Reducing mitochondrial bound hexokinase II mediates transition from non-injurious into injurious ischemia/reperfusion of the intact heart

Abstract

Ischemia/reperfusion (I/R) of the heart becomes injurious when duration of the ischemic insult exceeds a certain threshold (approximately ≥20 min). Mitochondrial bound hexokinase II (mtHKII) protects against I/R injury, with the amount of mtHKII correlating with injury. Here, we examine whether mtHKII can induce the transition from non-injurious to injurious I/R, by detaching HKII from mitochondria during a non-injurious I/R interval. Additionally, we examine possible underlying mechanisms (increased reactive oxygen species (ROS), increased oxygen consumption (MVO2) and decreased cardiac energetics) associated with this transition. Langendorff perfused rat hearts were treated for 20 min with saline, TAT-only or 200 nM TAT-HKII, a peptide that translocates HKII from mitochondria. Then, hearts were exposed to non-injurious 15-min ischemia, followed by 30-min reperfusion. I/R injury was determined by necrosis (LDH release) and cardiac mechanical recovery. ROS were measured by DHE fluorescence. Changes in cardiac respiratory activity (cardiac MVO2 and efficiency and mitochondrial oxygen tension (mitoPO2) using protoporphyrin IX) and cardiac energetics (ATP, PCr, ∆GATP) were determined following peptide treatment. When exposed to 15-min ischemia, control hearts had no necrosis and 85% recovery of function. Conversely, TAT-HKII treatment resulted in significant LDH release and reduced cardiac recovery (25%), indicating injurious I/R. This was associated with increased ROS during ischemia and reperfusion. TAT-HKII treatment reduced MVO2 and improved energetics (increased PCr) before ischemia, without affecting MVO2/RPP ratio or mitoPO2. In conclusion, a reduction in mtHKII turns non-injurious I/R into injurious I/R. Loss of mtHKII was associated with increased ROS during ischemia and reperfusion, but not with increased MVO2 or decreased cardiac energetics before damage occurs.

The role of succinate and ROS in reperfusion injury – A critical appraisal

“…Rat hearts were perfused with low concentrations (200 nM) of a membrane permeable peptide (TAT-HKII) that dissociates bound HK2 from mitochondria and which was without significant effect on normoxic hearts. However, after 15 min ischemia, when reperfusion of control hearts caused only reversible injury (impaired hemodynamic function), the presence of the TAT-HKII peptide caused irreversible injury (lactate dehydrogenase release) [151] .…”

Hexokinase-2 Glycolytic Overload in Diabetes and Ischemia–Reperfusion Injury

“…Recent research suggests that abnormal increase of HK2 plays a key role when control of glycolysis is impaired, as in unscheduled glycolysis. In this case, abnormal increase in G6P and downstream glycolytic intermediates are key players in the tissue-specific chronic pathogenesis of hyperglycemia associated with diabetes, ischemia-reperfusion injury, and potentially, in cell senescence [14][15][16]. We call this damaging condition glycolytic overload.…”

“…The extent of mitochondrial dissociation of HK2 during ischemia correlated with mitochondrial cytochrome c release and related cell death, reactive oxygen species (ROS) production, and infarct size on reperfusion [34]. A cell-permeable peptide containing the HK2 mitochondrial binding motif that induced displacement of HK2 from mitochondria, TAT-HK2 (TAT peptide, GRKKRRQRRRPQ, is from the transactivator of transcription of HIV), exacerbated cardiac reperfusion injury [15], suggesting that HK2 displacement from mitochondria is a contributing feature to the pathogenesis. HK2 protein was increased and increasingly displaced from mitochondria in ischemia-reperfusion injury of patients with diabetes.…”

NADPH-oxidase 2 is required for molecular adaptations to high-intensity interval training in skeletal muscle.

“…Hexokinase II is known to shuttle to and from mitochondria [49]. Interestingly, acute disruption of HKII binding to mitochondria, using a peptide-inhibitor in a perfused mouse heart ischemia-reperfusion injury model, markedly reduced cardiac recovery and increased ROS during ischemia and reperfusion [50]. Similar links have been established in other tissues, including skeletal muscle [51].…”