Recipient IgG Research Articles

P16.27: A Case of Cytomegalovirus (CMV) Esophagitis Without CMV Viremia After SARS-CoV-2 Infection in a Kidney Transplant Recipient While on CMV Prophylaxis

Introduction: CMV prophylaxis significantly reduces CMV infection after solid organ transplants, but patient may still present with CMV primary disease or reactivation during the first six months post-transplant. Although high CMV viral load is usually seen in tissue invasive disease, blood CMV PCR my not be able to diagnose gastrointestinal CMV disease as sensitivity of blood CMV PCR is lower in seropositive patients. CMV and SARS-CoV-2 co-infection has been reported particularly in elderly and immunocompromised patients. We report a unique case of CMV esophagitis without viremia diagnosed after recent SARS-CoV-2 infection in a kidney transplant recipient. Case: Twenty-eight year old Caucasian female with end stage kidney disease due to unknown etiology received negative crossmatch living related renal transplant from her sister. She received Basiliximab for induction and Tacrolimus and mycophenolic acid as maintenance immunosuppression. Patient had intermediate risk for CMV (Donor CMV IgG negative and recipient IgG positive) and was on Valganciclovir for prophylaxis as per the protocol. She had a stable allograft function with serum creatinine around 1-1.2 mg/dl. She developed SARS-CoV-2 infection with mild respiratory symptoms after 2 months of kidney transplant. Mycophenolic acid was held very briefly, and Prednisone 10 mg daily was added to the regimen. She reported chest pain, nausea and vomiting after resuming mycophenolic acid. Blood CMV PCR was undetectable and other laboratory studies were significant for anemia of 9.3 g/dl (baseline 10 g/dl), with stable renal and hepatic functions, without leucopenia or thrombocytopenia. Upper Gastrointestinal endoscopy revealed a large 4-5 cm circumferential ulceration in the mid esophagus with biopsy features suggesting viral cytopathic effect and rare CMV infected cells. Stomach and duodenum were normal. HSV and fungal stains were negative. Patient was started on oral valganciclovir induction dose and showed symptomatic recovery in a few weeks. Conclusion: High index of suspicion is necessary to diagnose CMV co-infection and/or reactivation in the settings of SARS-CoV-2 infection and the tissue biopsy is the key for diagnosis of the tissue invasive gastrointestinal disease.

CMV Pneumonitis After Discontinuation of Prophylaxis Therapy Guided by CMV T-cell Immunity Panel

<h3>Introduction</h3> CMV T-cell immunity panels (TCIP) may reliably qualify T-cell specific CMV immunity, however, utility within lung transplant recipients (LTR) is unclear. We describe one of several CMV pneumonitides occurring in LTR after a positive CMV TCIP. <h3>Case Report</h3> A 61yo bilateral LTR (CMV donor IgG+/recipient IgG-) for COPD was admitted for worsening dyspnea 17mo post-LT. During admission, he experienced persistent leukopenia requiring multiple granulocyte colony-stimulating factor doses, MMF discontinuation, and frequent bronchoscopy for clearance. Respiratory cultures were positive for <i>Aspergillus</i> species only. Despite drug modification, his WBC and ANC ranged 1-4K cells/µL and ∼1K cells/µL, respectively. Durable CMV seroconversion to IgG+ occurred 9.5mo post-LT. No prior CMV viremia occurred and only 2 episodes on bronchoalveolar lavage (BAL) viral shedding ranging 1243-2762 copies/mL, with the second episode occurring 3 months prior to current admission. During hospitalization, a CMV TCIP showed immunity [(reference range >0.20%) CD4+ CMV IFN-ɣ cells = 3.08%; CD8+ CMV IFN-ɣ = 10.64%; CD4+ control = 1.93%; CD8+ control = 8.28%]. Subsequently, valganciclovir (VGCV) was discontinued to reduce further bone marrow suppression. One month after VGCV cessation, routine plasma CMV PCR monitoring resulted 740 copies/mL with BAL 37114 copies/mL. Lung tissue pathology and cytology were not performed, however, given the high BAL viral load and persistent graft dysfunction, CMV pneumonitis was diagnosed and treatment with IV ganciclovir (GCV) was initiated. He received 21d of IV GCV treatment with clinical and virologic improvement, and now remains on lifelong VGCV secondary prophylaxis. <h3>Summary</h3> This case advises caution when interpreting CMV TCIP alone to guide duration of CMV prophylaxis in LTR. Pairing CMV TCIP assay with assessment of patient factors (i.e. age, recipient CMV IgG seroconversion, hypogammaglobulinemia) or need for serial monitoring requires further exploration.

Invasive Cytomegalovirus Disease in a Low Risk CMV Serostatus, D-/R- Lung Transplant Recipient

<h3>Introduction</h3> Cytomegalovirus (CMV) infection is a major cause of morbidity and mortality among transplant patients. Here, we report a case of lung transplant patient with low risk CMV serostatus (D-/R-) who developed invasive CMV disease. <h3>Case Report</h3> A 64-year-old lady with COPD underwent bilateral lung transplant. CMV serostatus was donor IgG negative / recipient IgG negative. Basiliximab was used for induction and was maintained on tacrolimus, mycophenolate and prednisone. Intraoperatively, she received 2 leucoreduced PRBC's. 15 days post-transplant she developed CMV viremia and was started on treatment dose valganciclovir. No mutations noted on the resistance panel. 2 weeks on therapy, there was no reduction in the CMV PCR. Chest CT showed a right pleural effusion which was exudative in nature. Microbiology testing was negative. Fluid cytology showed CMV cytopathic changes. Valganciclovir was changed to iv ganciclovir because of her GI absorption concerns. With 6 weeks of iv ganciclovir and monthly CMV Immune globulin, CMV PCR log values steadily declined. <h3>Summary</h3> Lung transplant patients with low risk CMV serostatus can have a minor risk of transfusion acquired (TA) or community acquired (CA) CMV infection. A single center retrospective study of low-risk organ transplant patients did not identify any cases of TA CMV infection in patients receiving leucoreduced PRBC. Another study showed 2.4% risk of TA CMV infection in patients receiving non-leucoreduced PRBC. CA CMV risk is greater than TA CMV, with the incidence being higher in children (3/100 patient years) compared to adults (0.46/100 patient years). Currently no guidelines exist to recommend CMV prophylaxis in low risk CMV serostatus transplant patients. Hence measures need to be taken to monitor such patients who are at increased risk for TA or CA CMV. Recipients whose IgG was negative during transplant work up should be retested at the time of transplant. CMV PCR monitoring should be considered if the donor or recipient received extensive transfusions.

Immunogenicity, safety, and reactogenicity of heterologous COVID-19 primary vaccination incorporating mRNA, viral-vector, and protein-adjuvant vaccines in the UK (Com-COV2): a single-blind, randomised, phase 2, non-inferiority trial

Given the importance of flexible use of different COVID-19 vaccines within the same schedule to facilitate rapid deployment, we studied mixed priming schedules incorporating an adenoviral-vectored vaccine (ChAdOx1 nCoV-19 [ChAd], AstraZeneca), two mRNA vaccines (BNT162b2 [BNT], Pfizer-BioNTech, and mRNA-1273 [m1273], Moderna) and a nanoparticle vaccine containing SARS-CoV-2 spike glycoprotein and Matrix-M adjuvant (NVX-CoV2373 [NVX], Novavax). Com-COV2 is a single-blind, randomised, non-inferiority trial in which adults aged 50 years and older, previously immunised with a single dose of ChAd or BNT in the community, were randomly assigned (in random blocks of three and six) within these cohorts in a 1:1:1 ratio to receive a second dose intramuscularly (8-12 weeks after the first dose) with the homologous vaccine, m1273, or NVX. The primary endpoint was the geometric mean ratio (GMR) of serum SARS-CoV-2 anti-spike IgG concentrations measured by ELISA in heterologous versus homologous schedules at 28 days after the second dose, with a non-inferiority criterion of the GMR above 0·63 for the one-sided 98·75% CI. The primary analysis was on the per-protocol population, who were seronegative at baseline. Safety analyses were done for all participants who received a dose of study vaccine. The trial is registered with ISRCTN, number 27841311. Between April 19 and May 14, 2021, 1072 participants were enrolled at a median of 9·4 weeks after receipt of a single dose of ChAd (n=540, 47% female) or BNT (n=532, 40% female). In ChAd-primed participants, geometric mean concentration (GMC) 28 days after a boost of SARS-CoV-2 anti-spike IgG in recipients of ChAd/m1273 (20 114 ELISA laboratory units [ELU]/mL [95% CI 18 160 to 22 279]) and ChAd/NVX (5597 ELU/mL [4756 to 6586]) was non-inferior to that of ChAd/ChAd recipients (1971 ELU/mL [1718 to 2262]) with a GMR of 10·2 (one-sided 98·75% CI 8·4 to ∞) for ChAd/m1273 and 2·8 (2·2 to ∞) for ChAd/NVX, compared with ChAd/ChAd. In BNT-primed participants, non-inferiority was shown for BNT/m1273 (GMC 22 978 ELU/mL [95% CI 20 597 to 25 636]) but not for BNT/NVX (8874 ELU/mL [7391 to 10 654]), compared with BNT/BNT (16 929 ELU/mL [15 025 to 19 075]) with a GMR of 1·3 (one-sided 98·75% CI 1·1 to ∞) for BNT/m1273 and 0·5 (0·4 to ∞) for BNT/NVX, compared with BNT/BNT; however, NVX still induced an 18-fold rise in GMC 28 days after vaccination. There were 15 serious adverse events, none considered related to immunisation. Heterologous second dosing with m1273, but not NVX, increased transient systemic reactogenicity compared with homologous schedules. Multiple vaccines are appropriate to complete primary immunisation following priming with BNT or ChAd, facilitating rapid vaccine deployment globally and supporting recognition of such schedules for vaccine certification. UK Vaccine Task Force, Coalition for Epidemic Preparedness Innovations (CEPI), and National Institute for Health Research. NVX vaccine was supplied for use in the trial by Novavax.

Early Cytomegalovirus Reactivation after Allogenic Bone Marrow Transplantation Is Associated with the Loss of Recipient-Derived Humoral Immunity and Is Reduced By IL-6 Inhibition

Background:Interleukin-6 (IL-6) promotes the differentiation of pathogenic T cells and represents a dominant pathway of cytokine-dysregulation and graft-versus-host disease (GVHD). Cytomegalovirus (CMV) reactivation is a common and life-threatening infectious complication following allogenic bone marrow transplantation (BMT). The role of T cell immunity is long-established, and our group recently demonstrated that CMV strain-specific immunoglobulins (Ig) are critical in preventing CMV reactivation in preclinical models (Martins JP et al, Science 2019).Methods and Results:In a phase I/II clinical trial (Kennedy GA et al, Lancet Onc. 2014), we observed unexpectedly low rates of CMV reactivation following IL-6 receptor (IL-6R) blockade with tocilizumab (TCZ) compared to historical control (8/35 vs 20/43, P < 0.05). We used our preclinical models of murine CMV (MCMV) reactivation to pursue the mechanisms. We transplanted bone marrow (BM) and T cells from naïve B6 donors into CMV-latently infected B6D2F1 recipients and monitored MCMV reactivation. Ablation of IL-6R in donor T cells (using B6 CD4 cre x IL-6R fl/fl transgenic mice; IL-6R -/-) significantly reduced MCMV reactivation 5 weeks after BMT, as measured by plasma viremia and infectious viral loads in the liver (Fig 1A).Next, we examined the contribution of donor T and B cells to the suppression of MCMV reactivation by IL-6R inhibition. At 2 - 3 weeks after BMT and prior to MCMV reactivation, cytokine (IFNɣ/TNF) secretion from CD4 + T cells (in response to MCMV infected DC) was not detectable; MCMV m38 tetramer + CD8 T cells were present at very low frequencies (< 0.1% of CD8 T cells) which were not altered by IL-6R ablation. The frequency of CXCR-5 +PD-1 + T FH cells was comparable or lower in IL-6R -/- vs wild-type (WT) T cells after BMT. Transplant of BM from B6.μMT mice (unable to generate mature B cells and plasma cells) did not increase MCMV viremia (Fig 1B), excluding a role for donor B cells. Thus, early MCMV reactivation in the presence of IL-6 is independent of MCMV-specific T cells and donor B cells.To study the effects of IL-6 on recipient-derived humoral immunity we quantified MCMV-specific IgG in plasma after BMT and found that levels were significantly higher in recipients of IL-6R -/- T cells (Fig 1C). Furthermore, MCMV-IgG levels in plasma correlated with MCMV viremia (r = - 0.72, P < 0.0001) and viral loads in liver (r = - 0.68, P < 0.0001). MCMV-IgG2a, which can only be generated by B6D2F1 recipients, was significantly higher in recipients of IL-6R -/- T cells, confirming that differences in humoral responses were of recipient-origin. To define relevant mechanisms, we examined the kinetics of IgG after BMT by monitoring the loss of murine IgG (transferred on day 0) in plasma (Fig 1D). Recipients of IL-6R -/- T cells showed significantly slower loss of IgG than recipients of WT T cells, with or without GVHD prophylaxis with cyclosporine (CSA). Thus, ablation of IL-6 signaling in donor T cells is associated with reduced loss of recipient IgG and protection from MCMV reactivation, an outcome which seems independent of the effects of IL-6 on GVHD.We correlated our findings with data from a recent randomized, placebo-controlled, double-blind phase III clinical trial of TCZ administration on Day -1 of BMT (Kennedy GA et al, Blood 2021). TCZ reduced CMV reactivation in at risk seropositive BMT recipients of volunteer unrelated donor grafts regardless of acute GVHD (all at risk recipients, TCZ vs. control: 12/26 vs 21/28, P = 0.03; at risk without grade 2-4 GVHD, TCZ vs control: 6/18 vs 9/13, P = 0.03). TCZ did not alter the frequency of B cells or the frequency and function of HCMV-specific CD8 + T cells, quantified by HCMV-pMHC tetramer-staining and HCMV peptide-specific cytokine (IFNγ/TNF) secretion. In contrast, levels of HCMV-specific IgG at day +30 in HCMV-seropositive recipients were significantly higher in the TCZ versus control group (Fig 1E). Consistent with our preclinical studies, the level of HCMV-IgG significantly and inversely correlated with early HCMV reactivation (within 35 days) after transplant.Conclusion:These data confirm the importance of recipient-derived humoral immunity in controlling early CMV reactivation in clinical BMT recipients. Critically, we demonstrate the ability of IL-6R blockade to maintain protective humoral responses until effective donor-derived adaptive immunity can be generated. [Display omitted] DisclosuresBoeckh: Merck: Consultancy, Research Funding; SymBio Pharmaceuticals: Consultancy; Helocyte: Consultancy; Evrys Bio: Consultancy; Moderna: Consultancy; Gilead: Consultancy, Research Funding; AlloVir: Consultancy; GSK: Consultancy. Hill: Generon corporation: Consultancy; NapaJun Pharma: Consultancy; Compass Therapeutics: Research Funding; Syndax Pharmaceuticals: Research Funding; Applied Molecular Transport: Research Funding; iTeos Therapeutics: Consultancy, Research Funding; Roche: Research Funding; Neoleukin Therapeutics: Consultancy.

Cytomegalovirus Disease in Renal Transplanted Patients: Prevalence, Determining Factors, and Influence on Graft and Patients Outcomes.

The prevalence and the factors related to cytomegalovirus (CMV) disease (CMVd) during the 1st year of renal transplantation (RTx) and the relationship between CMVd and early and long-term graft and RTx-patient (RTx-p) survival were evaluated. In 505 RTx-p, followed up for 8(5–11) years, data were recorded after 1-(T1) and 12-(T12) months of RTx. CMVd was defined either by CMV replication without clinical signs of disease (CMVr, 43%), or CMV replication with signs of disease (CMVs, 57%). During the 1st year of RTx, 45% of RTx-p had CMVd (CMVd+). CMVd+ patients were older than CMVd− patients. Female gender and Donor CMV-IgG+ (CMV IgG−D+)/recipient IgG- (CMV IgG−R-) status were more prevalent in CMVd+. At T1, CMVd+ had lower albumin, haemoglobin, and higher uric-acid and reactive C-protein than CMVd− and, at T1 and T12, received more steroids. Albumin-T1 was the unique factor in determining CMVd+, maintaining its significance also after the inclusion of IgG−D+/IgG−R− status to the model. CMVs had higher prevalence of CMV IgG-D+/IgG-R- than CMVr. CMVd, CMVr, and CMVs had no impact on graft loss (11% of RTx-p) and RTx-p death (8% of RTx-p). CMVd is highly prevalent during the 1st year of RTx. Albumin-T1 influences CMVd insurgence. CMVd did not impact on RTx and RTx-p loss.

Comparing the efficacy of three techniques to reduce isoagglutinin titers in AB0 incompatible kidney transplant recipients

ABO incompatible (ABOi) organ transplantation requires pre-transplant reduction of the recipient's IgG and IgM isoagglutinin titer against the donor to prevent hyperacute rejection. Over the past four years we primarily used unspecific IgG immunoadsorption (IA) for this purpose and combined this selectively with membrane filtration (IAc) to reduce IgM isoagglutinines. In patients with an initial IgG titer against donor below 1:64, plasma exchange (PE) was initiated.In this retrospective analysis covering January 2012 to August 2015 we compared how efficiently IgG and IgM isoagglutinines in a total of 22 ABOi kidney transplant recipients were reduced by either IA (n = 75 sessions), IAc (n = 14 sessions) or PE (n = 40 sessions).Median pre-treatment IgG isoagglutinin titers were 32 (4–4096) while IgM titers were 16 (1–256) respectively. Mean IgG reduction by either treatment modality was 1.3 ± 0.9 (IA), 1.8 ± 1.0 (IAc) and 2.6 ± 1.3 (PE) titer steps per session (p < 0.001 IA vs. PE; p < 0.04 PE vs. IAc). Mean IgM reduction was 0.6 ± 0.6 (IA), 1.8 ± 0.8 (IAc) and 2.4 ± 1.9 (PE) titer steps (p < 0.001 for both IA vs. PE and IA vs. IAc).Our data indicate that PE efficiently removed IgG- and IgM isoagglutinines. By processing only half the plasma volume per treatment PE was twice as effective as IA in terms of IgG-type isoagglutinin removal in our patient group. This is best explained by the presence of soluble AB0 antigens in the FFP used as plasma replacement. These advantages in efficacy have to be weighed against the potential hazards of PE. Combination of IA and plasma filtration effectively removes IgM-type and even enhances net IgG-type isoagglutinin elimination compared to IA alone. When trying to avoid PE, combined application of IA and IAc is a possible and effective way to reduce isoagglutinin titers before ABOi transplantation.

A prospective study of renal transplant recipients reveals an absence of primary JC polyomavirus infections

BackgroundBoth JC polyomavirus (JCPyV) and BK polyomavirus (BKPyV) are acquired at an early age. JCPyV causes progressive multifocal leukoencephalopathy and has been described in association with nephropathy. ObjectivesUrine and plasma samples from renal transplant recipients (RTRs) were examined for JCPyV to determine its involvement in causing infection and disease. Study designJCPyV testing was performed on 112 RTRs included in a randomised controlled study of steroid-sparing immunosuppressive regimens [1]. Urine and EDTA blood samples were collected pre- and post-transplantation and analysed for JCPyV using real-time PCR and sequencing to determine genotype and viral variation. Donor and recipient IgG antibody status to JCPyV was also determined. ResultsOverall, 13.3% of RTRs were positive for JCPyV of which one patient developed viraemia without viruria. JCPyV DNA was detected early following transplantation (defined as five days post transplantation) from recipients with donors that were positive for JCPyV IgG antibodies. No dual cases of JCPyV and BKPyV were observed. One patient sample had sequence duplication in the non-coding control region. ConclusionsLike BKPyV, JCPyV tends to occur early post transplantation but did not result in sustained viraemia. There was no deterioration of renal function in patients positive for JCPyV. As with other viruses, JCPyV donor serostatus was a risk factor for detection of JCPyV DNA. JCPyV appears to protect individuals from BKPyV infection, as recipients were twice as likely to develop BKPyV with a negative JCPyV donor.

Human and Porcine Islet Transplants in a New Double Capsule

Our goal is to improve islet transplantation for patients with Type I diabetes mellitus. We have studied encapsulated human and adult porcine islets (APIs) in diabetic NOD mice. In previous experiments with our single microcapsule, recipient IgG was detected on rejected donor islets within capsules. Therefore, we have developed a new double capsule that excludes IgG, postulating that double encapsulation will be more protective than single encapsulation, allowing less immunosuppression.

Unique processing pathways within recipient antigen-presenting cells determine IgG immunity against donor platelet MHC antigens

Recipient IgG immunity against leukoreduced donor platelets is dependent on indirect T-cell allorecognition and is suppressed in vivo by inhibitors (aminoguanidine, AMG) of inducible nitric oxide synthase (iNOS). To examine recipient processing pathways of donor platelet antigens, enriched macrophages (antigen-presenting cells [APC]) from BALB/c (H-2(d)) mice were pulsed with allogeneic C57BL/6 (H-2(b)) platelets and transfused weekly into naive BALB/c mice. Platelet-pulsed APC stimulated IgG antidonor antibody production in 45% of recipients by the second transfusion and in 100% by the sixth transfusion; this response was enhanced by pulsing in the presence of interferon-gamma. By the sixth transfusion, high-titer IgG1 (mean titer 4990) and IgG2a (1933) isotypes specific for donor major histocompatibility complex (MHC) class I antigens were detected. Platelet pulsing in the presence of AMG or colchicine significantly inhibited the ability of APC to stimulate IgG alloantibodies; only 50% (P <.005) and 20% (P <.0001) of recipients, respectively, produced antibodies by the sixth transfusion. AMG inhibition was reversed by the addition of L-arginine, the substrate for iNOS. In contrast, pulsing in the presence of chloroquine, the proteasome inhibitory peptide MG115, or Brefeldin A enhanced APC immunity (70-100% of recipients antibody positive by the second transfusion [P <.05]); these agents allowed the pulsed APC to stimulate IgG2a but inhibited IgG1 production and this correlated with a reduction in serum interleukin (IL)-4 levels. The results suggest that for donor platelet antigens to stimulate IgG alloantibodies, recipient APC use the essential generation of nitric oxide and a noncytosolic, pH-independent processing pathway, which can be exploited as an effective immunotherapy target to further inhibit alloimmunization against leukoreduced platelets. (Blood. 2000;95:1735-1742)

Flow cytometric crossmatching in renal transplantation - outcome after five years

Abstract The association of a positive flow cytometric crossmatch between recipient IgG directed against donor T lymphocytes and poor out come is well described in renal transplantation. Until now, no long-term follow-up on such patients has been available. A total of 117 renal transplant patients were followed up for a period of 5 years. Of these, 21 were known to have donor T cell-directed IgG and 5 had B lympho cyte-directed IgG. Both groups of patients with these antibodies had a significantly poorer outcome at 5 years than did the group of patients without IgG (P < 0.0001 Han del Maenzel test). Patients with antibody detected preoperatively were tested again, either at the time of graft failure or at 5 years posttrans plantation. The sera were tested against stored donor cells and the intensity of surface IgG compared with the preoperative levels. In those recipients who lost their grafts, the levels increased in 60 % of cases but those that retained their grafts also had an increase in levels of do nor-directed antibody in 50 % of cases. The changing levels of anti body therefore appeared to have little relevance to outcome. However, when IgG isotypes were considered, for those who experienced graft failure and also had a γ3 isotype, a rise in IgG was demonstrated in all cases. Conversely, successful grafts with γ3 had a decline in levels between preoperative and 5–year sam ples in three of the four cases (p not significant).

Flow cytometric crossmatching in renal transplantation--outcome after five years.

The association of a positive flow cytometric crossmatch between recipient IgG directed against donor T lymphocytes and poor outcome is well described in renal transplantation. Until now, no long-term follow-up on such patients has been available. A total of 117 renal transplant patients were followed up for a period of 5 years. Of these, 21 were known to have donor T cell-directed IgG and 5 had B lymphocyte-directed IgG. Both groups of patients with these antibodies had a significantly poorer outcome at 5 years than did the group of patients without IgG (P < 0.0001 Handel Maenzel test). Patients with antibody detected preoperatively were tested again, either at the time of graft failure or at 5 years posttransplantation. The sera were tested against stored donor cells and the intensity of surface IgG compared with the preoperative levels. In those recipients who lost their grafts, the levels increased in 60% of cases but those that retained their grafts also had an increase in levels of donor-directed antibody in 50% of cases. The changing levels of antibody therefore appeared to have little relevance to outcome. However, when IgG isotypes were considered, for those who experienced graft failure and also had a gamma 3 isotype, a rise in IgG was demonstrated in all cases. Conversely, successful grafts with gamma 3 had a decline in levels between preoperative and 5-year samples in three of the four cases (p not significant).

Flow cytometric crossmatching in renal transplantation—the long-term outcome

The association of a positive flow cytometric crossmatch between recipient IgG directed against donor T lymphocytes and poor outcome is well described in renal transplantation. Until now no long-term follow-up on such patients has been available. In this study, 117 renal transplant patients were followed up for a period of 5 years. Of these 21 were known to have donor T cell directed IgG and five had B lymphocyte directed IgG. Both groups of patients with these antibodies had a significantly poorer outcome at 5 years than did the group of patients without IgG ( p < 0.0001, Handel Maenzel test). Patients with antibody detected preoperatively were tested again either at the time of graft failure or at 5 years post-transplantation. The sera were tested against stored donor cells and the intensity of surface IgG compared with the preoperative levels. In those recipients who lost their grafts the levels increased in 60% of cases, but those who retained their grafts also had an increase in levels of donor directed antibody in 50% of cases. The changing levels of antibody therefore appeared to have little relevance to outcome. However, when IgG isotypes were considered, in those who experienced graft failure and also had a γ3 isotype, a rise in IgG was demonstrated in all cases. Conversely, successful grafts with γ3 had a decline in levels between preoperative and 5-year samples in three of the four cases (not significant).

A retrospective flow cytometric crossmatch study in transplant recipients with autoreactive lymphocytotoxic antibodies

Our previous data shows renal transplant recipients with autoreactive lymphocytotoxic antibodies to have a reduced transplant survival when compared to patients without autoantibodies. This could have been due to the presence of weak IgG antibodies inhibited by the dithiothreitol used to remove IgM antibodies in the pretransplant cytotoxicity crossmatch. That possibility was investigated in a retrospective study of 52 recipients of 57 renal transplants who were recrossmatched using a more sensitive flow cytometry crossmatch (FCXM) to detect recipient IgG antibodies to donor T and/or B cell splenic lymphocytes. Fourteen of the 57 (24%) transplants failed. Six losses were within the 1st month posttransplant and four of these were immunological failures. None of the transplant failures had a positive pretransplant FCXM. These results showed that the recipients with autoantibodies did not have pretransplant IgG anti-donor antibodies. The transplant failures did not, therefore, relate to the presence of antibodies undetected by the dithiothreitol-treated cytotoxicity crossmatch.

The association between a positive T and B cell flow cytometry crossmatch and renal transplant failure

Between January 1989 and June 1991 329 consecutive cadaveric renal transplants were carried out at this centre and, of those, 36 (10.9%) have failed. In order to assess whether the use of flow cytometry crossmatching (FCXM) would have predicted these failures, we carried out a retrospective T and B cell FCXM study comparing the failure group with a control group of 30 recipients carefully selected from patients transplanted during the same period. The number of first and second transplants in the control and failure groups was 25,5 and 23,8 respectively with six of the controls and three of the failures having panel reactive lymphocytotoxic antibodies > 50%. Stored donor material was available for 31 of the 36 failures. Two colour FCXM was performed using R-phycoerythrin-conjugated antihuman CD3 and antihuman CD19 to identify T and B cells respectively. For each recipient, three pretransplant and one post-transplant sera were tested against lymphocytes from the recipient's kidney donor. A fluorescein isothiocyanate conjugated F(ab′) 2 rabbit antihuman IgG was used to detect recipient IgG alloantibodies bound to donor T and/or B cells. There were no T or B cell FCXM positive (+) results in the control group whereas 11 of the 31 (35%) failures overall and nine of the 23 (39%) who failed within three months were T and B cell FCXM+ pretransplant ( p = 0.0002). Seven of the nine FCXM+ results in the early failure goup were with historic sera and two with historic and current sera. The latter two were the only early failures also FCXM+ post-transplant, suggesting a direct relationship between the antibodies detected by FCXM and transplant failure. Both of these cases could be considered ‘high risk’; one was a retransplant and the other a sensitized multiparous patient. Data on rejection episodes was available for 20 of the 23 early failures. For the 13 FCXM - recipients the mean number of rejection episodes was 0.8 compared with a significantly higher mean of 2.6 for the seven FCXM+ recipients ( p = 0.02). There was no association between FCXM results and immediate nonfunction of the transplants. On the basis of our results we recommend FCXM as essential prior to renal transplantation for high-risk patients such as the retransplant and the sensitized patient who were FCXM+ pre- and post-transplant in this study.

Inhibition of the Mixed Lymphocyte Culture Response by Antibody Following Successful Human Renal Transplantation

Immunoglobulin G, appearing after several months in the serum of a recipient of a successful kidney transplant from a closely matched sibling donor, was demonstrated to progressively inhibit unidirectional mixed lymphocyte cultures when donor lymphocytes were used either in responding or stimulating cell populations. The active recipient IgG had no effect in cultures in which donor cells were not used, nor did IgG obtained from other individuals show nonspecific inhibitory effects on cultures containing donor cells. It is suggested that the MLC inhibitory immunoglobulin may serve an immunoregulatory function after renal transplantation.