Hyperglycemia Primes Cells for Programmed Cell Death Shift in a Glycolysis-Dependent Manner

Necroptosis is a RIP1-dependent programmed cell death (PCD) pathway distinct from apoptosis. Downstream effector pathways of necroptosis include formation of advanced glycation end products (AGEs) and reactive oxygen species (ROS), both of which are influenced by glycolysis, suggesting that increased cellular glucose may prime necroptosis. Here, we establish that exposure to hyperglycemic levels of glucose enhances necroptosis in U937 monocytes. Pharmacologic or siRNA inhibition of RIP1 prevented the enhanced death, confirming it as RIP1-dependent necroptosis. Hyperglycemia induces a shift from apoptotic death to necroptosis in response to the extrinsic apoptotic stimulus TNFα. Increased membrane permeability and decreased phosphatidylserine exposure were seen in the hyperglycemic condition as compared to euglycemia. Additionally, cell death was shown to be caspase dependent under euglycemic conditions and shifted to RIP1 dependency in hyperglycemic conditions. Total levels of RIP1, RIP3, and MLKL increased following treatment with high levels of glucose suggesting a potential positive feedback mechanism. Hyperglycemic enhancement of necroptosis depends upon glycolysis with AGEs and ROS playing a role. Furthermore, the hyperglycemic state resulted in dysregulated calcium flux in a ROS dependent manner. Hyperglycemia resulted in increased infarct size in a mouse model of brain hypoxia-ischemia injury. The increased infarct size was prevented by inhibition of RIP1 with nec-1s, strongly suggesting that increased necroptosis accounts for exacerbation of this injury in conditions of hyperglycemia. This work reveals that hyperglycemia represents a condition in which cells are extraordinarily susceptible to necroptosis, that local glucose levels alter the balance of PCD pathways, and that clinically relevant outcomes may depend on glucose-mediated effects on PCD.