Abstracts - DGPRÄC

Abstracts
P35 � Angiogenic Response in Matrix Metalloproteinase-
19-Deficient Mice: An Intravital Analysis in Dorsal Skinfold
Chamber
Ring A 1,2 , Goertz O 2 , Muhr G 1 , Steinau H-U 2 , Steinsräßer L 2 , Sedlacek R 3,4 , Langer St 2
1 Department of Surgery, Trauma Center, University Hospital Bergmannsheil Bochum, 2 Department of
Plastic Surgery and Hand Surgery, Burn Center, Sarcoma Reference Center, University Hospital
Bergmannsheil Bochum, 3 Institute of Biochemistry, Faculty of Medicine, Christian-Albrechts-University
Kiel, 4 Department of Transgenic Models of Diseases, Institute of Molecular Genetics, University of Prague,
Czech Republic
Background: Zinc-dependent matrix metalloproteinases are supposed to
play an important role in developmental processes. These enzymes
mediate the restructuring of extracellular matrix, and control the activity
of gowth factors, chemikines and cytokines.
To investigate the impact of MMP-19-deficiency on tumor-induced angiogenesis
in vivo, the angiogenic response of mutant mice lacking MMP-19
was analyzed in dorsal skinfold chamber by means of intravital fluorescence
microscopy.
Materials and methods: Mice genetically deficient in MMP-19 and corresponding
wild type were used. Following chamber implantation, murine
melanoma cells were seeded on the top of striated skin muscle layer.
Tumor-induced angiogenesis was analyzed. Visualization of new vessel
growth was performed using intravital flourescence microscopy. Functional
vessel density (FVD) reflecting the neoformation rate of perfused
microvessel network, served as a parameter for vascularization. The
experiment was conducted in accordance with the German law for the
protection and the welfare of laboratory animals.
Results: Tumor growth and formation of new microvasculature occured in
both groups. Tumor cells induced the develompent of vessel sprouts that
rapidly infiltrated the seeded muscle and invaded the host tissue by remodelling
the mature microvasculature. However, the neoformation of
tumor-induced vasculature was comparatively increased in MMP-19deficient
animals. Here, the FVD was found significantly higher on day
12.
Conclusion: Based on intravital dynamic analysis of vessel growth and quantification
of FVD, we demonstrated that host MMP-19-deficiency is assosiated
with an increased tumor-induced angiogenic response. This finding
P36 � In Vivo Reconstruction of Nitric oxide induced
Desensitization of NO/cGMP Signalling Pathways in Dorsal
Skinfold Chamber
Ring A 1,2 , Goertz O 2 , Mullershausen F 3,4 , Koesling D 3 , Muhr G 1 , Steinau H-U 2 ,
Steinsträßer L 2 , Langer St 2
1 2 Department of Surgery, Trauma Center, University Hospital Bergmannsheil Bochum, Department of
Plastic and Hand Surgery, Burn Center, Sarcoma Reference Center, University Hospital Bergmannsheil
Bochum, 3Department of Pharmacology and Toxicology, Faculty of Medicine, Ruhr-University Bochumy,
4G-Protein-Coupled Receptors Expertise Program, Novartis Institutes for Biomedical Research, Novartis
Pharma AG Basel
Background: The NO/cGMP pathway plays a crucial role in regulation of
tissue perfusion. The use of NO-donors in reconstructive surgery is supposed
to be a pharmacological option for protecting ischemic tissue during
critical perfusion conditions. However, an NO-induced desensitization
of cGMP-mediated relaxation has been reported in isolated tissue. To
examine whether a similar phenomenon can be detected in vivo, we ana-
39. Jahrestagung der Deutschen Gesellschaft der Plastischen, Rekonstruktiven und Ästhetischen Chirurgen
13. Jahrestagung der Vereinigung der Deutschen Ästhetisch-Plastischen Chirurgen
lyzed relaxations of microvessels in response to repeated administration
of NO in dorsal skinfold chamber.
Materials and methods: The investigations were performed by means of dynamic
intravital flourescence microscopy in the dorsal skinfold chamber of
balb/c mice.
First, a maximal preconstricted mirovasculature was produced by incubation
the skin muscle with a vasoconstrictor, the 5-Hydroxytryptamine
(5-HT). Subsequently, relaxation was induced by
applying an NO-donor, the S-Nitrosoglutathione(GSNO), to the contracted
vessels. Following buffer exchange, constriction and relaxation were
repeated. Drugs were given topically into the chamber, directly onto the
skin muscle. Special interest was given to arterioles. The response of
microvasculature to the topical administration of vasoactive drugs was
determined as the change of the diameter of arterioles and quantified
using standard software. The experiment was conducted in accordance
with the German law for the protection and the welfare of laboratory animals.
Results: NO-induced relaxation of preconstricted arterioles was reduced
after the repetitive application of NO-donor. The short pretreatment with
NO entailed a reduced relaxation of arterioles in response to following
NO applications. The absolute change in vessel diameter induced by
GSNO was significantly reduce d from 21 to 16 micrometers after the first
and the second application, respectively. However, the data revealed also
a noticeable reduction of the constricting activity of 5-HT during the
second application, indicating a possible desensitization of the 5-HT response
or and/or neuronal cempensatory mechanisms.
Conclusion: The cGMP-mediated relaxation of microvessels was quantified
by means of intravital microscopy using the dorsal skinfold chamber
model. The phenomenon of NO-induced desensitization was reconstructed
and visualized for the first time within an intact in vivo system.
This finding indicates that NO-induced desensitization may play an
important role during NO-treatment in critical perfused tissue.
P 37 � Das angiogenetische Potential des non-viral
gentransfizierten Natriuretischen Peptids Typ-C (CNP) im
Wundmodell der SKH-1 HR/-Maus
Kühnl A, Pelisek J, Goertz O, Eckstein H-H, Jauch K-W, Hatz R, Steinau H-U, Langer St
BG Universitätskliniken Bergmannsheil, Bochum
Für CNP wurde ein proliferationssteigender Effekt auf Endothelzellen
sowie ein angiogenetisches Potential im ischämischen Muskelgewebe
nachgewiesen. Untersuchungen in wunden sind bisher noch nicht durchgeführt
worden. Ziel dieser Studie war die Etablierung einer neuen
Methode zum dermalen, non-viralen Gentransfers von CNP in einem
Wundmodell der SKH1 harrlosen Maus sowie die Untersuchung des therapeutischen
Potentials des CNP auf die dermale Wundheilung.
Material und Methoden: Als Plasmidgerüst diente der DsRed-Monomer-C1
Reportervektor in den das therapeutische Gen CNP einkloniert wurde.
In-vitro wurden Mäusefibroblasten unter Verwendung von linearem
Polyethylenimin (PEI) mit oben genanntem Plasmid transfiziert. In-vivo
erfolgte vor bzw. nach Setzen einer standardisierten Wunde am Ohr der
SKH-1 Maus die Applikation des Plasmids, entweder durch intradermale
Applikation (10 µl Hamilton 26 G) (Gruppe 1, n=4), durch subkutane
Injektion in den Wundrand (Gruppe 2, n=6) oder durch einfache Benetzung
der Wunde (Gruppe 3, n=6). Der Expressionsnachweis des rot
fuoreszierenden Reporterproteins sowie des therapeutischen Gens
erfolgte in-vitro und Fluoreszenzmikroskopie in-vivo (IFM) durch
Immunhistologie und RT-PCR. Die dynamischen mikrozirkulatorischen
Parameter (Funktionelle Kapillardichte) wurden anhand von online
Videosequenzen untersucht.
74 Plastische Chirurgie 8 (Suppl. 1): 74 (2008)