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Biomedical subjects

J M Wells

Publications and source records attributed to J M Wells.

At least 55 records · Page 3Linked to original sources
PubMed1995

Identification of exopolysaccharides produced by fluorescent pseudomonads associated with commercial mushroom (Agaricus bisporus) production.

The acidic exopolysaccharides (EPSs) from 63 strains of mushroom production-associated fluorescent pseudomonads which were mucoid on Pseudomonas agar F medium (PAF) were isolated, partially purified, and characterized. The strains were originally isolated from discolored lesion which developed postharvest on mushroom (Agaricus bisporus) caps or from commercial lots of mushroom casing medium. An acidic galactoglucan, previously named marginalan, was produced by mucoid strains of the saprophyte Pseudomonas putida and the majority of mucoid strains of saprophytic P. fluorescens (biovars III and V) isolated from casing medium. One biovar II strain (J1) of P. fluorescens produced alginate, a copolymer of mannuronic and guluronic acids, and one strain (H13) produced an apparently unique EPS containing neutral and amino sugars. Of 10 strains of the pathogen "P. gingeri," the causal agent of mushroom ginger blotch, 8 gave mucoid growth on PAF. The "P. gingeri" EPS also was unique in containing both neutral sugar and glucuronic acid. Mucoid, weakly virulent strains of "P. reactans" produced either alginate or marginalan. All 10 strains of the pathogen P. tolaasii, the causal agent of brown blotch of mushrooms were nonnmucoid on PAF. Production of EPS by these 10 strains plus the 2 nonmucoid strains of "P. gingeri" also was negative on several additional solid media as well as in two broth media tested. The results support our previous studies indicating that fluorescent pseudomonads are a rich source of novel EPSs.

Alginates↗
PubMed1995

Secretion of biologically active murine interleukin-2 by Lactococcus lactis subsp. lactis.

Secretion of functional recombinant murine interleukin-2 (mIL2) by Lactococcus lactis was achieved by fusion of the sequence encoding mature mIL2 to the secretion signal leader of the lactococcal usp45 gene placed under transcriptional control of the phage T7 promoter-T7 RNA polymerase expression system. The recombinant mature mIL2 was one of only a few proteins which accumulated in the growth medium. Sequence analysis revealed correct processing at the first amino acid of the mature protein. A T-cell proliferation assay showed that the recombinant protein has the same specific biological activity as mIL2 obtained from a natural source.

Amino Acid Sequence↗
PubMed1994

A small family of elements with long inverted repeats is located near sites of developmentally regulated DNA rearrangement in Tetrahymena thermophila.

Extensive DNA rearrangement occurs during the development of the somatic macronucleus from the germ line micronucleus in ciliated protozoans. The micronuclear junctions and the macronuclear product of a developmentally regulated DNA rearrangement in Tetrahymena thermophila, Tlr1, have been cloned. The intrachromosomal rearrangement joins sequences that are separated by more than 13 kb in the micronucleus with the elimination of moderately repeated micronucleus-specific DNA sequences. There is a long, 825-bp, inverted repeat near the micronuclear junctions. The inverted repeat contains two different 19-bp tandem repeats. The 19-bp repeats are associated with each other and with DNA rearrangements at seven locations in the micronuclear genome. Southern blot analysis is consistent with the occurrence of the 19-bp repeats within pairs of larger repeated sequences. Another family member was isolated. The 19-mers in that clone are also in close proximity to a rearrangement junction. We propose that the 19-mers define a small family of developmentally regulated DNA rearrangements having elements with long inverted repeats near the junction sites. We discuss the possibility that transposable elements evolve by capture of molecular machinery required for essential cellular functions.

Animals↗
PubMed1994

Aprotinin, a Kunitz-type protease inhibitor, stimulates skeletal muscle differentiation.

Aprotinin, a Kunitz-type inhibitor of serine proteases, stimulates myotube formation by mouse G8-1 and C2C12 skeletal muscle myoblasts. This stimulation of morphological differentiation is accompanied by accumulation of myogenin transcripts and production of muscle-specific proteins. In contrast, active TGF beta prevents differentiation of G8-1 and C2C12 myoblasts. When active TGF beta and aprotinin are both added to myoblast cultures, differentiation is inhibited, suggesting the active growth factor acts downstream of the protease inhibitor. TGF beta is found in serum as a latent, dimeric propolypeptide that is cleaved by limited proteolysis to release the biologically active carboxy-terminal dimer. To address the possibility that aprotinin may effect myogenesis by inhibiting proteolytic activation of latent TGF beta, levels of the endogenous growth factor were measured in differentiating myoblast cultures. Latent TGF beta is rapidly depleted from control cultures within 24 hours of plating, but remains relatively stable in aprotinin-treated cultures. Consistent with this, aprotinin-treated cultures have reduced levels of active TGF beta. These data indicate that Kunitz-domain containing protease inhibitors may help orchestrate the onset of myogenesis, possibly by regulating the activity of TGF beta-like molecules.

Amyloid beta-Protein Precursor↗
PubMed1993

Improved cloning vectors and transformation procedure for Lactococcus lactis.

Four shuttle vectors (pMIG 1, 2, 2H and 3) have been constructed based on the broad host-range plasmid pCK1. All the pMIG vectors possess a multiple cloning site containing 12 or more unique restriction enzyme sites, and are stably maintained at either high or low copy number in Lactococcus lactis and in Escherichia coli. By cloning the E. coli pUC replicon into one of these vectors a plasmid was constructed which can replicate to high copy number in recA strains of E. coli. The broad host-range of the pCK1 replicon may enable these cloning vectors to be used in a number of Gram-positive bacteria. One of these vectors was used to optimize an electroporation procedure for transformation of a commonly used plasmid-cured strain MG1363 of L. lactis which routinely yielded 1 x 10(7) to 5 x 10(7) transformants micrograms-1 supercoiled DNA using stored, snap-frozen cells. This transformation efficiency was obtained by growing the cells in medium containing the cell wall weakening agent glycine, to an upper limit of 2.5% w/v. Although growth of L. lactis strain MG1363 was inhibited by the use of 0.5 mol l-1 sucrose as an osmotic stabilizer, the presence of sucrose in the electroporation buffer was critical for high transformation efficiency. Other variables which were tested for their effect on the efficiency of transformation were cell concentration, DNA concentration, pulse time and field strength. These results provide a model procedure which can be followed to optimize conditions for the genetic transformation of various strains of L. lactis.

Base Sequence↗
PubMed1993

Lactococcus lactis: high-level expression of tetanus toxin fragment C and protection against lethal challenge.

To determine if the food-grade bacterium Lactococcus lactis holds promise as a vaccine antigen delivery vector we have investigated whether this bacterium can be made to produce high levels of a heterologous protein antigen. A regulated expression system has been developed which may be generally suitable for the expression of foreign antigens (and other proteins) in L. lactis. The system utilizes the fast-acting T7 RNA polymerase to transcribe target genes, and provides the first example of the successful use of this polymerase in a Gram-positive bacterium. When the performance of the expression system was characterized using tetanus toxin fragment C (TTFC) up to 22% of soluble cell protein was routinely obtained as TTFC. Mice immunized subcutaneously with L. lactis expressing TTFC were protected from lethal challenge with tetanus toxin. These results show for the first time that L. lactis is able to express substantial quantities of a heterologous protein antigen and that this organism can present this antigen to the immune system in an immunogenic form.

Animals↗
PubMed1993

A model system for the investigation of heterologous protein secretion pathways in Lactococcus lactis.

The capacity of recombinant strains of Lactococcus lactis to secrete a heterologous protein was investigated by constructing two expression-secretion vectors (pLET2 and pLET3) for use with a lactococcal gene expression system driven by the highly active T7 RNA polymerase. The vectors incorporated different lactococcal secretion leaders and translation initiation sequences. When tetanus toxin fragment C (TTFC) was used as a test protein, the quantities of TTFC produced by the pLET2-TTFC strain exceeded the rate of secretion of TTFC into the growth medium. However, nearly all of the soluble TTFC associated with the cell (3.4%) was translocated through the cell membrane. The pLET3-TTFC strain did not accumulate TTFC intracellularly and exhibited growth characteristics and viability identical to the growth characteristics and viability of the control strain. This strain secreted approximately 2.9 mg of TTFC per liter into the growth medium after 6 h of growth under test tube conditions. Our results indicate that L. lactis is capable of secreting substantial amounts of heterologous protein and also confirm the findings of other workers that the cell wall may serve as a functional barrier to the diffusion of some secreted proteins into the growth medium.

Amino Acid Sequence↗
PubMed1993

Preimplantation mouse embryos express a cell surface receptor for tissue-plasminogen activator.

The serine protease tissue-plasminogen activator (t-PA) has previously been shown to be intracellular in mouse secondary oocytes and extracellular in fertilized eggs. Here we demonstrate that extracellular t-PA activity is bound to the surface of the fertilized egg. The level of t-PA activity associated with preimplantation mouse embryos decreases in the 2-cell stage embryo, then increases in 4-cell and morula stage embryos. However, morulae grown in culture from fertilized eggs lack t-PA activity but are able to bind exogenously added mouse t-PA. Additionally, northern analysis indicates that preimplantation embryos do not contain detectable levels of t-PA mRNA. Therefore, the enzyme activity associated with 4-cell and morula stage embryos in vivo is derived from t-PA present in the oviduct lumen that binds the embryo, and not from protein produced from translation of embryonic mRNA. The binding activity is species and protein specific in that neither mouse urokinase-type plasminogen activator (u-PA) nor human t-PA bind to cultured morulae. Furthermore, binding activity is dose-dependent and saturable, and does not require the active site of t-PA. These data indicate that a cell surface-specific t-PA-binding activity exists in the preimplantation mouse embryo and may localize function and concentrate the proteolytic activity of t-PA in early mouse development.

Animals↗
PubMed1992

Multiple transduction as a means of preserving ganciclovir chemosensitivity in sarcoma cells carrying retrovirally transduced herpes thymidine kinase genes.

The potential value of retroviral gene transfer as a means of targeting therapeutic genes to neoplastic cells is threatened by the tendency of occasional cells to lose transduced genes or their expression. To determine whether this threat could be reduced by transducing multiple copies of a therapeutic gene, we compared 8 murine sarcoma sublines carrying from 1 to 6 copies of a retrovirally transduced herpes simplex virus thymidine kinase gene, which sensitizes cells to ganciclovir (GCV). When variability consequent to differences in vector integration site was excluded, increased copy number was associated with an increase in GCV sensitivity and a major reduction in the frequency of GCV-resistant mutants. The results suggest a potential means of preserving the efficacy of future antineoplastic gene therapy strategies.

Animals↗
PubMed1992

Pharmacokinetically designed double-blind placebo-controlled study of nortriptyline in 6- to 12-year-olds with major depressive disorder.

A random assignment, double-blind, placebo-controlled study of nortriptyline in 50 prepubertal 6- to 12-year-olds with Research Diagnostic Criteria and DSM-III major depressive disorder was performed. The protocol included a 2-week placebo wash-out phase and an 8-week double-blind, placebo-controlled phase with weekly plasma level monitoring. Active subjects had their plasma level pharmacokinetically placed at 80 +/- 20 ng/ml by using previously developed tables to determine the starting dose from a plasma level 24 hours after a single dose administered at baseline. The mean plasma level was 89.9 ng/ml. The study population was severely depressed, had a chronic, unremitting course of long duration before the study, had a high percentage of family histories with affective disorder, alcoholism and suicidality, and had a high rate of comorbidity. None of the subjects had ever received tricyclic antidepressants before this study. There was a poor rate of response in both treatment groups (30.8% active, 16.7% placebo). Active subjects did not evidence the anticholinergic side effects reported in adult samples. The implications of these findings for future pharmacotherapy studies of depressed children are discussed.

Child↗
PubMed1992

Differentiation of free-living Anabaena and Nostoc cyanobacteria on the basis of fatty acid composition.

The cellular fatty acids of free-living, nitrogen-fixing cyanobacteria belonging to the genera Anabaena and Nostoc were analyzed to differentiate the genera. The fatty acid compositions of 10 Anabaena strains and 10 Nostoc strains that were grown for 12 days on BG-11o medium were determined by gas-liquid chromatography-mass spectroscopy. Of the 53 fatty acids detected, 17 were major components; the average level for each of these 17 fatty acids was at least 0.9% of the total fatty acids (in at least one of the genera). These fatty acids included (with mean percentages in the Anabaena and Nostoc strains, respectively) the saturated fatty acids 16:0 (30.55 and 23.23%) and 18:0 (0.77 and 1.27%); several unsaturated fatty acids, including 14:1 cis-7 (2.50 and 0.11%), 14:1 cis-9 (3.10 and 3.41%), a polyunsaturated 16-carbon (sites undetermined) fatty acid with an equivalent chain length of 15.30 (1.20 and 1.03%), 16:4 cis-4 (0.95 and 0.87%), 16:3 cis-6 (2.16 and 1.51%), 16:1 cis-7 (1.44 and 0.36%), 16:1 cis-9 (6.53 and 18.76%), 16:1 trans-9 (4.02 and 1.35%), 16:1 cis-11 (1.62 and 0.42%), 18:2 cis-9 (10.16 and 12.44%), 18:3 cis-9 (18.19 and 17.25%), 18:1 cis-9 (4.01 and 5.10%), and 18:1 trans-9 (0.92 and 1.94%); and the branched-chain fatty acids iso-16:0 (2.50 and 1.14%) and iso-15:1 (0.34 and 2.05%).(ABSTRACT TRUNCATED AT 250 WORDS)

Anabaena↗
PubMed1992

Comparison of four methods to assess body composition in black and white athletes.

Four methods of assessing body composition were compared in 55 black and 35 white, Division I, American football players. Percent body fat (%BF) was estimated with hydrostatic weighing at residual volume, corrected for race; seven-site skinfolds (7 SF), corrected for race; bioelectrical impedance analysis (BIA); and near-infrared spectrophotometry (NIR). Percent body fat with HW in blacks (mean = 14.7%) and whites (19.7%) did not differ (P > .05) from %BF with 7 SF (blacks, 14.7%; whites, 19.0%). In relation to HW, BIA significantly (P < .05) overpredicted (blacks: 20.1%, SEE = 3.2%; whites: 22.3%, SEE = 4.3%) and NIR underpredicted %BF (blacks: 12.6%, SEE = 3.9%; whites: 17.7%, SEE = 3.6%). The contribution of BIA variables (resistance, phase angle, conductance) and NIR optical density to predict %BF was trivial compared to body mass index. It appears that race may not substantially influence %BF prediction by NIR and BIA. It was concluded that when considering the cost and expertise required with NIR and BIA, SF measurements appear to be a superior alternative for rapid and accurate body composition assessment of athletes, independent of race.

Adolescent↗
PubMed1992

Effects of taper on swim power, stroke distance, and performance.

Competitive swimmers progressively reduce training volume or "taper" prior to an important competition in an effort to improve performance capabilities. The purpose of the current study was to determine the effects of taper upon factors associated with swim performance. Twelve intercollegiate swimmers were tested before and after taper in preparation for their season-ending meet. Power during a tethered sprint swim increased significantly (P < 0.05) by approximately 5% with taper. No significant changes occurred in distance per stroke, oxygen consumption, and post-exercise blood lactate level during a 182.9-m submaximal swim with taper. Five swimmers were additionally tested after shaving exposed body hair upon completion of taper. Swim power did not increase further with hair removal. In contrast, shaving significantly increased distance per stroke (P < 0.05) by approximately 5%. These data indicate that reduced training specifically improves swim power; however, removing exposed body hair after taper may additionally enhance performance capabilities by increasing distance per stroke.

Adult↗
PubMed1991

The effect of warm-up on responses to intense exercise.

The purpose of this study was to determine if prior physical activity (warm-up) affected physiological responses to intense exercise. Eight highly trained collegiate swimmers performed a paced 365.8-m (440 yds) intense swim (mean +/- SE, 94.4 +/- 3.3% VO2max) 5 min after the following warm-up conditions: trial N, no warm-up; trial S, an intensity-specific interval set (4 x 45.7 m with one-min rest intervals at the intense swim pace); trial M, a mild-intensity, long-duration swim (1371.6 m at 64.7 +/- 3.3% VO2max); and trial MS, a mild-intensity, long-duration swim (1188.7 m at the same pace as trial M) followed by the intensity-specific interval set (trial S). When comparing trial N with trials M and MS, stroke distance (m/stroke) was significantly (p less than 0.05) lower during the last 91.4 m of the intense, paced swim and 3-, 5-, 8- and 10-min recovery blood lactate levels and one-minute recovery heart rates were significantly elevated (p less than 0.05). There was no significant difference (p greater than 0.05) in stroke distance during the final 91.4 m of the intense swim between trials S and N. There were no significant differences for any variables between trials M and MS. These results suggest that a warm-up consisting of mild-intensity, long-duration exercise was beneficial compared to no warm-up and that intensity-specific exercise was not a vital component of warm-up. Although performance was not directly measured, these data demonstrate the benefit of warm-up.

Adult↗