Presence of donor specific anti-HLA antibodies (DSA) is a negative predictor of graft survival in kidney transplantation. Moreover, the presence of any anti-HLA antibodies without accessing their specificity increases the risk of kidney graft failure [
There are conflicting data regarding impact of anti-MICA antibodies on acute rejection episodes and kidney graft survival [
The 14th International HLA and Immunogenetics Workshop Prospective Chronic Rejection Project was an international collaborative study of 45 transplant centers to assess impact of anti-HLA and anti-MICA antibodies detected after transplant on chronic graft failure. The 1-year follow-up data published in 2007 showed significant impact of anti-HLA antibodies on kidney graft survival [
We present 7-year follow-up data and in-depth clinical and pathological analysis of 123 liver transplant recipients from our center participating in this project.
Blood samples were collected from 123 liver transplant recipients who were at least 6 months posttransplant between September and November 2005 during the patients’ routine visits in the Outpatient Department of Transplantation Institute Medical University of Warsaw (MUW). Time from transplantation to blood sample collection was different for each patient. Liver transplantations were performed in the Department of General, Transplant and Liver Surgery MUW, between June 1999 and January 2005 in most cases. Only 4 transplantation cases were performed earlier: 1 in 1995, 1 in 1996, and 2 cases in 1997. All liver transplants were blood compatible but were performed without preoperative cross-match test. Patient death, graft failure, clinical and biopsy-proven rejection episodes, and liver function tests were recorded prospectively during the 7-year follow-up period. Causes of patients’ deaths were arbitrary divided as “nonimmunological” (cancer, cardiovascular diseases, and HCV related cirrhosis) and “immunological.” Retrospective data concerning liver disease before transplantation, clinical and biopsy proven rejection episodes before entrance to the study, type of immunosuppression (basiliximab induction, type, and number of immunosuppressive drugs), and HBV and HCV infection status were taken from patients’ medical records. Liver diseases before transplantation were arbitrary divided as “immunological” (autoimmune hepatitis, primary sclerosing cholangitis, and primary biliary cirrhosis) and “nonimmunological.” Acute graft rejection episode during first 6 months after transplantation was categorized as early rejection. Clinically suspected acute graft rejection episodes were diagnosed based on sharp elevation of liver enzymes which normalized after treatment with methylprednisolone pulses and/or with an increase in tacrolimus dose. HBV infection was diagnosed on the basis of repeated presence of anti-HBc antibodies and HCV infection diagnosis was based on positive anti-HCV antibodies immunoenzymatic assay.
Blood samples were tested for the presence of anti-HLA class I and II antibodies using Luminex kits (One Lambda, Inc., Canoga Park) and anti-MICA antibodies were tested using Luminex assays with MICA *001, *002, *004, *007, *012, *018, *019, and *027 antigens purified from recombinant cell line coated onto Luminex beads in Terasaki Foundation Laboratory (Los Angeles, CA). Mean fluorescence intensity (MFI) 1500–2999 was considered weak positive and 3000 or greater strong positive for MICA 001–027 antigens, except for MICA 019 antigen (weak positive 2500–3999 and strong positive 4000 or greater).
Statistical significance of differences among groups was assessed by chi-square or Fisher’s exact test. Additional analysis of the odds ratios was done by logistic model. The Wilcoxon rank-sum test was used for analysis of nonnormal data.
Anti-HLA and anti-MICA antibody results and full medical reports were available for 123 liver transplant recipients (61 women, 62 men). Three patients were lost to followup and were not included in 7-year survival analysis. Mean age at entry to the study (day of blood sample collection) was 44 (19–68) years and mean time between liver transplantation and blood collection for anti-HLA and anti-MICA antibodies was 34 (7–174) months (1 patient 125 and 1 patient 174 months). Primary liver diseases were hepatitis C (HCV) in 29 cases, toxic liver disease in 19, autoimmune hepatitis (AIH) in 12, primary biliary cirrhosis (PBC) in 12, primary sclerosing cholangitis (PSC) in 10, hepatitis B (HBV) in 9, and others in 32 cases (Wilson disease, Budd-Chiari syndrome, hemochromatosis, and cholangial malformations). Patient characteristics are listed in Table
Patients characteristics.
Anti-HLA positive | Anti-HLA negative | Anti-MICA positive | Anti-MICA negative | |
---|---|---|---|---|
Number | 33 | 90 | 37 | 86 |
Mean age (years) | 47 | 47 | 44 | 47 |
Sex F/M | 16/17 | 42/48 | 18/18 | 41/46 |
Primary liver disease* | ||||
|
5 | 7 | 5 | 7 |
|
4 | 8 | 3 | 9 |
|
2 | 8 | 2 | 8 |
|
7 | 22 | 8 | 21 |
|
2 | 7 | 2 | 7 |
|
6 | 13 | 4 | 15 |
|
7 | 25 | 12 | 20 |
Mean time since LT to blood collection (months) | 24 | 26 | 34 | 25 |
*AIH: autoimmune hepatitis; PBC: primary biliary cirrhosis; PSC: primary sclerosing cholangitis; HCV: hepatitis C virus; HBV: hepatitis B virus, LT: liver transplant.
Thirty-two patients had titers of anti-HLA antibodies (8 with both classes I and II, 16 with only class I, and 8 with only class II). Mean age of patients at the time of blood collection in anti-HLA positive group was 43 (21–60) years versus 44 (19–68) years in anti-HLA negative group and time after transplantation was 32 (7–101) months and 35 (8–174) months, respectively (
Correlation between presence of anti-HLA antibodies and time after liver transplantation.
Thirty-seven patients with anti-MICA positive antibodies included 10 patients with weak positive and 27 patients with strong positive antibodies. Mean age of patients at the time of blood collection was 44 (20–68) years in anti-MICA positive group versus 42 (19–68) years in the anti-MICA negative group (
Twenty-seven patients died during the 7-year study period. Progressive graft failure was the main cause in 16 cases whereas other medical conditions like malignancies, neuroinfection or cardiovascular disorders were the main cause of mortality in 11 patients. No retransplantations were performed in this group during the study period. The only predictors of longer patients survival in the whole group were younger age at transplantation (
Survival of patients in anti-HLA positive and anti-HLA negative groups. Graft loss occurred in 7 (23%) patients in the anti-HLA positive group and 20 (22%) in the anti-HLA negative group (
Survival of patients in anti-MICA positive and anti-MICA negative groups. Graft loss occurred in 8 patients in anti-MICA positive group (22%) and 19 (23%) in anti-MICA negative group (
Biopsy-proven early AGR were diagnosed in 23 cases and early AGR was clinically suspected in 24 cases based on retrospective data. Basiliximab induction was a negative predictor of both all and biopsy-proven cases of early AGR (
No correlations were detected between presence of anti-HLA antibodies (neither all cases nor HLA I and HLA II separately) or anti-MICA antibodies (neither positive nor only strong positive cases) with early or late AGR. In contrast, presence of any antibodies (anti-HLA or anti-MICA antibodies) correlated with late AGR (
Liver function test was performed every 3 months during patients’ routine visits in Outpatients Department. The anti-HLA positive versus anti-HLA negative groups did not differ in serum bilirubin concentration, alanine aminotransferase (ALAT), or gamma-glutamyl transpeptidase (GGTP) activity at the end of the 7-year study. Similarly, these variables did not differ between the anti-MICA positive and anti-MICA negative groups. However, mean bilirubin serum concentration was significantly higher in a group of patients with late acute graft rejection (
We analyzed the impact of anti-HLA and anti-MICA antibodies on liver graft failure and function at the 7-year followup and also the correlation between presence of these antibodies and several clinical and pathological data in 123 AB0 compatible liver transplant recipients.
Anti-HLA antibodies may be detected in early period after liver transplantation due to massive blood transfusion usually required during surgical procedures, so patients at least 6 months after operation were included in this study. Moreover, the percentage of patients with positive anti-HLA antibodies changed in a time-dependent manner in study group, suggesting that we observed true recipients’ humoral reactions to donor antigens rather than presence of preformed antibodies. Taner et al. showed that preformed donor specific antibodies (DSA) tend to disappear after liver transplantation in most cases but if persist complement activation in the liver allograft could be found in biopsy but it did not impact graft function at 1-year after transplantation [
In our analyses, no significant correlations were detected between presence of anti-HLA/anti-MICA antibodies and factors which could potentially activate humoral immune system response, like primary autoimmune liver diseases, minimalization of immunosuppression (HCV and HBV infection or HCC in explanted liver), or type and number of immunosuppressive drugs. The presence of anti-HLA (generally and separately in HLA I and II classes) and anti-MICA antibodies did not significantly correlate with 7-year graft survival or acute graft rejection in either early or late posttransplant period. In contrast, the presence of any antibodies (anti-HLA or anti-MICA antibodies) significantly correlated with late graft rejection. This observation suggests that increased activation of humoral immune system could be involved in late rejection episodes and agrees with results of a prospective study showing that DSA were found significantly more in liver transplant recipients with biopsy proven chronic rejection (36 of 39 (96%)) than in patients without rejection (24 of 39 (61%);
Limitations of this study included that antibodies were screened only once and at different time points after-transplantation for each patient in the study group; however it was caused by the methodology of the Workshop. Secondly, the presence of anti-HLA antibodies generally, but not DSA, was tested; however, availability of single-antigen Luminex assays was limited when the study was started. We are planning to reevaluate remaining patients sera for the presence of DSA.
Presence of anti-HLA or anti-MICA antibodies did not significantly correlate with inferior graft survival nor with increased incidence of acute graft rejection. In contrast, higher humoral immune system activity defined as presence of any antibodies (anti-HLA or anti-MICA antibodies) was a significant predictor of late graft rejection.
The authors declare that there is no conflict of interests regarding the publication of this paper.
The authors want to thank Dr. Paul Terasaki and Dr. Mikki Ozawa for their help in experimental procedures of this study.