Analysis of Cell Death in Meckel's Chondrocytes In Vitro Induced by Heat-Shock Treatment

To clarify whether in vitro cell death in Meckel's chondrocytes induced by heat-stress treatment involves apoptosis or necrosis, we performed morphological examinations using immunohistochemistry including BrdU-incorporation, the TUNEL method, and light and electron microscopy. Cell death was induced in Meckel's chondrocytes isolated enzymatically from 17-day embryonic mice, by exposure to temperatures 40-60°C. On exposure t0 sublethal temperatures of 40-45°C, the incorporation of BrdU was facilitated, and TUNEL-positive apoptotic cells appeared with high frequency. At the ultrastructural level, apoptotic cell death was characterized by chromatin condensation, nuclear segmentation, and the formation of apoptotic bodies. In contrast, cell death induced by lethal temperature (50-60°C) presented decreases in TUNEL-positive cells and cell incorporating BrdU. High temperature-treated cells showed apparent disruption of the membrane of cell organelles. Immunostaining for heat shock proteins (HSP) -27 and-70 revealed that these proteins are secreted continuously at lower temperatures, but have a tendency toward decrease on exposure to lethal temperatures. Our results suggest that Meckel's chondrocytes demonstrate apoptosis at sublethal temperatures, but undergo necrosis at lethal temperatures.