Investigations Into the Development and Consequences of Red Blood Cell Alloimmunization Público

Abstract

Red blood cell transfusions can be a life saving therapy for patients with severe anemia, but they are not without risks. The generation of alloantibodies to RBC antigens can have serious adverse outcomes, including causing acute hemolytic transfusion reactions. In addition to transfusion, alloantibodies derived against RBCs during pregnancy can lead to hemolytic disease of the fetus and newborn, a condition with severe and sometimes fatal consequences for the developing fetus.

With the goal of better understanding red cell alloimmunization and incompatible RBC clearance, we generated a murine model with RBC-specific expression of the human KEL glycoprotein, which has been implicated in causing severe hemolytic transfusion reactions and HDFN. Transfusion of KEL RBCs into WT recipients consistently results in an anti-KEL antibody response, which is enhanced by recipient inflammation. In addition, we demonstrated that maternal exposure during pregnancy to fetal RBCs expressing paternally derived KEL antigens can similarly result in anti-KEL alloimmunization. Both transfusion and pregnancy-induced anti-KEL alloantibodies are boostable upon repeat exposure and are capable of binding to and causing antibody-mediated destruction of KEL RBCs. This destruction is accompanied by a systemic inflammatory response following subsequent transfusion or by decreased fetal survival in the case of pregnancy.

We further investigated the mechanisms of alloimmunization and found that removal of the spleen and more specifically, the marginal zone B-cells within the spleen, prevented alloimmunization to KEL RBCs in WT recipients. We also analyzed clearance patterns of KEL RBCs in immunized recipients and discovered a potential role for the complement protein C3 in clearance of incompatible RBCs as well as in antigen modulation, which may help to protect the remaining transfused RBCs from further immune mediated destruction.

These data have enhanced our understanding of the mechanisms of both the induction of anti-KEL alloantibodies as well as their clinical consequences in transfusion and pregnancy settings alike. This is the first murine model with the ability to recapitulate the essential features of a human response to a RBC antigen and will likely serve as a platform for the future development of targeted therapies to prevent or minimize the dangers of RBC alloimmunization.

Table of Contents

TABLE OF CONTENTS

Chapter 1 - Introduction............................................................................................ 1

History of Blood in Medicine................................................................................. 2

Blood in ancient culture and medicine........................................................ 2

The first blood transfusions............................................................................ 4

The first human to human transfusions....................................................... 6

Transfusion in the early 20^th Century............................................................ 8

Transfusion medicine in the last 50 years................................................... 10

Red Blood Cell Biology and Modern Transfusion Medicine.......................... 11

Red blood cell structure and function.......................................................... 11

Blood group antigens...................................................................................... 12

The Kell Blood Group System....................................................................... 15

Benefits and risks of blood transfusion........................................................ 17

The Immunology of Transfusion Medicine........................................................ 20

Alloimmunization to red cell antigens......................................................... 20

The role of the spleen in alloimmunization................................................ 24

Consequences of red cell alloimmunization.............................................. 26

Hemolytic disease of the fetus and newborn.............................................. 28

Immune destruction of red blood cells........................................................ 30

Escape from immune-mediated clearance................................................ 33

Animal Models of Red Cell Alloimmunization.................................................. 35

Summary................................................................................................................. 37

References ............................................................................................................. 40

Chapter 2 - Transfusion of murine RBCs expressing the human KEL glycoprotein induces clinically significant alloantibodies 50

Abstract.......................................................................................................................... 51

Introduction.................................................................................................................... 52

Materials and Methods................................................................................................ 56

Results ......................................................................................................................... 59

Discussion.................................................................................................................... 68

References.................................................................................................................... 72

Figure Legends............................................................................................................ 75

Chapter 3 - Depletion of marginal zone B-cells inhibits red blood cell alloimmunization 84

Abstract.......................................................................................................................... 85

Introduction.................................................................................................................... 86

Materials and Methods................................................................................................ 89

Results ......................................................................................................................... 92

Discussion.................................................................................................................... 99

References.................................................................................................................... 104

Figure Legends............................................................................................................ 107

Chapter 4 - A novel role for C3 in antibody-induced red blood cell clearance and antigen modulation 118

Abstract.......................................................................................................................... 119

Introduction.................................................................................................................... 120

Materials and Methods................................................................................................ 123

Results........................................................................................................................... 127

Discussion.................................................................................................................... 133

References.................................................................................................................... 139

Figure Legends............................................................................................................ 141

Chapter 5 - Alloantibodies to a paternally derived RBC Kell antigen lead to hemolytic disease of the fetus/newborn in a murine model 152

Abstract.......................................................................................................................... 153

Introduction.................................................................................................................... 154

Materials and Methods................................................................................................ 157

Results .......................................................................................................................... 161

Discussion.................................................................................................................... 170

References.................................................................................................................... 175

Figure Legends............................................................................................................ 178

Chapter 6- Discussion.............................................................................................. 189

Summary....................................................................................................................... 190

Discussion.................................................................................................................... 196

Red blood cells are unique as cells and immunogens.................................. 196

The use of transgenic mouse models to study red cell alloimmunization. 198

The role of the spleen in the generation of the immune response to RBC antigens 201

Mechanisms of clearance or survival of transfused RBCs............................. 205

Implications of an animal model for hemolytic disease of the fetus and newborn 206

Limitations of the Kell transgenic model........................................................... 208

Conclusions.................................................................................................................. 209

References.................................................................................................................... 211

About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Degree	PhD
Submission	Dissertation
Language	English
Research Field	Health Sciences, Immunology
Palavra-chave	Pregnancy Erythrocyte Red Blood Cell Transfusion Alloimmunization
Committee Chair / Thesis Advisor	Ford, Mandy L, Emory University
Committee Members	Evavold, Brian, Emory University Iwakoshi, Neal N., Emory University Denning, Timothy, Emory University Hendrickson, Jeanne, Emory University

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