Serological responses to cytomegalovirus during renal transplant rejection.

We studied the role played by CMV in kidney transplant rejection by recording serological responses to CMV replication in cadaver graft recipients and recording clinical graft rejection by monitoring acute changes in renal function and the appearance of antidonor lymphocyte antibody (anti-Dab). If CMV plays a significant role in rejection, clinical rejection should correlate with CMV activity; if CMV does not play a role, clinical rejection would be likely to correlate with anti-Dab but not necessarily with CMV activity. We selected retrospectively 18 rejectors and 18 nonrejectors by clinical criteria and assayed for anti-Dab (by fluorescence-activated flow cytometry) and CMV antibody (by complement fixation and Western blot for IgG and IgM) over a 3-6-month period after transplantation. Primary CMV infection occurred in 8 of 12 (67%) CMV seronegative graft recipients and reactivation or reinfection occurred in 16 of 24 (67%); 12 of 14 (86%) rejectors developed anti-Dab compared with 2 of 18 (11%) nonrejectors (P less than 0.00001). Active CMV infection occurred in 11 of 18 (61.1%) rejectors and 13 of 18 (72.2%) nonrejectors (P = 0.36), and in 8 of 15 (53.3%) of those who developed anti-Dab and 12 of 17 (70.6%) of those who did not (P = 0.26). The results show no evidence to link CMV activity with kidney graft rejection.

Interleukin-2 (rIL-2)-induced lymphokine-activated killer (LAK) cells and their precursors express the VGO1 antigen.

Precursor and effector cells of recombinant interleukin-2 (r-IL-2)-induced lymphokine-activated killer (LAK) activity were investigated for their expression of VGO1. Peripheral blood lymphocytes (PBL) from normal donors were purified and separated in a FACS 420 into VGO1+- and VGO1- cell fractions before and after culture for 96 hr with 100 U/ml of r-IL-2. Their lytic activity against K 562 and Daudi cells was measured in a 51Cr release assay. The majority, if not all, of the LAK effector and precursor cells was VGO1+ lymphocytes. The expression of VGO1 by LAK precursor cells remained stable under the culture conditions used in our experiments. VGO1- lymphocytes cultured with r-IL-2 demonstrated neither LAK-induced activity nor expression of VGO1 antigen.

Characterization of peripheral blood mononuclear cells expressing the VG01 antigen: their NK and memory function.

This study describes VG01, a lymphocyte activation antigen of approximately 40 kd and the functional characteristics of VG01 circulating peripheral blood lymphocytes (PBL). We have previously described that VG01 is expressed by monocytes, the majority of CD16 lymphocytes, and a small fraction of circulating T lymphocytes. It is also newly expressed by T cells upon activation with mitogens and alloantigens and allows for the selection and enrichment in vitro of allospecific primed T lymphocytes. In this manuscript, we confirm that most, if not all, the NK activity resides in the VG01-enriched population. We also demonstrate that the memory response to PPD and Candida in previously sensitized normal individuals is found within the VG01+ cell fraction. In contrast, the proliferative response to PHA is significantly greater in the VG01-depleted PBL, but both cell fractions have an overall adequate proliferative response to this mitogen. The absence of proliferation to PPD and Candida by VG01-PBL is not due to differences in the kinetics of response because this cell fraction was almost devoid of responding cells when proliferation to the antigens was measured at different time intervals. Anti-VG01 has no effect in the in vitro lymphocyte proliferative response to antigens and mitogens. VG01 appears to be a lymphocyte activation antigen that includes circulating T memory cells and that at least in these cells appears to be permanently rather than transiently expressed.

A novel lymphocyte activation antigen present on allospecific primed lymphocytes and defined by the monoclonal antibody VG01.

This study describes VG01, a monoclonal antibody to a novel lymphocyte activation antigen. The cells reacting with VG01 were first characterized by indirect immunofluorescence and fluorescence-activated cell sorter analysis. The antigen defined by VG01 was expressed on circulating T lymphocytes (23.8 +/- 9.0) and on the majority of large granular lymphocytes and monocytes (greater than 90%), while granulocytes stained weakly. All other cells tested so far--thymocytes, hematological malignancies and several cell lines--were negative. Upon activation with mitogens or allogeneic cells more than 90% of the transformed lymphocytes became VG01-positive. The kinetic studies demonstrated that the antigen defined by VG01 lags the expression of the IL-2 receptor by 48 hr and remains present for up to 18 days after stimulation in culture. Thus, this antigen does not seem to be involved in the initial steps of T cell activation and its expression continues after cell division has stopped. Assays with purified sorted populations showed that lymphocyte proliferation to mitogens and in mixed lymphocyte culture came primarily from the VG01- population. The responding cells became VG01+ during activation--and when alloantigen-primed lymphocytes were sorted and restimulated with the original stimulators, all the in vitro memory response came from the VG01+ population. Thus, VG01 antibody is efficient in selecting for alloreactive T lymphocytes in vitro and has potential for selective modulation of immune responses. Its reactivity with activated T cells and NK cells may also help define a common functional program for these two cell types, adding to the understanding of their mechanism of action and/or their origin.

Persistent synovial lymphocyte responses to cytomegalovirus antigen in some patients with rheumatoid arthritis.

Synovial lymphocytes from 6 of 40 patients with rheumatoid arthritis responded to cytomegalovirus antigen stimulation. 3H-thymidine uptakes were more than 3 times greater than were those of the responses to 13 other microbial antigens. Similar results were obtained in 1 patient on 7 occasions over 17 months, and in the 5 other patients on each of 2 occasions. In 3 of the 6 patients, synovial lymphocyte responses to cytomegalovirus antigen were markedly different from simultaneous peripheral blood lymphocyte responses.

Allogeneic bone marrow transplantation using partially-matched related donors.

Six patients with advanced leukemia and one with severe combined immunodeficiency syndrome received allogeneic bone marrow transplantation (BMT) from HLA haploidentical donors who were compatible for one or two loci on the non-shared chromosome. Methotrexate was used for prophylaxis against acute graft-versus-host disease (GVHD). All patients engrafted but developed moderate to severe acute GVHD, and three patients died. Two leukemic patients relapsed but two others survived and were free of disease 403 and 936 days post-transplant. BMT using related partially matched donors may result in durable engraftment and long-term survivors. The implications of using such donors in expanding the application of BMT are discussed.

Immunotherapy of primary immunological aborters: rationale for the use of pooled cryopreserved purified normal peripheral blood mononuclear cells.

Patients with recurrent spontaneous abortions have been successfully treated in many centers with third-party immunization directed to a putative TLX antigen system. This immunotherapy requires the screening of a large number of donors to match the patients' red blood cell (RBC) phenotype and has the potential risks associated with transfusions from 30 to 50 donors. Our modified approach to third-party immunization is to use irradiated frozen-stored purified lymphocytes pooled from five normal donors. Mononuclear cells from normal donors are obtained in a cell separator. After sedimentation and Ficoll-Hypaque separation, the cells are stored in liquid N2. The RBC depletion of the final preparation is of the order of 5 to 6 logs, theoretically decreasing the need for RBC phenotyping except for the Rh system. Using a highly sensitive fluorescence-activated cell sorter technique and an ADCC assay, we found that ABH, Rh, Fya Fyb, Jka Jkb, MNS, and Kell antigens are either not expressed by cryopreserved human mononuclear cells, or, if so, they are below the level of detection of these highly sensitive assays. We conclude that the use of pooled frozen mononuclear cells is an adequate alternative for immunotherapy. It decreases the transfusion risks associated with exposure to a large number of donors and the need for RBC phenotyping, making this modality of treatment more accessible.

Differences in immunoglobulin subclasses between dye exclusion and 51Cr release complement-dependent lymphocytotoxicity assays.

Paradoxical differences previously noted between lymphocytotoxicity detected by dye exclusion at room temperature (CDCE) or by 51Cr release (CDC51Cr) at 37 degrees C in maternal antipaternal complement-dependent lymphocytotoxicity have suggested that CDCE and CDC51Cr at 37 degrees C, but not at 20 degrees C, may detect different immunological antibody-antigen interactions. Reactions in the two test systems against the same target cells were compared in sera from known immune dialysis patients, secondary aborting women, and refractory platelet recipients before and after heat treatment of sera, absorption with solid-phase heparin, anti-light-chain augmentation, and the addition of murine monoclonal anti-IgG subclass antibodies. The results demonstrate significant differences between the two tests using the same target and sera. Further, the results imply the presence of an inhibitor and an inhibitor of inhibitor in sera. The involvement of different immunoglobulin subclasses was shown in the two tests. These data demonstrate the necessity for further study of the nature of the differences in the mechanisms of these clinically important antibody-detecting systems.

Adoptive immunotherapy for acute non-lymphocytic leukaemia: a long-term follow up.

Immune cells cultured in vitro against autologous blast cells were infused twice in 8 patients with ANLL after achieving complete remission. 3 of the 8 patients remained in first remission for 74-94 months and they appeared to be cured of their leukaemia. These 3 patients received significantly higher numbers of in vitro sensitized immune cells as well as having a better recovery of the in vitro cultured cells. The other 5 patients relapsed within a year of diagnosis; 3 of them had had organomegaly and a preceding myeloproliferative disorder at diagnosis. None of 20 ANLL patients in complete remission treated simultaneously with similar chemotherapy at our institution remained in first complete remission, nor were long-term survivors. Adoptive immunotherapy with in vitro sensitized immune cells may be effective treatment in acute leukaemia. The therapeutic potential of infusion of in vitro stimulated autologous immune cells requires further investigation.

Fibroblast inhibition: a new and treatable cause of prosthetic graft failure.

We have encountered two patients who had repeated false aneurysms, seromas, and unseated grafts which presented about 18 months after aortobifemoral grafting. An inhibitor of fibroblast growth was detected in each patient's serum by in vitro dilution tests. Normal ingrowth of fibrous tissue resumed after plasmapheresis, leaving the patients clinically stable for up to 3 1/2 years after treatment. Criteria for the diagnosis of fibroblast inhibition are an unseated prosthesis which simulates infection, no bacterial growth on carefully studied cultures, and inhibition of fibroblast growth by the patient's serum in vitro. The diagnosis should be entertained in the differential diagnosis of graft infection when an unseated graft is encountered in association with a late seroma or false aneurysm. Plasmapheresis removed the inhibitor. Fibroblast inhibition has not been reported previously.

Hypomagnesemia associated with gentamicin therapy.

Two patients developed symptomatic hypomagnesemia most likely secondary to gentamicin therapy. Both patients were on regular doses of the antibiotic and had hypokalemia and hypocalcemia secondary to hypomagnesemia. Mechanisms postulated for the occurrence of hypomagnesemia secondary to gentamicin therapy include drug-induced hyperaldosteronism and tubular toxicity. Patients on gentamicin therapy should have serum magnesium, calcium, and potassium levels monitored routinely.

Coccidioidomycosis tenosynovitis: case report and review of the literature.

Coccidioidomycosis tenosynovitis is an unusual rheumatological manifestation of Coccidioides immitis infection. We report a case in a 46-year-old man with leukemia. The literature is reviewed and the treatment discussed.

Neonate with Down's syndrome and transient congenital leukemia. In vitro studies.

A white female neonate with clinical manifestations of Down's syndrome was found to have a peripheral white count of 74,000/mm3 with 65% blast forms. Bone marrow aspirate revealed 36% blasts. On chromosomal analysis of bone marrow and peripheral blood, two cell lines were found, one with trisomy 21 and another with pentasomy 21. Colony cultures on the patient's bone marrow cells revealed an adequate growth pattern of CFU/GM. Cells were reactive with myeloblast alloantisera. At the age of 3 weeks, peripheral counts were within normal limits as was morphology of the repeat bone marrow aspirate. Chromosome analysis at that time showed complete absence of pentasomy 21 cell line, and repeat cell colony studies revealed normal growth, differentiation, and maturation of CFU/GM. Maternal serum totally inhibited colony growth of the patient's remission marrow and allogeneic blast cells. This serum inhibitor actually disappeared 1 month postpartum. The patient has had no recurrence of blast cells up to the present age of 3 years.

In vitro cell growth in neonates with Down's syndrome and transient myeloproliferative disorder.

Bone marrow and peripheral blood cells from three newborns with Down's syndrome and transient myeloproliferative disorders were cultured in vitro. In the methylcellulose semiliquid system, normal colony formation with maturation and differentiation into granulocytes and monocyte-macrophages were observed in all three patients. This is different from the growth pattern usually seen in acute nonlymphocytic leukemia. A maternal serum inhibitor of both normal allogeneic GM-CFU and blast cell growth was also demonstrated, but its role in pathogenesis is uncertain. Normal in vitro granulopoiesis may help to differentiate the transient myeloproliferative syndrome from congenital leukemia in newborns with Down's syndrome.