Beschreibung:
<jats:p>Glycine protects hepatocytes from injuries caused by toxins, hypoxia and ischemia. However, the mechanisms by which glycine exerts its cytoprotection remain elusive. In excitable cells like neurons, glycine binding to its receptor can trigger influx of chloride. Although the existence and molecular composition of a glycine receptor in non‐excitable cells is not clear, cell lines with no expression of glycine receptor proteins have either attenuated or no protection by glycine (Pan <jats:italic>et al.</jats:italic>, <jats:italic>Biochem. J.</jats:italic> 390, 447). Another model has been proposed that protection is through blockage of non‐specific plasma membrane pores by glycine (Nishimura and Lemasters, <jats:italic>Cell Death Differ.</jats:italic> 8, 850). To further understand the mechanism of glycine protection, we studied killing of rat hepatocytes by carbonyl cyanide m‐chlorophenyl hydrazone (CCCP), a mitochondrial uncoupler. After CCCP (15 μM), hepatocytes started to die in a progressive manner after a latency time of 15–95 min. After 6 h, necrotic cell death approached 100% by the criterion of nuclei staining with propidium iodide (PI). Glycine (3 mM) strongly protected rat hepatocytes against CCCP toxicity: cell killing was both delayed and decreased to approximately 30% over 6 h. After expression of a genetically encoded chloride biosensor, no significant difference in chloride concentration was observed with and without glycine after CCCP. However, glycine caused a dramatic change in cell death behavior. In the absence of glycine, hepatocytes died in a membrane‐bursting manner, and fluorescence of the chloride biosensor rapidly disappeared, indicating leakage of cellular contents. In the presence of glycine, hepatocytes began to shrink substantially, beginning at approximately the same time necrotic cell death would otherwise have occurred. After CCCP in the absence of glycine, plasma membrane staining by Alexa488‐Annexin V occurred simultaneously with PI staining of nuclei. With CCCP plus glycine, hepatocytes first stained by Annexin V as shrinkage began and only hours later by PI. Rhodamine‐dextran labeling of lysosomes showed that lysosomal leakage accompanied cell shrinkage and Annexin V labeling in the presence of glycine. These findings indicate that features of apoptotic cell death (Annexin V labeling, cell shrinkage) emerge when onset of necrotic cell death is suppressed by glycine in ATP‐depleted hepatocytes. Lysosomal permeabilization may promote the apparent activation of apoptotic pathways.</jats:p>