James C. Cicciarelli

Comparison of renal allograft outcomes in combined liver-kidney transplantation versus subsequent kidney transplantation in liver transplant recipients: Analysis of UNOS Database.

Background. There may be an allograft-enhancing effect by the liver on the renal allograft in the setting of simultaneous combined liver-kidney transplantation (CLKT) from the same donor. This study was performed to investigate whether an existing liver allograft could protect a kidney allograft from immunologic injury due to histoincompatibility in liver transplant recipients who received sequential kidney transplantation (KALT). Methods. Using the United Network for Organ Sharing database covering January 1996 to December 2003, outcomes of 352 KALT were compared to 1,136 CLKT. Incidence of acute and chronic rejection and rejection-free renal graft survival was compared between two groups. Results. Renal half-life of KALT allografts was shorter than CLKT group (6.6±0.9 vs. 11.7±1.3 years, P<0.001). Incidence of chronic rejection in KALT group was higher than CLKT group (4.6 vs. 1.2%, P<0.001). One and three-year rejection-free renal graft survival of KALT and CLKT groups were different (77% and 67% KALT vs. 85% and 78% CLKT, respectively; P<0.001). Among human leukocyte antigen mismatched and sensitized patients, rejection-free renal graft survival of KALT group was inferior to the CLKT group (75% at 1 year and 61% 3 years vs. 86% at 1 year and 79% 3 years, P<0.001). Conclusion. Liver allograft provided renal graft immunoprotection if both organs are transplanted simultaneously (immunogenetic identity), but not for kidneys transplanted subsequently.

Hyperacute and acute kidney graft rejection due to antibodies against B cells

Because of the perception of its uncertain clinical significance, the B cell crossmatch is not universally performed before renal transplantation. Even though sporadic cases of hyperacute rejection associated with B cell antibodies have been reported, doubts remain in light of other studies suggesting no effect on graft survival. This report describes 4 cases of graft rejection (3 hyperacute and 1 acute) that occurred in patients with anti-B-cell antibodies specific against donor HLA-DR or DQ antigens. Absence of anti-donor class I antibodies was confirmed in all cases by 2-color flow cytometry. Strong evidence for an antibody-mediated mechanism was found in one patient with anti-class I and anti-class II antibodies in serum transplanted with a class II mismatched kidney. In this case, only anti-class II antibodies were recovered in the eluate of the nephrectomy specimen. These four cases were compiled from three different institutions over a four-year period, which confirms the infrequent occurrence of these events. While anti-class II antibodies may not always be detrimental for graft survival, these results also confirm that they have the potential to cause hyperacute or acute graft loss. We conclude that the information provided by the B cell crossmatch should be available at the time that a decision to proceed with a renal transplant is made.

Risk factors for short- and long-term survival of primary cadaveric renal allografts in pediatric recipients: a UNOS analysis.

Background. Pediatric kidney graft survival rates have improved in the United States. This study evaluates early and late risk factors for cadaveric graft loss in pediatric recipients. Methods. From January 1994 to December 2002, 2,597 primary cadaveric kidney-alone transplants (donor age 5-45 years, recipient age 2–20 years) were reported to the United Network for Organ Sharing (UNOS). The analysis includes follow-up information based on OPTN data as of October 14, 2003. Odds ratio of early graft loss and relative risk of late graft loss are estimated using logistic regression and Cox proportional hazards model, respectively. Results. Graft survival rates significantly improved during 1999–2002 (95% and 79% at 1-year and 3-years, respectively) compared with those of 1994–1998 (88% and 76% at 1-year and 3-years, respectively) (log rank P = 0.02). After adjusting for other variables, the factors that significantly affected early transplant outcome adversely within 3 months posttransplant were prolonged cold ischemia time (>36 hours, odds ratio [OR] = 3.38 vs. 0–36 hours) and young recipient age (2–5 years old, OR = 2.02 vs. 6–12 years). Beyond 3 months, significant risk factors were African-American recipients (relative risk [RR] = 1.93 vs. others), teenage recipients (13–20 yrs, RR = 1.50 vs. 6–12 yrs), and patients with focal glomerulosclerosis (FGS) (RR = 1.27 vs. others). Conclusions. The short-term graft survival rate of pediatric cadaveric kidney transplants has significantly improved, yet the long-term outcome has changed little. The long-term outcomes for teenagers (13–20 yrs), patients with FGS, and African-Americans lag significantly behind other groups. In order to improve long-term graft survival in these high-risk patients, newer preventive or treatment strategies must be developed.

The effect of zero HLA class I and II mismatching in cyclosporine-treated kidney transplant patients

The effect of HLA matching on one-year first cadaver donor graft survival rates between best and worst matches was 6% (P < 0.001) for A, B; 7% (P < 0.001) for DR; 9% (P < 0.001) for A, DR; 15% (P < 0.001) for B, DR; and 17% (P < 0.001) for A, B, DE. For second cadaver donor grafts, the differences were comparable. Analysis of the cyclosporine-treated patients separately yielded similar results: 5% (NS) for A, B; 7% (P < 0.001) for DR; 13% (P < 0.001) for A, DR; 16% (P < 0.001) for B, DR; and 18% (P < 0.001) for A, B, DR. The most significant effect of matching was achieved by zero mismatching B and DR antigens. The one-year graft survival for patients with zero A, B, DR mismatch was 88% with cyclosporine. Without cyclosporine, zero mismatched A, B, DR grafts survive at 84%; this difference is not statistically significant. Zero mismatching for class I and II antigens (that is, A, DR or B, DR with cyclosporine) gives one-year graft survivals of 84% and 87%, respectively. The zero mismatching HLA class I and II antigen effect is lost ‘when even one antigen is mismatched. Transfusions improved the one-year graft survival 10% in cyclosporine-treated patients, but not in those “who were not treated with cyclosporine. Seventy-one patients transfused with more than 4 units of blood, zero B, DR mismatched, and treated with cyclosporine had a 91% one-year graft survival. Recipient pool sizes for obtaining zero A, B, DR or B, DR mismatched donors are calculated. Zero A, B, DR mismatched patients can be transplanted at a 19% frequency with a 10,000 recipient pool. The success rate for zero mismatching of class I and class II antigens indicates that kidney sharing and large recipient pool sizes are a reasonable policy.

Kidney transplantation in children and adolescents: an analysis of United Network for Organ Sharing Database.

Specific pediatric allocation schemes can not only lead to minimization of waiting time, but also to better clinical outcomes for children with end-stage renal disease. The outcome of 4125 deceased donor kidney transplants (DDKT) aged 5-35 years were compared with those of 6456 living donor kidney transplants (LDKT) using univariate and multivariate Cox regression analyses. Unadjusted graft survival rates of DDKT were significantly lower than those of LDKT (hazards ratio [HR] = 1.53; P < .001). Chronic rejection was reported in 416 (10.1%) of 4125 in the DDKT group compared with 537 (8.3%) of 6456 in the LDKT group (P < .001). Among African American recipients, 67 (3.4%) grafts were lost due to noncompliance as a contributory cause of failure compared with 126 (1.5%) among other races (P < .001). A significantly lower incidence of noncompliance was observed in children (0.9%) compared with adolescents (2.2% in ages 10-14; P < .001) and high teens (2.0% in ages 15-20; P < .001). Multivariate analysis showed that adjusted graft survival rates of LDKT were superior to DDKT (HR = 1.22; P < .001) after adjusting for recipient race, recipient age, regraft status, and HLA mismatch. The differences of long-term graft survival rates between DDKT and LDKT have not been reduced (4% at 1 year, 10% at 3 years, and 12% at 5 years for unadjusted survival rates and 3% at 1 year, 6% at 3 years, and 9% at 5 years adjusted survival rates). In our analysis presented here the difference in graft survival between LDKT and DDKT has doubled compared with earlier analysis. Therefore, we recommend LDKT whenever possible as a first choice for pediatric transplant recipients.

Impact of hepatitis C infection on outcomes after heart transplantation.

Background. Prevalence of hepatitis C infection (HCV) among heart transplant (OHT) recipients ranges from 7% to 18%. Despite the paucity of data regarding the outcomes of heart transplant recipients who are HCV positive before transplant, many transplant centers are declining to perform OHT in HCV-seropositive patients. Methods. We assessed the clinical outcome of HCV-seropositive compared with HCV-seronegative heart transplant recipients using the Organ Procurement and Transplant Network/the United Network for Organ Sharing database. Between January 1, 2000, and December 31, 2005, 224 HCV-seropositive and 10,406 HCV-seronegative recipients who received HCV-seronegative donor organs were identified. Results. Overall patient survival rates of HCV-seropositive recipients were significantly lower than those of HCV-seronegative recipients (84.8% at 1 year, 77.1% 3 years, 68.9% 5 years for HCV-seropositive group vs. 87.9% at 1 year, 80.7% 3 years, and 74.1% 5 years for HCV-negative recipients, log rank P=0.036). However, adjusted relative risk of recipient HCV-seropositive versus HCV-seronegative status did not reach to statistical significance level (relative risk=1.23 with P=0.087) after adjusting for other donor and recipient factors. Causes of death among HCV-seropositive and HCV-seronegative groups were similar. Cumulative incidence of an acute rejection episode in the first year after transplantation among HCV-seropositive recipients was 35.7% versus 32.6% HCV-seronegative recipients (P=0.32). Conclusions. A more rational approach should be developed for the management of HCV-seropositive heart transplant candidates. Carefully selected HCV-seropositive patients should not be excluded from OHT.

Blood transfusions and HLA matching--an either/or situation in cadaveric renal transplantation.

Cyclosporine-treated recipients of primary cadaver donor renal transplants had a one-year graft survival rate of 79% if they received pretransplant blood transfusions (n = 5308). The one-year survival rate for nontransfused recipients (n = 709) was significantly lower at 69% (P less than 0.001). The transfusion effect was larger in black recipients (a 17% difference) than in white recipients (5%). The effect was also larger in recipients of grafts poorly matched for HLA-A, B, -B, DR, or -DR antigens than in recipients of well-matched grafts. Transfusions did not significantly improve graft survival in recipients with zero or one HLA-A, B or -B, DR, or zero -DR-mismatched grafts. However, transfusions accounted for increases of 10%, 14%, and 17% in patients receiving grafts mismatched at 2, 3, or 4 HLA-B, DR antigens, respectively. Several factors including cyclosporine and HLA matching have contributed to improving graft survival rates in nontransfused recipients. Sensitization was noted in 20% of transfused patients awaiting primary renal transplants in Southern California, as compared with 10% in transplanted patients, suggesting a tendency to transplant nonsensitized patients. Of the sensitized patients, 75% were female. Based on these data, we suggest that high survival of primary kidney allografts in the cyclosporine era can best be maintained by the continued use of pretransplant transfusions for the majority of recipients--or, alternatively, by HLA matching for patients who are at higher risk of becoming sensitized.

Effects of mycophenolic acid on highly sensitized patients awaiting kidney transplant

BACKGROUND 10-30% of dialysis population awaiting renal transplantation is sensitized. Mycophenolic acid (MPA) has been shown to reduce panel reactive antibody (PRA) formation in kidney transplant recipients. Our aim was to investigate whether MPA could effectively reduce anti-HLA antibody levels and allow successful transplantation. METHODS A total of 40 highly sensitized patients were treated orally with MPA. All patients had T-cell PRA values greater than 30% (73% of patients were ≥ 75%). The PRAs, T-cell/B-cell flow cytometry crossmatch (FCXM) mean channel shift (MCS), patient/graft survival, acute rejection, and serum creatinine (SCr) were recorded. RESULTS All 40 patients showed a decrease in PRA levels. Eighteen of the 40 patients (40%) received a transplant. All four living donor recipients converted to a negative crossmatch. There was a significant decrease in FCXM MCS in all 18 transplanted patients. The mean SCr at 24 months was 1.00 ± 0.25mg/dL. Five patients (28%) experienced acute rejection. The overall one year actuarial patient and graft survival were 94% and 88%, respectively. CONCLUSIONS These findings suggest that MPA therapy is effective in reducing PRAs and increases the likelihood of successful transplantation in sensitized recipients in a potentially simpler and more cost effective manner than the current regimens employed.

Prospects for Designing ‘Universal’ Stem Cell Lines

Successful transplantation of conventional tissues between individuals requires matching of human leukocyte associated antigens (HLA), in order to prevent rejection. Although the same principles apply to tissues differentiated from embryonic stem (ES) cells, recent advances in gene delivery and genetic regulation have raised the prospect of engineering grafts with reduced levels of HLA expression. This strategy may mitigate the effects of extensive HLA polymorphism which restricts the availability of suitable donors and necessitates the maintenance of large donor registries. Here, we discuss the potential of employing RNA interference (RNAi) to knockdown HLA expression, enabling allogeneic cells to evade immune recognition. We discuss how lentivirus-mediated delivery of short hairpin RNAs (shRNA) targeting pan-class I and allele-specific HLA achieves efficient, dose-dependent reduction in surface HLA expression in human cells. Thus, by combining genetic engineering and regenerative medicine, RNAi-induced silencing of HLA expression has the potential to create histocompatibility-enhanced and, perhaps even, “universally” compatible cellular grafts.

1016-LBP: RosetteSep HLA – An exceptional method for isolation of lymphocytes from peripheral blood

Aim The isolation of a sufficient quantity of lymphocytes from deceased donor peripheral blood can be problematic. As an OPO HLA lab that serves six different transplant centers, we routinely perform both complement dependent cytotoxicity (CDC) and flow cytometric crossmatching (FCXM) for 15 potential renal recipients plus additional non-renal recipients. Historically, we have used T/B Lympho-Kwik to isolate lymphocytes for FCXM, but this reagent is no longer available. As a replacement method, we adopted a protocol using CD14 and CD15 magnetic beads to negatively select non-lymphocyte cells. However, the lymphocyte yield and purity was frequently insufficient, so we sought to validate a method using RosetteSep HLA Total Lymphocyte Enrichment Cocktail. Methods RosetteSep contains bispecific antibodies which crosslink unwanted cells to RBCs, forming rosettes, which are removed by centrifugation. Buffy coats were pulled from ACD blood tubes and lymphocytes were isolated by centrifugation over IsoPrep and negative selection with CD14/CD15 Dynabeads or RosetteSep. The isolated lymphocytes were counted on a hemacytometer and either used directly for FCXM or further separated by positive selection of B cells using Class II Dynabeads for CDC testing. Results The average cell yield for the RosetteSep isolations was nearly double that of the CD14/CD15 isolations, lymphocyte purity was >95%, and there was an increase in the ratio of B cells to T cells. The viability and CDC reaction of the B cells was comparable between the two methods. The results of the FCXM were also comparable between the two methods, both in terms of T and B cell MFI with negative serum (NS) and T and B cell MCS with patient sera. The increased background on B cells with NS historically seen when using Lympho–Kwik was not observed with RosetteSep. Conclusions Our method using RosetteSep is cost efficient, fast, easy to perform, and significantly increases the yield of lymphocytes compared to CD14/CD15 magnetic bead negative selection.