Untersuchungen zur Bedeutung der Interaktion zwischen Tumor- und Stromazellen bei der Invasivitaet von Plattenepithelkarzinomen des Kopf- und Hals-Bereiches anhand der Expressionsverteilung von Matrix Metalloproteinase-9

Die Inzidenz von Plattenepithelkarzinomen des Kopf-Hals-Bereiches (HNSCC) hat in den letzten Jahren stark zugenommen. Diese Tumorentität zeichnet sich durch einen invasiven Charakter, die Neigung zur frühzeitigen Metastasierung und eine schlechte Prognose aus. Um neue Therapiekonzepte zu erarbeiten...

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Bibliographic Details
Main Author: Kulle, Carolin
Contributors: Mandic, Robert (Dr.) (Thesis advisor)
Format: Dissertation
Published: Philipps-Universität Marburg 2008
Hals- Nasen- und Ohrenheilkunde
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Table of Contents: The incidence of head and neck squamous cell carcinomas has steadily increased during the last years. This tumor entity is characertized by its invasive character, its tendency for early metastasis and its poor prognosis. To achieve an optimal treatment one needs to understand the complex biological mechanisms of invasion and metastasis. The family of zinc-dependent endopeptidases, the matrix metalloproteinases, represent a group of proteins which are capable of remodelling the extra cellular matrix. They contribute to the intra- and extravasation and to the local invasion. In this thesis the co-expression of several MMPs in HNSCC-cell lines and tissue samples was investigated using western blot and immunohistochemical methods. The tumor-stroma-interference of tumor tissues as well as HNSCC-cell lines was regarded with special interest by means of immunohistochemistry and immunozytochemistry. In addition, the influence of the in vitro invasion on the MMP-9 expression and activity was examined. For this purpose co-cultures of two cell lines were applied. The gelatine zymography was used to determine the activity of the gelatinase MMP-9. The in vitro invasion of cell lines was quantified by means of electrical resistance breakdown assay (ERBA). The experiment has been carried out on 12 HNSCC-cell lines and 15 HNSCCtissue samples. The expression of MMP-1, -3, -7, -9, -11 und -15 in HNSCC-cell lines and tissue samles was shown by immunohistochemical staining and WB analysis. The immunohistochemical analysis revealed MMP expression in tumor cells as well as in fibroblasts from the surrounding tissue. Healthy oral mucosa served as control tissue and displayed in immunohistochemical staining less MMP expression compared to the analyzed tumor tissues. Immunohistochemical staining with MMP-9 clearly indicated an increased expression at the tumor-stroma-interference. Based on this the role of the gelatinase MMP-9 in HNSCC was further investigated. Immunozytochemical analysis of the tumor-stroma contact zone, which was imitated using a co-culture of fibroblasts and tumor cells in vitro, followed. Partially NIH-3T3 fibroblasts expressed MMP-9 stronger than tumor cell lines. The immunofluorescence staining sporadically detected an increased MMP-9 expression at the tumor-stroma-interference site. However, WB analysis did not show an increased MMP-9 expression after co-culturing the HNSCCcell lines with NIH-3T3. In addition, the zymography revealed no significant gelatinolytic activity in any of the samples. The invasiveness of single cell-lines was compared to invasiveness of co-cultured cells by the means of ERBA. The co-culture with fibroblasts did not result in a change of invasiveness. It was observed that HNSCC-cell lines altogether grew less invasive in vitro than in vivo. The gained results suggest that an induction of the MMP-9 expression at the invasive tumor front is present in vivo. This observation indicates an interaction between tumor cells and the surrounding tissue, which seems to play a role in the MMP secretion. However, it could not be managed to induce the MMP-9 expression in cell lines by means of in vitro invasion. Presumably, numerous factors from the surrounding tissue play a role in the invasion, metastasis and in the induction of the MMP-9 expression, which cannot be copied sufficiently by co-culture. The present study contributed to the elucidation of the importance of MMPs in HNSCC. In summary, MMPs are expressed in HNSCC-cell lines as well as in tumor tissues. A difference in the MMP expression between tumor- and control tissue with a greater expression in tumor tissues could be observed. Furthermore, MMP-9 production seems to be intensified at the tumor margins. This can be interpreted as an evidence for an interaction between tumor cells and the surrounding tissue. The degradation of the adjacent tissue and tumor growth takes place at the aggressive tumor front. The influence of the MMP-9 expression at the invasive tumor front could - although only sporadically - be shown for co-culture of fibroblasts with tumor cells. This circumstance implies that the interaction between multiple factors from the tumor cells themselves as well as from the surrounding tissue is essential for invasion and metastasis. In order to improve diagnostic tools and therapies it is essential to explore the tumorstoma interference in further studies. The development of a feasible, lifelike model suitable for HNSCC, which is capabale of mimicing the invasion of tumor cells into surrounding healthy tissue, is desirable to increase our understanding of this process.