Similar Inhibition of the Mitochondrial Permeability Transition Pore Opening by Intralipid and Cyclosporine-A after Ischemia Reperfusion

Abstract

It has been shown that intralipid(ILP) protects the heart against ischemia reperfusion (I/R) injury. However the mechanism of its action is not clear. Here we investigated whether ILP-induced cardioprotection is mediated by inhibition of the mitochondria permeability transition pore (mPTP) opening. We compared the effect of ILP with cyclosporine A (CsA), a well known inhibitor of mPTP. Isolated mouse hearts were subjected to 20 minutes of global ischemia followed by 10 minutes reperfusion with i)Krebs Henseleit buffer (CTRL),ii) additional 1% ILP or iii)1.5µM CsA.The hearts which were not subjected to ischemia/reperfusion served as sham. The calcium retention capacity (CRC) was measured in isolated cardiac mitochondria in the absence or after addition of 2 µM of CsA in the cuvette. DHE staining of the heart tissue sections was used to measure the production of reactive oxygen species (ROS). The CRC was significantly lower in CTRL compared to sham (1.5±0.2 vs 3.7±0.2µM/mg protein, p<0.05). However, the treatment with ILP or CsA significantly improved the CRC compared to CTRL(2.8±0.1 in ILP, 2.6±0.3 µM/mg protein in CsA). Addition of CsA directly in the cuvette, resulted in a similar significant increase in CRC between ILP and CsA groups (4.5±0.3 vs. 4.2±0.5µM/mg protein, p>0.05). However, the increase in CRC in CTRL and sham groups after addition of CsA in vitro were much higher (2.2 and 1.6 fold increase separately). ROS production was significantly lower in ILP and CsA group compared to CTRL (normalized to CTRL, 1.00±0.03 in CTRL vs. 0.57±0.04 in ILP, p<0.05). In conclusion, intralipid inhibits the opening of the mPTP in a similar fashion as CsA via a CypD-dependent mechanisms. This inhibition resulted in decreased sensitivity of mPTP to calcium overload and reduction of ROS production.