Sanne de Bruin

130 Chapter 5 Glycolysis and Luebering-Rapoport shunt The glycolysis and Luebering-Rapoport shunt are important pathways for the genera- tion of ATP and 2,3-DPG respectively. However, both ATP as well as 2,3-DPG are rapidly depleted during storage. This is more pronounced than during in vivo aging, especially for 2,3-DPG levels. Throughout storage 2,3-DPG levels rapidly decline up to 99% com- pared to baseline level 35–38 while in vivo 2,3-DPG levels decline up to 42% in the oldest cell fraction 21,22,26 . Due to the importance of these metabolites, new storage techniques aim to improve this, for instance by changing the composition of the additive solutions. In this review a distinction is made between non-alkaline additive solutions (e.g. SAGM, AS-1, AS-5) and alkaline additive solutions (e.g. AS-7 and PAGGGM). The latter are devel- oped to maintain ATP and 2,3-DPG levels better during cold storage. In non-alkaline additive solutions glycolysis is active during the first two weeks after which activity decreases resulting in lower ATP levels at the end of storage 35,38,39 . Glu- cose is rapidly converted into glucose-6-phosphate after uptake into the cells. After this, glucose-6-phosphate is converted via multiple steps into 2 molecules of glycer- aldehyde-3-phosphate. In the first week of storage, these early glycolysis substrates increase but are almost completely depleted after two weeks 40 . In the second phase of the glycolysis, glyceraldehyde-3-phosphate is converted via five steps into pyruvate with a yield of 2 ATP molecules. In non-alkaline additive solutions, the glycolysis inter- mediates in these conversions: 1,3-diphosphoglycerate (1,3-DPG), 3-phosphoglycerate, 2-phosphoglycerate and phosphoenolpyruvate are all significantly decreased at the end of storage while pyruvate and lactate are accumulating 36,40 . Initially ATP levels remain constant during the first two-three weeks, but after this period ATP levels decline ac- companied by increased levels of ATP breakdown products 35,41 . In non-alkaline storage solutions, 2,3-DPG levels decrease rapidly within two weeks with 99% compared to baseline level 36,37 while in vivo 2,3-DPG levels declined up to 42% in the oldest cell fraction 21,22,26 . The current opinion is that decreased glucose flux through the glycolysis results in less substrate for the Luebering-Rapoport shunt and thus less 2,3-DPG production. A second explanation could be increased breakdown of 2,3-DPG. There are indications that diphosphoglycerate mutase, the enzyme responsible for the conversion from 1,3-DPG into 2,3-DPG, is also able to catalyze the reverse reaction. Ac- cording to an in silico model, 2,3-DPG could be reconverted by this enzyme to 1,3-DPG 42 . However, this is not yet confirmed in vivo or in vitro . Thus ATP and 2,3-DPG are depleted during storage in non-alkaline additive solutions. The current hypothesis is that this is due to decreased pH caused by lactate accumu-