The consequences of antibody binding to red blood cells in alloantibody responses Público

Mener, Amanda (Summer 2018)

Permanent URL: https://etd.library.emory.edu/concern/etds/47429921g?locale=pt-BR
Published

Abstract

Red blood cell (RBC) transfusions provide a life-saving intervention in the setting of hematologic conditions and trauma. However, it is not without risk. Repeated RBC transfusions can lead to formation of alloantibodies against allogeneically distinct antigens on the surface of transfused RBCs. These RBC alloantibodies increase the risk of hemolytic transfusion reactions, or in the case of pregnancy, hemolytic disease of the fetus and newborn, as well as causes difficulty in finding compatible RBCs for future transfusions. The mechanism(s) of how RBC alloantigens induce alloantibodies and furthermore, the immunologic consequences of antibody binding to RBCs following transfusion of RBCs, remain poorly understood. Currently, there is only one pharmacologic agent available to actively prevent the development of RBC alloantibodies, Rh immune globulin (RhIg), which consists of pooled plasma from donors purposely exposed to RhD+ RBCs. The ability of passive administration of antibodies to prevent the de novo development of alloantibodies is termed antibody-mediated immunosuppression (AMIS). However, the mechanism of how AMIS occurs is not well-understood. Furthermore, with the decreasing availability of plasma donors and the ethical limitations in purposefully alloimmunizing patients against RBC alloantigens other than RhD, there is a critical need to better understand the mechanism(s) of AMIS in order to develop alternatives to RhIg. Given previous studies suggesting that decreases in the level of surface antigen, termed antigen modulation, may account for one mechanism of AMIS, we utilized a murine model of RBC alloimmunization, in which RBCs express a fusion protein consisting of hen egg lysozyme (HEL), ovalbumin (OVA) and human Duffy (HOD RBCs). Through exposure of mice to HOD RBCs in the presence or absence of anti-HEL monoclonal antibodies, we explore how monoclonal antibody-induced changes in the level of the target RBC antigen impacts the development of an anti-HOD alloantibody response. Additionally, through utilizing RBCs that express both the clinically-relevant HOD and KEL RBC antigens on the same RBC, we also investigate whether AMIS is antigen-specific and whether this antigen-specific AMIS occurs through antigen modulation.

Furthermore, as RhD and HOD RBCs do not induce complement fixation and complement has been shown to be important in the development of antibody responses, we next sought to determine how complement impacts the development of RBC alloantibodies utilizing a model of RBC alloimmunization that induces complement fixation, the KEL RBC antigen. Unlike previous studies describing complement as an adjuvant in the development of an antibody response, we actually observe that mice deficient in complement component 3 (C3 KO) exhibited increased alloantibody responses against the KEL antigen due to a decrease in antibody-induced antigenic changes on the RBC surface, indicating that complement negatively regulates the development of an antibody response to the KEL RBC antigen.

   Finally, given the differences in IgG subclass development observed between C3-deficient and wild type mice in response to KEL RBC transfusion and the involvement of CD4+ T cells in regulating class switching, we explore the role of CD4+ T cells in the development of the alloantibody response to KEL. We find that in the presence of C3, mice develop alloantibody responses to KEL even in the pharmacologic or genetic absence of CD4+ T cells. However, C3 KO and mice deficient in complement receptors 1 and 2 are completely dependent on CD4+ T cells to develop an anti-KEL IgG response, suggesting that C3 fixation and interaction with complement receptors regulates whether the antibody response to KEL is T-independent or T-dependent. 

Taken together, these findings suggest that the consequences of antibody binding to the RBC surface, acquired either through passive administration of antibody, in the case of AMIS, or following de novo antibody development, include both complement-independent and -dependent antigen modulation. This antigen modulation impacts not only the subsequent development of an alloantibody response to RBC antigens, but also the immunologic pathways involved. These findings contribute to our understanding regarding how the mechanism of alloantibody development to RBC antigens may vary by the individual antigen.

Table of Contents

Chapter 1: Introduction

A brief history of transfusion medicine

Antibody-mediated immunosuppression

Introduction to complement and historical perspective

Complement and humoral immunity

Complement’s role in T cell immunity

Figure and Figure Legend

References

1-71

2-6

6-18

18-28

28-43

43-49

50

51-71

Chapter 2: Antibody-mediated immunosuppression can result from RBC antigen loss independent of Fcg receptors in mice

Abstract

Introduction

Materials and Methods

Results

Discussion

References

Figures and Figure Legends

72-115

73-74

75-77

78-80

81-87

88-92

93-104

105-115

Chapter 3: Antibody-Mediated Immunosuppression by Antigen Modulation is Antigen-Specific

Abstract

Introduction

Materials and Methods

Results

Discussion

References

Figures and Figure Legends

116-155

118-119

120-122

123-125

126-132

133-135

136-144

145-155

Chapter 4: Complement component 3 Negatively Regulates Antibody Response by Modulation of RBC Antigen

Abstract

Introduction

Materials and Methods

Results

Discussion

References

Figures and Figure Legends

156-223

157

168-160

161-167

168-177

178-187

188-205

206-223

Chapter 5: Complement serves as a switch between CD4+ T cell independent and dependent RBC antibody responses

Abstract

Introduction

Materials and Methods

Results

Discussion

References

Figures and Figure Legends

224-284

225

226-228

229-234

235-244

245-250

251-267

268-284

Chapter 6: Discussion

           Summary

Antibody-mediated immunosuppression

Complement fixation and RBC alloantibody development

Negative regulatory role of complement and implications for vaccination

Complement as a regulator of T cell independent antibody responses

General conclusions

Figures and Figure Legends

References

285-329

286-287

287-292

292-297

297-299

299-306

306-308

309-312

313-329

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