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Biomedical subjects

R Davies

Publications and source records attributed to R Davies.

At least 163 records · Page 9Linked to original sources
PubMed1994

The primary and secondary metabolism of aflatoxin B1 by rat hepatocytes cultured on matrigel.

Rat hepatocytes on culturing rapidly lose the ability to metabolize many xenobiotics including the mycotoxin aflatoxin B1 (AFB1). Rat hepatocytes cultured for 5 days on plastic retain < 10% of the initial cytochrome P450 content compared with 34% in cells grown on matrigel alone and 74% in cells cultured on matrigel plus phenobarbital (PB). Although microsomal activation of AFB1 was preserved in matrigel cultured hepatocytes and increased by PB in all the culturing conditions examined, the AFB1 macromolecular binding capacity was reduced to < 10% of that present in the initial cultures. This lack of correlation between cytochrome P450 levels, microsomal activation, and AFB1 binding involved cytosolic glutathione-S-transferase activity (GST). This increased in matrigel cultured cells and was further enhanced by PB. Western blotting of the cytosols showed that the expression of the alpha class GST subunit Yc2 correlated with AFB1-GSH conjugate formation. By contrast, expression of the alpha class GST Ya-type subunit showed no similar close correlation. The expression of pi class GST Yf subunit, which is associated with dedifferentiation in hepatocytes, was suppressed by culturing on matrigel. The results obtained using hepatocytes cultured on matrigel indicate that despite limitations, this comprises a potentially useful model for the in vivo situation.

Aflatoxin B1↗
PubMed1994

Acid-base changes following urinary tract reconstruction for continent diversion and orthotopic bladder replacement.

A prospective determination of serum electrolytes, arterial blood gases, urinalysis and urine cultures was done in 31 patients who underwent a successful continent urinary reservoir or orthotopic bladder replacement. The patients who underwent reconstruction with a long detubularized intestinal segment (group 1-50 cm. long) demonstrated the greatest tendency for metabolic hyperchloremic acidosis (35.2%). In group 2 (patients with an orthotopic bladder replacement) only 1 individual (16.7%) had hyperchloremia, which proved to be the sole metabolic derangement encountered. In group 3 (individuals with a continent gastroileac reservoir) 2 patients (25%) had a slight tendency for compensated and asymptomatic alkalosis. Urinalyses and urine cultures in groups 1 and 2 demonstrated a trend toward urine alkalinity (52.1%) and asymptomatic bacteriuria (74%), respectively. On the contrary, among the patients undergoing a gastroileac reservoir (group 3), mild urinary acidity (pH between 5 and 6) was demonstrated in 4 (50%), while asymptomatic bacteriuria was present in 3 (37.5%). In this group symptomatic urinary acidity and/or ulceration of the ileal component has not occurred to date. Metabolic hyperchloremic acidosis predominates when longer colonic segments are used for reservoir construction. This abnormality is magnified in patients in whom an accessory small bowel was resected. The majority of the gastroileac reservoir patients showed electrolytic neutrality. With our surgical technique, the gastric secretory properties predominate over those of the ileum. The differences in homeostatic findings with the use of these varieties of bowel segments suggest that we could modify the final electrolytic environment by using different combinations of bowel and bowel length.

Acid-Base Imbalance↗
PubMed1994

Analysis of cytokine gene expression in Graves' disease and multinodular goiter.

Cytokines have a central role in the generation of an autoimmune response and can directly affect the target organ. In Graves' disease, both the infiltrating mononuclear cells and the thyroid follicular cells produce certain cytokines, but the relative contribution of each is unclear, and there are conflicting data on the exact profile of cytokines expressed within the thyroid. To clarify these issues, we used the method of reverse transcription-polymerase chain reaction amplification to analyze cytokine gene expression by intrathyroidal lymphocytes (ITL) and purified thyroid follicular cells (TFC) from six patients with Graves' disease. All ITL samples were positive for interleukin-1 alpha (IL-1 alpha), IL-1 beta, IL-6, IL-8, IL-10, and tumor necrosis factor-alpha (TNF alpha) messenger ribonucleic acids (mRNAs). Four samples were positive for IL-2 mRNA, and of these, three were also positive for interferon-gamma (IFN gamma). All TFC samples contained IL-6 and IL-8 mRNAs, even after depletion of CD3-positive T-cells. One TFC sample was additionally positive for IL-10 and TNF alpha mRNAs, and in the case of IL-10, this signal was not eliminated by CD3-positive T-cell depletion. IL-4 was not detected in any sample of ITL, TFC, or whole tissue. Semiquantitative analysis showed that the ITL fraction represented the major source of IL-6, IL-8, and TNF alpha mRNAs. By contrast, only three of five multinodular goiter samples were positive for IL-1 alpha mRNA; of these, two were also positive for IL-6, and 1 was positive for IL-8 mRNA. One multinodular goiter sample was positive for IL-8 mRNA alone, but IL-2, IL-4, IL-10, and TNF alpha mRNAS were not detected. These results suggest that although the TFC themselves may express certain cytokines, the ITL population represents the most important source of cytokine production in Graves' thyroid glands. The presence of IL-2, IFN-gamma, and TNF alpha and the absence of IL-4 mRNA in samples of ITL indicate a pattern of cytokine production that most closely resembles that of the TH1 helper T-cell subset. Given the etiological role of thyroid-stimulating antibodies in Graves' disease, the production of which is likely to depend upon TH2 helper T-cell function, it is perhaps surprising that the TH1 subset appears to predominate. It is possible that IL-10 is important in stimulating intrathyroidal autoantibody production, and this cytokine may also play a role in inhibiting cell-mediated thyroid injury in Graves' disease.

Base Sequence↗
PubMed1994

An investigation of the ability of TSH and Graves' immunoglobulin G to increase intracellular calcium in human thyroid cells, rat FRTL-5 thyroid cells and eukaryotic cells transfected with the human TSH receptor.

The purpose of this study was to determine if immunoglobulin G preparations (IgGs) from patients with Graves' disease can increase intracellular calcium in thyroid cells, as has been reported for TSH. Both TSH and Graves' IgGs (prepared by protein G affinity chromatography) increased calcium in a range of thyroid cells; however, the response seen, using Fura-2-loaded coverslips of cell monolayers, varied considerably. Chinese hamster ovary (CHO/JPO9) cells transfected with a high number of human TSH receptors showed the greatest response: TSH (10 mU/ml) increased calcium in 46% of experiments and 18 out of 25 (72%) Graves' IgGs increased calcium at 0.1 mg/ml (significantly greater, P < 0.001, than for control IgGs where cells responded to 2 out of 13 preparations). Rat FRTL-5 cells only responded to TSH in 22% of experiments and to 2 out of 8 (25%) of Graves' IgGs. Similarly, human thyroid cells responded to TSH in 22% of experiments and to 2 out of 9 (22%) of Graves' IgGs. (When studying cyclic AMP responses in JPO9 cells, much higher concentrations of Graves' IgGs were required (1-3 mg/ml). However, higher concentrations (0.3 mg/ml) of both Graves' IgGs, and to a lesser extent of control IgGs, were capable of increasing calcium in cells both with and without TSH receptors (control CHO cells and normal human dermal fibroblasts). We conclude that relatively low concentrations of patient IgGs can be distinguished from control IgGs in JPO9 cells on the basis of their ability to increase calcium, but that additionally all IgG preparations possibly contain another factor which can increase calcium in a range of cells independent of the presence of the TSH receptor.

Adolescent↗
PubMed1994

The next generation: poor compliance with risk factor guidelines in the children of parents with premature coronary heart disease.

OBJECTIVES: The offspring of individuals with premature coronary heart disease are themselves at increased risk for myocardial infarction before the age of 55. Consensus panels have recommended that all such offspring undergo an evaluation of cardiovascular risk, including cholesterol testing. METHODS: To examine self-reported rates of cardiovascular risk factor assessment in this population, we conducted a telephone survey of 318 Canadian adults with premature coronary heart disease and of one offspring from 298 (94%) of the 318 families. RESULTS: The median age of the offspring was 20 years (range 2 to 39 y). Among the 219 late adolescent and young adult offspring, only 97 (44%) reported having had a blood cholesterol measurement during the preceding 3 years. Thirty-seven percent reported being current smokers, 31% were overweight, and 30% exercised fewer than three times per week. Men were less likely than women to report having had their blood pressure measured in the preceding year (57% vs 80%). CONCLUSION: These low rates of cardiac risk factor assessment families of patients with premature coronary heart disease represent missed opportunities for primary prevention. More effective strategies to prevent atherosclerosis in this population are needed.

Adolescent↗
PubMed1993

IL-2 receptor-positive intrathyroidal lymphocytes in Graves' disease. Analysis of V alpha transcript microheterogeneity.

There has been considerable interest recently in the possible restriction of the TCR repertoire in autoimmune disorders, because such restriction would have important therapeutic implications. Reports of restriction of the TCR V alpha but not V beta repertoire in the thyroid in Graves' disease could not be repeated in an earlier study. Using RNA derived from matched peripheral blood, thyroid tissue, intrathyroidal lymphocytes (ITL), and IL-2R+ and IL-2R- subpopulations of ITL from Graves' patients, we conducted reverse transcription polymerase chain reaction/Southern blot analysis of TCR V alpha family usage. No evidence was found for V alpha restriction in the IL-2R+ subpopulation of ITL from eight patients, one of whom was operated on within 1 mo of diagnosis. We have further analyzed samples from seven of these patients by resolution on denaturing polyacrylamide gels. Typically, a single dominant band was amplified with surrounding minor bands in a normal distribution. Dominant and minor species were both the result of amplification from in frame V alpha transcripts, and the minor bands were separated in size by increments of 3 bp. We found no evidence for reduced heterogeneity of the V alpha transcripts in ITL or IL-2R+ ITL populations relative to peripheral blood in the vast majority of samples. This therefore suggests that there is little difference between the blood lymphocyte population and the activated T cell population in the thyroid in patients with Graves' disease, and indicates that in autoimmune thyroid disease, this method of analysis is not sufficient to distinguish between autoreactive and bystander T cell populations.

Amino Acid Sequence↗
PubMed1993

Increased membrane permeability of apoptotic thymocytes: a flow cytometric study.

We have recently developed a method for the separation and quantification of viable apoptotic cells without the need for permeabilisation or fixation of the cells. The method is based on the observation that apoptotic rat thymocytes fluoresce more brightly than normal cells after a brief incubation with the DNA binding dye, Hoechst 33342. In order to understand these differences, we have investigated the uptake of Hoechst 33342 by normal and apoptotic thymocytes. By staining with fluorescein diacetate, we have shown that the efflux of fluorescein from apoptotic cells is more rapid than that from normal thymocytes. This result demonstrated an increase in the permeability of the plasma membrane of the apoptotic thymocytes and it is this change which probably results in the more rapid uptake of Hoechst 33342. The data also revealed two populations of apoptotic thymocytes.

Animals↗
PubMed1993

HIV-salivary gland disease. Salivary scintiscanning with technetium pertechnetate.

The salivary disease in two patients with human immunodeficiency virus infection was investigated by technetium pertechnetate scintiscanning. Although there was good histologic evidence of benign lymphoepithelial disease, scintiscanning failed to delineate any salivary lesions. Technetium pertechnetate scintiscanning seems to be of little value in the detailed investigation of salivary disease in human immunodeficiency virus infection, though gallium scanning can help. Fine needle aspiration or biopsy remain the main diagnostic tools.

AIDS-Related Opportunistic Infections↗
PubMed1993

The gastroileoileal pouch: an alternative continent urinary reservoir for patients with short bowel, acidosis and/or extensive pelvic radiation.

We report on 6 patients who underwent a new type of continent urinary diversion: the gastroileoileal reservoir. These are a select group of patients who presented with the short bowel syndrome, acidosis, borderline diarrhea and/or severe pelvic radiation, which precluded the use of terminal ileum and the ileocecal segment. Considering these factors, and based on the different functional properties of the stomach as well as the need for a large reservoir, a segment of stomach and proximal ileum was used to construct the reservoir. Four patients have been followed for at least 6 months, with the longest followup being 12 months. Temporary dysphagia requiring hydrogen blockers developed in 1 patient. Results indicate excellent function of the continent urinary system, lack of metabolic complications, absent diarrhea and excellent patient tolerance. This procedure could be a useful alternative in some difficult clinical situations when continent urinary diversion is desirable.

Acidosis↗
PubMed1993

Analysis of fibroblast-stimulating activity in IgG from patients with Graves' dermopathy.

Conflicting results have been reported on the effect of IgGs from patients with Graves' dermopathy on dermal fibroblast function. We have analyzed 14 dermopathy IgGs prepared by protein G affinity chromatography. These caused FRTL-5 thyroid cells to synthesize significantly greater amounts of glycosaminoglycans (GAG) and protein than IgGs from normal controls (p < 0.05). [3H]Thymidine incorporation was also significantly increased (p < 0.05) and there was a significant correlation between all three parameters and the ability of these IgGs to stimulate iodide incorporation (p < 0.001). Dermopathy and control IgGs caused a modest increase in GAG synthesis by dermal fibroblasts but there was no significant difference between the two types of IgG. Increased GAG production was detectable in both culture medium and the extracellular matrix, and there was no difference between fibroblasts derived from the arm and leg. Conditioned medium from thyroid cells treated with dermopathy or control IgGs caused a greater increase in GAG production than the IgGs alone, but again there was no difference between the two sources of IgG. Neither control nor dermopathy IgG affected fibroblast protein synthesis or [3H]thymidine incorporation. Our results argue against a role for circulating IgGs in mediating Graves' dermopathy and make it unlikely that the TSH receptor antibodies are acting directly on a functional receptor expressed by dermal fibroblasts in this disorder.

Cell Line↗
PubMed1993

Efficiency of breathing systems A and D in the Carden Ventmasta ventilator.

We have compared the efficiency of the enclosed Magill attachment (System A) and the co-axial System D (Bain) in the Carden Ventmasta ventilator using both systems, each under five different ventilatory conditions, in each of five anaesthetized patients. Efficiency was assessed in terms of the effective alveolar ventilation as a fraction of the fresh gas flow. For System A, efficiency increased from a mean of 0.37 when the total ventilation was only 50% of the fresh gas flow, to a mean of 0.74 when ventilation was 2.3 times the fresh gas flow. The efficiency was substantially and significantly less with System D: 75% of that for System A at the smaller total ventilation (95% CI 65-85%) and 65% at the larger (95% CI 59-71%). A critical examination is made of conflicting definitions and terminology of the efficiency of breathing systems.

Adult↗
PubMed1993

Antioxidants can delay liver cell maturation which in turn affects gamma-glutamyltranspeptidase expression.

In normal rats just before weaning the majority of hepatocytes are mononucleated diploids, but within days the number of binucleated cells reaches a peak (approximately 50%) before declining again and there is a steady shift of diploid to tetraploid nuclei. When weanling rats were exposed to ethoxyquin (EQ), the conversion of 2N nuclei to 4N and 8N nuclei as measured by flow cytometry was slowed down. The rapid rise in the number of binucleate cells was also delayed, although the long-term effect was an increased number compared with age-matched controls. It appeared that when EQ was present in the diet, significant numbers of diploid hepatocytes undergoing DNA synthesis also underwent mitosis and cytokinesis giving rise to new diploid hepatocytes. However, many hepatocytes from animals maintained on a control diet did not undergo cytokinesis. Thus the slower 'conversion' of 2N to 4N nuclei in treated hepatocytes was due in part to promotion of cytokinesis in diploid cells undergoing DNA synthesis. The ploidy of a cell would be expected to affect gene expression. EQ is a very potent inducer of gamma-glutamyltranspeptidase (GGT), but expression depended on the age of the animals, the length of treatment time and apparently the ploidy status of the liver. In weanling rats treated with EQ for 7 days, > 80% of the hepatocytes expressed GGT, while in 42 day old rats similarly treated < 50% were positive for this enzyme. GGT expression was closely correlated with the percentage of 2N nuclei present in hepatocytes, suggesting that it was more easily induced in cells containing these nuclei than in those containing nuclei of higher ploidy. Although butylated hydroxytoluene (BHT), at the same concentration in the diet, had a similar negative effect on weight gain as did EQ, it had no effect on ploidy, nor did it induce GGT to the same extent as EQ.

Animals↗
PubMed1993

Epidural catheters. Breaking and extraction forces.

The forces required to break and to remove catheters from epidural spaces were investigated. Provided the patient's back is fully flexed and a slow steady pull is applied to the epidural catheter, the extraction forces required at both thoracic and lumbar levels are well below the minimum force required to break the same catheters.

Analgesia, Epidural↗
PubMed1993

No restriction of intrathyroidal T cell receptor V alpha families in the thyroid of Graves' disease.

Recently it has been reported that the intrathyroidal T cells in Graves' disease display restriction in V alpha T cell receptor (TcR) gene family usage, although this is not found with TcR V beta gene families in the same individuals. We have performed a qualitative analysis of TcR V alpha family usage in 12 patients with Graves' disease by reverse transcription and polymerase chain reaction (PCR) amplification of RNA extracted from isolated, unstimulated intrathyroidal lymphocytes and from snap-frozen whole thyroid specimens. No restriction was observed, with 10-15 V alpha gene families being amplified in all cases. The pattern of usage was similar to that in peripheral blood lymphocytes derived from normal subjects (n = 3) and from patients with Graves' disease (n = 3), as well as that present in the thyroids of patients with non-autoimmune toxic multinodular goitre (n = 4). These results indicated that there is no marked restriction of the unselected intrathyroidal T cell population in patients with Graves' disease who have been treated with antithyroid drugs.

Cell Line↗
PubMed1993

The gamma delta T cell repertoire in Graves' disease and multinodular goitre.

gamma delta T cells are a subset of T cells with unknown function, and restriction of the gamma delta T cell receptor (TCR) repertoire has been described in rheumatoid arthritis and multiple sclerosis. Elevated numbers of gamma delta T cells have been reported in the peripheral blood and thyroids of patients with Graves' disease. We have carried out flow cytometric analysis on peripheral blood mononuclear cells (PBMC) and intrathyroidal lymphocytes (ITL) from 12 patients with Graves' disease and nine patients with multinodular goitre (MNG), a thyroid disease of unknown etiology. There was no significant difference between the proportion of gamma delta T cells in the PBMC of Graves' and MNG patients, nor between the PBMC and ITL populations in either patient group. We have also carried out polymerase chain reaction amplification on RNA prepared from matched PBMC, ITL and the activated (CD25+) subset of ITL using six TCR V delta-family specific primers. Although there were differences in the amounts of each V delta transcript amplified from PBMC and ITL, there was no difference between the two patient groups. No consistent differences were therefore found between the gamma delta T cell populations in Graves' and MNG patients, arguing against the direct involvement of this T cell subset in the pathogenesis of Graves' disease.

Blotting, Southern↗
PubMed1993

Human papillomavirus type 16 E7 associates with a histone H1 kinase and with p107 through sequences necessary for transformation.

The transforming function of human papillomavirus type 16 (HPV16) E7 has been shown to depend on activities additional to the ability to bind RB. In this paper we describe two further properties of E7 which may also contribute to transformation, an association with a histone H1 kinase at the G2/M phase of the cell cycle and an ability to bind the RB-related protein p107. The region of E7 identified previously as important for RB binding was found to be involved in the association with the kinase and complex formation with p107, although analysis of E7 point mutants within this region revealed a difference in the precise sequence requirement for RB and p107 binding. Association with the kinase activity correlated with the ability to bind RB, but the restriction of the kinase association to the G2/M phase of the cell cycle implies that this activity might not be directly mediated by RB binding. Since kinase-binding-deficient E7 mutants are also transformation defective, this may represent an independent function of E7 which plays a role in the G2/M phase of the cell cycle.

Cell Line↗