Regulation of dendritic cell function by nutrients [Elektronische Ressource] / von Nguyen Thi Xuan

Regulation of Dendritic Cell Function
by Nutrients

der Fakultät für Biologie
der EBERHARD KARLS UNIVERSITÄT TÜBINGEN
zur Erlangung des Grades eines Doktors
der Naturwissenschaften

von

NguyenThi Xuan
aus Thai Binh, Vietnam

vorgelegte

DISSERTATION

2009



























Tag der mündlichen Prüfung: 02.11.2009
Dekan: Prof. Dr. Hanspeter Mallot
1. Berichterstatter: Prof. Dr. Florian Lang
2. Berichterstatter: Fritz Götz

ACKNOWLEDGEMENTS

It is with immense pleasure, I am thankful very much to my
research guide Prof. Dr. Florian Lang, for his scientific guidance,
critical review, support throughout my Ph.D. work at Institut of
Physiology, University of Tübingen, Germany.
I am very grateful to Dr. Shumilina Ekaterina for her timely
help, support, thought provoking suggestions and stimulating
discussions during the progress of the work. I would like to thank
Prof. Dr. Fritz Götz for giving me an opportunity to present the
dissertation at the Faculty of Biology, Eberhard Karls Universität
Tübingen, Germany.
I am very thankful to my friends and my colleagues Ms. Matzner
Nicole, Dr. Zemtsova Irina, Mr. Mahmud Hasan, Mr Diwakar
Bobbala, Mr Qadri Syed for their help, suggestions, friendship and
support. I also thanks to my colleagues Ms. Zhang Ying, Mrs. Efi
Faber, Mr. Uwe Schueler, Mr. Rexhepaj Rexhep and Mr. Peter for
their support.
This task never completes without the mention of my loving
parents, my brother and sister who have blessed me in every aspect of
my life.
I dedicate my thesis to my husband Dr. Quach Duc Tin and my
daughter Quach Huong Linh for being a constant source of inspiration
for my work. I am greatly indebted for his support, constructive
criticism and endless love in my life.
iii

TABLE OF CONTENTS

ACKNOWLEDGEMENTS ...................................................................................................... iii 
TABLE OF CONTENTS .......................................................................................................... iv 
ABBREVIATIONS ................................................................................................................. viii 
ABSTRACT ............................................................................................................................... x 
Z U S A M M E N F A SS U N G ........................................................................................................... xi 
1.  INTRODUCTION .............................................................................................................. 1 
1.1.  Adaptive immune system and innate immunity .............................................................. 1 
1.1.1.  Innate immune system ................................................................................................. 1 
1.1.2.  Adaptive immune system ............................................................................................ 2 
1.2.  Nature of DCs .................................................................................................................. 4 
1.3.  Interaction of DCs with NK, B and T cells ..................................................................... 6 
1.4.  Tolerance immunity and Th1/Th2 balance drived by DCs ............................................. 7 
1.5.  Toll-like receptors (TLRs) .............................................................................................. 8 
1.6.  Subsets of conventional mouse DCs ............................................................................... 9 
1.7.  Migration and Phagocytosis .......................................................................................... 11 
1.8.  The mucosal immune system ........................................................................................ 11 
1.9.  Mucosal dendritic cells .................................................................................................. 12 
1.10.  Cell death ................................................................................................................... 13 
1.10.1.  Intrinsic apoptosis pathway .................................................................................... 13 
1.10.2.  Extrinsic apoptosis pathway .................................................................................. 14 
1.10.3.  Sphingomyelinase pathway ................................................................................... 14 
1.10.4.  Regulation of apoptosis by ceramide ..................................................................... 15 
1.10.5.  The Bcl-2 family: inhibitors and promoters of apoptosis ...................................... 15 
1.10.6.  Caspase activity ..................................................................................................... 16 
1.11.  Nutrients .................................................................................................................... 16 
1.11.1.  Thymoquinone ....................................................................................................... 17 
1.11.2.  Gum Arabic ............................................................................................................ 17 
2+1.11.3.  Zn ........................................................................................................................ 18 
1.11.4.  Xanthohumol .......................................................................................................... 19 
1.11.5.  Thymol ................................................................................................................... 19 
2.  AIMS OF THE STUDY ................................................................................................... 20 
3.  MATERIAL AND METHODS ....................................................................................... 20 
3.1.  Equipment ..................................................................................................................... 20 
3.2.  Mice ............................................................................................................................... 21 
3.3.  Chemicals ...................................................................................................................... 21 
3.4.  DC culturing .................................................................................................................. 22 
3.5.  Immunostaining and flow cytometry ............................................................................ 23 
3.6.  Cytokine quantification in cell supernatants ................................................................. 23 
3.7.  DC phagocytosis assay .................................................................................................. 23 
3.8.  Immunoblotting ............................................................................................................. 24 
3.9.  Isolation of T and B lymphocytes from the spleen ....................................................... 24 
3.10.  Phosphatidyl residue translocation ............................................................................ 25 
3.11.  Ceramide formation ................................................................................................... 25 
3.12.  DNA fragmentation ................................................................................................... 25 
3.13.  Measurements of mitochodrial membrane potential ................................................. 26 
3.14.  Caspase 8 and Caspase 3 activation assay ................................................................. 26 
3.15.  Immunocytochemistry ............................................................................................... 26 
iv

3.16.  Statistics ..................................................................................................................... 27 
4.  RESULTS ......................................................................................................................... 27 
4.1.  Thymoquinone .............................................................................................................. 27 
4.1.1.  Thymoquinone inhibits maturation of LPS- stimulated DCs. ................................... 27 
4.1.2.  Thymoquinone impairs cytokine secretion by LPS- stimulated DCs. ....................... 29 
4.1.3.  Thymoquinone enhances percentage of annexin V-binding DCs. ............................ 30 
4.2.  Gum Arabic ................................................................................................................... 31 
4.2.1.  GA enhances bo