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Biomedical subjects

V Jackson

Publications and source records attributed to V Jackson.

At least 19 recordsLinked to original sources
PubMed2005

Food safety knowledge of consumers and the microbiological and temperature status of their refrigerators.

The objectives of this study were to examine domestic food safety knowledge levels of consumers, establish the levels and incidence of bacterial contamination and operational temperatures in domestic refrigerators, and identify areas in which consumer food safety education is necessary in Ireland. A food safety knowledge questionnaire applied to a representative sample of households (n = 1,020) throughout the island of Ireland found the gaps in consumer food safety knowledge. Analysis of swab samples (n = 900) recovered from the domestic refrigerators in these households showed average total viable counts of 7.1 log CFU/cm2 and average total coliform counts of 4.0 log CFU/cm2. Analysis of swab samples also detected the incidence of Staphylococcus aureus (41%), Escherichia coli (6%), Salmonella enterica (7%), Listeria monocytogenes (6%), and Yersinia enterocolitica (2%). Campylobacter jejuni and E. coli O157:H7 were not detected in domestic refrigerators. The temperature profiles of a subset of the sampled refrigerators (100) were monitored for 72 h, and 59% were found to operate, on average, at temperatures above the recommended 5 degrees C. Knowledge and temperature survey results varied considerably, but consumers who scored better in terms of basic food safety knowledge had reduced levels of bacterial contamination in their refrigerators and reported a reduced incidence of food-associated illnesses. This study confirms the effect of basic food hygiene knowledge on hygienic practice and identifies specific areasfor emphasis in the development and delivery of effective food safety risk communication messages to consumers.

Adolescent↗
PubMed2000

In vitro studies on the maintenance of transcription-induced stress by histones and polyamines.

Several factors were evaluated to determine their role in facilitating the presence of transcription-induced stresses in a circular DNA. Transcription was done with T7 RNA polymerase in the presence of E. coli topoisomerase I and closed circular DNA. Positive stress was observed in hypotonic conditions or when one of the polyamines, spermidine or spermine, were present. Polycations such as polylysine, polyarginine, histone H1, histones H2A-H2B, and protamine were observed to induce minimal positive stress. It is known that polyamines influence DNA structure by causing both self-association and sequence-specific structural alterations (polyamine-induced localized bending). Experimental evidence indicates that the likely cause of the positive stress is the induced bending. In order to evaluate protein-mediated bending, transcription was done on nucleosomes. A minimum of three nucleosomes on a DNA of 6055 bp was sufficient to generate very high levels of positive stress. Histones H3-H4 in the absence of H2A-H2B were responsible for this effect. Since these histones by themselves are able to maintain negative coils on DNA, it is concluded that protein-mediated bending is yet another mechanism for placing rotational restriction on DNA. The bending of DNA by either polyamines or histones is an effective mechanism for promoting transcription-induced stresses at physiological ionic strength.

Animals↗
PubMed1999

Formaldehyde cross-linking for studying nucleosomal dynamics.

Methods are described for the utilization of formaldehyde as a reversible cross-linking agent for the characterization of protein-protein and protein-DNA interactions. The methods include a description of procedures to: (1) isolate and characterize transcriptionally active chromatin from cells cross-linked with formaldehyde; (2) study histone mobility during replication and transcription by the characterization of the formaldehyde-cross-linked histone octamer that is isolated from cells labeled with density-labeled amino acids; and (3) cross-link the in vitro reconstituted histone-DNA complex in order to maintain its structural state during subsequent characterization. Included in these methods are procedures for a second dimensional analysis of protein-protein cross-links in which the monomer components are electrophoretically resolved in the second dimension. The methods also include procedures to selectively reverse protein-DNA cross-links while maintaining the protein-protein cross-links. Potential artifacts are also discussed; i.e., data are presented which indicate that the helical pitch of DNA can be altered if the ionic strength is not properly controlled. The stability of the cross-linked nucleosome in the presence of altered pH or salt/urea concentrations is described in order to indicate that there are limitations to procedures that can be used for the subsequent characterization of the cross-linked complex.

Cross-Linking Reagents↗
PubMed1998

A midwife to all.

Explore the source record for details and available documents.

Delivery Rooms↗
PubMed1997

Measurement of the frequency of histone displacement during the in vitro transcription of nucleosomes: RNA is a competitor for these histones.

Transcription through tandemly arranged nucleosomes was studied to determine the frequency at which the nucleosomes would disrupt and cause displacement of the associated histones to a competitor DNA. In order to more effectively preserve topological effects, the template that was used in the in vitro transcription system was a large covalently, closed circular plasmid (8.9 kb). The plasmid contained two promoters for T7 RNA polymerase, each separated by 4.4 kb, and transcription was done in the presence of topoisomerase I at physiological ionic strength. Nucleosome disruption was observed at an approximate frequency of 1 in 4 nucleosomes such that after several rounds of transcription on the plasmid 80% of the nucleosomes were disrupted. Unexpectedly, all four histones were found associated with the RNA rather than the competitor DNA. The histones bound the competitor DNA only after removal of the RNA by RNase A treatment. By analyzing the topological state of the competitor DNA, it was observed that the majority of the histones that were displaced from the RNA were able to re-form nucleosomes. Additional experiments were done to determine the reasons for the preferential binding of histones to the newly synthesized RNA. It was found that the large molecular weight RNA binds histones with an approximate 100-fold greater affinity relative to DNA when at physiological ionic strength. Within the cell, this high-affinity binding would be expected to require cellular mechanisms to regulate the interaction of RNA with histones. The relatively high frequency of displacement of all four histones during transcription is higher than what is observed in vivo and suggests that additional factors are needed to regulate this displacement. These observations are discussed and compared with previous studies that have examined the process of transcription through nucleosomes.

Animals↗
PubMed1997

Effects of spermine and its cytotoxic analogs on nucleosome formation on topologically stressed DNA in vitro.

We investigated the effects of the polyamine spermine and two of its cytotoxic analogs 1,11-bis(ethylamino)-4,8-diazaundecane (BE-3-3-3) and 1,19-bis(ethylamino)-5,10,15-tirazanonadecane (BE-4-4-4-4) on the formation of nucleosomes on negatively and positively supercoiled DNA in vitro. Histones H2A, H2B, H3 and H4 were reconstituted onto DNA to form nucleosomes and the polyamines were added either before or after histone addition. The structural state of the nucleosome was monitored by analyzing the DNA topoisomers that were present after topoisomerase I treatment. Although polyamines induced DNA aggregation to various degrees. high concentrations of topoisomerase I were able to relax the aggregated DNA and the helical pitch was found to be unaltered in the aggregates. When histones were associated with negatively coiled DNA, the polyamine-induced aggregation did not alter nucleosome structure. The induced aggregate did inhibit nucleosomal transitions when examined on positively coiled DNA. BE-4-4-4-4 was most effective and BE-3-3-3 least effective. These analogs were also extremely effective in inhibiting histone deposition onto DNA. A potential mechanism for the action of these analogs is both to inhibit histone deposition during DNA replication and also disrupt nucleosomal dynamics due to aberrant chromatin condensation. These results also suggest that BE-4-4-4-4 and BE-3-3-3 may produce their cytotoxic effect through slightly different mechanisms.

Animals↗
PubMed1996

Percutaneous core biopsy of the breast: effect of operator experience and number of samples on diagnostic accuracy.

OBJECTIVE: The purpose of our study was to assess the degree of operator experience and the number of core biopsy samples required to achieve an accurate histologic diagnosis for each of five common mammographically defined lesions, using percutaneous core breast biopsy performed on a dedicated prone biopsy table. SUBJECTS AND METHODS: A prospective multisite study was performed that involved nine institutions (academic and private) with experienced breast radiologists and the use of dedicated prone biopsy table units with digital assistance and standardized protocol. Asymptomatic women evaluated during a 2-year study period were assigned a mammographic diagnosis reported in a manner prescribed by the American College of Radiology Breast Imaging Reporting and Data System lexicon. Mammographic lesions evaluated included masses, masses with calcifications, clustered calcifications, focal asymmetries, and architectural distortions. Where histologic diagnosis was indicated, core biopsy was performed with five individual samples obtained and sequentially analyzed. Two hundred thirty patients had immediate excisional biopsy, the results of which provided the basis for a statistical analysis to compare the accuracy of each sequential core biopsy sample with surgical results. Statistical analysis was also done to ascertain the accuracy of core biopsy diagnosis as a function of operator experience. RESULTS: Trends toward increasing accuracy were observed by increasing the number of core biopsies for each of five types of mammographically defined lesions, especially for clustered calcifications. Statistically significant increased accuracy was observed when the number of biopsies was increased beyond one (p = .003). Trends toward increased accuracy with more experience were observed for all lesions, especially for calcifications. Of the 230 lesions studied with immediate surgical validation, more than 80% of all lesions except clustered calcifications (75%) were diagnosed on the basis of two core biopsies; accuracy after five biopsies was 98% for masses, 91% for calcifications, 100% for masses with calcification, 100% for focal asymmetries, and 86% for architectural distortions. CONCLUSIONS: Accuracy of diagnosis based on the results of percutaneous core breast biopsy improved with an increase in the number of core biopsy samples obtained for any given lesion seen on mammograms and with increased experience in performing the procedure. Five samples yielded an overall diagnostic accuracy of 97%. Familiarity with expected accuracy from this procedure for different mammographic lesions and following increased experience may assist physicians in planning patient management.

Analysis of Variance↗
PubMed1996

Anti-Holley detected in a primary immune response.

Anti-Holley (Hy) has been reported as an IgG antibody occurring in previously transfused or multiparous black patients. In this case anti- Hy was identified in a 16-year-old black, primigravida female admitted at 32 weeks gestation because of premature rupture of the membranes. On admission, her blood type was determined to be A2B, D-positive and an antibody screen was negative. A second antibody screen, performed 4 days later, was positive in all three cells. Anti-Hy was subsequently identified. The antibody was reactive at room temperature, 37 degrees C, and in the antiglobulin phase. IgG and IgM components of anti-Hy were demonstrated in the maternal serum, documenting a primary immune response. This resulted in serologic findings not previously described for anti-Hy. A direct antiglobulin test on the newborn red cells was negative and there was no clinical evidence of hemolytic disease of the newborn (HDN). A monocyte monolayer assay performed with maternal serum yielded negative results. Recent scientific information has resulted in the placement of Hy in the Dombrock blood group system. Alloantibodies to Dombrock system antigens have not been associated with severe HDN.

Journal Article↗
PubMed1995

Preferential binding of histones H3 and H4 to highly positively coiled DNA.

The interaction of histones H3 and H4 with highly positively coiled DNA has been studied. To carry out this study, it was necessary to develop a protocol for the preparation of large quantities of highly positively coiled DNA. Such preparations were obtained by maximizing transcription-induced stress in cells containing PBR322 and then applying a short-term treatment with novobiocin, which inhibits gyrase. Fractionation on CsCl-EtBr gradients gave PBR322 plasmids with topological states ranging from +0.15 to -0.043 superhelical density. In competition experiments between negatively and positively coiled DNA, histones H3,H4 preferentially bound the positively coiled DNA when the superhelical density was greater than +0.10. This preference was shown on the basis of sedimentation rate on sucrose gradients, selective aggregation by H3,H4 binding, and cross-linking experiments in which the histone-DNA content was characterized on CsCl density gradients. An analysis of the DNA helical pitch in 1.1 M NaCl, a condition in which moderately positively coiled DNA preferentially binds H3,H4 indicated that the preferential binding may be due to a decrease in helical pitch that approximates 0.06 bp/turn. Consistent with this observation is the ability of histones H3,H4 to transiently hold this altered pitch in the presence of topoisomerase I. A possible explanation for this preference is based on the known observation that histones overwind the DNA helix when in a nucleosome. These data provide an explanation for the in vivo observation that histones H3,H4 are rarely displaced during the transcription process. These observations are discussed with regard to mechanisms for transcription through nucleosomes.

Animals↗
PubMed1995

Ty1-copia group retrotransposon sequences in amphibia and reptilia.

We have isolated sequences belonging to Ty1-copia group retrotransposons from the genomes of an amphibian (Pyxicephalus adspersa) and three reptiles (Conolophus subscristatus, Amblyrynchus cristatus and Pytas mucosus). Two different sequences were found in the amphibian (Tpa1 and Tpa2). Each is present in several copies per genome and absent from the genomes of two other amphibian species. The C. subcristatus sequence Tcs1 is present in multiple copies in both its host genome (Galapagos land iguana) and the genome of the related Galapagos marine iguana (A. cristatus). There is little or no polymorphism in Tcs1 insertions between different individual animals, suggesting that this sequence is not transposing rapidly in either iguana genome. The P. mucosus sequence Tpm1 shows a discontinuous distribution in snake species, suggesting that it has either been lost from many lineages during vertical germline transmission or has been transferred horizontally in some snake species. Phylogenetic comparisons of all these sequences with each other and with other members of this retrotransposon group from other animals and plants show that sequences within a particular vertebrate species are most closely related to each other, consistent with a vertical transmission model for their evolution.

Amino Acid Sequence↗
PubMed1994

The rapid transfer and selective association of histones H2A and H2B onto negatively coiled DNA at physiological ionic strength.

The interaction of histones H2A and H2B with closed circular DNA maintained in either a positive or negatively coiled state has been studied. The methods involved separation of complexes by sucrose gradients and by agarose gel electrophoresis. Histones H2A and H2B have a 10-fold preference for DNA that contains performed negative coils as compared to DNA that is positively coiled or topologically unrestrained. It was also observed that histones H2A and H2B, when initially reconstituted onto positively coiled DNA, transfer very efficiently to negatively coiled DNA at 100 mM NaCl. Transfer is virtually complete within 5 s at 4 degrees C. By utilizing a cross-linking agent (disuccinimidyl suberate), it was found that histones H2A and H2B could be cross-linked into a tetrameric structure when on negatively coiled DNA but not on positively coiled DNA. It is concluded that protein-protein interactions facilitate the transfer. The importance of these protein-protein interactions is further demonstrated in experiments in which H3,H4 were included, and, in this instance, the strong interaction of H2A,H2B with H3,H4 negates the preference of H2A,H2B for negatively coiled DNA. These observations are discussed with regard to the mobility of histones during the transcription process during which transcription-induced stresses may be produced.

Animals↗
PubMed1994

Dynamics of the interactions of histones H2A,H2B and H3,H4 with torsionally stressed DNA.

The interactions of histones H2A,H2B and H3,H4 with closed circular DNA maintained in either a positively or negatively coiled state have been studied. The interactions were assayed by measuring the rate at which negative stress was stored in the DNA by the histones and by the salt concentration sufficient to cause dissociation on sucrose gradients. Additional experiments were performed in which DNAs of substantially different molecular weights and opposite topological states were mixed with the histones in order to study histone mobility under varied conditions. This mobility was characterized by separating the complexes on sucrose gradients and by analyzing the DNA's topological state after topoisomerase I treatment. Histones H3,H4 were found to differ substantially from histones H2A,H2B with regard to the DNA topology with which they prefer to interact. The results are consistent with a model in which transcription-induced positive stress in advance of the RNA polymerase unfolds the nucleosome to facilitate the release of H2A,H2B. The data are also consistent with a model in which histones H3,H4 remain associated with the DNA during polymerase passage and serve as a nucleation site for the reassociation of H2A,H2B. The rapid production of transcription-induced negative stress in the wake of a polymerase would have substantial importance in facilitating the reassociation of histones H2A,H2B.

Biomechanical Phenomena↗
PubMed1993

Influence of positive stress on nucleosome assembly.

Nucleosome assembly on closed circular DNA has been studied using reconstitution protocols which involve salt gradient dialysis in the presence or absence of urea. The closed circular DNA contained either +3 or -14 supercoils. The nucleosomal content of the DNA was assayed by measuring the superhelical content in the presence of topoisomerase I as a function of time and temperature and by measuring the DNA fragment size after DNase I digestion. Positively coiled DNA appears to contain an altered "open" nucleosome structure as compared to the negatively coiled DNA. Formaldehyde fixation of the reconstituted complexes was also done in an attempt to freeze the nucleosomal state prior to topoisomerase I or DNase I addition. The data from these additional experiments are consistent with a conclusion that an altered nucleosomal structure exists on positively supercoiled DNA. Control experiments are included to verify the efficiency of formaldehyde fixation in preserving the structure in the reconstituted complexes. The physiological relevance of positive stress and its effect on nucleosome structure are discussed.

Cold Temperature↗
PubMed1993

Vaccination with irradiated tumor cells engineered to secrete murine granulocyte-macrophage colony-stimulating factor stimulates potent, specific, and long-lasting anti-tumor immunity.

To compare the ability of different cytokines and other molecules to enhance the immunogenicity of tumor cells, we generated 10 retroviruses encoding potential immunomodulators and studied the vaccination properties of murine tumor cells transduced by the viruses. Using a B16 melanoma model, in which irradiated tumor cells alone do not stimulate significant anti-tumor immunity, we found that irradiated tumor cells expressing murine granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulated potent, long-lasting, and specific anti-tumor immunity, requiring both CD4+ and CD8+ cells. Irradiated cells expressing interleukins 4 and 6 also stimulated detectable, but weaker, activity. In contrast to the B16 system, we found that in a number of other tumor models, the levels of anti-tumor immunity reported previously in cytokine gene transfer studies involving live, transduced cells could be achieved through the use of irradiated cells alone. Nevertheless, manipulation of the vaccine or challenge doses made it possible to demonstrate the activity of murine GM-CSF in those systems as well. Overall, our results have important implications for the clinical use of genetically modified tumor cells as therapeutic cancer vaccines.

Animals↗
PubMed1990

Studies on rates of nucleosome formation with DNA under stress.

Rates of nucleosome formation on DNA with various superhelical topologies were determined using an in vitro deposition system. The core histones, H3, H2B, H2A, and H4 were reconstituted with polyglutamic acid (a deposition vehicle) and added to closed-circular DNAs. Aliquots were taken during the incubation, added to excess topoisomerase I, and incubated for 20 s to determine the number of negative coils constrained into nucleosome structure by the histones. When negatively supercoiled DNA was used, nucleosomes were formed virtually instantaneously in a temperature-independent process with all the negative coils being trapped into nucleosome structure. When relaxed DNA (no negative coils) was used, nucleosome formation was severely retarded. If topoisomerase I was maintained in the total incubation of histones with the relaxed DNA, nucleosome formation occurred with a measurable rate in a temperature-dependent process (activation energy of 17 kcal/mol). Similar observations were made when the method of reconstitution of histones with DNA involved salt gradient dialysis. The data indicates that the lack of preformed negative coils is a rate-limiting step in nucleosome formation and that when positive coils are also maintained within the circular DNA, these positive coils prevent nucleosome formation. The effect of DNA topology on nucleosome formation is discussed with regard to its physiological relevance in replication and transcription processes.

Animals↗
PubMed1990

In vitro evidence that transcription-induced stress causes nucleosome dissolution and regeneration.

During transcription, positive and negative superhelical stresses are generated on a DNA template which could potentially affect nucleosomal structure. When transcription was performed on a closed circular plasmid containing nucleosomes, using T7 RNA polymerase and topoisomerase I, nucleosomal structure was lost from the DNA. Nucleosome content was assayed by analyzing both the topological state of the DNA and the nuclease-resistant fragments produced by micrococcal nuclease and DNase I treatment. This nucleosome dissolution required positive superhelical stress as evidenced by the requirement that the extended RNA transcript remain associated with the polymerase during the transcription process. Rates of transcription were found to be independent of whether the nucleosomes dissolved. When transcription was performed in the absence of topoisomerase I, nucleosome reformation occurred very rapidly. This observation suggests that negative superhelical stress, induced in the wake of polymerase action, facilitates nucleosome reformation.

Animals↗
PubMed1990

In vivo studies on the dynamics of histone-DNA interaction: evidence for nucleosome dissolution during replication and transcription and a low level of dissolution independent of both.

Histones have been both radiolabeled and density-labeled with amino acids in vivo to determine the dynamics of histone-DNA and histone-histone interactions at the replication fork and on active genes. Proteins were uniformly labeled and subsequently chased for three cell generations. During the chase period, H3,H4 tetramers dissociated from the H2A,H2B dimers to re-form nucleosomes with the corresponding nondense histones synthesized during the chase period. These data suggest that the prereplicative nucleosomes are dissolved during advancement of the replication fork with release of associated histones in the form of the H3,H4 tetramers and H2A,H2B dimers. Experiments that involve density labeling of cells in the presence of actinomycin D indicate that the dynamic exchange of H2A,H2B that has been previously described [Jackson, V. (1987) Biochemistry 26, 2315-2324] is partially dependent on RNA polymerase movement. These results provide indirect evidence that nucleosome dissolution occurs during transcription. When deposition during replication and transcription is inhibited by simultaneous treatment of cells with cytosine arabinoside and actinomycin D, the majority of the newly synthesized histones are unable to deposit into nucleosome structure. The low level of deposition that is observed has characteristics similar to the deposition of uH2A and uH2B, and it is proposed that conjugation of H2A and H2B by ubiquitin occurs when these proteins are in a free pool within the nucleus. The new H3,H4 tetramers and new H2A,H2B dimers when prevented from depositing are not stable. New and old H3 and H4 intermix to form hybrid tetramers, and a similar intermixing is observed for the H2A,H2B dimers. A model is presented to describe the dynamics of histone-DNA interactions during replication and transcription.

Animals↗