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J Thorac Cardiovasc Surg 1994;108:182-184
© 1994 Mosby, Inc.
LETTERS TO THE EDITOR |
Departments of Pathology and Anatomy
George Washington University Medical Center
Washington, D.C. 20037
Departments of Neonatology,
Pathology and Laboratory Medicine
Children's Hospital
Washington, D.C.
To the Editor:
Posttransfusion graft-versus-host disease (GVHD) is an infrequent but serious complication of blood transfusion with a mortality rate of 84% or greater.
1 Posttransfusion GVHD has been described in preterm and term neonates, in children with congenital immune deficiency, in patients with hematologic and nonhematologic malignant diseases, in both autologous and allogeneic bone marrow transplant recipients, and in an infant undergoing extracorporeal membrane oxygenation (ECMO). 2 A very high degree of suspicion is required to make the diagnosis of posttransfusion GVHD. Both underreporting and overreporting of posttransfusion GVHD are likely occurring because of the lack of uniform confirmatory criteria in establishing the diagnosis. We report the case of a child presumed to have posttransfusion GVHD on the basis of very strong clinical and histopathologic evidence. Amplified fragment length polymorphism (FLP) analysis was successfully used to rule out what appeared to be a bonafide case of posttransfusion GVHD.
A 2
-year-old boy with complex congenital heart disease was admitted to Children's Hospital for postoperative ECMO support. He underwent a Blalock-Taussig procedure in early infancy and 3 days before admission underwent a Glenn procedure. His postoperative course was complicated by pneumonia caused by Hemophilus influenzaas well as adult respiratory distress syndrome. The patient was supported by ECMO because conventional ventilatory assistance had failed. During the first 12 days of ECMO, the patient required 21 red cell transfusions and 64 units of platelets. Only those red cell transfusions derived from the nursery inventory for small volume transfusions of 25 ml or less were irradiated per a preexisting protocol. Six units of packed red cells and 64 platelet units were transfused and not irradiated for two cardiac bypass procedures and ECMO, as well as post-ECMO thrombocytopenia. On day 12, an erythematous maculopapular rash appeared on the abdomen and subsequently spread to the remainder of the face and body. Three days after the rash appeared, a significant gastrointestinal hemorrhage developed. Alanine aminotransferase and aspartate aminotransferase blood concentrations were mildly elevated. A skin biopsy suggested the diagnosis of GVHD. Amplified FLP and HLA antigen typing on peripheral blood and an abdominal skin biopsy specimen were obtained. Results were compared with peripheral blood specimens from both his biologic parents.
Blood cultures at this time were positive for Candida tropicalis. He subsequently had massive gastrointestinal hemorrhage and hemoperitoneum. Because of continued cardiac dysfunction and pulmonary failure, ECMO support was terminated. The patient died on day 22 of his hospitalization and an autopsy was performed.
High-molecular-weight deoxyribonucleic acid (DNA) was obtained from heparinized peripheral blood from the patient and both biologic parents. Skin fibroblasts from the patient were analyzed in a similar manner. Peripheral blood mononuclear cells and neutrophils were separately isolated by Ficoll-Hypaque density gradient centrifugation. The blood and fibroblast DNA were then amplified in the D1S80, apolipoprotein B, Co12A1, and pYNZ22 regions using the polymerase chain reaction. Products from the reactions were separated by electrophoresis on polyacrylamide gels, stained with ethidium bromide to visualize the bands, and photographed. Because of the lack of a pretransfusion sample, laboratory standards of sensitivity consisted of diluting DNA from normal individuals at 1% and 5% final concentrations. Histopathologic analysis and HLA typing were performed by standard methods.
Microscopic examination of the skin biopsy obtained on day 12 of hospitalization revealed an intact epidermis with basal cell vacuolar degeneration (Fig. 1). Single cell necrosis (apoptosis) and intraepidermal lymphocytes surrounding the apoptotic cells (satellitosis) were also noted. The papillary dermis was infiltrated by a mononuclear cell population and an occasional colloid body was seen. The final autopsy diagnostic evaluation revealed that the patient had situs inversus totalis and complex congenital heart disease. Evidence of widespread disseminated intravascular coagulation and multiple pulmonary hemorrhages was present, likely caused by the patient's sepsis. No evidence for posttransfusion GVHD was seen in sections obtained from the gastrointestinal tract, liver, and bone marrow.
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3 In these reports, the technique was able to confirm the presence of third party–mediated GVHD in patients receiving blood transfusions, and it appeared to be a satisfactory alternative to HLA typing. We included amplified FLP analysis in the evaluation of a 2 year old male child undergoing ECMO. Although infants undergoing ECMO do not appear to be immunosuppressed by the procedure, we 4 have recently reported that they do not show any evidence of B-cell or T-cell activation after concomitant blood transfusions. This is in contrast to what is seen in adults undergoing cardiopulmonary bypass procedures. 5 Therefore, our patient could have had an altered immune response to blood transfusion, possibly placing him at higher risk for posttransfusion GVHD. We were further concerned that posttransfusion GVHD had developed because the skin biopsy results were highly suggestive. The presence of single cell dyskeratosis, satellitosis, and basal cell vacuolar degeneration in an at-risk patient, although not pathognomonic of posttransfusion GVHD, is virtually diagnostic. However, distinction of GVHD from a drug reaction, erythema multiforme, spongiotic dermatitis, and viral eruptions may be impossible. One or more of the aforementioned disorders could have been responsible for the histopathologic changes seen in the skin biopsy. Determining the correct diagnosis in this setting is crucial because posttransfusion GVHD is not treatable and has a high mortality as compared with the other disease entities listed.
Sensitivity and accuracy of amplified FLP analysis is likely to be different with respect to HLA analysis; the latter technique may not always be useful. In fact, in addition to HLA analysis on peripheral blood lymphocytes, patient fibroblast cultures may be necessary for the possibility of posttransfusion GVHD to be fully evaluated. Our data suggest that amplified FLP analysis can be used as an alternative to HLA assays for the evaluation of posttransfusion GVHD. HLA analysis in this setting has been shown to have a lower sensitivity with respect to Southern blot analysis with FLP. FLP analysis alone had a sensitivity of 1% detection of allogeneic DNA. When used in conjunction with polymerase chain reaction for the detection of residual leukemic cells, sensitivity was in the range of 1 in a total of 10 5 cells. A sensitivity of 1% as illustrated in our investigation of posttransfusion GVHD appears to be an excellent detection limit, in that no evidence of posttransfusion GVHD was seen at autopsy. However, because the minimum number of circulating allogeneic lymphocytes necessary to elicit posttransfusion GVHD is currently not known, future studies are needed to establish optimum sensitivity of this analysis in this pathologic setting.
References
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