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Biomedical subjects

G M Smith

Publications and source records attributed to G M Smith.

At least 127 records · Page 7Linked to original sources
PubMed1991

High-resolution 1H- and 15N-NMR studies of Rhodospirillum rubrum cytochrome c2.

Rhodospirillum rubrum cytochrome c2 was uniformly enriched in 15N and studied by 1H- and 15N-NMR spectroscopy. Relaxation and NOE data allowed determination of the rotational correlation time and indicated more rapid side-chain motion in the native protein and increased segmental motion in the base-denatured protein. The pi nitrogen of the ligand histidine and the indolic nitrogen of the invariant tryptophan both remain protonated and act as proton-donors in hydrogen bonds over a wide pH range and therefore do not contribute to pH-related changes in the midpoint potential. pK values identified by numerous methods in the ferrocytochrome at pH 6.9 and in the ferricytochrome at pH 6.2 arise from His-42. At pH values below the pK, the imidazolium group participates in a salt bridge or in a hydrogen bond with the carboxylate group of the inner propionate of the heme. Loss of the proton causes a local conformational change which alters the midpoint potential. The pK values of the amino terminus and lysines were also determined from pH titrations monitored by 15N-NMR. Similar titrations of the ferricytochrome monitored by 1H-NMR showed structural heterogeneity in that the resonance of heme ring methyl 8 split into a doublet as the pH was raised.

Cytochrome c Group↗
PubMed1991

Immature type-1 astrocytes suppress glial scar formation, are motile and interact with blood vessels.

Previous studies have shown that immature but not mature astrocytes have the capacity to suppress glial scar formation and enhance axon outgrowth when transplanted into the adult mouse brain. We report here that glial scar formation is suppressed following transplantation of purified immature but not mature cultured type-1 rat cortical astrocytes into the adult rat brain. To examine the fate of transplanted cells, cultured astrocytes were labeled with either fluorescent beads or BSA-conjugated colloidal gold and traced after transplantation using both light and electron microscopy. While both immature and mature astrocytes survived transplantation, mature astrocytes appeared more susceptible to phagocytosis by cells of the immune system than immature astrocytes. Furthermore, while mature astrocytes were restricted to the region of the implant, immature astrocytes migrated into the surrounding CNS and became closely associated with host blood vessels. Such blood vessels were impermeable to the diffusion of systematically applied Evans blue dye. To determine whether immature astrocytes were intrinsically more motile than mature astrocytes, their rate of translocation was compared in vitro. Immature astrocytes translocated more than twice as fast as mature astrocytes. This ability of immature astrocytes to translocate throughout the host CNS and become associated with blood vessels may be a major factor in their ability to suppress glial scar formation in the adult animal.

Animals↗
PubMed1991

Circulating phospholipase A2 activity associated with sepsis and septic shock is indistinguishable from that associated with rheumatoid arthritis.

Elevation of circulating phospholipase A2 (PLA2) activity is associated with sepsis and septic shock. Elevated levels of PLA2 activity also are seen in association with chronic inflammatory disorders such as rheumatoid arthritis. The relationship between these phospholipases is unclear. We have developed a highly specific enzyme-linked immunosorbent assay (ELISA) capable of measuring human synovial PLA2 in plasma, using monoclonal antibodies raised to recombinant synovial PLA2. This ELISA has been used to quantitate circulating PLA2 levels in patients clinically diagnosed with sepsis. These elevated levels positively correlated with the elevation seen in plasma PLA2 enzyme activity. The antibodies also have been used to purify immunoreactive PLA2 from plasma of patients with sepsis, thus enabling characterization of the purified protein by amino-terminal sequence analysis. We conclude from this study that the increase in PLA2 activity seen in association with sepsis and septic shock results from a dramatic elevation in levels of a circulating PLA2 enzyme. This inflammatory PLA2 is indistinguishable, both immunologically and chemically, from that associated with rheumatoid arthritis. Therapeutic agents directed towards inhibition of this inflammatory PLA2 enzyme may have utility in the treatment of both chronic and acute inflammatory disease.

Adolescent↗
PubMed1991

Effect of impression tray design and impression technique upon the accuracy of stone casts produced from a putty-wash polyvinyl siloxane impression material.

This study examined the accuracy of stone casts produced from impressions taken in stock polycarbonate trays, some of which had been strengthened with autopolymerizing polymethyl methacrylate resin. Three techniques were used to make the impression of an acrylic master model of the mandibular arch on which two extracoronal preparations for bridgework and one intracoronal inlay preparation had been carried out. Each preparation had been indented with a reference point for later measurement. The impression material was a putty-wash polyvinyl siloxane material. Five impressions were taken for each type of tray for each impression technique and these were cast in die-stone after 24 h. The distances between the points were measured with a reflex microscope and the means determined for each design of tray. The mean difference between casts produced from the various tray designs and the acrylic master model were determined for each of the distances between the three measuring points for the various impression techniques. Statistical analysis showed that, with the polycarbonate stock trays, there were significant differences between some of the modifications and between them and the acrylic model, for the three distances (P less than 0.05). These differences were limited to one measurement for one design of tray for each of the two-stage impression methods. With the one-stage technique the unreinforced tray and those reinforced with acrylic, over the heels and anteriorly, and the barred design were statistically significantly different from the acrylic model for measurement A-B.(ABSTRACT TRUNCATED AT 250 WORDS)

Adhesives↗
PubMed1991

Bactericidal effects of amoxycillin/clavulanic acid against a Legionella pneumophila pneumonia in the weanling rat.

Amoxycillin/clavulanic acid and clavulanic acid have been previously reported to demonstrate bactericidal activity in tissue culture studies against intracellular Legionella pneumophila. A rat model of legionellosis was therefore developed for the purpose of assessing the efficacy of these agents against L. pneumophila in vivo. Therapy by the subcutaneous route was started 12 h after infection when the majority of the bacteria observed in lavage fluid were residing in alveolar macrophages. Treatment with amoxycillin was ineffective in reducing the bacterial counts of L. pneumophila in lung homogenates whereas amoxycillin/clavulanic acid displayed bactericidal effects of the same order as the control antibiotic, erythromycin. Further in-vivo studies are planned to assess the clinical relevance of these findings.

Age Factors↗
PubMed1991

Prognostic indicators for the development of AIDS in HIV antibody positive haemophiliac patients: results of a three-year longitudinal study.

In February 1986, 40 out of 75 adult patients with haemophilia A attending St. James's University Hospital were human immunodeficiency virus (HIV) antibody positive. Over a three-year period these patients were prospectively studied with regard to possible prognostic indicators for the development of the acquired immune deficiency syndrome (AIDS). Using the Centres for Disease Control (CDC) classification of HIV infection, 17 patients (42.5%) developed group 4 disease during this time, giving an actuarial three-year progression rate of 44%, and 5 patients (12.5%) died. The following parameters measured at recruitment were found independently to predict progression to AIDS: a serum beta 2-m level of greater than 3.5 mg/l, (chi 2 = 15.95, P less than 0.001), a serum IgA level of greater than 4.5 milligram(s) (chi 2 = 6.08, P less than 0.02) and p24 antigenaemia (chi 2 = 5.7, P less than 0.05). The actuarial three-year progression rate in those patients abnormal by two or more of these parameters was 100% (n = 7), compared to only 7% in patients who were normal by all three values (n = 15). CD4+ lymphocyte counts and CD4+:CD8+ ratios were significantly lower in HIV positive compared with HIV negative patients (P less than 0.01), but did not predict the development of AIDS.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Occlusal abnormalities, pericranial muscle and joint tenderness and tooth wear in a group of migraine patients.

Seventy-two migraine sufferers, whose attacks normally begin during or soon after waking from sleep, were compared with 37 age- and sex-matched controls to establish whether signs of mandibular dysfunction, occlusal discrepancies and known clenching or grinding habits were any more frequent among the former group. Temporomandibular joint tenderness or pain history, masticatory muscle tenderness and known clenching or grinding habits were found to occur at significantly increased levels in the migraine sufferers, with two-thirds of these patients reporting a parafunctional habit. Occlusal abnormalities, including non-working side or protrusive interferences and slides of greater than 1 mm to the intercuspal position, were found to occur at similar frequencies in the two groups. Assessment of wear of the occlusal surfaces also showed no difference, suggesting that any nocturnal habit with a role in migraine is more likely to be clenching in nature. In conclusion, evidence was found to support an aetiological role for nocturnal tooth clenching or grinding in migraine characterized by attacks that start predominantly during sleep or soon after waking, but no evidence of a link with occlusal factors was found in these patients.

Adolescent↗
PubMed1991

Bactericidal effects of ticarcillin-clavulanic acid against Legionella pneumophila pneumonia in immunocompromised weanling rats.

A model of acute Legionella pneumophila pneumonia in neutropenic weanling rats was developed as a means of assessing the efficacies in vivo of the beta-lactams ticarcillin, ticarcillin-clavulanic acid, and clavulanic acid, agents active against the organism in vitro. Weanling rats were dosed with cyclophosphamide 3 days before and immediately prior to infection by intrabronchial intubation with L. pneumophila. The bacteria persisted in the lungs of untreated animals at high counts (5.0 to 7.0 log10 CFU/g of lung tissue) for up to 168 h after infection, and the histological characteristics of the infection were similar to those of the disease in humans. Transmission electron micrography revealed the presence of L. pneumophila multiplying within alveolar macrophages. Therapy with ticarcillin was ineffective in reducing the bacterial numbers in the lung tissue, whereas ticarcillin-clavulanic acid and clavulanic acid were active, producing bactericidal effects similar to those of erythromycin. The ticarcillin-clavulanic acid combination was significantly more efficacious (P less than 0.01) than corresponding doses of clavulanic acid alone. Synergistic activity between ticarcillin and clavulanic acid against L. pneumophila has been demonstrated in vivo, and the combination showed activity similar to that of erythromycin.

Animals↗
PubMed1991

Comparison of the effects of FK-506, cyclosporin A and rapamycin on IL-2 production.

The immunosuppressive compounds FK-506, cyclosporin A (CsA) and rapamycin inhibit both the human and mouse mixed lymphocyte reactions (MLR) with IC50s of 2-5 x 10(-10) M for FK-506 and rapamycin and 10(-8) M for CsA. FK-506 and CsA were also potent inhibitors of A23187/PMA-stimulated IL-2 production by Jurkat and HuT-78 cells but had no effect on the response of mouse CTLL cells to IL-2. IC50 values for inhibition of IL-2 production closely matched those for inhibition of the MLR and both drugs were active only during the first 4-6 hr following stimulation. In contrast, rapamycin was a poor inhibitor of IL-2 production, although it inhibited cellular responses to IL-2. The IC50 values for these two activities indicated that neither alone accounted for rapamycin inhibition of the MLR. FK-506 and CsA affected IL-2 gene transcription in Jurkat cells by the same mechanism. Both inhibited the appearance of the transcription factor, NFAT, whereas rapamycin did not. The appearance of another transcription factor, NFK beta, was unaffected by all three drugs. The effects of FK-506 and CsA on IL-2 gene expression, therefore, are similar even though the two drugs act through distinct cytosolic receptors.

Anti-Bacterial Agents↗
PubMed1990

Assignments of 15N and 1H NMR resonances and a neutral pH ionization in Rhodospirillum rubrum cytochrome c2.

The phi NH proton and 15N resonances of the ligand histidine of Rhodospirillum rubrum fericytochrome c2 are found at 14.7 and 184 ppm, respectively, contradicting the proposal that this proton is absent in the R. rubrum ferricytochrome. Substitution of the deuterium atom for this proton causes small upfield shifts of the phi nitrogen in both oxidation states, indicating that the phi NH-peptide carboxyl hydrogen bond is not substantially weakened by the substitution. The proton and 15N resonances of the indolic NH group of the invariant tryptophan-62 and numerous proton resonances of the heme and extraheme ligands in the spectrum of the ferricytochrome are also assigned. An ionization in the ferrocytochrome occurring at neutral pH is assigned to the single nonligand histidine. This attribution is supported by the direct measurement of the ionization by NOE difference spectroscopy and by comparative structural arguments involving closely related cytochromes and chemically modified cytochromes.

Ammonium Chloride↗
PubMed1990

Characterization of pH-dependent conformational heterogeneity in Rhodospirillum rubrum cytochrome c2 using 15N and 1H NMR.

The 15N-enriched ferricytochrome c2 from Rhodospirillum rubrum has been studied by 15N and 1H NMR spectroscopy as a function of pH. The 15N resonances of the heme and ligand tau nitrogen are broadened beyond detection because of paramagnetic relaxation. The 15N resonance of the ligand histidine phi nitrogen was unambiguously identified at 184 ppm (pH 5.6). The 15N resonances of the single nonligand histidine are observed only at low pH, as in the ferrocytochrome because of the severe broadening caused by tautomerization. The dependence of the 15N and 1H spectra of the ferricytochrome on pH indicated that the ligand histidine tau NH does not dissociate in the neutral pH range and is involved in a hydrogen bond, similar to that in the reduced state. Because neither deprotonated nor non-hydrogen-bonded forms of the ligand histidine are observed in the spectra of either oxidation state, the participation of such forms in producing heterogeneous populations having different electronic g tensors is ruled out. Transitions having pKa's of 6.2, 8.6, and 9.2 are observed in the ferricytochrome. The localized conformational change around the omega loops is observed in the neutral pH range, as in the ferrocytochrome. Structural heterogeneity leads to multiple resonances of the heme ring methyl at position 8. The exchange rate between the conformations is temperature dependent. The transition with a pKa of 6.2 is assigned to the His-42 imidazole group. The displacement of the ligand methionine, which occurs with a pKa of 9.2, causes gross conformational change near the heme center.(ABSTRACT TRUNCATED AT 250 WORDS)

Cytochrome c Group↗
PubMed1990

Maturation of astrocytes in vitro alters the extent and molecular basis of neurite outgrowth.

In the developing mammalian central nervous system astrocytes have been proposed as an important substrate for axon growth. In the adult central nervous system following injury, astrocytes are a major component of the gliotic response which has been proposed to block axon growth. Experimental transplantation studies using cultured astrocytes have suggested that immature but not mature cultured astrocytes have the capacity to support axon outgrowth when transplanted into the adult rodent CNS. These observations suggest that astrocyte maturation is accompanied by changes in the functional capacity of these cells to support axon outgrowth. To determine whether this functional change reflects an intrisic astrocyte property, the extent and molecular bases of neurite outgrowth from embryonic rat cortical and chick retinal neurons on cultures of purified immature and mature astrocytes have been compared in vitro. The rate and extent of neurite outgrowth from both neuronal populations are consistently greater over the surface of immature than over the surface of mature astrocytes. Furthermore, antibodies to NCAM and G4/L1 significantly reduce neurite outgrowth on immature but not mature astrocytes, while antibodies to the integrin B1 receptor reduced outgrowth on both immature and, to a lesser extent, mature astrocytes. These results suggest that in vitro mature astrocytes have a reduced capacity and different molecular bases for supporting neurite outgrowth compared to immature astrocytes and are consistent with the proposal that functional changes during astrocyte maturation may partially contribute to regulating axon growth in the mammalian CNS.

Animals↗
PubMed1990

Astrocyte-polymer implants promote regeneration of dorsal root fibers into the adult mammalian spinal cord.

To overcome obstacles to the regeneration of crushed dorsal root fibers at the dorsal root entry zone, we have employed specially designed Millipore implants coated with embryonic astrocytes to serve as a substrate for axonal growth. This strategy was successful in promoting the growth of crushed dorsal root axons into the grey matter of the adult mammalian spinal cord in a small number of animals. Fiber ingrowth into the spinal cord was closely associated with the surface of the polymer implant. In addition, unique terminal arbor malformations, not normally present, were seen in several animals. A consistent finding was the presence of a limited inflammatory response in regions immediately adjacent to the implant where axons penetrate the spinal cord. Our findings suggest that providing the dorsal root entry zone with an embryonic milieu can stimulate a limited amount of axonal regeneration into the adult mammalian spinal cord.

Animals↗
PubMed1990

A comparison of the regeneration potential of dorsal root fibers into gray or white matter of the adult rat spinal cord.

To assess the role of white matter inhibition as a barrier to neurite outgrowth in vivo, we unilaterally transected three consecutive lumbar dorsal roots (L4-L6), incised the spinal cord, and transplanted the peripheral stump of L4 either medially onto the white matter of the dorsal columns or laterally, just superficial to the gray matter of the dorsal horn at the level of L5. Three weeks to seven months later, the translocated root was retransected, and its central stump was anterogradely labeled with HRP. The staining pattern demonstrated that regenerating sensory axons had entered the spinal cord from both medially and laterally placed roots. Axonal staining from medially placed dorsal roots (onto the white matter of the dorsal columns) was sparse and limited to the white matter. Staining of laterally placed roots revealed a small subpopulation of regenerating axons which had entered the gray matter and formed terminal arbors. Successful axonal regeneration into the gray matter, albeit minimal, was associated with a localized and limited inflammatory response near the sites of axonal ingrowth.

Animals↗
PubMed1990

Structure and function of G protein coupled receptors.

The G protein coupled receptors (GPC-Rs) comprise a large superfamily of genes encoding numerous receptors which all show common structural features, e.g., seven putative membrane spanning domains. Their biological functions are extremely diverse, ranging from vision and olfaction to neuronal and endocrine signaling. The GPC-Rs couple via multiple G proteins to a growing number of recognized second messenger pathway, e.g., cAMP and phosphatidyl inositol turnover. This review summarizes our current knowledge of the molecular mechanisms by which the GPC-Rs activate second messenger systems, and it addresses their regulation and structure.

Amino Acid Sequence↗
PubMed1990

Occlusion and reperfusion-induced arrhythmias in rats: involvement of platelets and effects of calcium antagonists.

The role of blood platelets in ischemia- and reperfusion-induced arrhythmias and the efficacy of three calcium blocking drugs (verapamil, diltiazem, and nicardipine) in preventing the arrhythmias were investigated. Using anesthetized rats, we measured platelet count (Pc) continuously in vivo with a Technicon autocounter. Thromboxane B2 (TxB2) and 6-keto-PGF1 alpha levels in blood from coronary sinus were determined by radioimmunoassay (RIA). Myocardial ischemia and arrhythmias were monitored from lead I ECG during and after occlusion of the left anterior descending coronary artery (LAD) for 7 min. Ischemia-induced arrhythmias were mainly ventricular ectopic contractions (VECs), whereas reperfusion produced VECs, ventricular tachycardia (VT), and reversible and irreversible ventricular fibrillation (VF). Both ischemia and reperfusion decreased platelet count and increased TxB2 level in blood from the coronary sinus. The effects of the CEBs were determined at two dose levels (0.1 and 0.3 mg/kg). Each calcium entry blocker (CEB), at both dose levels, significantly inhibited ischemia-induced arrhythmias. Verapamil and diltiazem significantly reduced reperfusion-induced VECs, prevented VT and irreversible VF, and reduced the number of animals with reversible VF. Nicardipine in preventing arrhythmias was not very effective at either dose. The CEBs also inhibited both ischemia- and reperfusion-induced decreases in PC with a moderate increase (up to 7%) as compared with levels in sham-operated controls. The CEBs also significantly reduced TxB2 levels in blood from the coronary sinus. These results indicate that ischemia and postischemic reperfusion both induce platelet aggregation in rats. Aggregating platelets release biologically active substances including thromboxane A2 (TxA2) which exacerbates existing ischemia and facilitates generation of arrhythmias. CEBs inhibit platelet aggregation and TxA2 release and enhance PGI2 synthesis, thereby preventing arrhythmias.

6-Ketoprostaglandin F1 alpha↗
PubMed1990

Characterizing auditory neurons using the Wigner and Rihacek distributions: a comparison.

Because of their dynamic properties, most sounds can best be characterized in the combined frequency-time (FT) domain. Powerful frequency-time characterizations are the Wigner distribution function (WDF) and the Rihacek energy density function (RDF). In the present paper several new concepts are introduced such as using the WDF to characterize the tuning of auditory neurons under wideband noise stimulation and a new method to quantify phase lock of auditory neurons to a wideband noise. No appreciable differences were found between the WDF and RDF in narrow-band signal representations. However, the differences between the WDF and RDF increase as the bandwidth of the signal increases. When signals are buried in uncorrelated background noise, the average FT function of these signals may be obtained through averaging the FT functions for each signal plus noise segment. The WDF takes at least a factor 2 more in time to compute than the RDF. The FT functions can be used to characterize (linear) filters by averaging FT functions of input-noise segments that precede threshold crossings of the filter's output signal. Both the WDF and the RDF were used to characterize auditory neurons from the midbrain in anurans; the WDF always had a smaller bandwidth than the RDF. By comparing the spectrum of the reverse correlation function and the average spectrum of the noise segments preceding the spikes, a quantification of the amount of phase lock of the auditory neuron to the noise is obtained.

Acoustic Stimulation↗