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C J Smith

Publications and source records attributed to C J Smith.

At least 181 records · Page 10Linked to original sources
PubMed1993

Identification of a circular intermediate in the transfer and transposition of Tn4555, a mobilizable transposon from Bacteroides spp.

Transmissible cefoxitin (FX) resistance in Bacteroides vulgatus CLA341 was associated with the 12.5-kb, mobilizable transposon, Tn4555, which encoded the beta-lactamase gene cfxA. Transfer occurred by a conjugation-like mechanism, was stimulated by growth of donor cells with tetracycline (TC), and required the presence of a Bacteroides chromosomal Tcr element. Transconjugants resistant to either FX, TC, or both drugs were obtained, but only Fxr Tcr isolates could act as donors of Fxr in subsequent matings. Transfer of Fxr could be restored in Fxr Tcs strains by the introduction of a conjugal Tcr element from Bacteroides fragilis V479-1. A covalently closed circular DNA form of Tn4555 was observed in donor cells by Southern hybridization, and the levels of this circular transposon increased significantly in cells grown with TC. Both the cfxA gene and the Tn4555 mobilization region hybridized to the circular DNA, suggesting that this was a structurally intact transposon unit. Circular transposon DNA purified by CsCl-ethidium bromide density gradient centrifugation was used to transform Tcs B. fragilis 638, and Fxr transformants were obtained. Both the circular form and the integrated Tn4555 were observed in transformants, but the circular form was present at less than one copy per chromosomal equivalent. Examination of genomic DNA from Fxr transformants and transconjugants revealed that Tn4555 could insert at a wide variety of chromosomal sites. Multiple transposon insertions were present in many of the transconjugants, indicating that there was no specific barrier to the introduction of a second transposon copy.

Bacteroides↗
PubMed1993

Heat shock protein and the double insult theory for the development of insulin dependent diabetes.

Heat shock proteins (HSP) are the most widely conserved group of proteins in phylogeny and play an important role in infection and autoimmunity. HSP65 has been suggested as the primary antigen in insulin dependent diabetes while an alternative antigen glutamic acid decarboxylase (GAD), has similar amino acid sequences. A 'double insult theory' for the development of insulin dependent diabetes is suggested whereby a bacterial infection leads to the production of HSP antibody. If during a 'window of opportunity' this is followed by a viral infection of the islet cells this could, in certain histocompatibility locus antigen (HLA) groups only, lead to the production of HSPs on the cell surface and a destructive autoimmune reaction.

Animals↗
PubMed1993

A putative marker for human melanoma. A monoclonal antibody derived from the melanoma gene in the Xiphophorus melanoma model.

A MoAb was raised against a peptide corresponding to an exposed domain of the putative tyrosine kinase receptor protein encoded by Xmrk, a gene involved in melanoma formation and/or progression in the Xiphophorus fish melanoma model. The antibody reacts specifically with cells from human melanocytic lesions, ie, common acquired nevi, primary and metastatic melanoma biopsies. No reactivity with other cells, including normal melanocytes, was observed in the biopsies or with cells in biopsies from normal tissue (skin, liver, lung, spleen) and from other malignancies including those of neuroectodermal origin. The reactivity was very weak and variable in metastatic melanomas but very strong and characteristic of a receptor-type antigen in primary melanomas, a stage in melanoma progression in which cells have acquired metastasizing potential. It is suggested that the antigen recognized may be involved in growth promotion and represents the human equivalent of the fish melanoma gene product.

Animals↗
PubMed1992

In Indian country.

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Anecdotes as Topic↗
PubMed1992

The development of enzyme-linked immunosorbent assays (ELISA) for the catecholamines adrenalin and noradrenalin.

Monoclonal antibodies have been raised against adrenalin and noradrenalin and used as the basis of enzyme-linked immunosorbent assays (ELISA) to detect and estimate the concentrations of these catecholamines. Inhibition assays are described, with sensitive quantification in the range from 1 mg/ml to 100 pg/ml. Cross-reactivity assays reveal that neither assay is subject to interference by catecholamine metabolites at concentrations less than 100 ng/ml and 1 micrograms/ml respectively. Isolation and quantification of both catecholamines from clinical samples is discussed and the potential application of these ELISAs in a clinical setting is proposed.

Animals↗
PubMed1992

Molecular cloning and expression of human myocardial cGMP-inhibited cAMP phosphodiesterase.

We have cloned a cDNA for a myocardial cGMP-inhibited cAMP phosphodiesterase (cGI PDE) from a human heart cDNA library in lambda Zap II. The open reading frame [3.5 kilobases (kb)] of cDNA clone n.13.2 (7.7 kb) encodes a protein of 125 kDa. In Northern blots of total human ventricle RNA, a single mRNA species (8.3 kb) hybridized with a 4-kb EcoRI restriction fragment of clone n.13.2 cDNA (containing the entire open reading frame). The carboxyl-terminal region of the deduced amino acid sequence of the cGI PDE contains the putative catalytic domain conserved among mammalian PDE families. A partial cDNA clone, n.2, encoding a truncated, 54-kDa cGI PDE containing the conserved domain was expressed as a catalytically active fusion protein in Escherichia coli. cAMP hydrolytic activity was inhibited by cGMP and OPC 3911 but not by rolipram. Thus, this report provides direct proof that the conserved domain contains the catalytic core of cGI PDEs.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1992

Insulin and isoproterenol induce phosphorylation of the particulate cyclic GMP-inhibited, low Km cyclic AMP phosphodiesterase (cGI PDE) in 3T3-L1 adipocytes.

The cGI PDE in particulate fractions of differentiated adipocytes (but not control 3T3-L1 fibroblasts) was cross-reactive with a polyclonal antibody raised against the bovine adipose cGI PDE. The 3T3-L1 adipocyte cGI PDE is a 135 kDa protein which is phosphorylated in 32P-labeled cells in response to beta-agonist or insulin. These results indicate that the 3T3-L1 cGI PDE is similar in structure and hormonal regulation to the analogous enzyme in the rat adipocyte.

3T3 Cells↗
PubMed1992

Efficacy of topically applied ivermectin against sarcoptic mange (Sarcoptes scabiei var. bovis) of cattle.

A total of 12 adult cattle with sarcoptic mange were included in a study to confirm the efficacy of ivermectin applied topically at 500 micrograms/kg against Sarcoptes scabiei var. bovis. Animals were allocated by restrictive randomization according to mite counts to serve as untreated controls or were treated with a topical formulation of ivermectin applied along the backline on healthy skin. Mites were counted in epidermal scrapings made prior to treatment and at 7, 14, 21, 28, 42 and 56 days thereafter. Except for three mites recovered from one animal on day 28, no mites were found after day 14 over the remainder of the trial in scrapings from animals treated with ivermectin. Differences in the numbers of mites recovered from animals treated with ivermectin and those found on controls were significant (P less than 0.05) for counts carried out on days 14-56. Clinical signs of mange resolved following treatment, whereas the extent of lesions and severity of clinical signs increased in the control animals over the trial period.

Administration, Topical↗
PubMed1992

Exposure to ovarian steroids elicits a female pattern of plasma cortisol levels in castrated male macaques.

Our recent observations (1) that there is a difference in circadian patterns of plasma cortisol levels between male and female macaques and (2) that after gonadectomy these differences in the patterns and in the levels of cortisol were reduced prompted us to investigate how 17 beta-estradiol (E2) and progesterone affect cortisol secretion in orchidectomized male rhesus macaques. Five male macaques, castrated as adults, were implanted subcutaneously with segments of silastic tubing filled with E2 and with progesterone in a manner such that the levels and the sequence of these hormones mimicked those that occur during the menstrual cycle of intact female macaques. Since previous studies had shown that the difference in cortisol patterns was due to higher levels in females during the day, these studies were conducted from 0800 to 2000 hours. Blood samples were collected in an adjacent room at 15-minute intervals. Separate trials were conducted 2 weeks after E2 was implanted and levels were 110 +/- 14 pg/ml and again 2 weeks later after progesterone was implanted and E2 levels were 59 +/- 15 pg/ml; progesterone levels averaged 4.0 +/- 0.65 ng/ml. Mean plasma concentrations of cortisol (microgram/100 ml) for the 12-hour period were three-fold higher in orchidectomized males treated with E2 (17.2) and with E2 + progesterone (18.0) than in intact males (4.9). Levels in males treated with ovarian steroids were double that (8.5 micrograms/100 ml) observed for intact females.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Heterologous gene expression in Bacteroides fragilis.

Bacteroides fragilis and other gastrointestinal tract Bacteroides are unusual gram-negative eubacteria in that genes from other gram-negative eubacteria are not expressed when introduced into these organisms. To analyze gene expression in Bacteroides, expression vector and promoter probe (detection) vector systems were developed. The essential feature of the expression vector was the incorporation of a Bacteroides insertion sequence element, IS4351, which possesses promoter activity directed outward from its ends. Genes inserted into the multiple cloning site downstream from an IS4351 DNA fragment were readily expressed in B. fragilis. The chloramphenicol acetyltransferase (cat) structural gene from Tn9 was tested and conferred chloramphenicol resistance on B. fragilis. Both chloramphenicol resistance and CAT activity were shown to be dependent on the IS4351 promoters. Similar results were obtained with the Escherichia coli beta-glucuronidase gene (uidA) but activity was just 30% of the levels seen with cat. Two tetracycline resistance determinants, tetM from Streptococcus agalactiae and tetC from E. coli, also were examined. tetC did not result in detectable tetracycline resistance but the gram-positive tetM gene conferred high-level resistance to tetracycline and minocycline in Bacteroides hosts. Based on the cat results, promoter probe vectors containing the promoterless cat gene were constructed. These vectors were used to clone random B. fragilis promoters from partial genomic libraries and the recombinants displayed a range of CAT activities and chloramphenicol MICs in B. fragilis hosts. In addition, known E. coli promoters (Ptet, Ptac, Ptrc, Psyn, and P1P2rrnB) were tested for activity in B. fragilis. No chloramphenicol resistance or CAT activity was observed in B. fragilis with these promoters.

Bacteroides fragilis↗
PubMed1992

Graded work exposure to promote work return after severe hand trauma: a replicated study.

Fifty-one patients with posttraumatic stress disorder after work-related hand injuries were placed on a graded work exposure program to facilitate return to work. These patients consisted of an initial group of 25 patients and a replication group of 26 patients. The program returned 92% of the initial group and 88% of the replicated group to work with their previous employers. At 6-month follow-up, 88% of the initial group and 80.1% of the replication group were still working full-time at the jobs to which they had returned. All of the patients not working with their previous employer at follow-up had appraisal/projected flash-backs, which have previously been associated with a 90% failure to return to work. This intervention was successful with 73% of the patients experiencing such flashbacks. In conclusion, graded work exposure was an effective treatment to promote return to work for patients experiencing significant psychological symptomatology after severe hand injury.

Accidents, Occupational↗
PubMed1992

Activation of a cryptic streptomycin-resistance gene in the Bacteroides erm transposon, Tn4551.

Bacteroides compound transposons encoding erm resistance are highly homologous but previous studies have shown some divergence of Tn4551. Results presented here describe a novel Tn4551 streptomycin-resistance gene, aadS, that was phenotypically silent in wild-type Bacteroides. However, aadS expression could be activated by a trans-acting chromosomal mutation. The aadS-encoded peptide displayed significant homology to Gram-positive streptomycin-dependent adenyltransferases, and enzymatic analysis confirmed the production of this activity. Examination of the nucleotide sequence showed that 200 bp upstream of aadS, the DNA base composition changed abruptly from 31% G+C to 48% G+C. These two regions were demarcated by a DNA sequence with homology to the recombination hot spots reported for Tn21 and the Bacteroides ermFU gene and to sequences at the ends of the chromosomal Bacteroides conjugal element, XBU4422.

Amino Acid Sequence↗
PubMed1992

Analysis of rRNA restriction fragment length polymorphisms from Bacteroides spp. and Bacteroides fragilis isolates associated with diarrhea in humans and animals.

The Escherichia coli rRNA operon rrnB was used as a 32P-labeled hybridization probe in Southern blots of genomic DNAs from representative strains of the saccharolytic, gram-negative, obligate anaerobes of the genus Bacteroides. Control experiments with the B. fragilis type strain ATCC 25285 established that nearly identical rRNA fragment patterns were produced when either the E. coli rrnB gene probe or homologous rRNA isolated from B. fragilis was used as the probe. In addition, it was shown that a specific 16S or 23S rrnB gene probe also could be used to produce fragment patterns suitable for analysis. Thirty-one strains from 8 of the 10 recognized Bacteroides species were then examined. The resulting autoradiographs revealed specific fragment patterns for all but one (B. ovatus) of the species tested. Restriction fragment length polymorphisms were observed for many of the strains tested, but these differences did not hinder species classification. The five B. ovatus strains examined did not form a distinct group, and their rRNA fragment patterns displayed a marked heterogeneity. The same approach was applied to a unique set of enterotoxin-producing B. fragilis strains isolated from animals and humans with diarrhea. The results demonstrated that these strains were in fact B. fragilis and that they produce rRNA fragment patterns closely related to those of the type strain ATCC 25285. This set of strains did not appear to form a separate subgroup or genotype within the B. fragilis species, and there were no distinguishable restriction fragment length polymorphisms that could be used to specifically separate enterotoxin-producing strains from nonenterotoxigenic strains.

Animals↗
PubMed1992

Restraint inhibits luteinizing hormone and testosterone secretion in intact male rhesus macaques: effects of concurrent naloxone administration.

The present study investigates how restraint affects the hypothalamo-hypophysial adreno-cortical axis and the hypothalamo-hypophysial gonadal axis in intact, adult male rhesus macaques. Restraint was chosen because it is not physically painful or harmful to the animal, but rather serves both as a physical and psychological stressor. Blood samples were collected from a remote site at 15-min intervals beginning at 07.00 h from tethered adult male rhesus macaques. Each of 4 animals was subjected to 6 h of chair restraint after a 3-hour control period in the animals' home cage. Samples were collected for an additional 6 h at the end of the restraint period when the animal was returned to its home cage. Brief anesthesia with ketamine (administered through the indwelling catheter) facilitated transfer of the animals to and from the chair. Blood samples were collected from 4 undisturbed males to document LH and testosterone secretion throughout the day. Plasma ACTH and cortisol, measured as indexes of stress, were elevated within 15 min after initiation of restraint and remained elevated for most of the restraint period. Conversely, LH and testosterone began to fall immediately after restraint and remained suppressed for several hours after the animals were removed from restraint and returned to their home cage. Testosterone levels were more consistently inhibited than were LH levels, a reflection of the fact that in some animals, testosterone remained low after the return of pulsatile LH secretion. In studies with naloxone (Nx), the opiate receptor antagonist (5 mg bolus plus 5 mg/h) was given beginning either at the initiation of restraint (n = 2) or 2 h thereafter (n = 2), and continued until the end of the restraint period. With Nx treatment of the restrained animals, both ACTH and cortisol were elevated as in the controls and LH and testosterone secretion were significantly increased within 1-2 h. However, after the Nx treatment was terminated and the animals were returned to their home cages, plasma levels of LH and testosterone were not different from levels in restrained animals and were significantly less than levels in untreated animals. These data show that restraint is a potent stimulus for activation of the HPAC axis and inhibits both LH and testosterone release. The pathway through which restraint inhibits LH release probably includes endogenous opiate suppression of hypothalamic GnRH release since Nx partially blocks the effect of stress.

Adrenocorticotropic Hormone↗