Alternative techniques to osmium tetroxide to detect fat embolism in cetacean lungs

Abstract

The presence of fat emboli within pulmonary vessels has been described in cetaceans suffering a violent trauma, like ship collisions, or presenting a decompression like sickness in temporal and spatial association with antisubmarine sonar activities. Postfixation with osmium tetroxide in paraffin-embedded tissues is the technique traditionally used to histologically detect these fat emboli. However, its high toxicity together with the elimination of part of the emboli due to the washes with picric acid pose the need to look for different fixation and staining techniques. The objective of this work was to propose alternative techniques that allow an adequate diagnosis of fat emboli in addition to being less toxic. For this purpose, lungs from sperm whales killed by ship collisions were analyzed and treated with osmium tetroxide and two alternative techniques. These two techniques were: 1) Frozen tissue samples, which were preserved in PBS sucrose solution during weeks before being cut with a cryostat, 2) Fixation with chromic acid for tissues embedded in paraffin sections. Chromic acid is less toxic than osmium tetroxide and it has proven to be effective in diagnosing fat embolism in human crushing injuries and atherosclerosis. In both methods, lipids were stained with Oil Red O. Results showed that both techniques detect fat emboli successfully in the lungs and can be routinely used to diagnose fat embolism in cetaceans.