Zusammenfassung

171

Karpfen zu nutzen, mit neutrophilen Granulozyten aus humanem peripherem Blut durchgeführt. Die Ergebnisse zeigten, dass MβCD-induzierte Cholesteroldepletion zu einer gesteigerten NET Produktion führte. Die Behandlung mit dem Sphingolipid-Synthese-Inhibitor NB-DNJ hat bedeutenderweise keinen Effekt auf die NET Produktion. Zusätzlich zeigte sich, dass die MβCD-induzierte NET Produktion interessanterweise ROS unabhängig war. Demzufolge zeigten die Ergebnisse, das Cholesterol eine signifikante Rolle in den Mechanismen zur NET Produktion spielt.

Da wir bereits die Bedeutung der Lipide, insbesondere Cholesterol, in Immunreaktionen dargestellt haben, wollten wir die Lipidkomposition in Zellmembranen von Karpfen charakterisieren. Daher etablierten wir ein Protokoll zur Isolierung und Analyse von Membranlipiden und Lipid Rafts, wonach Membranfraktionen mit Triton X-100 isoliert wurden und die Lipidzusammensetzung durch Hochleistungs-Dünnschichtchromatographie (HPTLC) visualisiert wurde. Die Methode wurde durch Westernblot validiert, welches eine hohe Konzentration des Lipid Rafts assoziierten Flotillin Proteins in der Lipid Raft Fraktion zeigte. Höhere Konzentrationen von Cholesterol, freien Fettsäuren, Triglyceriden und Sphingomyelin wurden in den Lipid Raft Fraktionen detektiert, während in der DSM Fraktion höhere Konzentrationen von Phospholipiden gefunden wurden.

Schließlich wollten wir mithilfe der Cholesterolreduzierung und der Fettsäureanalyse, die in dieser Arbeit beschrieben werden, die Rolle von Lipid Rafts während einer CyHV-3 Infektion untersuchen. Die Ergebnisse zeigten, dass nach einer Cholesterol-Depletionsbehandlung mit MβCD von CCB-Zellen, der Viruseintritt verhindert wurde. Dies wurde durch ein geringeres Level von Viruskopien gekennzeichnet. Untersucht wurde dies durch qRT-PCR und Immunhistochemie.

Zusammenfassend zeigten die Ergebnisse, die in dieser Studie vorgestellt wurden, die

Wirt-Pathogen Interaktionen während einer bakteriellen und einer viralen Infektion. Weiterhin

können die hier vorgestellten Methoden für detailgenauere Studien der Wirt-Pathogen

Interaktion genutzt werden und die erhaltenen Erkenntnisse können bei der Identifizierung

neuer Ziele der Infektionsabwehr für neue Behandlungsmöglichkeiten oder prophylaktische

Strategien in der Aquakultur genutzt werden.

172

Affidavit

I herewith declare that I autonomously carried out the PhD-thesis entitled “Cell pathogen interactions in common carp (Cyprinus carpio L.): Studies on cell membranes and neutrophil responses”.

No third party assistance has been used. I did not receive any assistance in return for payment by consulting agencies or any other person. No one received any kind of payment for direct or indirect assistance in correlation to the content of the submitted thesis.

I conducted the project at the following institutions:

1 Fish Disease Research Unit, University of Veterinary Medicine Hannover, Germany

2 Department of Physiological chemistry, University of Veterinary Medicine Hannover, Germany

The thesis has not been submitted elsewhere for an exam, as thesis or for evaluation in a similar context.

I hereby affirm the above statements to be complete and true to the best of my knowledge.

_______________________________

[Date], signature

Acknowledgements

173 Acknowledgements

First of all I would like to thank my supervisor Prof. Dr. Steinhagen. It was a great project and I really enjoyed working under his supervision. From the first day when I arrived to the end of my PhD studies, Dieter always had time for me and wanted to make sure that I was surviving in Germany. Dieter also taught me a lot scientifically about fish biology and how to plan experiments and write manuscripts. I will always be thankful for the opportunity to work in the Fish Disease Research Group.

I would also like to thank Prof. Dr. Hassan Naim who adopted me into his group and converted me into a fish biochemist. I think that it was a very fruitful collaboration between the two groups and I was lucky to get the opportunity to learn a lot of new methods. I would also like to thank Prof. Dr. Hassan Naim for allowing me to work in the Department of Physiological Chemistry and for providing me with financial support towards the end of my PhD studies.

Thirdly, I would like to thank Prof. Dr. Bernd Schröder and Prof. Dr. Anaclet Ngezahayo. They gave lots of helpful tips and comments on my work, often from a different perspective which was very important.

I really appreciate all the help and advice that Dr. Maren von Köckritz-Blickwede gave me during the last 3 years of my PhD study. Maren always made time to help me and I really enjoyed working with her. I learnt a lot during these 3 years and she was a great unofficial supervisor.

I would also like to thank all my colleagues from both the fish hut and the biochemistry, especially my 2 officemates; Mikołaj and Hamdan, with whom I shared lots of scientific and philosophical discussions. Working with all of you has made my time here a lot more enjoyable! Finally Björn and Tanja, my two master students. I enjoyed working with both of you and I learnt a lot. I wish you both successful (hopefully scientific) careers in the future.

I would also like to the thank the EU for not just funding my PhD studies, but also for allowing me to participate in a large number of conferences and training courses, which have all helped me develop as a person and as a scientist. A also thank members of the NEMO group for the many successful collaborations and meetings that we shared together.

Also, I would like to thank the fraternity ‘Hannoversche Burschenschaft Teutonia’ for

allowing me to live there for the first 3 years of my stay in Germany. I got to live the student

lifestyle for another 3 years and was also able to practice a lot of the management skills that

174

I learnt during my PhD course. Importantly, I also made many friends who I will hopefully stay in contact with in the future.

I would also like to thank my friends back at home who I see too rarely and Sandra who has been very patient and supportive during the more stressful moments.

Last but not least, I would like to thank my family for their continuous support and belief

throughout all aspects of my life, and without them I would never have gotten this far. I also

always looked forward to their visits which gave me a break from the lab and put things into

perspective.