Distribution of TNFα-like proteins correlates with some regions of programmed cell death in the chick embryo

Early chick embryos have previously been shown to express tumor necrosis factor-alpha-cross-reactive proteins (TNFα-CRPs) in a developmentally regulated manner, thus implicating these proteins in programmed cell death and in tissue remodeling. In this study, cells undergoing DNA fragmentation have been identified, using terminal deoxynucleotide transferase (TdT) mediated dUTP-biotin nick-end-labeling (TUNEL), during the embryonic development of the chick, between stages 18 and 29. DNA fragmentation is indicative of cells undergoing programmed cell death. TUNEL-positive cells were identified in several well documented areas of programmed cell death, including the limb buds, the heart, spinal motoneurons, dorsal root ganglia, and the ventral horn of the neural tube. In addition, other areas of cell death were identified including the floor plate and the mesonephros. In several locations, a close correlation was noted between the presence of TUNEL-positive cells and regions of TNFα-immunoreactivity. These regions included the ventral horn and marginal zone of the neural tube, spinal motoneurons, paravertebral ganglia, parts of the myotome, mesenchyme of the body wall, and the mesonephros. In addition, using the TNFα-sensitive L929-8 bioassay it was shown that homogenate of stage 18 chick embryos is cytotoxic to L929-8 cells and that this toxicity can be reduced using neutralizing antibodies to mouse TNFα. This bioassay allowed us to estimate the mean concentration of TNFα-like activity in embryo homogenate, which is within the range of physiological (pg/ml) levels of TNFα found in other systems. These results suggest that proteins with TNFα-like activity may have a role in programmed cell death in some tissues during early chick embryo development.