The effect of fixation and histological preparation on coronary artery dimensions

Histometric data are necessary for quantifying the cellular and tissue structure which serves as a basis for the construction of mathematical and integrative models. However, fixation and histological processing of tissues such as dehydration, embedding, sectioning and staining can cause dimensional changes and tissue distortion. The present study was designed to analyze the effects of one widely used fixation and histological preparation protocol for muscle tissue on several morphometric parameters of the coronary arteries. A total of 110 left anterior descending (LAD) artery segments obtained from eight pig hearts were fixed in 6.25% glutaraldehyde and embedded in JB-4 solution. The dimensions of the fixed rings in the loaded and no-load states and histological sections were compared with those of the fresh tissue using Bland-Altman scatter diagrams; i.e., the percent differences between measurements in two different states of various morphometric parameters (inner and outer circumference and wall thickness and area) against their means. We found that vascular elastin cannot be fixed, as seen by the retraction of the vessel dimensions when the loading of the fixed ring was removed. This retraction is time dependent and can lead to significant differences in wall thickness. The differences in dimensions between the histological sections and the fresh tissues in the no-load state were small for the inner and outer diameter (5.6 and 5.2%, respectively) and wall thickness and area (13.4 and 13.1%, respectively). These results are important for establishing an accurate quantitative data base of histological structure that can be related to vascular function or dysfunction.