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R C Rees

Publications and source records attributed to R C Rees.

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

Preliminary evidence for genetic anticipation in Crohn's disease.

BACKGROUND: The term genetic anticipation is used to describe earlier onset of disease, increased severity, or both, in succeeding generations of families affected by a particular disease. This process has been linked with expanded genomic trinucleotide repeat regions in some neurological disorders. Crohn's disease, an inflammatory bowel disorder, has genetic influences, which remain undefined. We studied pairs of two-generation first-degree relatives with Crohn's disease to seek evidence for genetic anticipation in this disorder. METHODS: Through retrospective review of the records of 552 patients treated for Crohn's disease at Johns Hopkins Hospital, we identified 27 pairs of two-generation first-degree relatives. We also studied 32 such pairs identified through a multicentre survey. After exploratory analyses by t tests, a generalised estimating equations approach was used to fit a marginal regression model. FINDINGS: The age at diagnosis was earlier in the younger member of the pair both in the Johns Hopkins Hospital dataset (18.9 [SE 6.9] vs 31.4 [12.0] years) and in the multicentre survey dataset (16.9 [7.4] vs 33.1 [11.9] years). The regression model confirmed the findings: the best-fitting model for the Johns Hopkins Hospital data showed an average 10.8 (SE 4.2) year difference in age at diagnosis between parent and child; that for the multicentre data showed an average difference of 15.1 (1.5) years. There was evidence of a further difference between the second and third generations. Disease was more extensive in the offspring than in the parent for 15 of the 27 pairs at Johns Hopkins Hospital; in 13 of these pairs the affected parent was the father. INTERPRETATION: The evidence of a lower age at diagnosis and a greater extent of disease in the younger member of two-generation pairs affected by Crohn's disease, as well as the association with paternal transmission, suggest that genetic anticipation does occur in Crohn's disease. A search for triplet repeat regions is warranted.

Adolescent↗
PubMed1996

Alterations in endothelial cell proteinase and inhibitor polarized secretion following treatment with interleukin-1, phorbol ester, and human melanoma cell conditioned medium.

Polarized secretion of matrix metalloproteinases and plasminogen activators by monkey aortic endothelial cells was studied in vitro, using transwell inserts. The endothelial cells constitutively expressed matrix metalloproteinase-2, tissue inhibitors of metalloproteinases 1 and 2, urokinase, and tissue plasminogen activator, all with basal preference. Matrix metalloproteinase-9 activity was induced by phorbol 12-myristate 13-acetate (apical), interleukin-1 alpha (basal), and by conditioned medium from DX3 human melanoma cells (basal). The DX3 melanoma conditioned medium also stimulated basal secretion of matrix metalloproteinase-2, urokinase, tissue plasminogen activator, and tissue inhibitors of metalloproteinases. The rise in proteolytic activity in the basal direction was reflected by increased capacity to degrade subendothelial basement membrane type IV collagen, shown immunohistologically, using monkey kidney tissue sections and basement membrane deposited by endothelial cells into the transwell membrane. Thus, IL-1 alpha and DX3 melanoma conditioned medium can stimulate endothelial cells in vitro to concentrate secretion of proteinases spatially onto the underlying basement membrane. We suggest that the stimulation of endothelial cell proteinase activity by tumor cells may facilitate tumor cell extravasation.

Animals↗
PubMed1996

Identification of peptide epitopes of MAGE-1, -2, -3 that demonstrate HLA-A3-specific binding.

The MAGE gene family of tumour antigens are expressed in a wide variety of human cancers. We have identified 43 nonamer peptide sequences, from MAGE-1, -2 and -3 proteins that contain binding motifs for HLA-A3 MHC class I molecules. The T2 cell line, transfected with the cDNA for the HLA-A3 gene, was used in a MHC class I stabilisation assay performed at 37 degrees C and 26 degrees C. At 37 degrees C, 2 peptides were identified that stabilised HLA-A3 with high affinity (fluorescence ratio, FR > 1.5), 4 peptides with low affinity (FR 1.11-1.49) and 31 peptides that did not stabilise this HLA haplotype (FR < 1.1). At 26 degrees C, 12 peptides were identified that stabilised HLA-A3 with high affinity, 8 peptides with low affinity and 17 peptides that did not stabilise this HLA haplotype. Two peptides stabilised HLA-A3 at both temperatures. Small changes in one to three amino acids at positions distinct from the anchor residues altered peptide affinity. Data were compared to a similar study in which a peptide competition assay was used to investigate MAGE-1 peptide binding to several HLA haplotypes. This study demonstrates that anchor residues do not accurately predict peptide binding to specific HLA haplotypes, changes in one to three amino acids at positions distinct from anchor residues influence peptide binding and alternative methods of determining peptide binding yield different results. We are currently investigating the ability of these peptides to induce antitumour cytotoxic T lymphocyte activity as they may be of potential therapeutic value.

Amino Acid Sequence↗
PubMed1996

Detection of circulating prostate-specific antigen-positive cells in patients with prostate cancer by flow cytometry and reverse transcription polymerase chain reaction.

The presence of prostate-specific antigen (PSA)-positive cells has previously been demonstrated in the peripheral blood of prostate cancer patients by flow cytometry (FC), but the identity of these cells has not been established. In this study, the reverse transcriptase polymerase chain reaction (RT-PCR) was compared with analytical FC in an attempt to detect and characterise these cells. Peripheral blood was obtained from 12 patients with newly diagnosed and untreated prostate cancer and five controls. Nine of the 12 patients with prostate cancer (75%) had circulating PSA-positive cells as shown by FC. Only one of those patients (11.1%) was found to express PSA mRNA by RT-PCR. The absence of PSA mRNA in the majority of samples showing PSA-positive cells suggests that they do not represent haematogenous micrometastases. PSA-positive cells in the blood could represent monocytes that express PSA, either following binding/phagocytosis of free serum PSA or phagocytosis of tumour cells.

Base Sequence↗
PubMed1996

The effect of TGF-beta 1 and TGF-beta 2 on the proliferation of human Tenon's capsule fibroblasts in tissue culture.

This study examines the effect of transforming growth factor beta 1 and 2 (TGF-beta 1 and TGF-beta 2) on the proliferation of human Tenon's capsule fibroblasts in tissue culture. Both TGF-beta 1 and TGF-beta 2 were shown to stimulate proliferation at 1000 and 100 pg/ml concentrations. We discuss the significance of these findings with respect to wound healing following trabeculectomy, and postulate why some eyes are more at risk of failure than others.

Cell Division↗
PubMed1996

Analysis of the colonization of unirradiated and irradiated SCID mice by human lymphoma and non-malignant lymphoid cells.

We have evaluated the severe combined immunodeficient (SCID) mouse as an in-vivo model for the study of non-Hodgkin's lymphomas (NHL). Characterization of the immune system of the animals in our SCID mouse colony was carried out to assess the numbers of lymphoid cells present, to determine natural killer (NK) cell activity as a function of age and to examine the histology of the lymphoid organs. In this study four human NHL established cell lines (Daudi, Namalwa, U937, MC116), lymphoma cells from four fresh NHL biopsies and normal peripheral blood mononuclear cells (PBMC) and bone marrow cells were investigated, after intraperitoneal injection into the mice. The presence of the human NHL cells in the peritoneum and spleen was assessed by FACS analysis. The colonization potential was investigated in a range of tissues by polymerase chain reaction (PCR) amplification of human repetitive sequences. These studies revealed clear differences in the abilities of the NHL cell types to colonize the SCID mice. Namalwa, Daudi and U937 cells demonstrated the highest efficiency of colonization and readily formed tumours, whereas MC116, the NHL biopsy cell populations and the non-malignant lymphoid cells showed little ability to survive and colonize other tissues in the SCID mice. Whole body irradiation of the SCID mice appeared to improve the survival of human PBMC, NHL biopsy cells and MC116 cells in the peritoneum, but had little effect on their colonization potential. The significance of these studies is discussed.

Animals↗
PubMed1995

Tyrosine phosphorylation and activation of STAT5, STAT3, and Janus kinases by interleukins 2 and 15.

The cytokines interleukin 2 (IL-2) and IL-15 have similar biological effects on T cells and bind common hematopoietin receptor subunits. Pathways that involve Janus kinases (JAKs) and signal transducers and activators of transcription (STATs) have been shown to be important for hematopoietin receptor signaling. In this study we identify the STAT proteins activated by IL-2 and IL-15 in human T cells. IL-2 and IL-15 rapidly induced the tyrosine phosphorylation of STAT3 and STAT5, and DNA-binding complexes containing STAT3 and STAT5 were rapidly activated by these cytokines in T cells. IL-4 induced tyrosine phosphorylation and activation of STAT3 but not STAT5. JAK1 and JAK3 were tyrosine-phosphorylated in response to IL-2 and IL-15. Hence, the JAK and STAT molecules that are activated in response to IL-2 and IL-15 are similar but differ from those induced by IL-4. These observations identify the STAT proteins activated by IL-2 and IL-15 and therefore define signaling pathways by which these T-cell growth factors may regulate gene transcription.

Base Sequence↗
PubMed1995

Thrombopoietin (TPO) induces tyrosine phosphorylation and activation of STAT5 and STAT3.

The growth and differentiation of megakaryocytes are regulated by thrombopoietin (TPO), a recently characterized cytokine which exerts its effects via a member of the hematopoietin receptor superfamily, c-Mpl. Since many cytokines which bind hematopoietin receptors activate the STAT family of transcription factors, we investigated whether STAT proteins were activated by TPO. TPO induced the formation of a DNA-binding complex recognizing a known STAT-binding sequence. STAT5 was a major component of this DNA-binding complex, and STAT5 was tyrosine phosphorylated in response to TPO. Additionally, TPO-induced the tyrosine phosphorylation and DNA-binding activity of STAT3. Together with the recent demonstration of JAK2 activation in response to TPO, the data presented here define a rapid signaling pathway likely to be important in TPO-induced gene regulation.

Base Sequence↗
PubMed1995

Interleukin 12 induces tyrosine phosphorylation and activation of STAT4 in human lymphocytes.

Interleukin 12 (IL-12) is an important immunoregulatory cytokine whose receptor is a member of the hematopoietin receptor superfamily. We have recently demonstrated that stimulation of human T and natural killer cells with IL-12 induces tyrosine phosphorylation of the Janus family tyrosine kinase JAK2 and Tyk2, implicating these kinases in the immediate biochemical response to IL-12. Recently, transcription factors known as STATs (signal transducers and activators of transcription) have been shown to be tyrosine phosphorylated and activated in response to a number of cytokines that bind hematopoietin receptors and activate JAK kinases. In this report we demonstrate that IL-12 induces tyrosine phosphorylation of a recently identified STAT family member, STAT4, and show that STAT4 expression is regulated by T-cell activation. Furthermore, we show that IL-12 stimulates formation of a DNA-binding complex that recognizes a DNA sequence previously shown to bind STAT proteins and that this complex contains STAT4. These data, and the recent demonstration of JAK phosphorylation by IL-12, identify a rapid signal-transduction pathway likely to mediate IL-12-induced gene expression.

Base Sequence↗
PubMed1995

Interleukin 12 (IL-12) induces tyrosine phosphorylation of JAK2 and TYK2: differential use of Janus family tyrosine kinases by IL-2 and IL-12.

Interleukin (IL-12) has many effects on the function of natural killer and T cells, and is important in the control of cell-mediated immunity. IL-2 and IL-12 display many similar activities, yet each also induces a distinct set of responses. A human IL-12 receptor subunit has recently been cloned and, like the IL-2R beta and IL-2R gamma, is a member of the hematopoietic receptor superfamily; however, the molecular mechanisms of IL-12 action are unknown. In this report we show that IL-12 and IL-2 induce tyrosine phosphorylation of distinct members of the Janus (JAK) family of protein tyrosine kinases in human T lymphocytes. IL-12, but not IL-2, stimulates the tyrosine phosphorylation of TYK2 and JAK2, whereas JAK1 and JAK3, which are phosphorylated in response to IL-2, are not phosphorylated after IL-12 treatment. The use of distinct but related JAK family tyrosine kinases by IL-12 and IL-2 may provide a biochemical basis for their different biological activities.

Humans↗
PubMed1995

The effect of topical cholinergic medications on human Tenon's capsule fibroblasts in tissue culture.

BACKGROUND: The prior long-term use of topical anti-glaucoma medications has been suggested as an adverse factor for the outcome of trabeculectomy. The mechanism of action is not known. This study investigates the effect of cholinergic medications on the proliferation and viability of human Tenon's capsule fibroblasts in tissue culture to see whether this may be one mechanism by which increased surgical failure occurs. METHODS: We examined the effect of two commercial pilocarpine products, pure pilocarpine hydrochloride, and pure benzalkonium chloride, at various dilutions, on the proliferation (as assessed by 3H-thymidine uptake) and viability (assessed microscopically) of human Tenon's capsule fibroblasts in tissue culture. RESULTS: None of the tested compounds stimulated the proliferation of fibroblasts in tissue culture. Three of the compounds inhibited proliferation, and all compounds had toxic effects on cell morphology. At equivalent dilutions of commercially available concentration, pure benzalkonium chloride and the commercial product containing it appeared more toxic and antiproliferative to cells in culture than pure pilocarpine hydrochloride and the preservative-free commercial product. CONCLUSIONS: These results show an antiproliferative and toxic effect of medications on fibroblasts in tissue culture. They do not support the theory that topical medications directly stimulate fibroblast proliferation. The possible relevance of these results for the outcome of glaucoma surgery is discussed.

Aged↗
PubMed1995

Pilot study on the effect of topical adrenergic medications on human Tenon's capsule fibroblasts in tissue culture.

Recent publications have suggested that the long term use of topical antiglaucoma medications may be detrimental to the outcome of trabeculectomy. In order to investigate this further, the effect of several adrenergic agents and a preservative on the proliferation and viability of human Tenon's capsule fibroblasts in tissue culture were examined. The following compounds were tested: adrenaline (Eppy 1% and pure adrenaline base 1%, Smith & Nephew Pharmaceuticals Ltd); dipivefrine hydrochloride (Propine 0.1% and pure dipivefrine hydrochloride 0.1% Allergan Ltd and Allergan Pharmaceuticals (Ireland) Ltd); benzalkonium chloride (pure benzalkonium chloride 0.01%, Sigma Chemical Company Ltd); the two adrenaline based preparations were also tested in the presence of an antioxidant. None of the tested compounds stimulated the proliferation of fibroblasts. The commercial products tested, their pure compounds, and the preservative all inhibited proliferation and had toxic effects on the cells. In the presence of antioxidant, commercial Eppy and pure adrenaline base appeared to have less effect on proliferation and toxicity. These findings are discussed with reference to the outcome of trabeculectomy.

Adrenergic Agents↗
PubMed1995

Effect of TNF, IL-1, and IL-6 on the proliferation of human Tenon's capsule fibroblasts in tissue culture.

Trabeculectomy is a commonly performed procedure for primary open angle glaucoma and is successful in the majority of cases. However, certain factors including aphakia, previous surgery, secondary glaucomas, ethnic origin, and the long term use of topical antiglaucoma medications may be associated with a reduced success rate. The mechanism (or mechanisms) which influence clinical outcome following trabeculectomy remain elusive. Alterations in the composition of the conjunctiva or aqueous humour may be partly responsible for this effect, and this could be mediated by cytokines. In this study we found that tumour necrosis factor (TNF), and interleukin 1 (IL-1) were capable of stimulating the proliferation of Tenon's capsule fibroblasts in tissue culture. Interleukin 6 (IL-6) did not appear to have any effect. The relevance of this to wound healing following trabeculectomy is discussed.

Aged↗
PubMed1995

Cytokine modulation of adhesion molecule expression on human retinal pigment epithelial cells.

PURPOSE: The purpose of this study was to assess qualitatively the expression of adhesion molecules by human retinal pigment epithelium (RPE) and to study their regulation by inflammatory cytokines. These molecular events and the role played by inflammatory cytokines are important for selective lymphocyte trafficking into the eye during uveitis. METHODS: Expression of intracellular adhesion molecule-1 (ICAM-1), endothelial leukocyte adhesion molecule-1 (ELAM-1), platelet endothelial cell adhesion molecule-1 (PECAM-1), and vascular adhesion molecule (VCAM-1) by early passage human RPE cells was assessed by flow cytometry. In addition, the regulation of the expression of these molecules by the inflammatory cytokines interleukin-1 beta (IL-1 beta), IL-6, tumor necrosis factor alpha (TNF alpha), and interferon gamma (IFN gamma) was determined. Reverse transcription-polymerase chain reaction (RT-PCR) was used to characterize further adhesion molecule expression. RESULTS: Flow cytometric analysis determined that ICAM-1 was constitutively expressed on RPE cell lines and that in the presence of TNF alpha, IFN gamma, and IL-1 beta, there was a median fold increase in expression of 4.4, 5.4, and 4.4, respectively. In contrast, flow cytometric analysis of ELAM-1, PECAM-1, and VCAM-1 indicated that these adhesion molecules were not constitutively expressed at the cell surface; only the expression of VCAM-1 was upregulated by the presence of cytokines. The results of RT-PCR on RPE cells indicated that mRNA for all the adhesion molecules was present constitutively in some RPE cultures. After activation with IFN gamma, TNF alpha, and IL-1 beta, RT-PCR analysis showed that the number of RPE cell lines expressing all the adhesion molecules increased. CONCLUSIONS: ICAM-1 expression is markedly upregulated by inflammatory cytokines. Although mRNA for other adhesion molecules is expressed in RPE cells and is enhanced by inflammatory cytokines, this does not necessarily reflect the cell surface protein expression. Thus, the expression of adhesion molecules by RPE cells, and the subsequent recruitment of specific leukocytes, may be determined by the local cytokine environment.

Base Sequence↗
PubMed1994

The detection of intracytoplasmic interleukin-2 in Jurkat E6.1 and human peripheral blood mononuclear cells using direct conjugate, two-colour, immunofluorescent flow cytometry.

The most commonly used approaches for the estimation of cytokine protein production involve the quantification of cytokines produced, and accumulated, in a complex body fluid or supernatant of cultured cells, by means of a bioassay or immunoassay, but these techniques do not permit an estimation of the frequency or phenotype of cytokine-producing cells. Traditional methods use immunohistochemical based techniques which can be difficult to perform and interpret, whereas flow cytometry has the advantage of objective assessment and standardisation and is less labour intensive. In this study we have established a rapid and sensitive technique for the simultaneous detection of intracellular IL-2 in conjunction with CD3. Polyclonal goat anti-IL-2 and control goat IgG were conjugated to FITC and then separated from free fluorochrome using column chromatography. PHA activated PBMC or Jurkat E6.1 cells were fixed in paraformaldehyde and permeabilised with saponin, followed by the addition of directly conjugated antibodies (FITC anti-IL-2 or PE anti-CD3) alone or in combination. Samples were then analysed using a flow cytometer and the percentage of dual labelled cells calculated. Several methods have been previously established for the detection of intracellular cytokines using flow cytometry and employing multiple layers of antibodies in the detection steps. By using direct conjugates the technique is less time consuming, requires fewer controls and can be used to examine cytokine production by identifiable cell phenotypes in a mixed cell population.

CD3 Complex↗
PubMed1994

Nm23 'anti-metastatic' gene product expression in colorectal carcinoma.

Nm23 is a putative metastasis suppressor gene and alterations in this gene have been reported in colorectal carcinomas suggestive of a role for nm23 in the dissemination of these tumours. In this study we used an affinity purified polyclonal antibody, ab-11, on formalin-fixed, paraffin-embedded sections of colorectal carcinomas from 46 patients in a three-stage avidin-biotin complex immunoperoxidase technique. Follow-up of these patients was until time of death or for 5 years, with a mean time of 31.2 months. Two observers scored the staining results from 1 to 3 according to the proportion of tumour cells positive. The association of nm23 staining with survival, sex, age, vascular invasion, and Dukes' stage was determined using Cox's regression model. The association of death from colorectal cancer and nm23 status reached marginal significance in this study (P = 0.0417). Moreover, there is some suggestion of a protective effect from nm23 as the relative risk of dying from colorectal cancer for each increment of nm23 positivity is 0.573 (95 per cent confidence limits 0.34-0.98).

Adenocarcinoma↗