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Biomedical subjects

K Williamson

Publications and source records attributed to K Williamson.

At least 55 records · Page 3Linked to original sources
PubMed1996

Requirements for interleukin-4-induced gene expression and functional characterization of Stat6.

Interleukin-4 (IL-4) stimulation leads to the activation of the signal transducer and activator of transcription 6 (Stat6). In this study, we present data relating to the functional properties of Stat6. Human embryonic kidney 293 cells were shown to be deficient of Stat6 yet express all other components of the IL-4 signaling cascade. This cell line was used for transient-transfection studies of wild-type and mutant Stat6 proteins. The wild-type protein was shown to activate a reporter construct carrying multiple copies of the IL-4 response element derived from the human immunoglobulin heavy-chain germ line epsilon promoter. Similarly, a truncated protein lacking 41 amino acids of the N terminus was fully active. However, removal of the C-terminal 186 amino acids completely abolished transcription activation. Amino acid substitutions were introduced into the putative DNA binding domain (VVI at positions 411 to 413), the SH2 domain (R-562), or the tyrosine (Y-641) which presumably becomes phosphorylated upon activation. All three of these Stat6 mutants were unable to activate transcription in 293 cells. Wild-type and mutant Stat6 derivatives were also expressed in insect cells, and purified proteins were analyzed in vitro for the ability to interact with both DNA and tyrosine-phosphorylated peptides derived from the IL-4 receptor alpha chain. Mutations within the DNA binding domain, the SH2 domain, or tyrosine 641 completely abolished DNA binding. In contrast, only the SH2 mutant failed to interact with tyrosine-phosphorylated peptides. The transdominant effects of all Stat6 derivatives were analyzed by using HepG2 cells, which express endogenous Stat6 protein. Differential effects were observed with various mutants, supporting the current model of the Jak/STAT activation cycle.

Amino Acid Sequence↗
PubMed1996

Three cases of nalbuphine hydrochloride dependence associated with anabolic steroid use.

Three case reports are presented of nalbuphine hydrochloride dependence meeting DSM IIIR and ICD10 criteria for opioid dependence. Nalbuphine hydrochloride is being obtained from illicit sources and used by those using performance enhancing drugs. In some cases this leads to opioid dependence. There is a potential risks of crossover between the misuse of drugs of performance and the misuse of psychoactive drugs by injection. Further research into the dependence potential of nalbuphine and the extent of the crossover between steroid misuse and other psychoactive drug misuse is required. The legal status of nalbuphine should be reviewed in the light of its availability on the black market.

Adult↗
PubMed1995

A high-resolution integrated physical, cytogenetic, and genetic map of human chromosome 11: distal p13 to proximal p15.1.

We describe a detailed physical map of human chromosome 11, extending from the distal part of p13 through the entirety of p14 to proximal p15.1. The primary level of mapping is based on chromosome breakpoints that divide the region into 20 intervals. At higher resolution YACs cover approximately 12 Mb of the region, and in many places overlapping cosmids are ordered in contiguous arrays. The map incorporates 18 known genes, including precise localization of the GTF2H1 gene encoding the 62-kDa subunit of TFIIH. We have also localized four expressed sequences of unknown function. The physical map incorporates genetic markers that allow relationships between physical and genetic distance to be examined, and similarly includes markers from a radiation hybrid map of 11. The cytogenetic location of cosmids has been examined on high-resolution banded chromosomes by fluorescence in situ hybridization, and FLpter values have been determined. The map therefore fully integrates physical, genic, genetic, and cytogenetic information and should provide a robust framework for the rapid and accurate assignment of new markers at a high level of resolution in this region of 11p.

Base Sequence↗
PubMed1995

Isolation of two new members of the NF-AT gene family and functional characterization of the NF-AT proteins.

The activation of cytokine genes in response to antigenic stimulation of T cells is mediated by NF-AT proteins. Previous studies have identified two NF-AT proteins, NF-ATp and NF-ATc, that are homologous within a 290 aa domain distantly related to the Rel domain. We have isolated two additional members of this gene family, NF-AT3 and NF-AT4, which encode proteins 65% identical to the other NF-AT proteins within the Rel domain. The four NF-AT genes are transcribed in different sets of tissues that included many sites of expression outside the immune system. The Rel homology domain is sufficient for DNA recognition and cooperative binding interactions with AP-1. Although other members of the Rel family bind DNA as dimers, NF-AT proteins are monomers in solution or bound to DNA. Transfection assays indicate that each of the four NF-AT proteins can activate the IL-2 promoter in T cells.

Amino Acid Sequence↗
PubMed1995

Nocturnal hypoxaemia in late pregnancy.

We have measured arterial oxygen saturation (SpO2) continuously overnight in 13 non-pregnant (NP), 13 pregnant normotensive (NPIH) and 15 pregnant patients with a diagnosis of pregnancy-induced hypertension (PIH). The two pregnant groups did not differ in duration of pregnancy (> 35 weeks) and none was in labour. There was no significant difference in age between these three groups. Mean SpO2 in group NP was 98.5% (range 97-99%). This was significantly higher than that in group NPIH (95.2 (91-98) %) and group PIH (94.9 (89-99) %). In seven pregnant patients, more than 20% of the recording was spent with an SpO2 < 90%. We conclude that a significant number of pregnant women (> 35 weeks' gestation) suffer from prolonged nocturnal hypoxaemia.

Adolescent↗
PubMed1994

Hydrochloric acid denaturation of colorectal tumour tissue infiltrated with bromodeoxyuridine.

The thymidine analogue, bromodeoxyuridine, is substituted into DNA during DNA synthesis and can be used to identify those cells which have passed through the S-phase of the cell cycle. Incorporated bromodeoxyuridine can be detected using anti-bromodeoxyuridine monoclonal antibodies which bind to the exposed bromodeoxyuridine in single-stranded DNA after a suitable denaturation step. Hydrochloric acid is the most commonly employed denaturation agent in bromodeoxyuridine monoclonal antibody methodologies. This preliminary study was to validate the hydrochloric acid denaturation step for colorectal tumour tissue infiltrated in vivo with bromodeoxyuridine. Standard immunohistochemistry and flow cytometric techniques using Bu20a were employed across a range of hydrochloric acid concentrations. Although high labelling indices were achieved using acid concentrations of 0.10 M HCl, the optimal hydrochloric acid concentration was not the same in all tumours. Sensitivity of DNA to hydrochloric acid denaturation should be carefully considered in bromodeoxyuridine methodologies using the monoclonal antibody Bu20a.

Antibodies, Monoclonal↗
PubMed1993

Morphometric analysis of colorectal cancer.

Thirteen nuclear and cellular morphometric variables were measured in 312 cases of colorectal adenocarcinoma. All variables, except nuclear shape factors, differed significantly (P < 0.001) between normal colorectal and tumor tissue. In adenocarcinomas, epithelial nuclei in well-differentiated mucosa tended to be elliptic, while those in poorly differentiated mucosa were more spheric. Increasing values of maximum nuclear and elliptic diameter were associated with progression from none to simple tubule configuration (P < 0.001), none to easily discerned nuclear polarity (P < 0.001), and expanding growth pattern (P < 0.001). Univariate survival analysis revealed that none of the morphometric variables was significantly related to patient survival. Multivariate regression analysis showed that no morphometric variable could add significantly to a model containing the variables of patient age, Dukes stage, and tumor differentiation. Morphometry may be useful in distinguishing malignant from normal tissue and degrees of differentiation, but it is of little prognostic value in colorectal adenocarcinoma.

Adenocarcinoma↗
PubMed1993

In vitro BrdUrd incorporation of colorectal tumour tissue.

This study describes a novel in vitro method for the incorporation of the thymidine analogue, bromodeoxyuridine (BrdUrd), in fresh colorectal tumour tissue. Disaggregation by pronase, collagenase and DNAse resulted in high cell yields of viable single cell suspensions, representative of the original tumour, which could be infiltrated with BrdUrd. A modified ELISA identified optimal incubation times and BrdUrd concentrations. This technique has been used in preliminary studies to investigate two important areas intrinsic in the analysis of BrdUrd colorectal cell proliferation data: 1) to determine the effects of the individual constituents of the cell culture media, in particular glutamine, on BrdUrd incorporation in suspensions of colorectal cells and 2) to examine the denaturation step. This method will have wide applicability in investigations of cell proliferation status in both normal and diseased tissue.

Bromodeoxyuridine↗
PubMed1993

DNA densitometry of colorectal cancer.

DNA analysis was assessed by densitometry for 281 cases of colorectal adenocarcinoma. Detection of aneuploidy in a single case rose from 65% if one, to 92.5% when three or more sections, were analysed. Although aneuploid tumours had significantly larger nuclear areas than near diploid tumours (p = 0.009), densitometric measurements showed no association with clinicopathological variables. DNA content determined by densitometry was compared with that from flow cytometry on 465 tissue sections from 241 cases. Aneuploidy assessed by flow cytometry was significantly associated with that determined by densitometry (p < 0.01 for all comparisons), ploidy state being similar in 381 sections (82%, kappa = 0.63, p < 0.001), and 187 cases (77.6%, kappa = 0.57, p < 0.001). Univariate survival analysis showed that DNA densitometric variables had no significant association with survival in (a) all cases, (b) cases without lymph node metastases, or (c) cases without distant metastases. Multivariate regression analysis of densitometric and clinicopathological variables identified Dukes's stage, patient age, and tumour differentiation as the combination of variables most closely related to survival. Densitometric measurement of DNA content could not significantly improve on the prognostic model containing these three variables. It is concluded that, although the assessment of DNA content by densitometry is comparable with that of flow cytometry, conventional histological variables remain the best predictors of prognosis in colorectal cancer.

Adenocarcinoma↗
PubMed1993

The role of flow cytometry in carcinoma of the colon and rectum.

Flow cytometry was performed upon 312 patients with adenocarcinoma of the colon and rectum, satisfactory results being obtained with 275 (108 diploid, 130 aneuploid and 37 tetraploid). The proportion of nondiploid instances increased from 28 percent if one, to 80 percent when six specimens were assessed per patient. Reproducibility of the technique showed substantial agreement in the assessment of deoxyribonucleic acid ploidy (Kappa value equals 0.74). Increasing values of cells in the diving (G2/M) phase of the cell cycle were associated with little lymphocytic tumor infiltration (p = 0.0002) and extensive tumor fibrosis (p = 0.003). Univariate survival analysis revealed that, although diploid tumors tended to have a better prognosis than nondiploid tumors (p = 0.06), no flow cytometric variable was significantly related to survival. Flow cytometry similarly was not of prognostic value in instances without lymph node metastases or without distant metastases. Multivariate regression analysis of flow cytometric and clinicopathologic variables identified Dukes' stage, patient age and tumor differentiation as the combination of variables most closely related to survival. No flow cytometric variable could significantly improve on the prognostic model containing these three variables. It is concluded that conventional histologic variables remain the best predictors of prognosis in carcinoma of the colon and rectum.

Adenocarcinoma↗
PubMed1988

Solubilization and reconstitution of the formylmethionylleucylphenylalanine receptor coupled to guanine nucleotide regulatory protein.

We describe the solubilization, resolution, and reconstitution of the formylmethionylleucylphenylalanine (fMet-Leu-Phe) receptor and guanine nucleotide regulatory proteins (G-proteins). The receptor was solubilized with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. Guanine nucleotides decreased the number of high-affinity binding sites and accelerated the rate of dissociation of the receptor-ligand complex, suggesting that the solubilized receptor remained coupled to endogenous G-proteins. The solubilized receptor was resolved from endogenous G-proteins by fractionation on a wheat germ agglutinin (WGA)-Sepharose 4B column. High-affinity [3H]fMet-Leu-Phe binding to the WGA-purified receptor was diminished and exhibited reduced guanine nucleotide sensitivity. High-affinity [3H]fMet-Leu-Phe binding and guanine nucleotide sensitivity were reconstituted upon the addition of purified brain G-proteins. Similar results were obtained when the receptor was reconstituted with brain G-proteins into phospholipid vesicles by gel filtration chromatography. In addition, we also demonstrated fMet-Leu-Phe-dependent GTP hydrolysis in the reconstituted vesicles. The results of this work indicate that coupling of the fMet-Leu-Phe receptor to G-proteins converts the receptor to a high-affinity binding state and that agonist produces activation of G-proteins. The resolution and functional reconstitution of this receptor should provide an important step toward the elucidation of the molecular mechanism of the fMet-Leu-Phe transduction system in neutrophils.

Adenine Nucleotides↗