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Biomedical subjects

M R James

Publications and source records attributed to M R James.

At least 73 records · Page 4Linked to original sources
PubMed1992

Cloning of a candidate gene for ataxia-telangiectasia group D.

Transfection, with a human cosmid clone library, of an ataxia-telangiectasia (AT) cell line (AT5BIVA) from complementation group D previously resulted in the isolation of a cell line (1B3) with partially restored resistance to ionizing radiation. We rescued the integrated cosmid sequences within 1B3 and obtained two cosmid clones that contained overlapping DNA from chromosomal region 11q23, previously shown to be the region containing the AT gene(s) from three complementation groups. Isolation of an apparently full-length 3.0-kb cDNA from a HeLa cell library demonstrated a previously unidentified gene (ATDC) within these cosmid clones. The transfected copy of the ATDC gene in 1B3 is truncated at the 3' end but is a complete transcription unit, because of the presence of SV40 termination sequences within the adjacent cosmid DNA. After further screening of cosmid clones from a chromosome 11 library, we identified contiguous DNA that contained the missing portion of the gene. Southern blot analysis indicated that the ATDC gene is present in a single copy in the human genome; however, RNA blot analysis revealed mRNA of several sizes (1.8, 2.6, 3.0, 4.7, and 5.7 kb) that varied among different cell lines. Because no large rearrangements were detected in AT5BIVA cells by Southern or RNA blot analysis, any alteration in the ATDC gene in this cell line would involve a point mutation or a small rearrangement. Transfection of the AT5BIVA cell line with one of the cosmids partially restored radioresistance. Analysis of 100 X-radiation hybrid cell lines containing various fragments from the chromosomal region 11q23 showed that the ATDC gene is closely linked to THY1. The ATDC gene therefore lies outside the linkage region predicted to contain the AT gene(s) for complementation groups A and C, indicating a separate locus for the AT complementation group D gene.

Ataxia Telangiectasia↗
PubMed1991

Pulse oximetry during apparent tonic-clonic seizures.

A pulse oximeter was used to monitor arterial oxygen saturations in 11 patients during apparent tonic-clonic seizures. 8 had a clinical diagnosis of genuine fits, 6 of whom showed striking falls in oxygen saturation during the seizures. 3 had a clinical diagnosis of pseudoseizures, none of whom had hypoxia during these episodes. Pulse oximetry during apparent tonic-clonic seizures may help to identify patients with low arterial oxygen tension who need immediate intervention.

Adolescent↗
PubMed1991

A study to compare the use of fusidic acid viscous eye drops and chloramphenicol eye ointment in an accident and emergency department.

A total of 300 patients attending an Accident and Emergency department with ophthalmic complaints needing topical antibiotic treatment were treated with fucithalmic or chloramphenicol (chloromycetin). Their compliance and the incidence of side effects were assessed. There was a 70.3% response rate to the questionnaire. Of the patients 51.3% using fucithalmic completed the 5 day course compared with 37.2% of those taking chloramphenicol. However, no patients were found to have suffered as a result of failing to complete the course. The incidence of side effects was similar in both groups.

Chloramphenicol↗
PubMed1991

Spinal fractures associated with ejection from jet aircraft: two case reports and a review.

Virtually any A&E department in the UK may see aircrew who have ejected. The case histories of two pilots who ejected and sustained spinal fractures are presented. Both had minimal back symptoms and walked from the site of landing. All ejectees should be considered to have a spinal fracture until proven otherwise radiographically. Other injuries that these patients may sustain during ejection are also discussed.

Acceleration↗
PubMed1991

X-ray reporting in accident and emergency departments--an area for improvements in efficiency.

All Accident and Emergency departments in England seeing more than 20,000 new patients each year were surveyed for the extent and mechanism of their radiological reporting service. A total of 190 of the 210 departments replied, 39% of these departments were not satisfied with the service available to them. A 'hot' reporting service was available in 2.6% of departments and 49% did not have films reported in under 48 h. The methods used to detect radiological abnormalities missed by the A&E doctor were investigated further. A total of 60% of departments had a traditional reporting system but 29% had introduced systems that required the A&E doctor to include his or her radiological diagnosis with the films prior to their being returned to the X-ray department for reporting. This allowed rapid audit of radiological assessment. Its introduction to all A&E departments is recommended.

Accidents↗
PubMed1990

Regulated expression of Epstein-Barr virus nuclear antigen 3-encoding gene carried on stable episomal vectors in human cells.

Recently a small number of Epstein-Barr viral (EBV) genes, characteristically expressed in latently infected, growth-transformed B-lymphocytes, have been cloned and several have been transiently expressed by DNA transfection. Here we demonstrate production of stable human cell lines containing episomal EBV vectors and expressing EBV nuclear antigen 3 from the adenovirus major late promoter or the mouse metallothionein promoter, which retains metal-regulation in the episomal state. This system has proved useful in an analysis of the role of these and other EBV genes implicated in immortalization and/or oncogenic transformation of human cells.

Antigens, Viral↗
PubMed1989

High recombination rate of an Epstein-Barr virus-simian virus 40 hybrid shuttle vector in human cells.

The stability of an Epstein-Barr virus (EBV)-simian virus 40 (SV40) hybrid shuttle vector, the p205-GTI plasmid, was analyzed in human cells during EBV- or SV40-type replication mode. When the p205-GTI plasmid was maintained as an episomal EBV vector in the human 293 cell line, no rearrangement was detected. To induce the SV40 replication mode, cells containing the episomal p205-GTI plasmid were either transfected with vectors carrying the T antigen gene or infected with SV40. Surprisingly, we observed both production and amplification of different classes of recombinant molecules. Particular types of modifications were found in most of the recombinants. The most striking rearrangement was a duplication of the promoter and enhancer regions of SV40 which was inserted in the thymidine kinase (TK) promoter. This recombination process involved a few bases of homology, and one of the recombination junctions implicated the GC boxes which constitute the essential components of the TK and SV40 early promoters. Our results suggest that a combination of a low level of base homology and a specific DNA sequence function (promoter and enhancer sites) leads to a very high level of recombinational activity during T-antigen-dependent plasmid replication.

Base Sequence↗
PubMed1989

Nurse practitioners in the accident and emergency department.

The theoretical management of walking wounded patients by experienced nurses was compared with that of middle-grade doctors. Out of 400 patients seen, 332 were assessed by the nurses. The theoretical management by the nurses of 298 of these patients was satisfactory. The nurses mismanaged 12 of the patients according to local practice. They also requested X-rays in 22 more patients than the doctors, none of whom returned with a fracture. Out of the patients suitable to be seen by the nurses 94% said that they would use a nurse practitioner system if introduced. The theoretical waiting-time saving for the patients during the trial was 11 min. The authors recommend that if nurse practitioners are introduced in this country there should be an adequate training programme along nationally debated guidelines.

Diagnostic Errors↗
PubMed1988

Relation between the human fibroblast strain 46BR and cell lines representative of Bloom's syndrome.

46BR is a human fibroblast strain derived from an immunodeficient young female of stunted growth. The diploid fibroblasts as well as a Simian Virus 40-transformed cell line are hypersensitive to killing by many DNA-damaging agents, exhibit a slightly increased level of spontaneous sister chromatid exchange, and show a defect in DNA ligation in vivo. 46BR is now shown to have abnormal DNA ligase I and is similar in this regard to cell lines derived from Bloom's syndrome patients. In a direct comparison, both 46BR and several Bloom's syndrome lines were found to be hypersensitive to the cytotoxic effect of simple alkylating agents, 46BR being more markedly sensitive. Bloom's syndrome lines do not exhibit the strong delay in joining of Okazaki fragments during DNA replication characteristic of 46BR. The cell line 46BR probably has a mutation in the gene encoding DNA ligase I different from those occurring in classical cases of Bloom's syndrome.

Benzamides↗
PubMed1988

Problems encountered when administering general anaesthetics in accident and emergency departments.

Junior anaesthetists in 75 English hospitals were surveyed for their views on whether administering general anaesthetics in A&E departments provoked more anxiety than in the main theatre, and if so what factors contributed to this. Of these anaesthetists, 71% were more apprehensive working in A&E departments than in main theatre; 91% felt that they were adequately experienced but despite this there was a marked decline in apprehension with increasing experience. Sixty eight per cent of the anaesthetists thought that their assistance was inadequate and only 28% had an Operating Department Assistant (ODA). Forty eight per cent said that the equipment was inadequate in either standard or maintenance and 40% said that some of the patients were unsuitable for day case anaesthesia. The authors recommend that anaesthetists performing general anaesthetics in A&E departments should be adequately experienced using equipment provided and maintained by the anaesthetic department and assisted by adequately trained nurses or ODAs.

Anesthesia, General↗
PubMed1987

Analysis of mutations occurring during replication of a SV40 shuttle vector in mammalian cells.

To analyse mutations that arise in mammalian cells we have used a SV40::plasmid shuttle vector containing a portion of the E. coli lacZ gene. We have found that following transfection into monkey Cos-7 cell mutations are not detected in the recovered plasmids at 24 h post transfection, but are found at 48 h post transfection, after the onset of DNA replication. Analysis of the mutant plasmids shows that in almost all cases the mutant phenotype is caused by a deletion or rearrangement of the lacZ gene in the shuttle vector.

Animals↗
PubMed1987

An immortalized xeroderma pigmentosum, group C, cell line which replicates SV40 shuttle vectors.

We have established and characterized an immortalized xeroderma pigmentosum (XP), group C, cell line. Transformation of the human fibroblasts was carried out with a recombinant plasmid, pLAS-wt, containing SV40 DNA encompassing the entire early region with a defective origin of DNA replication. The transformed XP cell line, XP4PA-SVwt, and the normal transformed fibroblasts AS3-SVwt, both express SV40 T antigen together with enhanced levels of the transformation-associated cellular protein, p53. XP4PA-SVwt retains the XP UV-repair defective phenotype as demonstrated by low levels of unscheduled DNA synthesis and by the reduced survival of irradiated SV40 virus. Analysis of cellular DNA shows a single major, stable, integration site of pLAS-wt in the XP4PA-SVwt cells. The T antigen in these cells supports efficiently the replication of SV40 based shuttle vectors and should prove suitable for the introduction, expression and selection of genes related to DNA repair and to the study of mutagenesis using defined molecular probes.

Cell Division↗
PubMed1987

SV40-based Escherichia coli shuttle vectors infectious for monkey cells.

We describe SV40-based Escherichia coli shuttle vectors which can be packaged as pseudovirions without excision of plasmid sequences and which can be rescued in bacteria. These vectors replicate and are transmitted as virus in monkey COS cells without requiring a helper virus. Extrachromosomal vector DNA isolated from infected cells can be rescued in E. coli, so that DNA alterations can be easily screened. Indeed, some of the constructions give rise to very stable plasmids with no detectable rearrangements after multiple lytic cycles in COS cells. The spontaneous mutation frequency measured in bacteria, on the lacO target, is smaller than those usually found with shuttle vectors. We also constructed an expression vector derived from one of our infectious viruses by inserting the chloramphenicol acetyl transferase gene, expressed from the SV40 early promoter, which is efficiently transduced to cells by infection. In this system, the shuttle virus combines the convenience of plasmid rescue and analysis in bacteria, with the advantages of infectious virus.

Acetyltransferases↗
PubMed1987

Inactivation of a transfected gene in human fibroblasts can occur by deletion, amplification, phenotypic switching, or methylation.

Plasmids containing the bacterial gpt gene under control of the simian virus 40 promoter were transfected into a simian virus 40-transformed human fibroblast line. Two transfectants, E2 and C10, which contain stably integrated single copies of the gpt gene, were isolated. These two lines produce Gpt- variants spontaneously with a frequency of about 10(-4). We carried out a detailed molecular analysis of the spectrum of alterations which gave rise to the Gpt- phenotype in these variants. DNA from 14 of 19 Gpt- derivatives of one of the cell lines (E2) contains deletions or rearrangements of gpt-containing sequences. In four of the remaining five lines, the Gpt- phenotype was correlated with reduced levels of expression rather than with changes in the gross structure of the gpt gene, and it was possible to reactivate the gpt gene. In one Gpt- line, gpt mRNA was present at normal levels, but no active enzyme was produced. Spontaneous Gpt- derivatives of the other cell line (C10) produced a completely different spectrum of alterations. Very few deletions were found, but several derivatives contained additional extrachromosomal gpt sequences, and, remarkably, in two other Gpt- lines, gpt-containing sequences were amplified more than 100-fold. The phenotypes of the majority of the Gpt- derivatives of C10 could be attributed to alterations in gene expression caused by methylation.

Cell Line↗