3 Material und Methoden
EP 4 3’ TTA CGT TGG TGG TCC AGT CGA T
5.3 Schlussfolgerung und Ausblick
Es konnte nachgewiesen werden, dass eine proliferative Wirkung an Keratinozyten über zwei verschiedene G-Protein-gekoppelte Rezeptoren, dem A2B- und dem P2Y2-Rezeptor, vermittelt werden kann. Dieser Effekt ist nicht nur auf die In-vitro-Ebene beschränkt, sondern wurde auch erfolgreich in einem Wundheilungsmodell in vivo nachvollzogen. Zu den von BURNSTOCK und WILLIAMS (2000) angesprochenen Indikationsgebieten für die Anwendung purinerger Substanzen ist evtl. die Behandlung von Wunden hinzuzufügen. Es bleibt weiteren Studien vorbehalten zu prüfen, ob sich diese therapeutische Perspektive eröffnet.
6 Zusammenfassung
Karin Lelieur (2004)
Untersuchungen zur Beeinflussung der Keratinozytenproliferation und der Wundheilung über Purinrezeptoren.
Ziel dieser Arbeit war es, die Rolle G-Protein-gekoppelter Purinrezeptoren bei der Regulierung des Keratinozytenwachstums an Mäusen zu untersuchen. Der erste Schritt bestand darin, das Rezeptor-Expressionsprofil in primären murinen Epidermalzellen und in der murinen Keratinozytenlinie MSC-P5 zu etablieren. Es konnte mittels RT-PCR festgestellt werden, dass murine Keratinozyten die Rezeptoren P2Y1, P2Y2, P2Y4 (nur primäre Epidermalzellen) und P2Y6 sowie A2A, A2B und A3 exprimieren. Die purinergen Agonisten ATP, UTP und NECA übten einen proliferationsfördernden Einfluss auf MSC-P5-Zellen aus.
Dieser Effekt konnte für ATP und UTP mit dem P2Y-Antagonisten Suramin und für NECA mit dem A2B-Antagonisten Enprofyllin blockiert werden. Der P2Y6-Agonist UDP zeigte keinen Einfluss auf das Keratinozytenwachstum, während der P2Y1-Agonist 2MeSATP einen geringgradigen proliferationshemmenden Einfluss zeigte. Ein Einfluss auf die Zelldifferenzierung (Keratinmuster) sowie auf die COX-Regulation konnte nicht gezeigt werden. Für Fibroblasten war berichtet worden, dass ATP über die Bildung von PGE2 das Wachstum fördert. Dies konnte in dieser Arbeit für die Keratinozyten nicht bestätigt werden.
Die proliferationsfördernde Wirkung von UTP und NECA wurde in vivo in einem murinen Modell der verzögerten Wundheilung überprüft. Die Behandlung der Wunden mit UTP oder NECA beschleunigte signifikant die Heilung. Der P2Y-Rezeptor Antagonist Suramin hob den UTP-, der A2B-Rezeptor Antagonist Enprofyllin den NECA-Effekt auf.
Purinerge Substanzen könnten sich damit evtl. als therapeutische Perspektive im Zusammenhang mit der Behandlung chronischer Wunden erweisen.
7 Summary
Karin Lelieur (2004)
The role of purinoceptors in keratinocyte proliferation and wound healing.
The influence of G-protein coupling purinoceptors on keratinocyte growth was examined in mice. First, the expression profile of the receptors was determined in both primary murine epidermal cells and the murine keratinocyte line MSC-P5. The RT-PCR technique demonstrated the presence of the receptors P2Y1, P2Y2, P2Y4 (only in primary epidermal cells) and P2Y6 as well as A2A, A2B and A3 in murine keratinocytes. The purinergic agonists ATP, UTP and NECA had a proliferative effect on the MSC-P5 cells. The mitogenic effect of ATP and UTP was inhibited by the non-selective P2Y receptor antagonist suramin, while the effect of NECA was inhibited by the selective A2B receptor antagonist enprofylline. The P2Y6
agonist UDP had no influence on keratinocyte growth, while the P2Y1 agonist 2MeSATP had a slight antiproliferative effect. An influence was neither observed on cell differentiation (keratin expression) nor on COX-regulation. For fibroblasts it had been reported that ATP enhances cell growth via the production of PGE2. For keratinocytes, this was not confirmed by this study.
The proliferative effect of UTP and NECA was also tested in vivo in a murine model of impaired wound healing. The treatment of the wounds with UTP or NECA significantly enhanced the impaired healing. The P2Y receptor antagonist suramin inhibited the effect of UTP, while the A2B receptor antagonist enprofylline inhibited the effect of NECA.
These studies demonstrates that purinergic substances could maybe offer a new therapeutic perspective in the treatment of chronic wounds.
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