American Journal of Hematology 44:168-171 (1993)

Graft Versus Host Anti-Rho(D) Following Minor Rh-Incompatible Orthotopic Liver Transplantation

Jong-Hoon Lee and Paul D. Mintz Departments of Pathology (J.-H.L., P.D.M.) and Internal Medicine (P.D.M.), University of Virginia Health Sciences Center, Charlottesville

Hemolysis caused by ABO antibodies after ABO-compatible, nonidentical solid organ transplantation has been previously reported. The passenger B lymphocytes within the donor organ presumably generate an acute, primarily red cell-directed graft vs. host (GVH) response. Graft survival may also be compromised. GVH Rh antibodies have also been described, primarily in renal transplants. Only three cases, twoonly in abstract form, have been reported thus far describing GVH Rh antibodies in liver transplant patients, to which we add a fourth. A 62-year-old blood group A Rho(D)-positive woman with cirrhosis underwent orthotopic liver transplantation from a group A Rho(D)-negative, previously Rho(D)-sensitized donor and subsequently developed acute, self-limited hemolysis re- quiring four units of packed red cells. Anti-Rho(D) was identified in both serum and red cell eluate. An antibody detection test, identification, and assessment of the antibody reactivity score from the pretransplant donor specimen may identify patients at risk for hemolysis due to GVH Rh antibodies.

Key words: donor-derived antibody, GVH disease, hemolysis

Ic’ 1993 Wiley-Liss, Inc.

INTRODUCTION

Over 100 cases of graft vs. host (GVH) antibodies directed against red cell antigens arising from solid organ transplants have been reported in the literature. The over- whelming majority have been ABO antibodies presum- ably arising from the passenger B lymphocytes contained in the ABO-unmatched organ. and the incidences of the GVH antibodies and hemolysis correlate with the size and lymphoid content of the transplanted organ [ I ] .

GVH Rh antibodies in solid organ transplantation have been reported much less frequently. We have identified 13 reported cases describing anti-Rho(D), anti-E, anti-c, and anti-e, and of these, ten have been associated with renal allografts and only three with transplanted livers. Ten of the thirteen cases were associated with hemolysis with direct antiglobulin tests reactive up to 6 months. Paradoxical Rho(D) antibody was demonstrated in all three liver transplant cases [ 1-41. We report a fourth case of GVH anti-Rho(D) arising from a group A-negative liver transplanted into a group A-positive recipient.

CASE REPORT

A 62-year-old blood group A-positive woman with cirrhosis underwent orthotopic liver transplantation from

0 1993 Wiley-Liss, Inc.

a group A-negative, previously sensitized donor. She had a history of alcohol abuse, and a serologic evaluation revealed antibodies to hepatitis B and C viruses as well as cytomegalovirus. Other laboratory studies showed the following: total bilirubin 0.8 mgidl (0.2-1.2), ammonia 32 pmol/liter (11-3S), PT 13.2 sec (10.4-12.6), hemat- ocrit 39%) (3547%), and a negative antibody detection test. Intraoperatively she required four units of Rho(D)- positive packed red cells, four units of fresh frozen plasma, one unit of single donor platelets, and 3,500 ml of crystalloids for an -2-liter blood loss. By the seventh postoperative day, intravenous cyclosporine A and meth- ylprednisolone were changed to oral formulations, and the patient was ambulatory. The 34% hematocrit on the ninth postoperative day steadily decreased to 22% over the next 48 hr, without a demonstrable source of bleed- ing. The patient was symptomatic only for a fever of 39°C. She was transfused with four units of Rho(D)-

Received for publicittion December 71. 1992; acccpted March I I , 1993.

Address reprint requeqts to .long-Hoon Lee, MD. Blood Bank and Tran5fusion Services. University of Virginia Health Sciences Center, Medical Center Box 286. Charlottesville, VA 22908.

GVH Anti-Rho(D) in Liver Transplantation 169

TABLE 1. Antibody Score in Relation to Clinical and Laboratory Data’

POD PT TBIL HCT LDH IAT DAT Titer Score ______ ~ -~~~~ __---- ~ ~ ~ ~ _ _ __-_ ~ ~ _ _ _ _ _ _

0” 13 0.5, 40 Neg 1 IS 0.7 31 2386 7h 12 1 . 1 32 I + Neg 9 I 1 2. I 34 552

10 I I I .8 21 538 3+ 3 f I :32 41 1 I‘ I 1 2.4 22 443 I 2d I I 2.6 26 765 13 12 1.8 23 496 3+ 2 + 1:32 44 15’ I 1 29 567 20 12 I .2 32 I90 26 3 + I + I :37- 44 31 I . 3 3.5 I68 33 3+ I + 1:16 36 41 3 + I + 1:16 36 52 2+ w + I :8 2 h

*Anti-Rho(D) were confirmed froni serum on day\ 7, 10. and 52 and l‘roni eluate on day 10. POD. postoperative day; PT. prothronibin time in seconds (10.412.6) ; TBIL. total bilirubin in nigidl (0.2-1.2); HCT. hematocrit in percent; LDH, lactate dehydrogenase i n Uiliter ( 2 3 W S 0 ) ; IAT, indirect antiglobulin test (antibody detection test): IIAT. direct antiglobulin test, polyspecific. “lntraoperatively transfused four units of Rho(D)-positive packed red cells: started on intravenous cyclosporine A and iiiethylprzdnisoloiie hlmmunosupprrssivc agents changed to oral formulations ‘Fever of 38.9”C. “Transfused 2 units of Rho(D)-negative packed red cells. ‘Transfused 2 units of Rho(D)-negative packed red ccllb; discharged home

negative packed red cells, recovered uneventfully, and was discharged on day 15 with a 34% hematocrit.

MATERIALS AND METHODS

Iinniunohematologic tests were performed on post- transplant blood samples from days 7, 10, 13,26. 33,41, and 52, and the results were correlated with clinical and laboratory data. Standard methods and reagents were used according to the manufacturers’ instructions as fol- lows: indirect antiglobulin test with serial dilutions (Im- mucor Inc., Norcross, GA), direct antiglobulin test (polyspecific anti-IgG/C3d Coornbs reagent; Ortho Diag- nostic Systems, Inc., Raritan, NJ), acid elution (Elukit 11; Gamma Biologicals, Inc., Houston, TX), and antibody identification (ten cell panel; Immucor). The reaction strength and the antibody titer were combined into an antibody score as a single measure of antibody reactivity as outline in the Technical Manual 151.

RESULTS

The test results in relation to the clinical and laboratory data are shown in Table 1. The anti-Rho(D) was detected in serum on day 7 and in eluate on day 10. Hemolysis, clinically apparent by day 10 and necessitating four units of packed red cells, stabilized over the next 10 days. The

anti-Rho(D) persisted at low levels of detection, and both the DAT and the IAT (titer 1:8, score 26) were positive at day 52.

DISCUSSION

The survival advantage of adult ABO-identical grafts over ABO-compatible grafts in liver transplantation has been described in the literature, but a similar decrease in survival in pediatric ABO-compatible, nonidentical liver transplants has not been observed [6]. In addition, sensi- tized passenger B lymphocytes contained within a trans- planted liver are thought to generate donor-derived ABO antibodies directed at all ABO antigens as a limited but acute GVH response [ I ] . Furthermore, in ABO-compati- ble, nonidentical solid organ transplantation, hemolysis due to minor ABO incompatibility has been shown to occur with increasing frequency in the order kidney, heart, liver, and heart-lung transplants as reviewed by Ramsey [ I ] . These data suggest that the total dose of incompatible donor lymphocytes may be the major deter- minant in solid organ GVH disease due to ABO antibod- ies. Such a phenomenon is most readily detected in the irnmunohematology laboratory through red cell compati- bility testing.

An analogous problem may exist involving the Rh blood group system. A limited number of cases describ-

170 Lee and Mintz

ing heniolysis due to passenger lymphocytes within Rho(D)-negative kidneys transplanted into Rho(D)-posi- tive recipients have been reported [ 1 1. For liver transplant patients, only three cases (two in abstract form only) have been thus far reported describing complications from a minor mismatch between an Rho(D)-negative donor and an Rho(D)-positive recipient [ 2 4 ] . In case I , a CcDee recipient of a liver from a group 0 Rho(D)-sensitized donor developed severe hemolysis and acute renal failure requiring 14 Rho(D)-negative red cell units along with splenectomy. Anti-Rho(D) and anti-E were found on postoperative day 10 and were still present at 6 weeks [2]. In case 2, a group AB-positive recipient of a liver from a group A Rho( D)-sensitized donor developed anti-Rho(D) with an antibody score of 59 [ 3 ] . Four units of red cells were transfused, presumably due to hemolysis. Only case 3 describes a primary GVH anti-Rho(D) response. A group 0-positive recipient of a liver from a group 0-neg- ative donor with a negative antibody detection test tran- siently developed a positive DAT without evidence of hemolysis [4]. To our knowledge, the current case repre- sents only the fourth to be reported and reaffirms the clinical and laboratory findings seen in the previous three cases.

Calhoun et al. [3] described the second case as a part of a larger study focused mainly on minor ABO-incompati- ble liver transplants. The authors concluded that GVH ABO antibodies occur in approximately one-third of sus- ceptible cases and tend to develop 8-10 days after trans- plantation. The donor ABO antibody score of 60 or higher showed a good correlation with antibody develop- ment, positive DAT, and red cell transfusion require- ments [3]. Other studies on the immunohematologic complications of ABO-unmatched liver transplants have shown increased hemolytic complications from donor de- rived antibodies in 0-to-A mismatches and with cyclos- porine therapy, but no other parameters, including donor/ recipient age, race. gender, secretor status, and rejection have been predictive. In view of the relatively high inci- dence (up to 56%) of GVH antibodies in ABO-un- matched liver transplants, many investigators have rec- ommended prophylactic transfusion of donor type red cells at surgery and postoperatively [ I ,8].

In minor Rh-incompatible organ transplantation, pre- vious sensitization may predict GVH anti-Rh-induced he- molysis. The duration of the GVH anti-Rh detection has varied from 2 to 6 months and may be actually longer than the 1 month typically seen in ABO-mismatched cases. Transfusion requirements due to hemolysis from GVH anti-ABO and anti-Rho(D) have been comparable [ 1 I . The limited number of minor Rh-incompatible liver transplant cases reviewed here are in agreement with these observations. The high incidence of hemolysis and the high red cell transfusion requirements seen thus far in the limited number of cases at risk for developing GVH

anti-Rho(D) may be a genuine trend and may be ex- plained on the basis of red cell antigen distribution within the body. Although the ABO system is most important in determining serologic red cell compatibility, the antigens of this system are widely expressed on glycolipids in plasma and on epithelial and endothelial cells, lympho- cytes, and platelets [9] and hence may adsorb and weaken the impact of the newly generated GVH ABO antibodies. The Rh antigens are probably limited to the red cells, and the GVH anti-Rho(D) response may be targeted only to the red cells. This antigen distribution disparity between the ABO and the Rh systems may explain the protracted and seemingly exaggerated hemolysis seen thus far in the few reported cases of minor Rh-incompatible organ trans- plants from previously sensitized donors. As with ABO- unmatched liver transplantation, prophylactic transfusion with donor type red cells may also be warranted in minor Rh-incompatible liver transplantation with grafts from previously sensitized donors.

Whether the lymphoid content of a donor liver is re- lated to the incidence of anti-Rho(D)- induced GVH dis- ease is unknown. Furthermore, the role of immunosup- pression in reducing the incidence of such an occurrence is also unclear.

Cyclosporine A, known to suppress a primary immune response to a greater extent than a secondary response [7], may be less effective against a secondary GVH anti- Rho(D) response from previously sensitized donor lym- phocytes than a primary response. Although definitive criteria characterizing GVH anti-Rho(D) in minor Rh- incompatible liver transplantation cannot be extracted from four scattered cases, we conclude the following: 1) the paradoxical antibody arises from passenger B lym- phocytes carried within the donor liver and represents a GVH disease; 2) previous sensitization of the donor is more likely to result in postoperative hemolysis; 3 ) the complications may range from subclinical laboratory evi- dence of red cell sensitization to severe hemolysis with associated morbidity and mortality; 4) an antibody detec- tion test, identification, and assessment of the antibody score from a pretransplant donor specimen may be help- ful in identifying cases at risk for hemolytic complica- tions and in recognizing GVH antibody-induced hemoly- sis; 5 ) prophylactic support with red cells of donor type at surgery and postoperatively may be warranted in selected cases; and 6) routine performance of these tests will en- hance our appreciation for GVH anti-Rh-induced hemo- lysis in solid organ transplantation.

ACKNOWLEDGMENTS

The authors thank Shelia A. Mosher, M.T.(ASCP), and Thomas W. Brady, M.T.(ASCP)S.B.B., for helpful technical assistance.

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GVH Anti-Rho(D) in Liver Transplantation 171

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