Abstract no.: 10 DNA fragmentation, but not caspase‐3 activation or PARP‐1 cleavage, combined with macrophage immunostaining as a tool to study phagocytosis of apoptotic cells in situ

Abstract

Efficient phagocytosis of cells undergoing apoptosis by macrophages is important to prevent immunological responses and development of chronic inflammatory disorders, such as systemic lupus erythematosus, cystic fibrosis or atherosclerosis. To study phagocytosis of apoptotic cells (AC) by macrophages in tissue, we validated different apoptosis markers (DNA fragmentation, caspase‐3 activation and cleavage of its substrate poly (ADP‐ribose) polymerase‐1) in combination with macrophage immunostaining. Human tonsils were used as a model because they show a high apoptosis frequency under physiological conditions as well as efficient phagocytosis of AC by macrophages. On the other hand, advanced human atherosclerotic plaques were examined since phagocytosis of AC in a plaque is severely impaired. Our results demonstrate that the presence of non‐phagocytized TUNEL‐positive AC represents a suitable marker for poor phagocytosis by macrophages in situ. Other markers for apoptosis, such as cleavage of caspase‐3 or PARP‐1, should not be used to assess phagocytosis efficiency, because activation of the caspase cascade and cleavage of their substrates can occur in AC when they have not yet been phagocytized by macrophages.