Physiological significance of pyruvate kinase isozymes in erythrocyte.

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Pyruvate kinase (PK) is a rate-limiting enzyme of the glycolytic pathway and PK deficiency is one of the most prevalent causes of hereditary non-spherocytic hemolytic anemia. The Pk-1^<slc> mouse is found to be a spontaneous mutant of the murine LR-type PK gene, and manifests hemolytic anemia and splenomegaly. We showed that apoptotic cells were notably increased in spleen of the Pk-1^<slc> by the TUNEL method, and that the erythroid-specific expression of normal PK gene ameliorated apoptosis in the genetic rescue experiment. Flow cytometric analysis showed that Annexin V / Ter119-double positive cells were significantly increased in the Pk-1slc mouse compared to wild type CBA mice, confirming apoptotic cells were of erythroid lineage. In this study, we examined a physiological role of glycolysis upon apoptosis of the erythroid cells using two Friend erythroleukemic cells, SLC and CBA.These cell lines have been establushed from the Pk-1^<slc> and a wild-type CBA mouse, respectively. As previously reported, apoptosis spontaneously occurred in the Friend cells and was inducible in both cells by glucose deprivation or 0.1-20mM 2-deoxyglucose (2DG). SLC cells were more susceptible to apoptosis by either shorter exposure to glucose deprivation or lower 2DG concentration. The detailed mechanism are still unclear, but we have shown that reactive oxygen species (ROS) might be accountable for these apoptotic changes.