Immunhistologische Untersuchung ausgewählter Matrixmetalloproteinasen (MMPs) und deren Inhibitoren in der neoplastisch veränderten Mamma und den assoziierten Lymphknoten von Hunden

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Following the literature section, addressing the morphology, epidemiology, etiology and pathogenetic factors of canine mammary tumours, the experimental design including the study groups is described. These groups include simple adenomas (n= 15) and complex adenomas (n= 19), benign mixed mammary tumours (n= 15), simple carcinomas (n= 19) and complex carcinomas (n= 17) and also physiological (n= 9) and hyperplastic tissues (n= 7). The paraffin sections were immunohistochemically investigated for MMP-7, MMP-11, MMP-12 and TIMP-3 in various structures and localisations. Therefore the avidin-biotin-peroxidase-complex-technique was used (performed by Dr. Vanja Paltian). For MMP-7 and MMP-12 there were applied polyclonal rabbit-anti-human-primary-antibodies and a biotinylated goat-anti-rabbit-secondary-antibody. For MMP-11 and TIMP-3 there were used monoclonal Mouse-anti-human-primary-antibodies and a biotinylated horse-anti-mouse-secondary-antibody. MMP-7: The analysis of the MMP-7 signal for all investigation groups depicted five outstanding structures: the myoepithelial cells, the inflammatory cells, the stromal cells and the blood vessel components “endothelial cells” and “media”. These structures showed the strongest immunohistological signal for MMP-7 in the group of complex adenomas and they were significantly or highly significantly different to the other investigation groups. This finding is in accordance to the literature, where the high expression level of the MMP-7 in stromal cells was also found, dependent on their distance to mammary tumours. This was explained by the induction of MMP-7 expression in stromal cells by tumour cells. The present study confirms these results. It is also well known, that stromal cells influence the expression of MMPs by tumour cells due to paracrine secretion of growth factors and cytokines. Therefore the high MMP-7 expression level of the complex adenomas in this study could maybe explained by induction by cells of the well organized activated stromal cells and capsule. This could be verified in further studies. MMP-11: The group of the benign mixed mammary tumours with its very distinct MMP-11 expression was highly significantly different to the other investigation groups. The alveolar und ductal epithelial cells, the myoepithelial and inflammatory cells, the cortex of the lymph nodes, the endothelial cells of the vessels and the stromal cells showed these outstanding differences in MMP-11 expression. In the vicinity of the tumour and distant to the tumour the benign mixed mammary tumours showed (highly-) significant expression differences in MMP-11 expression in endothelial and stromal cells compared to the other investigation groups. The benign mixed tumors were strongly MMP-11 positive in both localisations of these structures. For the epithelial and inflammatory cells, the strongest MMP-11 signal was depicted in the physiological mamma, followed by the periphery and centre of the tumour. In all tumour groups, especially the benign mixed tumours, there was a higher expression level in epithelial cells compared to the same cell population of the control group. This result was confirmed by others, who found a higher MMP-11 expression in tumour cells than in normal mammary tissue. The prominent expression in this study within metastasized lymph nodes of the simple carcinomas is according to publications describing MMP-11 expression in breast carcinomas with metastatic potential. The outstanding expression in benign mixed tumours is a surprising and not yet published result. There should be further investigations to clear a possible correlation between the expression of MMP-11 and this tumour type. MMP-12: the evaluation of the MMP-12 signal for all investigation groups showed six outstanding structures: the alveolar and myoepithelial cells, the inflammatory cells, the cortex and medulla of the associated lymph nodes and the media of the vessels. The highest MMP-12 level in these structures was found in the groups of the simple carcinomas and the benign mixed tumours. Their expression was highly significantly different to the weaker MMP-12 signals of the other investigated groups. In literature, the MMP-12 expression is associated with tumour associated macrophages, so called TAMs, which are correlated with decreased survival, tumour cell proliferation, angiogenesis in malignant mammary tumours and metastasis. Our results suggest this, as the inflammatory cells in metastasising simple carcinomas showed the highest MMP-12 expression. The control group displayed the strongest MMP-12 expression in the vessel media of all groups. It is known, that MMP-12 can influence the angiogenesis, but an outstanding expression level in the media of blood vessels is not described. A correlation between the MMP-12 signal in the media and angiogenesis is possible, but not proved. TIMP-3: The statistical evaluation for the immunohistological TIMP-3 signal showed highly significant or significant values for the alveolar, ductal and myoepithelial cells, the inflammatory cells, and the media of the vessels of the tumour groups. All tumour groups, but especially the simple carcinomas showed distinct signals for TIMP-3, whereas the control group expressed only weakly. In inflammatory cells, the simple carcinomas showed the highest TIMP-3 expression level. This is according to publications, which found prominent TIMP-3 signals in inflammatory cells within the tumour. Similarly to human studies, the canine mammary tumours seemed to be accompanied by inflammatory cells expressing TIMP-3. In general, in our study there is more TIMP-3 expression in malignant than in benign tumours. This may be due to the inhibitory effect of the enzyme. Thus the highest TIMP-3 values were found in the tumour periphery, the area of tumour growth. In conclusion, in canine mammary tumours the MMP and TIMP expression varies in localizations and structures. A correlation with the tumour type is possible. There should be further studies on canine samples to clear the possibilities of using MMPs or TIMPs in clinical tests or even therapies.

Ce résumé expose les affirmations des auteurs. BNTIC ne l’interprète pas comme une validation indépendante des résultats.