Tissue distribution and phenobarbital induction of target SLC- and ABC- transporters in cattle.

In veterinary pharmaco-toxicological sciences, few data about uptake and efflux drug transporters (DTs) expression and regulation phenomena have been published. In this study, the tissue distribution and transcriptional modulation of solute carrier (SLC) and ATP-binding cassette (ABC) DTs were investigated in cattle orally administered with phenobarbital (PB) by using a quantitative real-time RT-PCR approach. The criterion for target gene selection was the PB-responsiveness in human and rodent model species. All target DTs were expressed in the liver. Only two of the seven PB-responsive target DTs (SLCO1B3 and SLC10A1) were not constitutively expressed in cattle extra-hepatic tissues. The greatest number of DTs (SLCO2B1, ABCB1, ABCC2, ABCG2) were expressed in intestine and testis, followed by, adrenal gland (SLCO2B1, ABCB1, ABCG2), lung (ABCB1, ABCG2), kidney, and skeletal muscle (ABCG2). PB administration never altered DTs mRNA levels, except for an increase in hepatic ABCC2 mRNA and a down-regulation of renal ABCG2. Altogether, these results confirm only to some extent data obtained in humans and laboratory species; clearly, they should be considered a preliminary step for further molecular investigations about species-differences in DT gene expression and regulation as well as in DT expression and function.

Expression of matrix metalloproteinases, tissue inhibitors of metalloproteinases and vascular endothelial growth factor in canine mast cell tumours.

Degradation of the extracellular matrix and angiogenesis are associated with tumour invasion and metastasis in human and canine neoplasia. Matrix metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs) and vascular endothelial growth factor-A (VEGF-A) are key mediators of these respective processes. Mast cell tumour (MCT) is the most common malignant cutaneous tumour in dogs. MCTs are always considered potentially malignant, but their true metastatic potential is unknown. In the present study, samples from seven grade 1, 22 grade 2 and six grade 3 MCTs were subjected to quantitative real-time polymerase chain reaction and immunohistochemistry (IHC) to evaluate MMP-2, MMP-9, membrane-type 1 MMP (MT1-MMP), TIMP-2 and VEGF-A mRNA and protein expression. Gelatin zymography (GZ) was also performed to evaluate MMP-2 and MMP-9 activity. MMP-9 and VEGF-A mRNA increased with histological grade, while TIMP-2 decreased with increasing grade. Gene expression data obtained for MMP-9, VEGF-A and TIMP-2 were confirmed by IHC for evaluation of the respective proteins. In contrast, MMP-2 and MT1-MMP had variable, but similar, expression for both mRNA and protein. Despite the high variability observed, there was correlation between MMP-2 and MT1-MMP mRNA expression (r=+0.91, P<0.0001). The MMP-2:TIMP-2 and MMP-9:TIMP-1 mRNA ratios showed an imbalance between MMPs and their specific inhibitors in MCTs, which increased with the histological grade. Finally, the activities of both latent and active forms of MMP-2 and MMP-9 were evaluated by GZ and there were significant increases in their activities with increasing histological grade and immunohistochemical expression. This study demonstrates that MMP-9, TIMP-2 and VEGF-A expression is related to histological grade and suggests that these markers are possible indicators of malignancy and targets for therapeutic strategies.