Gene expression and immunohistochemical detection of ki67 and Bcl-2 in canine mast cell tumours.

Mast cell tumors (MCT) are one of the most frequently occurring malignant tumor in dogs, with variable biologic behaviours and response rates to chemotherapy. In this study, the prognostic significance of Ki67 and Bcl-2 expression has been investigated. Thirty MCTs, were surgically removed and fixed in 10% buffered formalin for histological evaluation and grading determination. Biopsies of each tumor and its surgical margins were sampled and stored in RNA-later solution. Immunohistochemistry was performed against Ki67 and Bcl-2 proteins, and positive cells were counted in a total of 1,000 cells. Total RNA from biopsies was isolated. Real Time RT-PCR assays for canine Ki67, Bcl-2 and 2 housekeeping genes were set up by using UPL probes and validated for relative quantification. Finally, gene expression profiles were measured in tumor and margin biopsies. Ki67 was induced more than 2-fold (up to 50-fold) towards control canine tissue in 15 cases, while Bcl-2 was generally inhibited. Ki67 gene expression was significantly correlated with protein detection by immunohistochemistry, while this association was not demonstrated for Bcl-2. Bcl-2 protein expression in neoplastic cells was correlated with tumor grading, being significantly higher in grade 2 MCT. No correlation was observed between Bcl-2 gene expression and tumor grade. Further investigations are required to better understand the role of these proteins in tumor progression.