Abstract
A major complication of cardiac surgery, such as coronary artery bypass grafting, is the unavoidable imposition of ischaemic episodes followed by reperfusion. The ischaemia– reperfusion injury (IRI) produced can exacerbate myocardial damage and loss of contractile function. Calcium/calmodulin-dependent protein kinase II (CaMKIIδ) activation during IRI is heightened due to increased acidity and ROS production leading to subsequent autonomous activity. oCOm21 is a highly unstable species that rapidly decomposes to release carbon monoxide (CO) under physiological pH conditions. Our laboratory has previously demonstrated micromolar CO’s cardio-protective effects in vitro and in vivo/ex-vivo experiments. This present study builds on our wider research investigating, for the first time, the effect of oCOm21 on CaMKIIδ function, with the aim of evaluating the prophylactic potential of CO in IRI. This study conducted CaMKIIδ sensitivity assays within a FRET biosensor transfected HEK 293 cell lysate (Camui lysate). High concentrations of oCOm21 (30µM) significantly attenuated CaMKIIδ activity within Camui lysate experiments when compared to control (P < 0.05). No significant (P > 0.05) differences were observed between therapeutically relevant oCOm21 concentrations (3 & 10µM) when compared to the relevant controls. Gaseous CO (10µM) also failed to produce any significant (P > 0.05) attenuation of Camui opening. The oCOm21 byproduct (BP-21) at 3, 10 & 30µM produced similar effects to same concentrations of oCOm21. These experiments suggest that the attenuation of Camui activity was associated with a direct effect of BP-21 on the Camui sensor, and not to CO release. A protocol for CaMKIIδ sensitivity assays within whole cell lines was investigated within HEK293 and AC16 cell lines to ascertain potential oCOm21 induced upstream influences over CaMKIIδ activity. These preliminary findings act as a strong incentive for further studies to be conducted on CaMKIIδ with oCOm21.