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Biomedical subjects

J M Clark

Publications and source records attributed to J M Clark.

At least 181 records · Page 10Linked to original sources
PubMed1987

Novel blunt-end addition reactions catalyzed by DNA polymerase I of Escherichia coli.

DNA polymerase I (Klenow fragment) of Escherichia coli catalyzes the addition of deoxynucleotides to 3' hydroxyl termini of blunt-ended DNA fragments. The product of the reaction, which we call +1 addition, is found only in very low yield under conditions that permit editing by the 3'----5' exonuclease activity of the wild-type polymerase. A mutant form of the Klenow fragment that lacks detectable 3'----5' exonuclease activity shows an elevated accumulation of the +1 addition product. The mutant enzyme can use any one of the four dNTPs to carry out the reaction when each precursor is provided individually. However, in the presence of all four dNTPs the addition of dATP is strongly preferred. Suppression of the editing function of the wild-type polymerase through the use of high concentrations of exogenous deoxynucleoside monophosphates also results in a significant increase in the amount of +1 addition product formed. The presence of a high dNMP concentration also alters the specificity of the nucleotide addition reaction carried out by the wild-type enzyme. Thus, in addition to dATP, the dNTP which is complementary to the exogenous deoxynucleoside monophosphate, is also used in the +1 addition reaction. A similar effect of dNMPs on the specificity of nucleotide addition was obtained with the mutant Klenow fragment. These results define two pathways for the +1 addition reaction: one that does not require coding information from the DNA template and a second in which coding information is provided by the exogenous dNMP.

Base Sequence↗
PubMed1987

Functional effects of cis-thymine glycol lesions on DNA synthesis in vitro.

Thymine glycol, a DNA lesion produced by ionizing radiation, has been introduced site specifically at high frequency into a synthetic oligonucleotide by chemical oxidation of the single thymine residue within the sequence. The lesion-containing template was then annealed to a complementary synthetic primer and used to study the effects of cis-thymine glycol lesions on DNA polymerase function in vitro. Synthesis by polymerase I (Klenow fragment), T4 DNA polymerase, and polymerase alpha 2 was arrested quantitatively at the site of the lesion. AMV reverse transcriptase was less inhibited and was able to synthesize past a significant fraction of the lesions. Changing the template base immediately 5' to thymine glycol from A to C did not significantly alter the pattern of synthesis arrest for any of the polymerases. The correct nucleotide, dAMP, was inserted opposite the lesion more than 90% of the time by all four polymerases, suggesting that thymine glycol forms a reasonably stable base pair with adenine. However, the 3'-5' exonuclease activity of polymerase I removed a 3'-terminal dAMP residue more rapidly from an A . thymine glycol base pair than from an A.T base pair. These results suggest that increased nucleotide turnover at the site of the lesion contributes to the inhibitory effects of thymine glycol lesions on DNA synthesis in vitro, at least for polymerases such as polymerase I that have intrinsic or associated editing exonuclease functions.

Base Sequence↗
PubMed1987

Modeling and molecular mechanical studies of the cis-thymine glycol radiation damage lesion in DNA.

Computer graphics and energy minimization techniques were used to construct a model of DNA containing cis-thymine glycol, an oxidation product of thymine formed in DNA by ionizing radiation. The model simulated an experimental DNA substrate used to study the effects of this lesion on DNA synthesis in vitro. The results derived from the model indicate that cis-thymine glycol lesions introduce localized perturbations of DNA structure. Specifically the model shows that interactions with the neighboring base pair on the 5' side are significantly destabilized by thymine glycol whereas interactions with the 3' base pair are stabilized by the lesion. The magnitude of these effects is modulated by the nucleotide sequence around the lesion, particularly by the nature of the base on the 3' side. The base pair formed between adenine and thymine glycol is energetically stable and shows minimal distortion, suggesting that this lesion retains the ability to direct the insertion of the correct nucleotide during DNA synthesis.

Base Sequence↗
PubMed1987

The action of two classes of pyrethroids on the inhibition of brain Na-Ca and Ca + Mg ATP hydrolyzing activities of the American cockroach.

Ca2+-stimulated ATP hydrolyzing activities (i.e. Na-Ca ATP hydrolysis and Ca + Mg ATP hydrolysis) measured in cockroach brain tissue were highly sensitive to the action of pyrethroid insecticides under in vitro conditions. Non-cyano-containing pyrethroids inhibited Na-Ca ATP hydrolysis to a greater extent than their cyano-containing counterparts. The reverse is true for pyrethroid action on Ca + Mg ATP hydrolysis. Nonmitochondrial Ca + Mg ATP hydrolysis of disrupted synaptosomes was the most sensitive activity examined. Ca2+-stimulated ATP hydrolyzing activities were inhibited in cockroaches poisoned by permethrin in vivo. In vivo poisoning occurred in the presence of a similar amount of bound [14C]permethrin which had been determined to cause a substantial amount of inhibition to Ca2+-stimulated ATP hydrolyzing activities in vitro.

Adenosine Triphosphatases↗
PubMed1987

The relationship of neck orientation to the shape of the proximal femur.

Neck angles and various dimensions measured from 70 normal cadaver femora and from clinical radiographs of 120 normal hips were analyzed. The neck-shaft angle and the inclination of the medial trabecular system varied over a range of approximately 35 degrees. The location of the femoral head center in relation to the femoral shaft and to the trochanters was related to neck orientation. As the neck became vertical, horizontal offset of the head center from the shaft decreased and its vertical level relative to that of the trochanter increased. The abductor muscle lever arm was measured with two methods. Values obtained when assuming that the joint resultant force follows the medial trabeculae demonstrated the closest relationship to the distance from head center to body midline. The normal shape of the proximal femur is variable, and variations in the absence of intrinsic bone disease appear to reflect adaptation to physiologic variations in the line of action of muscle forces.

Adult↗
PubMed1987

In vivo evaluation of tigemonam, a novel oral monobactam.

Tigemonam, a new monobactam with excellent activity against gram-negative bacteria, was evaluated for in vivo efficacy and absorption after oral administration to laboratory animals. Tigemonam is absorbed when administered orally to mice and dogs. In a variety of gram-negative systemic infections in mice, orally administered tigemonam was efficacious in all infections studied. Comparison drugs such as amoxicillin, cephalexin, and cefaclor were less efficacious, especially in infections caused by beta-lactamase-producing organisms. In localized infections, tigemonam also demonstrated excellent in vivo activity. In acute pyelonephritis in mice caused by Escherichia coli or Proteus sp., tigemonam was very effective. In a rat lung model with Klebsiella pneumoniae, tigemonam was active with a median effective dose of 46 mg/kg compared with 160 mg/kg for cefaclor and over 200 mg/kg for amoxicillin. Tigemonam was well absorbed in laboratory animals and with its excellent gram-negative spectrum of activity should prove of value in oral antibiotic therapy in humans.

Acute Disease↗
PubMed1987

Hyperbaric hyperoxia reversibly inhibits erythrocyte phospholipid fatty acid turnover.

The present study is one component of a comprehensive investigation of oxygen tolerance of tissues and organs in normal human subjects. The focus of this study was the acylation of membrane phospholipid in situ by erythrocytes. Activation of exogenous [9,10-3H]oleic acid to acyl thioester and transesterification of the acyl thioester into phospholipid by intact human erythrocytes incubated in vitro decreased 30% after exposure of 10 human subjects to hyperbaric hyperoxia (100% O2, 3 ATA, 3.5 h). Partial recovery of activity could be detected when additional cells were obtained from these subjects and assayed in vitro 24 h after cessation of exposure. No significant change in membrane phospholipid fatty acid composition was detected under these conditions. The reduced glutathione content of intact erythrocytes increased by 15% after hyperbaric hyperoxia and remained elevated 24 h after exposure. In isolated membranes prepared from the same cells activation of [9,10-3H]oleic acid to acyl thioester and its transesterification into phospholipid did not change after hyperoxia. Since the ability of intact cells to replace oxidized fatty acids in membrane phospholipids via deacylation and reacylation in situ may be necessary for the maintenance of membrane integrity during exposure to oxidative stress, the decrease in [9,10-3H]oleic acid incorporation by human erythrocytes detected in vitro after hyperbaric hyperoxia in vivo may reflect an early event in the pathogenesis of oxygen-induced cellular injury and may be a useful index for assessment of the tolerance of tissues to hyperoxia.

Adult↗
PubMed1987

Mariel refugees: six years after.

In 1983, the economic situation of Mariel Cubans could be summarized as abysmal. Those without a job represented close to 1/2 of the sample; the unemployment rate amounted to 27%. A study conducted by Johns Hopkins University in collaboration with Miami-Dade Community College and Florida International University has investigated whether Mariels remain a group apart within the broader Cuban community or whether they have melted into the rest of the community. The study followed a large sample of Mariel refugees living in the Miami area over a period of several years. A sample of 514 Mariel men and women were interviewed in 1983 and were reinterviewed during 1985-1986. The authors conclude that, as a group, Mariel refugees have made rapid progress toward integration into the South Florida economy. There has been a rapid decline in unemployment during the past 2 years and a rapid shift into self-employment. There is still a sizable gap in labor force participation and earnings between this group and the pre-Mariel Cuban population. Mariel incorporation into South Florida society has taken place almost completely through their absorption into the pre-existing Cuban community; there is tension, however, as Mariel refugees see themselves as more discriminated against by fellow Cubans than by outside Anglos. This minority-within-a-minority syndrome is likely to underlie the reported willingness of many to leave the US if conditions in Cuba were to change for the better. Despite these problems, the majority of Mariel refugees would come again to the US if they had to make the choice anew and declare themselves satisfied with their present lives. Within Dade County, the more positive indicators of economic advancement and general adaptation are found among refugees in the cities of Miami and Hialeah. The most problematic economic situation and the greatest alienation from their surroundings is detected among refugees living elsewhere, primarily in Miami Beach. Results indicate that official and private programs targeted on this refugee group should give priority to 4 aspects: 1) support of small entrepreneurship through credit and training facilities to buttress the widespread efforts in this direction; 2) provision of English language courses and help to overcome extreme language deficiencies; 3) promotion of the reunification of the Mariel refugee families who were separated against their will; and 4) additional efforts by Cuban-American organizations to combat lingering prejudice against Mariel refugees.

Americas↗
PubMed1987

Anatomy of the abductor muscles of the hip as studied by computed tomography.

Computed tomography provides a non-invasive method for studying anatomy in live subjects. The abductor muscles of the hip were studied by using computed tomography to describe their position, size, and orientation. The outlines of the muscles were clearly identifiable; patterns that are associated with trauma, arthritis, and so on, differed from normal. The patterns in eighteen normal hips in twelve adults were mapped in detail. The cross-sectional areas of the abductor muscles were measured and vectors representing the total pull of the abductor muscle in the frontal and sagittal planes were constructed for each hip. The variations in individual and composite muscular anatomy were recorded. The inclination of the axis of the abductor muscle ranged from 17 to 26 degrees (standard deviation, 2.9 degrees) in the frontal plane and from -2 to 14 degrees (standard deviation, 4.2 degrees) in the sagittal plane.

Adult↗
PubMed1987

Prosthetic replacement for chronic unreduced dislocations of the shoulder.

Seven patients with chronic dislocations of the shoulder were treated by humeral head or total shoulder replacement. All patients had significant humeral head damage. Three dislocations were posterior and four were anterior. All had been unreduced for at least two months. A Neer prosthesis was used in each patient and the version of the humeral component was altered by 30 degrees-50 degrees away from the direction of the dislocation. Only a sling and swathe was used postoperatively, and the patient was allowed early motion in a limited-goals physical therapy program. There were no redislocations and all patients were improved over their preoperative status. Five patients achieved a good result and two had a fair result based on a 100-unit rating system. One transient axillary nerve palsy occurred, but there were no other complications. Prosthetic replacement for chronic unreduced shoulder dislocations is a reliable procedure when severe humeral head damage exists.

Adult↗
PubMed1986

Characterization of the ATP-promoted aspect of Na+-Ca2+ exchange present in squid retinal nerve axolemma.

Using an in vitro system which consists of an axolemma-rich vesicle fraction prepared from squid retinal nerve fibers, an Na+-Ca2+ exchange process has been characterized and appears identical with that reported in squid giant axon. This exchange is absolutely dependent on the establishment of an Na+ gradient, shows monovalent and divalent cation specificity and is highly sensitive to monensin, A23187 and valinomycin but not to ouabain, digitoxigenin, vanadate, pentylenetetrazole, tetrodotoxin or tetraethylammonium. Furthermore, it was found that the exchange process is enhanced by the addition of ATP. This ATP-promoted aspects of Na+-Ca2+ exchange shares many similar characteristics with Na+-Ca2+ ATP hydrolysis and may indicate a common mechanism for both activities via a protein phosphorylation-dephosphorylation event.

Adenosine Triphosphate↗
PubMed1986

Thymine glycol lesions terminate chain elongation by DNA polymerase I in vitro.

Single-strand circular DNA from bacteriophage M13mp9 was chemically modified with osmium tetroxide to introduce specifically cis-thymine glycol lesions, a major type of DNA damage produced by ionizing radiation. An oligonucleotide primer was extended on damaged and undamaged templates using either the large fragment of E. coli pol I or T4 DNA polymerase. The reaction products were analysed by electrophoresis alongside a DNA sequence ladder. Synthesis on the damaged templates terminated at positions opposite thymine bases in the template. These results indicate that cis-thymine glycol lesions in single-strand DNA constitute blocks to synthesis by DNA polymerases in vitro. Surprisingly, replication halts after the correct nucleotide, dAMP, is inserted opposite the lesion. These results imply that the primary effect of the thymine glycol lesion is suppression of DNA synthesis and that the lesion is not a potent mutagen.

Base Sequence↗
PubMed1986

Effect of hyperbaric oxygen exposure on oxygen tension within the medullary canal in the rabbit tibial osteomyelitis model.

The effect on intramedullary oxygen tension of 100% oxygen exposure at 1, 2, 2.4, and 3 atm pressure was studied in 12 New Zealand white rabbits with chronic right tibial osteomyelitis. The model, modified from that described by others, incorporates a multipuncture silastic closure plug placed transcortically in the proximal tibial metaphysis through which platinum needle, polarographic electrode oxygen tension determinations can be made without repeat surgical exposure. In 40% of the control, left, noninfected tibial metaphyses the baseline oxygen tension with the animals breathing room air at sea level was suboptimal for leukocyte bacterial killing. This oxygen tension was depressed further in the infected right tibia. Medullary canal oxygen tension increased in response to hyperbaric oxygen exposure in both the infected and noninfected tibiae. Whereas the amount of the oxygen tension increase varied with the presence of infection and depth of dive, neither the time for oxygen tension to plateau nor the time required for return to baseline tension after completion of hyperbaric oxygen exposure varied with the presence of infection or depth of dive. After completion of hyperbaric oxygen exposure, the oxygen tension within the medullary canal returned to baseline within 15 min.

Animals↗
PubMed1986

Evaluation of aztreonam, cefoperazone, latamoxef and ceftazidime in the hamster colitis model.

Aztreonam, ceftazidime, cefoperazone and latamoxef (moxalactam) were evaluated in a hamster model for antibiotic-associated colitis. Aztreonam, a novel monocyclic beta-lactam antibiotic specifically directed against aerobic Gram-negative bacteria with limited activity against Gram-positive and anaerobic bacteria did not cause haemorrhagic caecitis and death in hamsters when administered intraperitioneally or orally. Quantitative caecal cultures showed no changes in the anaerobic caecal microflora of hamsters treated intraperitoneally with aztreonam and only a temporary decrease in anaerobic Gram-negative bacilli in hamsters treated orally. Clostridium difficile and its cytotoxin were not present in these animals. Parenteral administration of ceftazidime also did not affect the anaerobic caecal microflora or cause caecitis. However, when given orally, ceftazidime suppressed the anaerobic caecal microflora and a lethal C. difficile-induced caecitis developed. Latamoxef or cefoperazone given parenterally resulted in caecitis and death in five days with marked changes in the anaerobic caecal microflora and C. difficile present in the caecal contents of these animals. The lethal haemorrhagic caecitis observed in this study was indistinguishable from that seen in the clindamycin-induced colitis model in the hamster. The possible correlation of these findings to gastrointestinal disturbances as a complication of antibiotic therapy in humans is discussed.

Administration, Oral↗