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Biomedical subjects

R James

Publications and source records attributed to R James.

At least 199 records · Page 11Linked to original sources
PubMed1991

Cloning and characterization of the ColE7 plasmid.

The 6.2 kb ColE7-K317 plasmid was mapped and the DNA fragments of the colicin E7 operon subcloned into pUC18 and pUC19. The size of the functional colicin E7 operon deduced by subcloning was 2.3 kb. The colicin E7 gene product was purified by carboxymethylcellulose chromatography. Both colicin E7 and E9 were demonstrated to exhibit a non-specific DNAase-type activity by in vitro biological assay. The molecular mass of colicin E7 was 61 kDa, as determined by SDS-PAGE. From DNA sequence data, the estimated sizes of the E7 immunity protein and the E7 lysis protein were 9926 Da and 4847 Da, respectively. Comparison of restriction maps and DNA sequence data suggests that ColE7 and ColE2 are more closely related than other E colicin plasmids.

Amino Acid Sequence↗
PubMed1991

Identification of a gene, closely linked to dnaK, which is required for high-temperature growth of Escherichia coli.

We have constructed four deletion derivatives of the cloned dnaK gene. Plasmid pDD1, in which the last 10 amino acids of the DnaK protein have been replaced by three different amino acids derived from the pBR322 vector, was as effective as plasmid pKP31, from which it was derived, in restoring the ability of a dnaK null mutant, Escherichia coli BB1553, to plate lambda phage and to grow at high temperatures. The other three mutations, involving much larger deletions of the dnaK gene, did not restore the ability to plate lambda phage or the ability to grow at high temperatures. Plasmid pKUC2, which contains the whole dnaK gene and its promoters, was capable of restoring the ability of E. coli BB1553 to plate lambda phage but, surprisingly, it did not restore the ability to grow at high temperatures, even though it was shown that the DnaK protein was efficiently expressed in these cultures. By transposon mutagenesis and sub-cloning, we have shown the presence of a second gene in plasmid pKP31 which is required for high-temperature growth of E. coli BB1553. This gene, which we call htg A, is presumably also defective in the dnaK null mutant E. coli BB1553. We have also demonstrated that the inability of E. coli K756 to grow above 43.5 degrees C is complemented by sub-clones which contain the htg A gene, but not by plasmid pKUC2.

Amino Acid Sequence↗
PubMed1991

Investigation of the specificity of the interaction between colicin E9 and its immunity protein by site-directed mutagenesis.

Comparison of the amino acid sequences of the C-terminal domain of three DNAase type E colicins has identified six candidate specificity determinants for the interaction of these E colicins with their homologous immunity proteins. We have changed these candidate specificity determinants of colicin E9, using site-directed mutagenesis, to the corresponding amino-acids of colicin E8. A 'mutant' colicin E9, in which four of the six candidate specificity determinants have been changed, demonstrated colicin activity against Escherichia coli indicator strains which carried either the E8imm or the E9imm genes, indicative of a 'novel' E. colicin. After changing all six of the candidate specificity determinants, the resulting colicin E9 'mutant' exhibited a phenotype very similar to that of colicin E8.

Amino Acid Sequence↗
PubMed1990

Detection of colonic growth factors using a human colonic carcinoma cell line (LIM1215).

Although the colonic mucosa is one of the most rapidly proliferating epithelial tissues in the body, little is known about the factors that direct this proliferation. In this report we have studied the parameters of both a mitogenic and a clonogenic assay for detecting potential colonic growth factors (CGF). Using a colon carcinoma cell line (LIM1215), which has retained a number of the properties of normal colonic mucosa, we have assayed a range of mitogenic factors for CGF activity. 3H-thymidine incorporation by the LIM1215 cell line was stimulated by low concentrations of epidermal growth factor and basic fibroblast growth factor and, to a lesser extent, by higher concentrations of interleukin-1 and insulin-like growth factor 1. The cells did not respond to a range of other mitogens and lymphokines. Optimal clonogenic response in a soft-agar assay was obtained using a primary pituitary extract.

Animals↗
PubMed1990

Hydrodynamic deposition: a novel method of cell immobilization.

A novel method of cell immobilization is described. The cell support consists of ceramic microspheres of approximately 50-75 microns diameter. The spheres are hollow, having a wall thickness of 10-15 microns and one entrance (ca. 20 microns diameter). The walls are porous with a mean pore size of approximately 90 nm. When a cell suspension (of S. cerevisiae) is passed through a column of such particles, cells are immobilized. Conditions are devised such that the overwhelming majority of cells are held in the central cavity of the support and not between the particles. Provided turbulence is avoided, the distribution of cells along the column length in the steady state is rather homogeneous. The facts that (a) essentially all particles, regardless of orientation, entrap cells, and (b) nonporous particles also entrap cells with high efficiency, indicate that filtration effects are irrelevant and that heretofore unrecognized hydrodynamic forces are alone responsible for the cell immobilization. Cells can be immobilized to high biomass densities, while the hydrodynamic properties of columns containing such immobilized cells are excellent. We describe an on-line electronic method for the real-time measurement of immobilized cellular biomass. Cell growth (so recorded) and metabolism continue to occur in such particles at high rates. Using the glycolytic production of ethanol by S. cerevisiae as a model reaction, volumetric productivities as great as any published are obtained. Thus the "lobster-pot effect" or "hydrodynamic deposition" represents a novel, promising, and generally applicable method of cell immobilization.

Biotechnology↗
PubMed1990

Case study: disease prevention--new roles for nurses in rural Australia.

Nurses are taking a leading role in preventing disease on the North Coast of New South Wales. In addition to heart health programs, nurses are also involved in Early Childhood Clinics, immunization campaigns, pap smear and breast self-exam campaigns, nutrition education and walking-for-pleasure groups. It would be safe to say that without nurses making new roles for themselves, there would be very few disease prevention campaigns on the North Coast.

Community Health Nursing↗
PubMed1990

Transfers of frozen-thawed human embryos in cycles stimulated by HMG.

A total of 130 transfers of frozen-thawed (F-T) human embryos was carried out after moderate ovarian stimulation with human menopausal gonadotrophin (HMG). Embryos were replaced 3 days after the spontaneous luteinizing hormone (LH) surge or 4 days if ovulation was induced by human chorionic gonadotrophin (HCG). Embryos were thawed a few hours prior to transfer. One-hundred-and-twenty-three transfers were effective and 23 pregnancies were achieved. The rate of ongoing pregnancies per transfer was 17.9% (22/123). The survival rate of embryos originating from cycles stimulated by a combination of an LHRH analogue and HMG in a long protocol (LA-HMG protocol) was significantly lower when compared with the rate of embryos retrieved from clomiphene citrate-HMG (CC-HMG protocol) stimulated cycles (52 versus 67%, P less than 0.05). When fresh embryos originated from cycles stimulated with an LHRH analogue and HMG in a short protocol (SA-HMG protocol), the survival rate was not affected (59 versus 67%, NS). Although the difference was not significant, the ongoing pregnancy rate per transfer according to the three protocols from which the embryos originated seemed to be better with the SA-HMG protocol: 16% with the CC-HMG protocol, 14.5% with the LA-HMG protocol versus 27.6% with the SA-HMG protocol. The success rate was independent of the number of F-T transferred embryos if at least one embryo with 100% intact blastomeres was replaced.

Cryopreservation↗
PubMed1990

HLA DR, DP, DQ induction in human islet beta cells by the cytokine combination IFN-gamma + TNF-alpha.

Human islet beta cells do not express HLA Class II normally, yet, in the diabetic pancreas, beta cells are selectively positive for Class II and this may facilitate their recognition by T cells. It has been demonstrated that human beta cells can be induced to express Class II when cultured with IFN-gamma + TNF-alpha or IFN-gamma + TNF-beta. To assess whether or not they can be induced to express the products of the Class II subregions, DR, DP and DQ, human islet cultures from 10 pancreas were supplemented with the combination of IFN-gamma + TNF-alpha using MoAbs specific for DR, DP and DQ products, and antibodies to insulin and glucagon. The combination IFN-gamma + TNF-alpha (100-1000 U/ml each) was able to induce the expression of the three subregions in both beta and alpha cells. The induction of subregion expression followed the hierarchy DR greater than DQ greater than or equal to DP. The capability of beta cells to express all three Class II subregions supports the possibility that these cells can present their self antigens to T cells.

Gene Expression Regulation↗
PubMed1989

Thigh length versus knee length stockings in the prevention of deep vein thrombosis.

Above-knee graduated compression stockings are effective in preventing postoperative deep vein thrombosis, but are more expensive and less acceptable than below-knee stockings. One hundred and fourteen patients undergoing major abdominal surgery were randomly allocated to wear above-knee or below-knee graduated compression stockings. Deep vein thromboses were diagnosed by isotope uptake in three of 56 patients (5.4 per cent) in the above-knee group and one of 58 patients (1.7 per cent) in the below-knee group. These differences are not statistically significant. Results suggest that below-knee stockings are as effective as above-knee in the prevention of postoperative deep vein thrombosis.

Abdomen↗
PubMed1989

A clinical trial of a self-care approach to the management of chronic headache in general practice.

This paper reports a trial which assessed the clinical effectiveness of adding a behavioural self-management programme to the existing management of chronic headache by general practitioners (GPs). Eighty-seven chronic headache sufferers, referred to the study by 35 GPs, were randomly allocated to either a self-care group or a GP-control group. Headaches, drug usage, visits to health-care providers and time off work were self-monitored daily by all subjects for 4 weeks prior to intervention, for 4 weeks during intervention, and for 4 weeks immediately after intervention. Additionally, self-monitoring was carried out for two further 4-week periods, one at 6 months and one at 12 months post intervention. Headache records showed that the self-care program significantly enhanced GP management. This effect was well maintained. However, drug usage, visits to health-care providers and time off work did not differ significantly between the treatment and control groups. 'No-show' rates, defined as those referred by a GP but who did not attend, were high--largely due to time requirements of the self-care program. However, drop-out rates, defined as those who left the self-care groups were low. It was concluded that this behavioural self-management program was a clinically effective adjunct to general practice management of headache but its use is likely to be limited due to problems of patient enrollment.

Adult↗
PubMed1989

T-cell nuclei contain a protein that binds upstream of the murine granulocyte-macrophage colony-stimulating factor gene.

Stimulation of a murine T-cell line (O18A) by Con A has been shown to cause a large increase in the cytoplasmic granulocyte-macrophage colony-stimulating factor (GM-CSF) mRNA level. Using run-on transcription experiments in isolated nuclei, we have shown that some of this response is from enhanced transcription of the GM-CSF gene. Changes in the transcription rate of this gene can be seen as early as 30 min after adding the Con A. With a DNA fragment mobility-shift assay on nuclear extracts from these cells we detected a protein that binds upstream of the murine GM-CSF gene. Partial purification and concentration of this protein by DEAE-Sephacel and phosphocellulose chromatography enabled us to examine its interaction with DNA in more detail. Competition and methylation interference experiments have shown that the protein binds to the sequence 3'-TCCATCAAGGGG-5' (positions -90 to -82). This sequence is contained within a region found to be involved in regulating inducible GM-CSF transcription in a human T-cell line [Miyatake, S., Seiki, M., Yoshida, M. & Arai, K. (1988) Mol. Cell. Biol. 8, 5581-5587].

Animals↗
PubMed1989

An evolutionary relationship between the ColE5-099 and the ColE9-J plasmids revealed by nucleotide sequencing.

The nucleotide sequence of a 1124 bp fragment of the ColE5-099 plasmid which encodes colicin E5 immunity, a lys gene involved in colicin release from the host cell, and the 3' end of the colicin E5 structural gene has been determined. Open reading frames corresponding to the three genes have been located by analogy with similar sequences from other E colicin plasmids. The location of these open reading frames corresponds with the position of the genes as determined by subcloning and transposon mutagenesis. The amino acid sequence of the carboxy-terminal 107 amino acid residues of the colicin E5 gene shows no homology with any other E colicin, suggesting a different mode of action in killing sensitive cells. A comparison of the nucleotide sequence of this region of the ColE5-099 plasmid with that of the equivalent region of the ColE9-J plasmid suggests a close evolutionary relationship between these two plasmids.

Amino Acid Sequence↗