SIMULTANEOUS IN-SITU HYBRIDIZATION AND TUNEL TO IDENTIFY CELLS UNDERGOING APOPTOSIS

Apoptotic cells in tissue sections can be localized by in situ labelli ng of partly degraded DNA. Ina heterogeneous population of cells, howe ver, the specific identity of cell types undergoing apoptosis often ca nnot be reliably achieved at the light microscope level because of the marked alterations in cellular morphology that characterize apoptosis . In order to clearly specify cell types undergoing apoptosis, in situ end labelling has been coupled to immunohistochemistry. This method i s limited by the availability of antibodies that bind to cell-specific protein markers in tissue sections. In contrast, we describe a method that combines in situ end labelling with in situ hybridization, a tec hnique that specifies cell types based on mRNA expression. Taking adva ntage of the specific expression of surfactant protein C mRNA in type II alveolar epithelial cells, we demonstrate that this technique has t he ability to localize alveolar type II cells undergoing apoptosis in vivo after the intratracheal instillation of an antibody that activate s the cell surface Fas protein. The wide availability of cell-specific gene markers suggests that this method can be adapted to define cell types that undergo apoptosis during various physiological and patholog ical states in vivo.