Studies on the mechanism of cellular death—I. Enzymatic changes during early and late cardiac necroses

Abstract

A battery of 15 different histoenzymological tests were employed to study the enzymatic changes which occur during the death of dog myocardial cells induced by local surgically induced cardiac ischemia and in autopsy specimens of infarcted human myocardium. The sequence of events was observed at intervals from 1 to 480 hr. Succinoxidase activity was apparent both in the longitudinal and transverse axes of the myofibers. In the early necrotic phases, the network arrangement was disorganized and discrete mitochondria were observed at the edge of the infarcts. In later stages these mitochondria were associated with fat droplets. Esterase activity was located on the same particulate elements as succinoxidase but no interconnecting network was evident. In older infarcts these particles increased in size and occurred both within and at the margin of the infarcts. Esterase activities persisted for a longer period of time than did the DPN diaphorase or succinoxidase activity. Cytochrome oxidase activity persisted much longer than did either succinoxidase or DPN diaphorase activity. Acid phosphatase activity using a standard azo dye coupling technique was also particulate and increased during the early stages of an infarct. The particulate acid phosphatase activity was replaced by a smooth, homogeneous staining of the cytoplasm at about 24 hr. Acid phosphatase activity revealed by ASBI as substrate was localized in the lipofuscin inclusions. Leucine aminopeptidase activity was undetectable in early stages of cell death but reached very high persistent levels in the most advanced infarcts. ATPase activity was localized at the blood vessels as well as myocardial tissue elements. The activity increased from 4 to 64 hr after occlusion and it disappeared thereafter. ADPase and alkaline phosphatase activities demonstrated a disruption of the blood vessels in the moderately advanced infarcts. ADPase tests revealed a greater number of vascular elements than did the alkaline phosphatase methods. Leukocyte infiltration was observed at about 24 hr. Cells having positive leucine aminopeptidase activity as well as having acid phosphatase activity by the ASBI method were observed after 24 hr of infarction. Histiocytes, rich in acid phosphatase and esterase activities, appeared at about 144 hr and persisted thereafter. Human material revealed a comparable sequence of histoenzymatic events.