Ischemic preconditioning increases myocardial O-GlcNAc glycosylation

Abstract

Objectives. Through the hexosamine biosynthetic pathway (HBP) proteins are modified by O-linked-β-N-acetylglucosamine (O-GlcNAc), which acts as a stress sensor. Augmentation of O-GlcNAc confers cardioprotection against ischemia- reperfusion injury, but its role in ischemic preconditioning (IPC) is unknown. Azaserine and alloxan are unspecific blockers of the HBP and have been used to block the cardioprotective effects of O-GlcNAc. We hypothesized that IPC reduces infarct size and increases O-GlcNAc levels in hearts subjected to ischemia-reperfusion injury, and that these effects could be blocked by azaserine and alloxan. Design. Isolated rat hearts subjected to 40 min global ischemia and 120 min reperfusion were randomized to control, IPC, IPC + azaserine or alloxan, or control + azaserine or alloxan. The effects on infarct size, hemodynamic recovery, myocardial O-GlcNAc levels, and HBP enzyme activities were determined. Results. IPC reduced infarct size, increased O-GlcNAc levels, O-GlcNAc-transferase levels, and O-GlcNAc-transferase activity. Azaserine and alloxan did not block the effect of IPC on O-GlcNAc levels and O-GlcNAc-transferase activity. Conclusions. IPC increased O-GlcNAc levels though increased O-GlcNAc-transferase expression and activity. Azaserine and alloxan failed to block these effects presumably due to poor specificity and sensitivity of the blockers, and IPC-mediated cardioprotection may therefore still be dependent on O-GlcNAc.

Key words::

• alloxan
• azaserine
• O-GlcNAc
• ischemic preconditioning
• ischemia-reperfusion injury

Acknowledgements

We wish to thank the Core C4 (NHLBI P01HL107153) at the Johns Hopkins University School of Medicine for the gift of O-GlcNAcase antibody (345; G.W. Hart) and O-GlcNAc antibody (Natasha Zachara).

Funding

This work was supported by the Danish Research Council (11-108354), The Danish Strategic Research Council (11-1115818), and Institute of Clinical Medicine, Aarhus University, and American Heart Association (SD930162N to N.E.Z) and the National Heart Lung and Blood Institute (R21-HL-108003 and PO1HL107153 to N.E.Z.)

Declaration of interest: The authors report no declarations of interest. The authors alone are responsible for the content and writing of the paper.

We greatly appreciate the technical assistance from Anja Helveg Larsen and Casper Elkjær.