Biomedical subjects

D M Thompson

Publications and source records attributed to D M Thompson.

At least 19 recordsLinked to original sources

PubMed2004

Genetics of the DST-mediated mRNA decay pathway using a transgene-based selection.

mRNA sequences that control abundance, localization and translation initiation have been identified, yet the factors that recognize these sequences are largely unknown. In this report, a transgene-based strategy designed to isolate mutants of Arabidopsis thaliana that fail to recognize these sequences is described. In this strategy, a selectable gene and a screenable marker gene are put under the control of the sequence element being analysed and mutants are selected with altered abundance of the corresponding marker RNAs. The selection of mutants deficient in recognition of the DST (downstream) mRNA degradation signal is used as a test-case to illustrate some of the technical aspects that have facilitated success. Using this strategy, we report the isolation of a new mutant, dst3, deficient in the DST-mediated mRNA decay pathway. The targeted genetic strategy described circumvents certain technical limitations of biochemical approaches. Hence, it provides a means to investigate a variety of other mechanisms responsible for post-transcriptional regulation.

Hydrolysis↗

PubMed2001

Schwann cell response to micropatterned laminin surfaces.

In the peripheral nervous system, Schwann cells are closely associated with, and play key roles in, the development, maintenance, and regeneration of peripheral neurons. Following injury, Schwann cell orientation may also play a role in guiding regenerating axons. To aid in the investigation of these interactions between Schwann cells and growing neurites, we have developed a method of controlling Schwann cell placement and orientation in vitro by using microlithographically patterned laminin substrates, alternating 20-microm regions of laminin with bovine serum albumin (BSA) stripes. The Schwann cells predominantly attached and elongated on the laminin stripes and organized into multicellular aggregates that were oriented with the micropattern. A detailed analysis of Schwann cell aggregate orientation and shape demonstrated a strong dependence on time. At 1 h after seeding the cells, 70% of the aggregates were oriented with respect to the micropattern; 94% were oriented at 24 h. Variations in laminin concentration and seeding density were also investigated. The only significant differences in Schwann cell response occurred 1 h after seeding (the earliest time point the cultures were observed), and the main factor controlling the cellular orientation appeared to be the presence of the laminin-BSA interface. This ability to control cell orientation and placement provides a tool for future investigations of Schwann cell-neuronal interactions in vitro.

Animals↗

PubMed2000

Identification of BFN1, a bifunctional nuclease induced during leaf and stem senescence in Arabidopsis.

Nuclease I enzymes are responsible for the degradation of RNA and single-stranded DNA during several plant growth and developmental processes, including senescence. However, in the case of senescence the corresponding genes have not been reported. We describe the identification and characterization of BFN1 of Arabidopsis, and demonstrate that it is a senescence-associated nuclease I gene. BFN1 nuclease shows high similarity to the sequence of a barley nuclease induced during germination and a zinnia (Zinnia elegans) nuclease induced during xylogenesis. In transgenic plants overexpressing the BFN1 cDNA, a nuclease activity of about 38 kD was detected on both RNase and DNase activity gels. Levels of BFN1 mRNA were extremely low or undetectable in roots, leaves, and stems. In contrast, relatively high BFN1 mRNA levels were detected in flowers and during leaf and stem senescence. BFN1 nuclease activity was also induced during leaf and stem senescence. The strong response of the BFN1 gene to senescence indicated that it would be an excellent tool with which to study the mechanisms of senescence induction, as well as the role of the BFN1 enzyme in senescence using reverse genetic approaches in Arabidopsis.

Amino Acid Sequence↗

PubMed2000

Advanced techniques in foot and ankle reconstruction.

Complex deformities of the foot and ankle remain a difficult problem for even the most experienced surgeon. Many techniques are available to provide correction and no single one is appropriate for all cases. Identical deformities often can be treated with different techniques with equally successful outcomes. Each deformity is unique and the surgeon should be capable of using multiple techniques to provide the most appropriate treatment for the patient and the deformity. Simple deformities often can be handled with simple techniques but more complex problems often require more complex solutions. The techniques discussed here have worked well at the authors' institution but are undergoing constant reevaluation and occasional modification. It is important that the surgeon and the patient understand that with these techniques it is usually possible to provide a functional outcome but never possible to provide a normal foot or ankle. If appropriate goals are set, correction of these challenging deformities can be satisfying to surgeon and patient.

Ankle↗

PubMed1999

Insulin adjustment by a diabetes nurse educator improves glucose control in insulin-requiring diabetic patients: a randomized trial.

BACKGROUND: Diabetic patients taking insulin often have suboptimal glucose control, and standard methods of health care delivery are ineffective in improving such control. This study was undertaken to determine if insulin adjustment according to advice provided by telephone by a diabetes nurse educator could lead to better glucose control, as indicated by level of glycated hemoglobin (HbA1c). METHODS: The authors conducted a prospective randomized trial involving 46 insulin-requiring diabetic patients who had poor glucose control (HbA1c of 0.085 or more). Eligible patients were those already taking insulin and receiving endocrinologist-directed care through a diabetes centre and whose most recent HbA1c level was 0.085 or higher. The patients were randomly assigned to receive standard care or to have regular telephone contact with a diabetes nurse educator for advice about adjustment of insulin therapy. RESULTS: At baseline there was no statistically significant difference between the 2 groups in terms of HbA1c level (mean [and standard deviation] for standard-care group 0.094 [0.008] and for intervention group 0.096 [0.010]), age, sex, type or duration of diabetes, duration of insulin therapy or complications. After 6 months, the mean HbA1c level in the standard-care group was 0.089 (0.010), which was not significantly different from the mean level at baseline. However, the mean HbA1c level in the intervention group had fallen to 0.078 (0.008), which was significantly lower than both the level at baseline for that group (p < 0.001) and the level for the standard-care group at 6 months (p < 0.01). INTERPRETATION: Insulin adjustment according to advice from a diabetes nurse educator is an effective method of improving glucose control in insulin-requiring diabetic patients.

Blood Glucose↗

PubMed1999

Engineering a mouse balancer chromosome.

Balancer chromosomes are genetic reagents that are used in Drosophila melanogaster for stock maintenance and mutagenesis screens. Despite their utility, balancer chromosomes are rarely used in mice because they are difficult to generate using conventional methods. Here we describe the engineering of a mouse balancer chromosome with the Cre-loxP recombination system. The chromosome features a 24-centiMorgan (cM) inversion between Trp53 (also known as p53) and Wnt3 on mouse chromosome 11 that is recessive lethal and dominantly marked with a K14-Agouti transgene. When allelic to a wild-type chromosome, the inversion suppresses crossing over in the inversion interval, accompanied by elevated recombination in the flanking regions. The inversion functions as a balancer chromosome because it can be used to maintain a lethal mutation in the inversion interval as a self-sustaining trans-heterozygous stock. This strategy can be used to generate similar genetic reagents throughout the mouse genome. Engineering of visibly marked inversions and deficiencies is an important step toward functional analyses of the mouse genome and will facilitate large-scale mutagenesis programs.

Animals↗

PubMed1999

Extraction site--foundation for restorative excellence.

Preservation of the socket site architecture promotes better esthetics, easier prosthetic treatment, and restoration of the alveolar bone for implant or bridge replacement options. The purported simplicity of the simple extraction can be misleading if the bone housing or soft tissues have been ravaged by disease. This article provides guidelines for case selection, patient consultations, selection of appropriate materials, and a step-by-step methodology for completing successful extraction site therapy.

Alveoloplasty↗

PubMed1999

Extraction sites--foundations for restorative excellence.

Alveolar Bone Loss↗

PubMed1998

A comparison of laparoscopic ultrasound with digital fluorocholangiography for detecting choledocholithiasis during laparoscopic cholecystectomy.

BACKGROUND: Laparoscopic ultrasound is an alternative to operative cholangiogram for evaluation of the common bile duct (CBD) during laparoscopic cholecystectomy. It is a safe, fast, and reliable method for detecting choledocholithiasis. METHODS: We prospectively evaluated the sensitivity and specificity of laparoscopic ultrasound (LUS) and digital fluorocholangiogram (DFCG) in a three-phase study of 360 consecutive patients. RESULTS: In phase I, 140 patients undergoing laparoscopic cholecystectomy had LUS performed first, followed by DFCG. Thirteen patients had CBD calculi identified on LUS. Four patients with confirmed (two cases) or presumed (two cases) CBD calculi on DFCG were not identified on LUS. Thus, the specificity of LUS was 100%, whereas the sensitivity was 76.5%. DFCG had four false positives, for a sensitivity of 100% with a specificity of 96.7%. LUS was performed, on average, in 6.6 min, whereas DFCG required 10.9 min to perform. In phase II, the infusion of saline through a cystic duct catheter was performed in instances where the distal CBD could not be well seen. This maneuver distended the intrapancreatic portion of the CBD, allowing better visualization. Nine stones were identified on LUS in 78 patients, increasing the sensitivity to 100%. One false positive DCFG was encountered, resulting in a sensitivity of 100% and a specificity of 98.6%. In phase III, we performed routine LUS and used DFCG only in select cases. The sensitivity and specificity for LUS were 95.7% and 100%, respectively, whereas DFCG had a sensitivity of 95.2% and a specificity of 100%. One patient in phase III has returned 11 months post-op with a CBD stone. This was initially missed on LUS, DFCG, and postoperative ERCP. The sensitivity and specificity in all 360 patients were 90% and 100% for LUS and 98.1% and 98.1% for DFCG, respectively. A total of five CBD stones were missed by LUS, four early in the study (phase I). One missed on LUS in phase III was also missed by DFCG and ERCP. CONCLUSIONS: LUS is a reliable alternative to DFCG during laparoscopic cholecystectomy (LC). With experience, it is as sensitive as DFCG and more specific. It is more rapidly performed than cholangiography.

Adult↗

PubMed1997

Cytochrome P450 (CYP)9A1 in Heliothis virescens: the first member of a new CYP family.

A novel cytochrome P450 cDNA with its complete coding sequence and part or all of the 3' (77 nucleotides) and 5' (87 nucleotides) non-coding sequence was isolated from the tobacco budworm, Heliothis virescens (F). The 1763 nucleotide sequence encodes a protein of 532 amino acids which includes a hydrophobic N-terminal region and the highly conserved heme binding regions typical of P450s. Low sequence similarity to other P450 sequences and the presence of a thromboxane synthase-like insertion upstream from the I helix resulted in its assignment as the first member of family 9, i.e. CYP9A1. CYP9A1 is most similar to CYP3A1 from the rat (34.7% identity), but is also similar to the insect P450s from family 6, including CYP6B1v1 from Papilio polyxenes (33.3%), CYP6A2A from Drosophila melanogaster (32.4%), CYP6A3 from Musca domestica (31.7%) and CYP6B2 from Helicoverpa armigera (30.1%). Comparative Western and Northern blot studies indicate that expression of CYP9A1 in thiodicarb selected populations of tobacco budworm is associated with insecticide resistance. The pattern of restriction fragment length polymorphism (RELP) variation in offspring of single-pair matings demonstrated autosomal inheritance of CYP9A1 and enabled its assignment to linkage group 7. The coding region of CYP9A1 occupies no more than 10 kb in the tobacco budworm genome.

Amino Acid Sequence↗

PubMed1996

Mercuric ion reduction and resistance in transgenic Arabidopsis thaliana plants expressing a modified bacterial merA gene.

With global heavy metal contamination increasing, plants that can process heavy metals might provide efficient and ecologically sound approaches to sequestration and removal. Mercuric ion reductase, MerA, converts toxic Hg2+ to the less toxic, relatively inert metallic mercury (Hg0) The bacterial merA sequence is rich in CpG dinucleotides and has a highly skewed codon usage, both of which are particularly unfavorable to efficient expression in plants. We constructed a mutagenized merA sequence, merApe9, modifying the flanking region and 9% of the coding region and placing this sequence under control of plant regulatory elements. Transgenic Arabidopsis thaliana seeds expressing merApe9 germinated, and these seedlings grew, flowered, and set seed on medium containing HgCl2 concentrations of 25-100 microM (5-20 ppm), levels toxic to several controls. Transgenic merApe9 seedlings evolved considerable amounts of Hg0 relative to control plants. The rate of mercury evolution and the level of resistance were proportional to the steady-state mRNA level, confirming that resistance was due to expression of the MerApe9 enzyme. Plants and bacteria expressing merApe9 were also resistant to toxic levels of Au3+. These and other data suggest that there are potentially viable molecular genetic approaches to the phytoremediation of metal ion pollution.

Amino Acid Sequence↗

PubMed1996

Cloning of a putative juvenile hormone-responsive storage protein gene from the tobacco budworm, Heliothis virescens.

A cDNA clone with 78% amino acid identity to a basic juvenile hormone (JH)-suppressible hemolymph protein from the cabbage looper, Trichoplusia ni, was isolated from the tobacco budworm, Heliothis virescens. This clone was obtained upon screening a cDNA library derived from larval fat body of a pesticide resistant strain of H. virescens with a cDNA probe for Drosophila melanogaster glutathione S-transferase. By comparison with other insect storage proteins, this clone was predicted to be part of an approximately 2,300 nucleotide (nt) cDNA, of which 691 nt were isolated and sequenced. The partial cDNA clone hybridizes to a RNA of approximately 2,370 nt in H. virescens. Treatment with a juvenoid (2-[1-methyl-2-(4-phenoxyphenoxy)ethoxy] pyridine; pyriproxifen) leads to a decrease in RNA levels of this putative hemolymph storage protein in early fifth stadium larvae of H. virescens, prior to commitment. In contrast, treatment in late fifth stadium (after commitment to pupal development) leads to an increase in the RNA level of this JH-responsive gene. This is the first report of both induction and suppression of storage protein RNA levels in the same stadium. We have given this gene the designation Hv-SP4 (H. virescens, storage protein 4; accession no. U48594). Genetic segregation analysis of restriction fragment length polymorphisms (RFLPs) defined by Hv-SP4 has shown that it is the product of a single-copy, Mendelian, autosomal gene.

Amino Acid Sequence↗

PubMed1996

Mechanism of action and cloning of epoxide hydrolase from the cabbage looper, Trichoplusia ni.

The majority of the JH III epoxide hydrolase activity in last stadium day 3 (gate 1) wandering Trichoplusia ni was membrane bound with approximately 9% of the activity found in the cytosol. Both the microsomal and cytosolic JH epoxide hydrolases were stable, retaining 30% of their original activity after incubation at 4 degrees C for 15 days. 18O-labeled water underwent enzyme catalyzed regioselective addition to the least substituted C10 position of JH III. In multiple turnover reactions with JH epoxide hydrolase in 97.9% 18O-labeled water, only 91.3% 18O incorporation was observed. This is consistent with an SN2 reaction likely involving a carboxylate in the active site of JH epoxide hydrolase. The DNA amplification cloning of a fragment of a putative T. ni epoxide hydrolase is reported. The deduced amino acid sequence shares 67% similarity to the rat microsomal epoxide hydrolase.

Amino Acid Sequence↗

PubMed1996

In the absence of streptomycin, minoxidil potentiates the mitogenic effects of fetal calf serum, insulin-like growth factor 1, and platelet-derived growth factor on NIH 3T3 fibroblasts in a K+ channel-dependent fashion.

There is considerable evidence to suggest that the opening of K+ channels plays an important role in stimulating mitogenesis. K+ channel blockers have been shown to inhibit mitogenesis in vitro, mitogens increase cytosolic membrane K+ channel permeability, K+ channel openers stimulate hair growth in vivo, and the Ras/Raf signal transduction pathway induces K+ channel activity. Paradoxically, however, K+ channel openers such as minoxidil have been reported in vitro not to modulate, or even to inhibit, mitogenesis in a range of cell types. Only untherapeutic concentrations have stimulated mitogenesis. These experiments, however, appear to have been carried out in the presence of aminoglycoside antibiotics, which inhibit potassium channel activity. We now report that in the absence of aminoglycoside antibiotics, minoxidil at 10 microg/ml (0.05 mM) causes a significant stimulation of proliferation of NIH 3T3 fibroblasts maintained over a 10-d period in 5% fetal calf serum-supplemented medium. Further, we show that in the presence of 100 microg streptomycin per ml, minoxidil at 10 microg/ml produces an initial inhibition of proliferation, which apparently confirms, in NIH 3T3 fibroblasts, that the inhibition of mitogenesis by minoxidil in the presence of streptomycin is an artifact. The potentiation of NIH 3T3 cell growth by minoxidil can be attributed to the opening of potassium channels, because the potassium channel blocker tolbutamide (5 mM) or combinations of the blockers tolbutamide (1 mM)/tetraethylammonium (2 mM) or glibenclamide (1 microM)/apamin (10 nM) block the minoxidil-induced stimulation of growth. We also demonstrate that minoxidil is able to significantly potentiate the mitogenic effects of both platelet-derived growth factor and insulin-like growth factor 1 on NIH 3T3 fibroblasts in the presence of CPSR-2 (a cytokine free serum substitute). Thus we have shown that minoxidil potentiates the mitogenic effects of fetal calf serum in vitro on NIH 3T3 fibroblasts by opening potassium channels and is also able to potentiate the mitogenic effects of the growth factors platelet-derived growth factor and insulin-like growth factor 1.

3T3 Cells↗

PubMed1995

7-Chloro-3-methyl-3,4-dihydro-2H-1,2,4-benzothiadiazine S,S-dioxide (IDRA 21), a congener of aniracetam, potently abates pharmacologically induced cognitive impairments in patas monkeys.

We report here on the ability of IDRA 21 and aniracetam, two negative allosteric modulators of glutamate-induced DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor desensitization, to attenuate alprazolam-induced learning deficit in patas monkeys working in a complex behavioral task. In one component of a multiple schedule (repeated acquisition or "learning"), patas monkeys acquired a different four-response chain each session by responding sequentially on three keys in the presence of four discriminative stimuli (geometric forms or numerals). In the other component (performance) the four-response chain was the same each session. The response chain in each component was maintained by food presentation under a fixed-ratio schedule. When alprazolam (0.1 or 0.32 mg/kg p.o.) was administered alone, this full allosteric modulator of gamma-aminobutyric acid type A (GABAA) receptors produced large decreases in the response rate and accuracy in the learning component of the task. IDRA 21 (3 or 5.6 mg/kg p.o.) and aniracetam (30 mg/kg p.o.) administered 60 min before alprazolam, having no effect when given alone, antagonized the large disruptive effects of alprazolam on learning. From dose-response studies, it can be estimated that IDRA 21 is approximately 10-fold more potent than aniracetam in antagonizing alprazolam-induced learning deficit. We conclude that IDRA 21, a chemically unrelated pharmacological congener of aniracetam, improves learning deficit induced in patas monkeys by the increase of GABAergic tone elicited by alprazolam. Very likely IDRA 21 exerts its behavioral effects by antagonizing AMPA receptor desensitization.

Alprazolam↗

PubMed1995

The study of vocal fold vibratory patterns in patients with unilateral vocal fold paralysis before and after type I thyroplasty with or without arytenoid adduction.

Type I thyroplasty and arytenoid adduction have been used for the treatment of symptomatic unilateral vocal fold paralysis since the mid-1970s. To this date, the vibratory patterns of the vocal folds in patients with unilateral vocal fold paralysis undergoing thyroplasty have not been studied in depth. Abnormal vibration of the vocal fold mucosa often contributes to voice problems in persons diagnosed with vocal diseases or disorders. The preoperative and postoperative videostroboscopic vibratory patterns including mucosal wave, amplitude, glottic closure, and symmetry were studied in 12 patients who underwent unilateral type I thyroplasty separately or in combination with an arytenoid adduction. The observed changes and clinical implications are discussed.

Adult↗

PubMed1995

Transcriptional activation analysis by the chloramphenicol acetyl transferase (CAT) enzyme assay.

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Animals↗

PubMed1995

Whole mouth microbiota effects following subgingival delivery of sanguinarium.

An increased incidence of antibiotic-resistant bacteria and yeast overgrowth has been reported following various periodontal treatments. The objective of this study was to detect possible overgrowth of opportunistic bacteria and fungi as well as changes in normal microbiota after application of a biodegradable delivery system containing 5% sanguinarium (ABDS-S) to one quadrant in a split-mouth study. An oral hygiene quadrant served as a control. The ABDS-S treated and control periodontal sites as well as the saliva of 17 subjects were sampled prior to treatment, immediately after ABDS-S removal at 7 days, and again at 30 and 60 days. At Day 7 sanguinarium-resistant bacteria increased in both control and ABDS-S periodontal sites as well as in the saliva. Enteric Gram-negative bacilli in both control and ABDS-S periodontal sites were 2.2 to 3.4 log colony forming units higher at Day 7 compared to baseline. This overgrowth was transient in that levels became undetectable at Days 30 and 60. No such overgrowth was observed for C. albicans or other fungi, or for S. aureus or other staphylococci in any periodontal sites. Levels of Actinomyces increased at Days 30 and 60 in both control and ABDS-S sites as well as saliva. These changes strongly suggest that a 7 day ABDS-S treatment in one quadrant of the mouth led to significant microbiota changes in the treated and control quadrants as well as in the saliva. Future microbial studies involving antimicrobials delivered by local delivery systems must consider the crossover effects of treatment inherent in the split-mouth design.

Adult↗