Calculation of Myocardial Percentage Collagen, Mast Cell Count and Presence of Arteriosclerosis in Dogs With...

This study aimed to (1) determine the feasibility of calculating percentage myocardial collagen (PMC) in formalin-fixed, paraffin-embedded canine tissue samples, (2) explore relationships between PMC, mast cell (MC) count and myocardial arteriosclerosis/arteriolosclerosis (MA) and (3) calculate the sample size required to compare PMC between dogs with and without myxomatous mitral valve disease (MMVD).

Histological sections were prepared from retrospective samples of formalin-fixed, wax-embedded ventricular myocardial tissue from 15 dogs with, and seven dogs without, MMVD. Sections from each sample were stained with Masson's trichrome, haematoxylin and eosin (H&E) or toluidine blue. In each Masson's trichrome-stained section, digital images of 12 random fields were captured and the total image area and total collagen area were measured using computer-assisted morphometric analysis. Mean collagen area, mean total image area and mean PMC ([mean collagen area/mean total area] × 100) were calculated per sample. MCs were counted manually in toluidine blue-stained sections and the mean MC count was calculated from 10 fields. MA was identified by visual inspection of transversely cut vessels in each H&E-stained section.

No significant difference was detected between MMVD and control dogs in PMC (p = 0.063), MC (p = 0.476) or MA (p = 0.172). A sample size calculation suggests that data from at least 26 MMVD dogs and 26 controls are required to detect a difference in PMC. No significant association was detected between PMC and MC count (p = 0.606) in cases of MMVD.

The methods described are feasible and the sample size for a definitive study has been estimated.