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Biomedical subjects

B K Shenton

Publications and source records attributed to B K Shenton.

At least 73 records · Page 4Linked to original sources
PubMed1993

Administration of ATG according to the absolute T lymphocyte count during therapy for steroid-resistant rejection.

In renal transplantation, treatment of steroid-resistant rejection (SRR) with antithymocyte globulin (ATG) has been widely reported but over-immunosuppression remains a common problem. In the first ten patients (group 1) treated for SRR with rabbit ATG, three developed serious viral infections and two deaths occurred due to CMV pneumonitis. ATG was only omitted if thrombocytopenia or neutropenia occurred. In the next 17 patients (group 2) with SRR, ATG was administered according to the absolute T lymphocyte count. T lymphocytes were measured by flow cytometric analysis of CD3-labelled lymphocytes. ATG dosage was adjusted on a daily basis to keep the absolute T lymphocyte count under 50 cells/microliters. Administration of ATG according to the absolute T lymphocyte count resulted in a significant reduction in the mean dose of ATG given to the group 2 patients (P < 0.001). A significant decrease in the incidence of serious viral infections (P = 0.04) was achieved without reducing the ability of ATG to reverse the SRR (P = 0.29) or increasing the number of grafts lost at 1 year in the group 2 patients (P = 0.23).

Adult↗
PubMed1993

Production of immunosuppressive factors by a cultured tumour cell line and their effect on lymphocyte proliferation and cell cycle response.

Immunosuppression observed in patients with malignancy may be due to factors released by tumour cells. Medium conditioned by COLO 205 cells was found to inhibit mitogen-stimulated lymphocyte proliferation. Examination of CD25 and Class II MHC induction on PBMC incubated in complete, or COLO 205 conditioned, medium was not significant. The prevalence of lymphocytes in the S-phase of the cell cycle was enhanced after mitogenic stimulation and addition of COLO 205 conditioned medium. This was balanced by a concomitant fall in proportion of cells in the G0/G1 phases of the cell cycle. The immunosuppressive properties of COLO 205 conditioned medium was abrogated by heating to 60 degrees C for 30 min and by digestion with trypsin. Fractionation of the medium by gel filtration yielded two immunosuppressive fractions with relative molecular weights of 60,000 and below 20,000. It was concluded that cultured COLO 205 cells produce immunosuppressive protein/peptide factors which block cell proliferation during DNA synthesis. These factors fail to prevent upregulation of membrane-associated markers of cell activation.

Cell Cycle↗
PubMed1993

Renal allograft rejection: examination of adhesion blockade by antilymphocyte antibody drugs.

An assay was developed to investigate the binding of lymphocytes to cultured human renal epithelial cells. This binding was increased following lymphocyte activation by culture either with a polyclonal mitogen or with allogeneic stimulator cells. It was shown that such activation increased lymphocyte expression of the adhesion molecules CD2, LFA-1, and VLA-4. The ligand for each of these molecules was demonstrated on the surface of cultured renal epithelial cells. Polyclonal antilymphocyte antibody (ALA) preparations are used frequently to reverse intractable episodes of acute renal allograft rejection. It was demonstrated that such agents reduce the binding of activated lymphocytes to renal epithelial cells and subsequent cell lysis with a similar dose-response curve. Application of this assay may allow improved evaluation and titration of therapeutic antibody preparations. A range of monoclonal antibodies specific for components of the three adhesion molecule systems investigated in this work were added to lymphoid cell binding assays. It was found that combinations of these antibodies designed to interfere simultaneously with each of these adhesion interactions inhibited binding less well than the ALA preparation. It is likely that the superior inhibition of binding produced by ALA is due to the polyclonality of the antibodies which can block multiple epitopes on a wide range of potential adhesion molecules.

Antibodies, Monoclonal↗
PubMed1993

Renal allograft rejection: expression and function of VCAM-1 on tubular epithelial cells.

The interaction between vascular cell adhesion molecule-1 (VCAM-1) and very late antigen-4 (VLA-4) is known to play an important role in stabilizing the adhesion of lymphocytes to endothelial cells. Such cellular adhesion is crucial to many immunological processes including lymphocyte-mediated cell lysis. In this study the expression of VCAM-1 on renal tubular epithelial cells is demonstrated on biopsy sections recovered during acute renal allograft rejection. Experiments performed using epithelial cells cultured from renal tubules show that VCAM-1 is up-regulated by addition of the inflammatory cytokines tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). Combination of TNF-alpha and IFN-gamma synergized to induce high levels of VCAM-1 expression. Further experiments demonstrated that the cytokines produced by activated lymphocytes in mixed leucocyte culture also up-regulate expression of VCAM-1. Assays of the adhesion of lymphoid cells to cultured renal epithelial cells showed that cytokine pretreatment of the renal cells enhanced the binding of lymphoid cells. The proportion of bound lymphoid cells was significantly reduced by addition of an antibody capable of blocking the interaction of VCAM-1 with VLA-4. This result indicated that the VCAM-1 induced on renal epithelial cells by inflammatory cytokines is functionally capable of binding VLA-4, thereby enhancing the adhesion of potentially graft-damaging lymphoid cells.

Cell Adhesion↗
PubMed1993

Experimental renal transplantation: IgG antibodies which bind to graft tubular epithelial cells cannot be detected in the serum of recipient rats after pre-operative transfusion of donor blood.

Transfusion of donor blood 7 days prior to renal transplantation between certain rat strain combinations can delay subsequent graft rejection. The mechanism responsible for this increased graft tolerance remains largely unclear. In this paper it is demonstrated that IgG antibodies with affinity for cultured donor renal epithelial cells develop in the plasma of recipient PVG rats within 7 days of transplantation of a DA rat kidney. These IgG antibodies are not specific for kidney cell-restricted antigens as the activity can be completely absorbed using cells which express donor class-I MHC antigens. However, the IgG may contribute to the acute rejection of graft cells by activating antibody-dependent cell-mediated cytolytic (ADCC) mechanisms. Transfusion of donor blood 7 days prior to renal transplantation between DA and PVG rats prevented the development of IgG antibodies which bind to resting or cytokine-activated donor renal epithelial cells. Serum taken 7 days after the transplantation of animals which had been previously transfused with donor blood was not able to activate ADCC mechanisms of donor cells. The donor blood transfusion-mediated abrogation of IgG binding to graft cells may delay the onset of graft rejection by preventing antibody-dependent graft cell lysis.

Animals↗
PubMed1992

Flow cytometric crossmatching and outcome one year after renal transplantation.

Previous studies have shown that flow cytometric crossmatch assays can identify an at risk population in renal transplantation. We used the assay for recipient selection for 1 year. Recipients with donor T cell directed IgG were excluded from transplantation and those with B cell directed IgG were treated with increased immunosuppression. The transplants performed over this period (n = 126) were compared with an earlier series (n = 118) in which flow cytometric crossmatch results did not influence patient management. The results were evaluated for mortality and graft outcome at 3 months and 1 year. In addition, postoperative complications and duration of hospital stay were also assessed.

Antilymphocyte Serum↗
PubMed1992

Improved graft outcome and reduced complications due to flow cytometric crossmatching and DR matching in renal transplantation.

Several previous studies, including our own, have indicated that flow cytometric assays can identify an at-risk population of kidney transplant recipients. We used the assay for recipient selection for a period of twelve months. Recipients with donor T cell-directed IgG were excluded from transplantation and those with B cell-directed IgG were treated with increased immunosuppression. The transplants performed over this period (n = 126) were compared with an earlier series (n = 118) where, although the flow cytometric crossmatches were performed, the results did not influence patient management. In the series where the flow cytometric crossmatch was used in management, a lower failure rate was found at three months (P = 0.037 chi square), primary non-function was reduced (P less than 0.0001, Mann-Whitney), rejection episodes were reduced (P less than 0.0001, Mann-Whitney) and the hospital stay was shorter (P less than 0.0001, Student's t). The risk factors of ischemic times, panel reactivity, exposure to previous grafts and A/B locus matching were identical between the two groups. However DR matching was found to be higher in the series with the improved results (P less than 0.0001, Mann-Whitney). In view of the significant improvement in graft success and low complication rate, we intend to continue with the policy of recipient selection by flow cytometric crossmatching and DR matching.

Adult↗
PubMed1992

Renal allograft rejection: induction and function of adhesion molecules on cultured epithelial cells.

The interaction of graft-infiltrating immune cells with donor parenchymal cells is an important early event in allograft rejection. This binding is stabilized by interaction of antigen-independent 'adhesion' molecules expressed on the two cell types. As the level of expression of these molecules can be altered during inflammation, a series of experiments was performed to examine the effects of the inflammatory cytokines interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) on adhesion molecules expressed by cultured human renal tubular epithelial cells. These cells constitutively expressed ICAM-1 and LFA-3. Incubation with IFN-gamma increased expression of ICAM-1 but had no significant effect on expression of LFA-3 (P greater than 0.05). Incubation with TNF-alpha increased expression of both ICAM-1 and LFA-3; IFN-gamma synergized with TNF-alpha to further augment expression of these molecules. Peripheral blood lymphocytes (PBL) showed an enhanced binding to allogeneic renal epithelial cell monolayers which had been pretreated with IFN-gamma or TNF-alpha. MoAbs specific for ICAM-1 or its ligand LFA-1 inhibited adhesion of PBL to either IFN-gamma- or TNF-alpha-pretreated renal cells. By contrast, antibodies specific for LFA-3 or its ligand CD2 only significantly blocked PBL adhesion to renal cells which had been pretreated with TNF-alpha. Combination of antibodies specific for multiple components of the adhesion systems produced greater inhibition of adhesion than was produced by any single MoAb. These results suggest that the inflammatory cytokines IFN-gamma and TNF-alpha up-regulate expression of functional ICAM-1 and LFA-3 molecules which can augment the binding of potentially graft-damaging lymphoid cells to renal tubular epithelial cells.

Antigens, CD↗
PubMed1991

Node negative breast cancer: the prognostic value of DNA ploidy for long-term survival.

The DNA content of breast tumours from 170 patients who presented between 1978 and 1980 was measured by flow cytometry. The relationship between tumour ploidy and disease outcome was assessed and its association with other prognostic factors evaluated. Compared with those with diploid tumours, patients with aneuploid tumours had significantly earlier relapse and shorter survival (P less than 0.0001). Tumour ploidy was strongly related to grade (P less than 0.001), but there was no significant association between DNA ploidy and c-erb-B-2 expression, lymph node status or tumour size. In lymph node negative and c-erb-B-2 negative patients, aneuploid tumours were associated with a poorer prognosis (P less than 0.001) than diploid tumours. Multivariate analysis showed that tumour ploidy gave independent information on disease free and overall survival. Tumour ploidy may be used as an independent prognostic variable in patients with breast cancer and it may be helpful in defining patients within the node negative or c-erb-B-2 negative groups likely to have a poor outcome who might benefit from adjuvant treatment.

Adult↗
PubMed1991

The effect of improvements in cytometer sensitivity on the detection of CD5-positive B cells with dim fluorescence.

When antigen density on the surface of a cell population is low and variable, the percentage of that population determined to express the antigen (i.e., to be positively stained) depends directly on the sensitivity of the flow cytometer for resolving particles which are dimly fluorescent from those which are unstained. In this study, the sensitivity of a commercial flow cytometer has been improved by changes in the photomultiplier tube, the fluorescence filter, and the amount of stray light entering the fluorescence channel. In a model system with human lymphocytes, modifications to these factors increased the percent of the B-lymphocyte population found to express the CD5 antigen.

Antigens, Differentiation↗
PubMed1991

Renal epithelium: reversal of cytotoxic damage by addition of anti-thymocyte globulin.

A novel in vitro assay of renal epithelium tight junction function was used to assess the efficacy with which rabbit anti-thymocyte globulin (ATG) blocks epithelium damage mediated by lymphokine-activated killer (LAK) cells. It was found that LAK cells lysed renal epithelial cells poorly in standard chromium-release assays but that they caused a rapid, and almost total, reduction in trans-epithelium monolayer resistance, indicating tight junction failure and, hence, loss of tissue function. LAK cell-mediated cytolysis of the sensitive K562 cell line was completely blocked in the presence of ATG at a concentration of 200 micrograms/ml. Addition of ATG at this concentration to damaged renal cell monolayers in the presence of LAK cells allowed the trans-monolayer resistance to recover rapidly to levels approaching the values recorded before initial addition of LAK cells. On this basis it seems likely that the rapid restoration of renal function frequently observed after appropriate "rescue" therapy during episodes of acute rejection may reflect subtle changes in tissue function rather than recovery from widespread graft cell cytolysis.

Animals↗
PubMed1991

Transfusion-induced immunosuppression and red cell clearance.

While blood transfusion is increasingly implicated in the aetiology of tumour recurrence, the mechanism of this effect is unclear. Cancer-bearing patients are known to have factors in their sera which depress the function of normal lymphocytes. It is possible that blood transfusion accentuates this natural suppression. An animal model was therefore developed to study the effect of blood transfusion on humoral immunosuppressive activity and its possible relationship to red cell clearance. WAG rats given a transfusion of chromium-labelled allogeneic but blood-group compatible DA rat blood, developed significantly increased (P < 0.001) levels of lymphocyte suppressive factors in plasma (maximum at 7 days) which coincided with accelerated red cell clearance (t1/2 = 7 days). A transfusion of syngeneic WAG blood caused only a small transient increase in plasma suppression and red cells were cleared at a normal rate (t1/2 = 13 days) consistent with previous studies. However, when syngeneic WAG red cells were lysed and the red cell membranes infused there was a rapid increase in plasma suppression (P < 0.001), similar to but less prolonged than that achieved with allogeneic blood. The immunosuppressive effect of blood transfusion may result from accelerated clearance of allogeneic or damaged syngeneic red blood cells.

Animals↗
PubMed1991

Is voided urine suitable for flow cytometric DNA analysis?

Samples of bladder washings are frequently used to provide cellular material for flow cytometric DNA analysis. Since voided urine is a potential source of similar material, the aim of this study was to determine whether voided urine yields satisfactory specimens. We compared the qualitative results of flow cytometric DNA analysis of the cells in 53 specimens of voided urine and 109 samples of bladder washings; 45% (24/53) of urine specimens gave satisfactory DNA histograms compared with 93% (101/109) of bladder washings. This difference was apparent regardless of whether the bladder contained a tumour. It was concluded that bladder washings provide superior material for flow cytometric DNA analysis and that flow cytometric DNA analysis of freshly voided urine may have deficiencies which preclude its use in routine clinical practice.

DNA, Neoplasm↗
PubMed1991

Human endothelial cells: effect of TNF-alpha on peripheral blood mononuclear cell adhesion.

Human umbilical vein endothelial cells (HUVEC) were cultured and treated for varying periods with a range of concentrations of tumour necrosis factor-alpha (TNF-alpha). After this treatment the proportion of peripheral blood mononuclear cells (PBMC), previously depleted of plastic adherent cells, capable of binding to the endothelial cells was assessed. Few PBMC bound to HUVEC which had not been pretreated with TNF-alpha but up to 36% bound after pretreatment of the endothelial cells with TNF-alpha for 10 hr at a concentration of 10 U/ml. Phenotypic characterization of the adherent and non-adherent PBMC subpopulations revealed that natural killer (NK) cells (CD16+) and a proportion of memory helper T cells (CD4+ CD45RA-) bound to TNF-alpha pretreated HUVEC but that few naive helper T cells (CD4+ CD45RA+) showed similar binding. Cytotoxicity assays for NK activity were used to analyse functionally the adherent and non-adherent PBMC subpopulations. It was found that the cell subpopulation which did not adhere to TNF-alpha pretreated HUVEC mediated little lysis of K562 target cells. Conversely, the endothelial cell-adherent PBMC subpopulation produced active lysis supporting the phenotypic evidence that NK cells were concentrated within this subpopulation. These results suggest that TNF-alpha has a rapid and profound up-regulatory effect on the expression of adhesion molecules on the surface of HUVEC. Furthermore, it is apparent that these up-regulated adhesion molecules preferentially bind NK cells and a subset of memory helper T cells from the PBMC population.

CD4 Antigens↗