Origin of CD8_1hn+ effector and memory T cell subsets [Elektronische Ressource] / Christian Stemberger

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Informations

• Publié par: technische_universitat_munchen
• Publié le: 01 janvier 2008
• Nombre de lectures: 22
• Langue: Deutsch
• Poids de l'ouvrage: 4 Mo

Extrait

TECHNISCHE UNIVERSITÄT MÜNCHEN
Lehrstuhl für Genetik




+Origin of CD8 effector and memory T cell subsets


Christian Stemberger




Vollständiger Abdruck der von der Fakultät Wissenschaftszentrum Weihenstephan für
Ernährung, Landnutzung und Umwelt der Technischen Universität zur Erlangung des
akademischen Grades eines

Doktors der Naturwissenschaften

genehmigten Dissertation.



Vorsitzender: Univ.-Prof. Dr. S. Scherer
Prüfer der Dissertation: 1. Univ.-Prof. Dr. A. Gierl
2. Univ.-Prof. Dr. D. Busch
3. Univ.-Prof. Dr. D. Haller



Die Dissertation wurde am 03.07.2008 bei der Technischen Universität München eingereicht
und durch die Fakultät Wissenschaftszentrum Weihenstephan für Ernährung, Landnutzung
und Umwelt am 21.09.2008 angenommen.

.Parts of this thesis have already been published:

Stemberger, C., Huster, KM., Busch, DH. (2007).
Defining correlates of T cell protection against infection.
Discov Med 6(34), 148-152.

Stemberger, C., Neuenhahn, M., Buchholz, V. R. and Busch, D. H. (2007).
+Origin of CD8 effector and memory T cell subsets.
Cell Mol Immunol 4(6), 399-405.

Stemberger, C., Huster, K. M., Koffler, M., Anderl, F., Schiemann, M., Wagner, H. and
Busch, D. H. (2007)
A single naïve T cell precursor can develop into diverse effector and memory subsets.
Immunity 27(4), 985-997.

1 – TABLE OF CONTENTS
1 TABLE OF CONTENTS

1 TABLE OF CONTENTS.........................................................................................................3
2 INDEX OF FIGURES.............................................................................................................5
3 ABBREVIATIONS..................................................................................................................7
4 INTRODUCTION .................................................................................................................10
4.1 The immune system .....................................................................................................10
4.2 Adoptive immunity ........................................................................................................11
4.3 T cells ...........................................................................................................................12
+4.3.1 CD4 T cells...........................................................................................................12
+4.3.2 CD814
+ +4.3.3 CD4 T cell help for the generation of CD8 T cell responses...............................16
+4.4 CD8 T cell subsets and the concept of “job sharing”...................................................17
4.5 The protective capacity of memory subsets .................................................................20
4.6 Differentiation pathways, lineage relationship and “stemness” ....................................21
5 AIM OF THIS PhD THESIS .................................................................................................28
6 MATERIALS AND METHODS.............................................................................................30
6.1 Materials .......................................................................................................................30
6.1.1 Equipment .............................................................................................................30
6.1.2 Chemicals and reagents........................................................................................31
6.1.3 Buffers and Media32
6.1.4 Gels .......................................................................................................................34
6.1.5 Antibodies..............................................................................................................35
6.1.6 Peptides ................................................................................................................36
6.1.7 MHC multimers......................................................................................................36
6.1.8 MHC streptamers ..................................................................................................37
6.1.9 Artificial antigen presenting cells (aAPCs) ............................................................37
6.1.10 Mice37
6.1.11 Software38
6.2 Methods........................................................................................................................38
6.2.1 Preparation of cells from different organs .............................................................38
6.2.2 Production of MHC-I multimers / streptamers .......................................................39
6.2.3 Antibody and multimer staining .............................................................................40
6.2.4 CFSE staining .......................................................................................................41
6.2.5 Intracellular cytokine staining and degranulation assay ........................................41
31 – TABLE OF CONTENTS
6.2.6 FACS acquisition and analysis..............................................................................42
6.2.7 FACS sorting .........................................................................................................42
6.2.8 Stimulation of naïve T cells with aAPCs................................................................43
6.2.9 Listeria monocytogenes infection and immunization.............................................43
6.2.10 Adoptive cell transfers ...........................................................................................44
6.2.11 Measurement of bacterial load45
6.2.12 Microscopy ............................................................................................................45
6.2.13 Statistics ................................................................................................................46
7 RESULTS ............................................................................................................................47
7.1 Development of an adoptive transfer system for single cells........................................47
7.2 Diverse effector T cell populations arise from single naïve precursor cells..................51
+7.3 CD8 memory cells develop from single naïve precursor T cells .................................57
+7.4 Different CD8 memory subsets develop from single naïve precursor T cells..............60
7.5 Effector and memory subsets originate from the same single naïve precursor cells....64
+7.6 Transferred- and endogenous CD8 T cell subset diversification is synchronized
independently of immunization or infection .....................................................................67
7.7 “Latecomer” cells still have the potential to differentiate into diverse subsets..............70
7.8 The descendants of a single naive T cell confer protection against Listeria
monocytogenes ...............................................................................................................73
7.9 Plasticity of antigen-experienced memory OT-I T cell subsets.....................................75
8 DISCUSSION ......................................................................................................................78
8.1 When during an immune response is heterogeneity achieved?...................................79
8.2 Clinical relevance for the improvement of immunotherapies........................................84
+8.3 Development of CD4 T cell subsets............................................................................86
9 SUMMARY ..........................................................................................................................89
10 REFERENCES ....................................................................................................................91
11 ACKNOWLEDGMENTS ....................................................................................................101


42 – INDEX OF FIGURES
2 INDEX OF FIGURES

Figure 1: Models of T cell subset diversification. ........................................................................25
Figure 2: Naïve antigen specific T cells take a bi-directional lineage fate decision during the first
cell division. .........................................................................................................................27
Figure 3: Adoptive transfer of single T cells................................................................................47
Figure 4: Rapid migration to the spleen and lack of homeostatic proliferation early after
intraperitoneal injection of OT-I cells. ..................................................................................49
Figure 5: Principles of the single cell microinjection method. .....................................................50
Figure 6: Precision of single cell aspiration.................................................................................51
Figure 7: Phenotypical diversity of T cells expanded from a single naïve precursor cell during
the effector phase (day 7).......