H e m a t o lo g y E d u c a t io n - European Hematology Association

16 th Congress of the European Hematology Association
ple transfusions of buffy coat depleted hGPA RBCs fail
to lead to recipient anti-hGPA responses. 51 However,
transfusion of buffy coat depleted hGPA RBCs mixed
with unmethylated bacterial CpG dinucleotides (CpG
ODN) (followed by weekly transfusions of buffy coat
depleted hGPA RBCs in the absence of an agonist) leads
to presumed anti-hGPA alloimmunization in some
recipients. Similar to what has been observed in Tg-FYB
alloimmunized recipients transfused with Tg-FVB
RBCs, hGPA RBCs have a reduced circulatory half life in
hGPA immunized recipients.
Yu et al. have also reported that depletion of regulatory
T cells with anti-CD25 enhances anti-hGPA alloantibody
responses, introducing the idea that recipient regulatory
T cells play a role in RBC alloimmunization. 51
Subsequently, Bao et al. have studied regulatory T cell
status in hGPA alloimmunized “responder” and “nonresponder”
recipient mice, finding reduced in vitro and in
vivo Treg-suppressive activity in responders. 52
Furthermore, similar to the observation of “antibody
pair” formation in humans, 10 they have reported that
murine “responders” are more likely than “non-responders”
to develop additional alloantibodies to a second
immunogen (in this case Tg-FYB).
Besides being a useful model of RBC alloimmunization,
the hGPA model has been useful to study the fate
of incompatible RBCs. 53 Many monoclonal anti-hGPA
antibodies are available, and passive immunization with
IgG1 anti-hGPA monoclonal antibodies induces rapid
clearance of incompatible transfused hGPA RBCs. 54
Furthermore, this rapid clearance is accompanied by a
pro-inflammatory recipient cytokine storm. 55 Mechanistic
studies investigating the role of the spleen, complement
and activating FcgRs in this incompatible RBC clearance,
are ongoing.
Limitations of the hGPA model, present in any mouse
model utilizing donors that differ genetically from recipients,
include difficulties in distinguishing anti-hGPA
responses from anti-MHC responses. MHC-I is present
on multiple cell types in mice, including RBCs, and also
present on WBCs and platelets that may contaminate
RBC preparations. Thus, careful controls are necessary
to ensure observed alloantibody responses are in fact
against transgenic RBC antigens and not against MHC-
I. Accepting this limitation, a significant strength of the
hGPA model includes hGPA being an authentic human
blood group antigen. Furthermore, studies generated in
the hGPA system have served as a springboard for
ongoing translational experiments investigating the role
of regulatory T cells in human alloantibody responses.
Future mouse models of RBC alloimmunization
Future directions in murine alloimmunization studies
include the development of additional model systems
with advantages not held by existing models. This
includes the development of a model with donors and
recipients expressing an RBC specific antigen that differ
by a single amino acid, akin to human antithetic blood
group antigens (e.g., E/e). Additionally, an Rh(D) transgenic
animal, though technically challenging to develop,
would also be quite useful. To increase the analytical
power of such models, antigen specific CD4+ and
CD8+ transgenic T cell companion mice would ideally
be constructed in parallel, potentially along with
tetramer reagents and B cell transgenic mice. Besides
being useful in studying antibody responses and antibody
coated RBC clearance mechanisms, the development
of these additional mouse models could potentially
shed light on the mechanisms of action of prophylactic
anti-D (e.g., RhoGam) and could also lay the groundwork
for a mouse model of hemolytic disease of the
newborn.
Conclusions
Table 1. Clinically relevant questions raised by murine models of RBC alloimmunization.
In summary, much has been learned about RBC
alloimmunization over the past few decades, and mouse
models have proven useful as a supplement to existing
knowledge about human RBC antigens and anti-RBC
antibodies. Being cognizant of inherent limitations of
model systems in general and of mouse models in particular,
the Tg-FVB, mHEL, HOD, and hGPA mouse models
of RBC alloimmunization have each provided insight
into the immune sequelae of transfusion. These include
a better understanding of both donor and recipient factors
influencing rates of RBC alloimmunization, mechanisms
of anti-RBC antibody formation, and clearance
patterns of antibody coated RBCs. Furthermore, these
model systems have generated discrete hypotheses and
questions that are testable in humans (Table 1). For
example, ongoing human studies are investigating the
impact of regulatory T cells on RBC alloimmunization
response rates in patients with sickle cell disease.
Additionally, human clinical trials investigating the relationship
between recipient serum cytokine profiles or
gene polymorphisms and responder/non-responder status
have been proposed. 56 Lastly, the effects of stored
human RBCs are currently being examined in multiple
prospective randomized clinical trials. 57–60
1) Does the inflammatory status of the human recipient play a role in their immune response to transfused RBCs?
2) Do recipient gene polymorphisms influence RBC responder/non-responder status?
3) How do recipient regulatory T cells impact rates and degree of RBC alloimmunization?
4) Are older, stored human RBCs more immunogenic than freshly collected and transfused RBCs?
5) Are leukoreduced human RBCs less immunogenic than non-leukoreduced human RBCs, and if so what is the mechanism behind this?
6) Is a spleen required for RBC alloimmunization in humans? (realizing the presence of the spleen on initial exposure to the antigen may be critical)
7) Can humans be primed for RBC alloantibody responses by prior exposure to pathogens containing CD4+ T cell epitopes shared with RBC antigens?
8) How common is antigen loss in humans, and how often is it not recognized due to lack of awareness?
9) How are incompatible transfused RBCs cleared in humans, and how can sequelae of incompatible transfusions be minimized?
| 370 | Hematology Education: the education programme for the annual congress of the European Hematology Association | 2011; 5(1)