Effect of one-HLA-haplotype-matched and HLA-mismatched blood transfusions on recipient T lymphocyte allorepertoires

BACKGROUND: Pretransplant blood transfusion has a well-known beneficial effect on posttransplant graft survival. Recently, it has been proposed that the clinical benefit of transfusion is due to HLA-DR antigen sharing between the blood donor(s) and the recipient. Immunological studies have suggested that this might result from a functional deletion of donor-reactive cytotoxic T lymphocytes. METHODS: We investigated frequencies of alloreactive lymphocyte precursors with cytotoxic or interleukin-2-producing helper function by limiting dilution analysis in 10 renal dialysis patients before and after transfusion with fresh, allogeneic whole blood. Five patients received blood transfusions from donors matched for one HLA haplotype (or one HLA-B-DR antigen) and the other five patients received blood from fully HLA-mismatched donors. RESULTS: Contrary to some previous reports, frequency analysis of cytotoxic T lymphocyte precursors revealed no significant differences between the two treatment groups in terms of development of blood donor-specific hyporesponsiveness after transfusion. Split-well analysis of cytotoxic T lymphocyte precursors reactive with single-mismatched HLA antigens demonstrated that the effects of transfusion on alloreactive specificity are complex and may vary depending on the particular antigens mismatched between the recipient and blood donor. Analysis of donor-specific helper T lymphocyte precursor frequencies revealed a significant decrease of interleukin-2-producing cells 3 months after transfusion in the total patient population. This effect was most prominent in the recipients of HLA-mismatched blood, but it also exhibited some degree of nonspecificity, as frequencies of third-party reactive helper T lymphocyte precursors were also significantly reduced. CONCLUSIONS: Our overall results suggest that the degree of HLA matching between blood donor and recipient does not greatly influence the effect of blood transfusion on the T lymphocyte allorepertoire. The apparent induced down-regulation of helper T lymphocyte activity may play a role in the reported immunosuppressive effects of allogeneic blood transfusion.     

Modulation of the T cell compartment by blood transfusion. Effect on cytotoxic and helper T lymphocyte precursor frequencies and T cell receptor Vbeta usage

Recent data suggest that the favorable effect of pretransplant blood transfusion (BT) on transplant outcome depends on the HLA match. HLA-DR or haplotype shared transfusions lead to transplantation tolerance, and HLA-mismatched BT leads to immunization. The immunological mechanism involved is still unknown. To investigate the effect of HLA compatibility between blood donor and recipient on the T cell compartment, we determined the frequency of cytotoxic and helper T cell precursors specific for blood donor cells (n=20) and the T cell receptor Vbeta (TCRBV) repertoire of the CD4- and CD8-positive peripheral blood mononuclear cells before, at 2 weeks after, and at more than 10 weeks after BT (n=10). Patients had received one transfusion of a nonstored (<24 hr after withdrawal) erythrocyte concentrate without buffy coat containing on average 6x10(8) leukocytes. Eight patients shared an HLA-B and -DR antigen, nine patients shared one HLA-DR antigen, and three patients shared no HLA class II antigens with the blood donor. All patients showed a significant increase in both cytotoxic and helper T cell precursor frequencies against the blood donor 2 weeks after BT. In most patients, the frequencies reached pretransfusion levels again long after BT. In 5 of 10 patients, an expansion of one or more TCRBV families was observed in either the CD4 or CD8 compartment. This study demonstrates that BT, irrespective of the degree of HLA matching, induces activation of the T cell compartment. The degree of sharing of HLA antigens was not correlated with quantitative changes in cytotoxic T lymphocyte precursor or helper T lymphocyte precursor frequencies, or changes induced in the TCRBV repertoire. Cytotoxic and helper T lymphocyte precursor frequencies and TCRBV repertoire determined after BT do not give an indication for a state of tolerance prior to transplantation.     

Cytotoxic T lymphocyte changes after HLA-DR match and HLA-DR mismatch blood transfusions

In the present study, we analyzed transfusion-induced cytolytic T lymphocyte (CTL) changes in patients who received either a one-HLA-DR-match or a zero-HLA-DR-match pretransplant blood transfusion. Twenty-four nonimmunized naive patients awaiting kidney transplantation received one planned, HLA-typed blood transfusion. The frequencies of CTL precursors (CTLp) directed against blood donor cells and controls were evaluated before and sequentially at days 7, 28, and 60 after transfusion. Results showed that sharing one HLA-DR between donor and recipient did not prevent CTL sensitization. Indeed, (1) An increase of donor-specific CTLp frequencies was observed in 8 of 11 patients who received a zero HLA-DR match (71%), as well as in 9 of 13 patients who received a one-HLA-DR-match (69%) transfusion. (2) This increase occurred with similar kinetics in the two groups, as it was highly significant 7 days after transfusion (P=0.002 and P=0.0035 in the first and second groups, respectively) but transient; CTLp frequencies returned to pretransfusion levels by day 60 after transfusion in both groups. (3) Finally, the magnitude of this increase was similar in the DR-match and DR-mismatch groups. Thus, if it is confirmed, in clinical practice, that one DR match between the blood donor and the patient would improve the tolerance effect of pretransplant blood transfusion, this phenomenon is unlikely to be related to the prevention of CTL sensitization.     

Acute rejection of marrow grafts in patients transplanted from a partially mismatched related donor: clinical and immunologic characteristics

Bone marrow transplantation (BMT) from a partially mismatched related donor (PMRD) provides a treatment option for patients lacking a matched sibling donor. T lymphocyte depletion of the graft reduces the risk of severe graft-versus-host disease, but may increase the risk of graft failure. We evaluated the pattern of acute graft rejection in eight patients receiving PMRD BMT with respect to the conditioning therapy, diagnosis, age and sex of donor and recipient, degree of HLA mismatch, and peripheral blood immunophenotype at the time of graft failure. All grafts were partially depleted of T lymphocytes. Marrow grafts infused into patients who experienced acute rejection did not differ significantly in nucleated cell dose, degree of T lymphocyte depletion, T cell dose, or CFU-GM/kg infused, from those received by 31 patients who showed durable engraftment. In three of four patients who rejected their grafts, and had sufficient peripheral blood cells for immunophenotyping, a CD3+CD8+ T lymphocyte phenotype was predominant at the time of acute rejection. In one patient rejection was associated with a predominant population of CD3+CD4+ T lymphocytes. Rejection was significantly associated with chronic myelogeneous leukemia and in patients mismatched by more than two antigens.     

Fatal acute myocardial infarction during general anesthesia in a 7-yr-old boy associated with total intramural coronary arteries

We evaluated the change in the percentage of cells of donor origin in pleural fluid of 13 consecutive patients who underwent lung transplantation. Pleural fluid was sampled 2, 4, and 8 days after lung transplantation. DNA, which was extracted from the blood of donors and recipients and from the pleural fluid, was amplified using a polymerase chain reaction technique. The reaction products were electrophoresed, and bands indicating amplified human leukocyte antigen (HLA)-DR alleles were quantified by determining the area under the curve (AUC) by a densitometric analysis. HLA-DR alleles, which were present only in recipient cells (recipient allele), were analyzed and compared to HLA-DR alleles that were present only in donor cells (donor allele). A dilution study was first performed to provide a standard curve relating the percentage of donor and recipient cells in a mixture to their AUC. The AUC of the recipient alleles did not change significantly over the first 8 postoperative days. The AUC of the donor alleles was less on postoperative days 4 and 8 than on day 2 (p<0.05). The donor allele AUC on day 8 was <20% of the shared allele AUC, corresponding to <1% of all cells by the dilution study. We conclude that donor cells are rapidly cleared from the pleural space after lung transplantation, with <1% of cells of donor origin by postoperative day 8.     

Transient increase in circulating donor leukocytes after allogeneic transfusions in immunocompetent recipients compatible with donor cell proliferation

Donor leukocytes in therapeutic blood components are implicated in transfusion-related complications ranging from alloimmunization to graft-versus-host disease (GVHD) to viral transmission and reactivation. To further characterize the kinetics of donor leukocyte clearance after allogeneic transfusion, we developed allele-specific polymerase chain reaction (PCR) assays directed at a single-copy Y chromosome gene and HLA class II alleles. These assays enable sensitive detection and quantitation of donor leukocytes at concentrations ranging from one cell to greater than 1,000 cells per 125 microL of recipient blood. When applied to serial samples from five consecutive orthopedic surgery patients who met study criteria, we observed 99.9% clearance of donor leukocytes over the initial 2 days posttransfusion, followed by a transient, 1-log increase in circulating donor leukocytes on days 3 to 5. This phenomenon was reproduced in a canine transfusion model, where the transient donor leukocyte expansion phase was prevented by gamma irradiation of donor blood, and was not observed after transfusions into alloimmunized dogs. We hypothesize that this transient increase in circulating allogeneic donor cells represents one arm of an in vivo mixed lymphocyte reaction, with activated donor T lymphocytes proliferating in an abortive GVHD reaction to HLA-incompatible recipient cells. Further investigation of this phenomenon should provide insight into the mechanisms involved in donor-recipient leukocyte interactions posttransfusion and the relationship of these interactions to leukocyte-induced complications.     

The use of umbilical cord blood in mismatched related and unrelated hemopoietic stem cell transplantation

Over the past 6 years, umbilical cord blood has emerged as an efficacious alternative source of hematopoietic stem cells in related bone marrow transplantation. These encouraging results led us to extend this technology to the mismatched related and unrelated settings in three high-risk leukemic children lacking a matched-related donor for transplantation. Two of the three children also lacked identifiable donors through the National Marrow Donor Program, while the third was in relapse and did not have time to wait for completion of a search. The first child was transplanted with haploidentical umbilical cord blood-derived mononuclear cells from his sister, while the remaining two children were transplanted with partially mismatched, unseparated, unrelated umbilical cord blood banked through the Placental Blood Project at the New York Blood Center. All three children demonstrated trilineage engraftment with donor cells within 6 weeks of transplantation. The patient transplanted with haploidentical marrow developed grade 2 graft vs. host disease (GVHD), which was controlled with steroid and anti-thymocyte globulin (ATG) therapy. One of the two patients grafted with unrelated umbilical cord blood developed mild grade 1 GVHD of the skin, which rapidly cleared with steroid therapy. One patient remains alive, in good health and disease-free 12 months from transplantation.     

Changes in lung and chest wall properties with abdominal insufflation of carbon dioxide are immediately reversible

There is increasing interest in blood cell transplants (BCT) from normal donors as an alternative to BMT. Ten patients with relapsed or persistent leukemia after BMT received intensive cytotoxic conditioning followed by allogeneic BCT. Three BCT were from single-antigen mismatched donors; two of the corresponding recipients had rejected a BMT from the same donor. Two patients received BCT from a different donor (one matched, one single-antigen mismatched). The other six BCT were from the same, fully matched, bone marrow donors. Donors were given G-CSF to mobilize progenitor cells which were collected by a single 2-4 h leukapheresis. Methotrexate, CsA and folinic acid were used for GVHD prophylaxis for all transplants but CsA was discontinued sooner after BCT than after BMT. One patient died without engraftment having rejected a BMT from the same single-antigen mismatched donor 4 years previously. Nine patients had granulocyte recovery at a median of 14 days, up to 6 days faster than with their previous BMT. Platelet recovery was also 2-6 days faster than with BMT in four previously engrafting patients. Four patients died without platelet recovery after BCT within a year of BMT, three of treatment-related toxicity and one of relapse. Two patients developed grade II acute GVHD. Of six patients given BCT more than a year from BMT, four, all with acute leukemia, survive 7, 14, 29 and 29 months after BCT and one relapsed at 7 months. All four survivors developed chronic GVHD. These results indicate that BCT may be useful therapy for relapse occurring more than a year after BMT.     

Partially mismatched pediatric transplants with allogeneic CD34(+) blood cells from a related donor

This was a phase I, multi-center study of 13 pediatric patients (median age, 11 years) to evaluate toxicity, hematopoietic recovery, and graft-versus-host disease (GVHD) after allogeneic transplantation of enriched blood CD34(+) cells obtained from genotypically haploidentical but partially HLA-mismatched related donors (8 parents and 5 siblings). With regard to rejection, donor HLA disparity was 1 (5), 2 (6), or 3 loci (2). With regard to GVHD, recipient HLA disparity was 0 (1), 1 (3), 2 (8), or 3 (1). The patients suffered from acute myelogenous leukemia (6), chronic myelogenous leukemia (4), acute lymphoblastic leukemia (2), or hemolytic anemia plus immunodeficiency disorder (1). To reduce the risk of graft failure through the infusion of a large amount of stem cells, peripheral blood cells (PBC) were mobilized by recombinant granulocyte colony-stimulating factor (G-CSF; lenograstim, 10 microgram/kg/d for 5 days) and collected by 2 to 5 aphereses. To both enhance engraftment and reduce GVHD, CD34(+) cells were enriched using immunomagnetic procedures with the Baxter ISOLEX 300 system (Baxter Healthcare Corp, Irvine, CA) and cryopreserved. After variable cytoreductive regimens, a median of 7.7 (range, 2.2 to 14) x 10(6)/kg of CD34(+) cells and 1.03 (0.05 to 2.09) x 10(5)/kg CD3(+) cells were infused. Using Center-specific posttransplant supportive care and immunosuppressive GVHD prophylaxis, two patients experienced early death; one from veno-occlusive disease at day 17 and one from sepsis at day 18. Nine of 11 patients showed signs of engraftment; however, subsequent rejection was seen in 4 patients, 2 of whom had autologous recovery. Eight patients were evaluated in the early phase of marrow recovery. The median number of days to achieve an absolute granulocyte count of 0.5 x 10(9)/L was 14 (range, 9 to 20) and that to achieve a platelet count of 20 x 10(9)/L was 17.5 (range, 12 to 23). Donor chimerism persisted in five patients until death or current survival. All of the surviving patients with functioning-donor-type hematopoiesis were given total body irradiation. De novo acute GVHD (grades II and IV) was observed in two of the eight evaluated patients. Scheduled donor lymphocyte infusion (DLI), using the CD34(-) fraction, was administered to four patients, free of de novo acute GVHD, beginning between 28 to 43 days after transplant. Three of these patients developed acute GVHD (grades I, II, and IV). Cytomegalovirus infection was a major infectious complication but was successfully managed with gamma-globulin and gancyclovir treatment with or without additional DLI. Five patients are currently surviving, free of disease, with a follow-up ranging from 476 to 937 days. Each survivor has functioning hematopoiesis, three of donor origin and two of autologous origin. In conclusion, our results show that enriched blood CD34(+) cells from a mismatched haploidentical donor are a feasible alternative source of stem cells, but do not appear to ensure engraftment. Because none of the patients who were administered DLI survived, the therapeutic efficacy and safety of periodic DLI, as an integrated part of such transplants, needs to be clarified in further studies.     

Postoperative inflammatory response after autologous and allogeneic blood transfusion

BACKGROUND: Allogeneic blood transfusions cause immunosuppression. The aim of this study was to determine whether complement anaphylatoxins, cytokines, or both are released in the recipient, after blood transfusions in general, and after autologous blood transfusions in particular. METHODS: Thirty-one patients having total hip joint replacement surgery were randomized to receive either allogeneic red blood cells (n = 15) or predeposited autologous whole blood transfusion (n = 16). Plasma concentrations of the anaphylatoxins C3a and C5a, the terminal C5b-9 complement complex, and cytokines IL-6 and IL-8 in the recipients were repeatedly analyzed before, during, and after surgery. RESULTS: Significantly increased concentrations of IL-6 and IL-8 appeared in both groups, with a significantly greater increase in the autologous blood group. Patients in both groups developed a moderate but significant increase of C3a without a significant difference between them. C5a and terminal C5b-9 complement complex were not greatly changed. CONCLUSIONS: The study showed a greater increase in cytokine concentration after autologous blood transfusion than after allogeneic blood transfusion. The lower response in the latter may result from transfusion-induced suppression of cellular immunity.     

Epstein-Barr virus transmission from a blood donor to an organ transplant recipient with recovery of the same virus strain from the recipient's blood and oropharynx

A previous study (Savoie et al, Blood 83:2715, 1994) identified eight transplant patients who acquired Epstein-Barr virus (EBV) infection during the peritransplant period. Three of these patients subsequently developed B-cell lymphoproliferative disease within 4 months of transplantation. Among these, there was a 16-year-old liver transplant patient who was negative for EBV at the time of transplant and who received an EBV-negative organ. After transplant, this patient was transfused with 9 U of packed red blood cells. Eight of the donors were EBV-positive and one was EBV-negative. We succeeded in obtaining spontaneous lymphoblastoid cell lines (LCLs) from the blood of three of these donors, one of whom also yielded a cord-blood line established with his throat-wash EBV. Blood from a fourth donor did not yield an LCL, but his throat washing did have transforming activity when inoculated onto cord-blood leukocytes. We initially could establish spontaneous LCLs only from the recipient's blood. However, a throat-wash sample taken 11 weeks later did show transforming activity. The recipient was shown to have acquired the EBV infection from one of eight EBV-seropositive blood donors. Analysis of fragment length polymorphisms after polymerase chain reaction amplification of the EBV BamHI-K fragment was used to establish strain identity. Western blot analysis for existence of size polymorphisms in three classes of Epstein-Barr nuclear antigens (EBNA-1, EBNA-2, and EBNA-3) confirmed the DNA results. It is noteworthy that the blood donor responsible for transmitting his EBV strain to the recipient had experienced clinical infectious mononucleosis 15 months before donating blood. Our results may, thus, indicate a requirement for leukodepletion of blood destined for immunosuppressed EBV-negative patients. Finally, blood donors with a recent history of infectious mononucleosis should probably be identified so that their blood is not given to EBV-negative transplant patients.     

Stable mixed chimerism without relapse after related allogeneic umbilical cord blood transplantation in a child with severe aplastic anemia

Umbilical cord blood (UCB) cells from HLA-matched donors are used as an alternative to bone marrow for allogeneic transplantation and reports of successful UCB transplantation in patients with severe aplastic anemia (SAA) are scarce. SAA was discovered in a 4-year-old girl in February 1990. Transfusion support started in August 1990 and standard treatments were unsuccessful. The birth of an HLA-compatible brother in October 1993 permitted the cryopreservation of UCB. In December 1994 UCB transplantation was decided upon. No toxicity occurred. G-CSF was started at day 28. WBC and PMN reached 0.5 x 10(9)/l at days 33 and 37. RBC and platelet transfusion independence were reached at days 50 and 52. Mixed chimerism was demonstrated in blood cells at 1.5, 4 and 6 months after UCBT by molecular biology (VNTR). FISH studies yielded similar results at 15 and 18 months. Twenty months after UCBT, molecular biology showed full donor chimerism. Clinical follow-up (last follow-up: 32 months post transplant) is unremarkable. We suggest that CY and ATG may be a suitable regimen for related HLA-compatible UCBT in patients with SAA. Residual recipient cells can disappear even very late after UCBT, permitting the establishment of complete donor chimerism.     

Immunotherapy with donor leukocyte infusions for patients with relapsed acute myeloid leukemia following partially mismatched related donor bone marrow transplantation

Donor leukocyte infusions (DLI) were used to treat 2 patients with AML who relapsed within 4 months of treatment with partially mismatched related donor (PMRD) BMT representing 1-2 HLA-mismatches. No other form of cytoreductive therapy was given to these patients. Both patients developed GVHD (grade II-III) following DLI requiring steroid therapy. One of these patients went into complete remission following development of GVHD and immunophenotypic analysis of peripheral blood showed increased numbers of CD3+/CD8+ T cells, CD56+/CD8+ lymphokine activated killer (LAK) cells and CD16+/CD56+ natural killer (NK) cells expressing intermediate affinity IL-2 receptor P75. Unfortunately, the response was of short duration and the patient relapsed 8 weeks later ultimately resulting in death. The second patient did not show any response to DLI and died of progressive leukemia in conjunction with active GVHD. We conclude that DLI from PMRD carries a high risk for the development of GVHD and may have an anti-leukemia effect for relapsed AML. The anti-leukemic effect from PMRD DLI may be mediated by cytotoxic T lymphocytes, LAK cells and NK cells.     

Allogeneic transplantation combining mobilized blood and bone marrow in patients with refractory hematologic malignancies

BACKGROUND: Mobilized blood stem cells have been used successfully in autologous transplant recipients to reduce the complications of pancytopenia due to dose-intensive chemotherapy. Reports of cytokine-mobilized blood progenitor cells in allogeneic transplant recipients are rare. STUDY DESIGN AND METHODS: This is a pilot trial of six patients. Patients with advanced hematologic malignancy received bone marrow (median total 2.6 x 10(8) mononuclear cells/kg) followed by four daily transfusions of blood (median total 9.5 x 10(8) mononuclear cells/kg) from HLA-matched sibling donors who were mobilized with recombinant human granulocyte-colony-stimulating factor (5 micrograms/kg/day subcutaneously for 5 days). All patients received cyclosporine and prednisone for graft-versus-host disease (GVHD) prophylaxis. RESULTS: An absolute neutrophil count greater than 500 per mm3 was achieved on Day 12, and platelet transfusion independence was achieved on Day 16. The median day of hospital discharge was Day 23 after transplant. All patients achieved 100-percent donor cell engraftment. Acute > or = Grade III GVHD did not develop in any patients, but all patients developed Grade I (n = 4) or Grade II (n = 2) acute GVHD. Chronic extensive GVHD developed in four of six patients. One patient died of pneumonia 263 days after transplant while undergoing immune-suppressive therapy for chronic GVHD. CONCLUSION: The transfusion of blood stem cells in patients undergoing allogeneic bone marrow transplant is well tolerated soon after transplant, but the development of chronic GVHD may limit the general usage of unmanipulated blood stem cells.     

Transfusion of newly stored blood

It is shown that freshly stored stabilized blood differs in its effect on a recipient only on account of addition of stabilizors to it. The replacement effect due to transfusion of freshly preserved citrated blood is analogous to that noted in direct blood transfusion, however the response of the cardiovascular system inadequate for the transfused blood volume would restrict the rate and volume of transfusion. Freshly heparinized blood differs from donor blood by impaired coagulation factors, but it is identical in hemodynamic effect. Freshly preserved sorbent blood is mostly close in its efficacy to direct transfusions, since its content differs from native donor blood only in calcium content.     

DNA typing of recipient blood after massive transfusion

BACKGROUND: Transfusion is known to alter the recipient's blood group, protein allotype, and alloenzyme phenotype by the introduction of donor material and the dilution of his or her own cells and proteins. There is little information about typing the DNA of blood after transfusion. STUDY DESIGN AND METHODS: Ten adult patients who had undergone massive transfusion with white cell-containing blood components were studied. Pretransfusion and posttransfusion blood specimen DNA types were compared by using Southern blot and polymerase chain reaction (PCR) analyses. RESULTS: DNA types were identical in all 90 paired observations. CONCLUSION: These preliminary findings, in a limited number of patients, suggest that certain DNA methods may provide reliable typing of adult recipients after massive transfusion.     

Immunization of pigs against Streptococcus suis serotype 2 infection using a live avirulent strain

BACKGROUND: To reduce the mortality rate associated with liver transplantation, it is important to identify the risk factors for increased mortality among liver transplant recipients. It has been suggested that cytomegalovirus (CMV) infection is one such risk factor, but no studies have examined mortality rates associated with the CMV serologic status of the donor and recipient by using multivariate techniques. OBJECTIVE: To study the effect of CMV on 1-year mortality rates in orthotopic liver transplant recipients. DESIGN: Intention-to-treat analysis of a cohort. PATIENTS: 146 liver transplant recipients who were enrolled in a multicenter, randomized, placebo-controlled intervention trial. SETTING: Four university-affiliated transplantation centers. RESULTS: 1-year mortality rates for the four strata of donor and recipient CMV serologic status before transplantation were as follows: seronegative donor and recipient, 11%; seronegative donor and seropositive recipient, 22%; seropositive donor and recipient, 30%; and seropositive donor and seronegative recipient, 44% (P = 0.0091). Multivariate analysis using a time-dependent Cox proportional hazards model showed that retransplantation (relative risk, 4.6 [95% CI, 1.9 to 10.7]; P < 0.001); total number of units of blood products administered during transplantation (relative risk, 1.006 per unit [CI, 1.003 to 1.010]; P < 0.001); and presence of CMV disease (relative risk, 3.9 [CI, 1.8 to 8.5]; P < 0.001), invasive fungal disease (relative risk, 3.3 [CI, 1.5 to 7.1]; P = 0.0020), and bacteremia (relative risk, 2.5 [CI, 1.2 to 5.2]; P = 0.0136) were independently associated with higher mortality rates. If post-transplantation variables that were highly correlated with donor and recipient CMV serologic status were restricted from the model, donor and recipient CMV serologic status was the only pretransplantation variable independently associated with higher mortality rates (P = 0.002). CONCLUSION: Donor and recipient CMV serologic status is a significant pretransplantation determinant for death in liver transplant recipients.