methoxy-N-(3-sulfopropyl) quinolinium (SPQ) (Dyes were supplied by CALBIOCHEM, Nottingham, England). After determination of the resting membrane potential, the calvaria were placed under tension by retraction of a micromanipulator. The fluorescence was measured again. A statistically significant difference was found in the calculated potassium ion concentration (Mann-Whitney; p < 0.05). This affected the resting cell membrane potential by an average of 5.2 mV. This effect was blocked by the addition of a potassium channel blocker, tetraethylammonium (TEA).

Animals↗
PubMed1994

Pro-inflammatory and anti-inflammatory effects of the stable prostaglandin D2 analogue, ZK 118.182.

This study examined the pro- and anti-inflammatory effects of the stable prostaglandin (PG) D2 analogue, ZK 118.182 and the mechanism by which prostaglandins may exert their anti-inflammatory activity. Co-injected locally, ZK 118.182, like PGE2 and PGD2, dose-dependently increased plasma leakage induced by intradermal injection of bradykinin in rabbit skin. Infused i.v., ZK 118.182 (0.45 microgram/kg/min), a dose which did not affect systemic blood pressure, inhibited oedema formation in rabbit skin induced by the neutrophil-dependent agonists, formyl-methionyl-leucyl-phenylalanine (FMLP) and leukotriene B4 (LTB4). However, it did not modify plasma leakage induced by the neutrophil-independent mediators, bradykinin and platelet-activating factor (PAF). In contrast, neutrophil accumulation in response to LTB4 and FMLP was not affected in animals infused with ZK 118.182. In vitro, ZK 118.182, like PGE2 and PGD2 inhibited FMLP-induced superoxide anion (O2-) production by rabbit neutrophils. The compound, however, had minimal effects on O2- production induced by phorbol myristate acetate (PMA). ZK 118.182 inhibited to a small extent FMLP but not PMA-induced neutrophil adherence. These results show that depending on the route of administration, the PGD2 analogue, ZK 118.182, exhibits either pro- or anti-inflammatory effects. The anti-inflammatory effect may be related to the ability of the compound to inhibit increased microvascular permeability induced by neutrophil activation without interfering with neutrophil accumulation. This latter effect may be due to the analogue's capacity to suppress neutrophil secretion to a greater extent than neutrophil adherence.

Animals↗