Collagen modifications predictive of lymph node metastasis in dogs with carcinoma in mixed tumours.

Introduction: Mixed tumours in the canine mammary gland are the most common histological type in routine diagnosis. In general, these neoplasms have a favourable prognosis that does not evolve into metastatic disease. However, some cases develop into lymph node metastases and are associated with worse patient survival rates. Methods: Here is a retrospective study of 46 samples of primary mixed tumours of the canine mammary gland: 15 cases of benign mixed tumours (BMT), 16 cases of carcinoma in mixed tumours without lymph node metastasis (CMT), and 15 cases of carcinomas in mixed tumours with lymph node metastasis (CMTM). In addition, we selected 23 cases of normal mammary glands (NMT) for comparison. The samples were collected from biopsies performed during nodulectomy, simple mastectomy, regional mastectomy, or unilateral/bilateral radical mastectomy. We used multiphoton microscopy, second harmonic generation, and two-photon excited fluorescence, to evaluate the characteristics of collagen fibres and cellular components in biopsies stained with haematoxylin and eosin. We performed Ki67, ER, PR, and HER-2 immunostaining to define the immunophenotype and COX-2. We showed that carcinomas that evolved into metastatic disease (CMTM) present shorter and wavier collagen fibres as compared to CMT. Results and discussion: When compared to NMT and BMT the carcinomas present a smaller area of fibre coverage, a larger area of cellular coverage, and a larger number of individual fibres. Furthermore, we observed a correlation between the strong expression of COX-2 and a high rate of cell proliferation in carcinomas with a smaller area covered by cell fibres and a larger number of individual fibres. These findings highlight the fundamental role of collagen during tumour progression, especially in invasion and metastatic dissemination.