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Biomedical subjects

W J Burlingham

Publications and source records attributed to W J Burlingham.

At least 55 records · Page 3Linked to original sources
PubMed1991

Donor-specific cytotoxic T lymphocyte hyporesponsiveness following renal transplantation in patients pretreated with donor-specific transfusions.

Our purpose was to investigate the mechanism of the continuing beneficial effect of donor-specific transfusions in the cyclosporine era. We describe the development of donor-specific cytotoxic T lymphocyte hyporesponsiveness in peripheral blood lymphocytes obtained up to 2 years posttransplant in patients preconditioned with 3 DST plus azathioprine. In a group of 12 such patients, hyporesponsiveness developed gradually, becoming detectable in some patients as early as 1 month posttransplant and becoming statistically significant for the entire group at 9-12 months posttransplant. A complete specificity for donor alloantigens was seen in the hyporesponsiveness of some patients; in others, partial suppression of the response to a third party HLA-mismatched control was also seen. Although slight suppression of the mixed lymphocyte culture response was seen in some patients, overall there were no statistically significant differences in MLC responses to control or donor stimulators at any time point posttransplant as compared with pretransplant, pre-DST. The mechanism of donor-specific CTL hyporesponsiveness 2 years posttransplant was explored in one patient (HLA A1, 2, B 57, 60; DR 3, 6) who had received a 2-HLA haplotype-mismatched kidney transplant from her husband (HLA A2,--; B5, 8; DR4,--) following DST plus AZA pretreatment. Bulk culture CTL analysis showed specific nonresponsiveness to donor stimulators; however in the presence of exogenous recombinant IL-2, the antidonor response was restored to the level of pretransplant PBL. Limiting dilution analysis using recombinant IL-2 revealed equivalent precursor frequency of antidonor CTL in pre- and posttransplant PBL. These data suggest that the hyporesponsive PBL contained donor-specific CTL precursors but were deficient in helper function necessary for CTL maturation.

Blood Transfusion↗
PubMed1991

Multivariate analysis of donor-specific versus random transfusion protocols in haploidentical living-related transplants.

A total of 127 haploidentical living-related transplants have been performed at our institution since March 1986. A donor-specific transfusion plus azathioprine protocol was used until July 1988 (n = 74) and a random transfusion (RT) protocol without AZA used thereafter (n = 53) in an effort to decrease risk of recipient sensitization and reduce the burden on the prospective donor. All patients were given cyclosporine 8 mg/kg/day orally beginning 1 week prior to transplantation. Immunosuppression was similar in both groups and consisted of triple induction therapy with prednisone, CsA, and AZA. A positive T cell crossmatch eliminated the potential donor. Seven individuals (9.6%) were sensitized in the DST group and 1 (1.9%) in the RT group, leaving 67 and 52 patients in the two groups of the study, respectively. Groups were similar with respect to age, sex, history of pregnancy in female patients, peak and baseline panel-reactive antibody (PRA), DR match, and prior transplants. The groups differed slightly with respect to AB antigens shared, with an advantage in the RT group. Actuarial graft survival was not statistically significantly different between the two groups, with 2-year graft survival of 95% in the DST and 91% in the RT group (log rank, P = 0.16). Patients in the RT group had significantly more rejection episodes and had them sooner than their counterparts in the DST group. At the end of 1 year, 50% of patients in the DST group had at least 1 rejection episode, compared with 75% of patients in the RT group (P = 0.0008). Multivariate (Poisson) analysis of 10 variables was performed, with an overall model P-value of 0.0001. Only DST (P = 0.0001) and pregnancy (P = 0.015) were significant predictors of rejection episodes, both protective. The difference in rejection episodes and the timing with which they occur has not yet translated into a significant difference in graft survival between DST and RT groups.

Adult↗
PubMed1990

Adsorption of cytotoxic anti-HLA antibodies with HLA class I immunosorbant beads.

In an effort to generate HLA immunosorbants to specifically remove anti-HLA antibodies from sera of highly sensitized patients, we purified HLA proteins, covalently coupled them onto Sepharose, and adsorbed antisera from five patients with narrowly reactive cytotoxic anti-HLA antibodies and from one patient with broadly reactive antibodies. We found that an HLA-A2 immunosorbant depleted anti-HLA-A2 cytotoxic antibodies, but did not deplete anti-HLA-B7 or anti-HLA-B44 cytotoxic antibodies from the narrowly reactive patient sera. Patient S.C. developed high PRA (81%) with strong cytotoxicity against HLA-A1 and -A2 following rejection of an HLA-A1, -B57 mismatched kidney. We adsorbed his sera with five HLA immunosorbants including HLA-A2 and HLA-A1,28. We found that the HLA-A2 immunosorbant depleted antibodies to HLA-A2+ and HLA-B57+ cells but not to HLA-A1+ cells, while the HLA-A1,A28 immunosorbant depleted antibodies to both HLA-A1+ cells and to the HLA-A28 cross-reactive HLA-A2+ cells. Adsorption was specific for HLA-A alleles to which the patient was sensitized, since neither HLA-B-C immunosorbants (containing HLA-B7, -B8, -B13, -B27, or -B37 plus HLA-C gene products) nor the control immunosorbants (bovine serum albumin or diphtheria toxoid) depleted serum S.C. of cytotoxic anti-HLA antibodies. Our results indicate that HLA immunosorbants are stable to sequential cycles of adsorption and elution, and thus may be of future therapeutic value in treatment of sensitized patients.

Antibodies, Monoclonal↗
PubMed1989

Secretion and cell surface expression of IgG1 are impaired in human B lymphoblasts that lack HLA-A, -B, and -C antigens.

B-lymphoblastoid cell line (LCL) 721.221 lacks HLA-A, -B, and -C class I antigens and transcripts as a result of gamma-ray-induced mutations. LCL 721, from which mutant .221 was derived, produces membrane and secreted forms of IgG1(kappa). In contrast, IgG expression in .221 had these characteristics: (i) gamma 1 heavy chains were diminished by 98% but were detectable with chain-specific antibodies in cell lysates; (ii) kappa light chains were present at normal levels in cell lysates and free kappa chains were secreted; (iii) cell-surface-associated IgG and secreted IgG were absent. Mutants that had partially reduced amounts of class I antigens continued to secrete IgG; however, both the absolute amount of IgG secreted and the relative amount of kappa vs. intact IgG secreted were abnormal in such partially class I-deficient cells. The failure to export IgG and the deficiency of HLA-A, -B, and -C were not merely coincidental in mutant .221, since production of IgG was restored by transferring a functional HLA-A, -B, or -C gene into .221. Cell surface antigen expression of cloned HLA-A, -B, and -C transgenes introduced into .221 was comparable to that of the same genes in their normal chromosomal locations. These observations reveal a relation between production of HLA class I gene products and production of IgG.

Antibodies, Monoclonal↗
PubMed1989

Risk factors for sensitization by blood transfusions. Comparison of the UW/Madison and UC/San Francisco donor-specific transfusion experience.

We have tested the predictive model for risk of sensitization by donor-specific transfusions developed at the University of California, San Francisco for its applicability to the DST experience at UW/Madison. Patient sample sizes between the two groups were similar (n = 249 for UW/Madison, n = 261 for UCSF) and the two groups of patients had similar compositions in terms of mean age, ABO type, baseline panel-reactive antibody, and pregnancy rate. The two groups differed in that the UW/Madison group had a higher percentage of males, diabetic patients, previously transplanted patients, and 0 haplotype-matched (2 HLA-mismatched related and unrelated) recipients. In addition, all the UW/Madison patients received azathioprine (AZA) whereas only half the UCSF group was given AZA. Despite these differences, application of the UCSF model for prediction of sensitization by DST gave remarkably similar results in our patient population, with pregnancy, prior transplant, baseline PRA, and HLA antigen sharing giving similar odds ratios and P values. However, when female sex was included in the model along with the other variables, the significance of pregnancy risk disappeared. We have developed an alternative multivariate model using stepwise logistic regression that identifies baseline PRA greater than 40%, female sex, and prior transplant as significant risk factors for sensitization, while number of HLA (A, B, DR) antigens shared and increasing recipient age significantly decreased risk of sensitization by DST.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1988

A rapid assay for measuring both colony size and cytolytic activity of limiting dilution microcultures.

A combined 51Cr-release/MTT dye method is described for accurately measuring cytolytic activity and colony size in the same set of culture microwells. The method was applied to the study of cell-mediated lympholysis (CML) in limiting dilution analysis (LDA) cultures of human PBL from a renal transplant recipient and a healthy control. The results showed that the combined CML/MTT method could detect differences in lytic activity per cell in LDA cultures, and thus is a useful adjunct to standard precursor frequency analysis.

Cells, Cultured↗
PubMed1988

Improved renal allograft survival following donor-specific transfusions. III. Kinetics of mixed lymphocyte culture responses before and after transplantation.

The essence of the clonal deletion model of the donor-specific transfusion (DST) effect is synergy between DST-priming and post-transplant immunosuppression. Using a sensitive kinetics assay of the mixed lymphocyte culture (MLC) response to donor and third-party stimulators, we compared the responses of controls (non-transfused healthy individuals) with those of patients who either had no rejection episodes in the first week posttransplant (group 1) or who had DST-type (greater than 3d onset) rejection episodes (group 2). We found that group 2 patients had normal or above-normal MLC responses after DST plus azathioprine (AZA) pretransplant treatment, but had a reduced/delayed posttransplant anti-donor MLC following reversal of early rejections (P = .05 compared with controls). Group 1 patients had a nonspecifically reduced MLC after DST + AZA treatment (P less than .02 compared with controls), while posttransplant MLC responses showed a return to normal (pretransfusion) levels. These data suggest a synergy of DST with immunosuppressive drug that induced MLC hyporesponsiveness, but only in patients who received anti-lymphoblast globulin or a sustained high dose immunosuppression in the early posttransplant period.

Azathioprine↗
PubMed1988

Multivariate analysis of risk factors for sensitization and early rejection episodes in a donor-specific transfusion plus azathioprine protocol.

A total of 205 patients who underwent donor-specific transfusions with azathioprine (DST+AZA) treatment from 1982 to 1987 included 25 patients (12%) who developed antibodies cytotoxic to donor T cells (+T warm crossmatch). When we examined a series of 14 variables in relation to the risk of sensitization using a univariate analysis, panel-reactive antibody (PRA) pre-DST of greater than 40%, pregnancy, female sex, male-to-female DST, and HLA antigen match all were significant (P less than .05) risk factors. Of these variables, only PRA pre-DST greater than 40% proved to be statistically significant (P less than .002) in a multivariate logistic regression model (overall P less than .001). Of those patients who went on to transplant with the donor-specific kidney, 41/170 (24%) developed early on or before day 5 posttransplant) rejection episodes. Univariate analysis of 17 variables in relation to early (less than 5 days) rejection episodes indicated no significant variables. Nonetheless using a multivariate logistic regression model (overall P = .09), HLA match (P less than .004), perioperative transfusions (P less than .05), and azathioprine withdrawal for greater than 1 week during DST (P less than .05) all correlated with the incidence of early rejection episodes. In a smaller multivariate model involving 47 patients for whom MLC/blocking factor data were available, the presence of plasma blocking factor at the time of transplant was associated with a decreased incidence of early rejection (P = .07). We conclude that the factors that influence the rate of T+ sensitization are distinct from those that influence the rate of early rejection episodes in a DST+AZA protocol.

Antilymphocyte Serum↗
PubMed1988

Induction of antiidiotypic antibodies to donor HLA-A2 following blood transfusions in a highly sensitized HLA-A2+ recipient.

A patient (HLA-A2,3; B35,60) with end-stage renal disease and a high level of pretransfusion (t0) anti-HLA cytotoxic antibodies (60% positive to a random panel), but lacking cytotoxic antibodies against her HLA haploidentical sister (HLA-A2,3; B35,44), received 3 donor-specific transfusions (DST) from the latter: 200 cc fresh whole blood at biweekly intervals, while being treated with azathioprine (AZA, 1 mg/kg/day). Her serum remained negative for antidonor antibodies both by standard cytotoxicity assay and by immunofluorescence flow cytometry after DST + AZA treatment, and she experienced no acute rejection episodes following donor kidney transplantation. Microcytotoxicity inhibition tests were performed using standard HLA-typing sera as a source of Ab-1, and pre- and posttransfusion sera were added to serial dilutions of Ab-1 to test for the presence of Ab-2 (antiidiotype) to donor HLA class I specificities. Although both pre- and posttransfusion sera inhibited cytotoxicity toward HLA-A2 antigens expressed on recipient target cells, only posttransfusion serum was found to inhibit cytotoxicity against the HLA-A2 antigens expressed on donor target cells. Absorption of soluble HLA class I antigens present in pre- or posttransfusion sera removed the inhibition of cytotoxicity toward recipient HLA-A2 but did not affect the inhibition of cytotoxicity toward donor HLA-A2 by posttransfusion sera. The F(ab')2 fragment of the IgG fraction of posttransfusion sera contained the inhibitory activity, suggesting induction of Ab-2 toward idiotypes specific for donor HLA-A2 antigens encoded on the unshared haplotype.

Antibodies, Anti-Idiotypic↗
PubMed1988

What is known about blocking factors in renal allograft recipients.

A variety of substances present in serum or plasma, either at the time of renal transplant or during stable graft function in long-term recipients, may interfere with cell-mediated immune functions. In several cases the presence of serum blocking factors has been correlated with decreased graft vulnerability to acute rejection. The question is: are serum blocking factors important for the induction or maintenance of allograft tolerance or, alternatively, are they merely by-products of the tolerant state? Perhaps the most compelling case for an essential role of serum blocking factors can be made in instances where vigorous cell-mediated immune responses can be demonstrated in vitro, but anergy (for example, absence of DTH response) is seen in vivo. However, in renal transplant recipients the majority of studies that show the presence of receptor blocking antibodies or other immunoregulatory serum factors also found a decreased cell-mediated immune response in vitro. Thus, allograft tolerance would appear to involve multiple mechanisms as suggested elsewhere. Due to the recent discovery of the molecular structure of the T cell receptor 33,34, it is now possible to identify determinants associated with the binding of T cell receptor blocking antibodies in renal transplant recipients; for example, one should be able to determine if families of V beta genes are involved. Alternatively, it may be possible to identify polymorphic structures on T cells other than the alpha, beta receptor complex as ligands for blocking antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Anti-idiotypes and transfusions.

There is growing evidence for the existence of an idiotypic network in the mammalian response to alloantigens, in particular to MHC antigens. The direct evidence for induction of Ab-2 to particular donor HLA-specificities by DST is strong; in addition, there is recent evidence for Ab-2 to Ab-1 specific for constant regions of HLA-ABC antigens in the sera of multiply-transfused patients. On the other hand, evidence for induction of anti-TCR antibodies by DST is based primarily on functional specificity rather than biochemical evidence; the evidence available suggests that MLC BF induced by DST may be directed against V beta family or public idiotypes on recipient T cells, since the amount of inhibition obtained is weak and no evidence of "clonotypic" antibodies has been found in studies of T cell lines or lymphoblasts. Better molecular probes, including human monoclonal antibodies, T cell clones, and purified TCR are needed to screen for specific anti-idiotypes in transfused patients.

Animals↗