Use of an inbred mouse model system for studies of allogeneic transfusion-induced immunosuppression.

Two studies published recently in Blood' and Transfiision2 describe application of an inbred mouse model of allogeneic transfusion to investigate the role ofdonor leukocytesin inducingimmunosuppressive consequences of transfusion. Blajchman et al' showed a median fivefold increase in the frequency of pulmonary metastases at 21 days in C57/BI/J6 (H-2b) mice that were transfused with allogeneic (Balb/c [H-2d]) whole blood IO days before injection of a fibrosarcoma cell line, relative to the rate of metastases in mice pretransfused with either syngeneic or leukodepleted allogeneic blood. Moreover, spleen cells harvested from C57/BI/J6 recipient mice IO days after allogeneic transfusion and infused into other C57/BI/J6 mice could passively transfer the tumor growth-promoting effect. Gianotti et al? in studies of the effect of allogeneic transfusions on sepsis-related mortality, infused either leukocytes, red blood cells, or plasma from C3H/HeJ (H-2k) mice into Balb/c (H2d) mice. Five days later the mice were subjected to a 20% bum and gavage with I X Escherichia coli. The percentage of viable bacteria and the mortality rate were significantly higher in the group receiving allogeneic leukocytes relative to other allogeneic blood constituents or control (buffer or syngeneic blood) infusions. These data further implicate donor leukocytes as the primary culprit in transfusion-related complications, thereby lending additional support to the movement toward routine leukodepletion of cellular blood components.' Although our own research also implicatesallogeneic leukocytes in transfusion-induced immunosuppression: recent studies in our laboratory characterizing survival kinetics of donor leukocytes in transfused humans, dogs, and mice indicate that allogeneic transfusions between strains of inbred mice may not be a good model system for such investigations. To facilitate survival studies of transfused leukocytes, we developed quantitative, allele-specific polymerase chain reaction (PCR) assays directed at Y chromosome-specific sequences of humans, dogs, and mice.s.6 Based on in vitro mixing studies of male into female blood for each species, the assays detect as few as 1 to 5 male donor leukocytes per 50 pL ofrecipient blood containing IO'to IO6 female leukocytes. Quantitation is achieved by parallel amplification and autoradiographic image analysis of dilutions of male into female blood.' Samples with Y-specific signal exceeding the dynamic range of the assay are serially diluted and reevaluated. In preliminary studies involving four transfused orthopedic surgery patients and four outbred dogs, we observed greater than 99.9% clearance of infused allogeneic leukocytes within 48 hours of transfusion, and male donor cells were undetectable by 7 days posttransfusion (Fig IC). In contrast, we were surprised to find that in inbred mice, allogeneic donor leukocytes are not cleared from the recipient's circulation for weeks to months posttransfusion. Specifically, 150 pL (1/10 blood volume) of male C57B (H-2') mouse blood was transfused into each ofthree female Balb/c (H-2d) recipients, and three female C57B recipients were given male Balb/c blood. Recipient mice were sampled daily for 1 week, and then at 3 weeks, 5 weeks, and 3 months. In all cases, male donor cells persisted at concentrations in the I to IO cells/pL range (0. I % to 1% of total leukocytes) in the circulation of recipient mice for over 1 month posttransfusion, and donor cells were not cleared to undetectable levels ( < I ce11/50 pL) until 3 months (Fig I , A and B). Similar results were obtained after transfusion of male C3H (H-2') blood into female Balb/c recipients. Despite this persistence of allogeneic donor cells, there was no evidence of graft-versus-host disease in recipient mice. The failure of rapid clearance ofdonor cellsafter allogeneic transfusions of immunocompetent inbred mice was unexpected. AIthough low-level chimeras have been reported after bone marrow and solid organ transplantation of inbred mice and humans subjected to immunosuppressive regimens,&" inherent responsiveness to "non-self' HLA antigens of immunocompetent subjects should,