Congenital cardiac defects disrupt circulation and oxygenation due to structural changes in the heart. Tetralogy of Fallot is the most common cyanotic congenital heart disease, accounting for 5 to 7% of cases. Depending on cardiac abnormalities, infants with Fallot’s syndrome are classified as “pink babies” with normal saturation (>94%) or “blue babies” with severe oxygenation reduction (saturation as low as 70%), leading to cyanosis. We hypothesize that cardiomyocytes from patients exposed to low oxygen saturation exhibit increased levels of O-GlcNAcylation, enhancing their survival potential by modulating cellular metabolism and function.

To clarify the role of O-GlcNAc in the survival of these patients and to identify new key O-GlcNAcylated proteins in the response to hypoxia. Isolation and dissection of cardiac cells have been performed to observe at cellular level the impact of hyper O-GlcNAcylation. The unique access to sample allows us to validate the isolation of cardiomyocytes from normoxemic vs hypoxemic patients and assess the impact of hyper O-GlcNAc observed in hypoxemic patients on cellular function, particularly mitochondrial function.

After surgical resection, heart samples from infants are digested in a type II collagenase solution for 35 to 50 minutes to obtain dissociated and viable cardiomyocytes. Enzymatic digestion is stopped with a serum-rich medium. Next, gradual calcium increases are initiated. The viability of cardiomyocytes is assessed using Ionoptix, and cellular function after hypoxia is measured.

The acquisition of viable cardiomyocytes from heart samples allows us to expose them to hypoxia in order to assess their susceptibility to cell death and to understand mechanisms induced. The impact of O-GlcNAc modulation on proteins, including SUCLA2 and PSMD11, has been evaluated. Treatment with 10 μM NButGT effectively increased O-GlcNAc level in cardiomyocytes and may protect them to damages associated with hypoxia by the modulation of HIF-1a signaling pathway.

Increasing O-GlcNAc levels could improve cellular function and survival. From a therapeutic perspective, it could be beneficial to consider increasing O-GlcNAc levels in patients identified as being at risk during surgery, especially those who may develop complications related to hypoxia.