Sevoflurane postconditioning is not mediated by ferritin accumulation and cannot be rescued by simvastatin in isolated streptozotocin-induced diabetic rat hearts

Abstract

Sevoflurane postconditioning (sevo postC) is an attractive and amenable approach that can protect the myocardium against ischemia/reperfusion (I/R)-injury. Unlike ischemic preconditioning (IPC), sevo postC does not require additional induced ischemic periods to a heart that is already at risk. IPC was previously shown to generate myocardial protection against I/R-injury through regulation of iron homeostasis and de novo ferritin synthesis, a process found to be impaired in the diabetic state. The current study investigated whether alterations in iron homeostasis and ferritin mRNA and protein accumulation are also involved in the cardioprotective effects generated by sevo postC. It was also investigated whether the protective effects of sevo postC in the diabetic state can be salvaged by simvastatin, through inducing nitric oxide (NO) bioavailability/activity, in isolated streptozotocin (STZ)-induced diabetic hearts (DH). Isolated rat hearts from healthy Controls and diabetic animals were retrogradely perfused using the Langendorff configuration and subjected to prolonged ischemia and reperfusion, with and without (2.4 and 3.6%) sevo postC and/or pre-treatment with simvastatin (0.5 mg/kg). Sevo postC significantly reduced infarct size and improved myocardial function in healthy Controls but not in isolated DH. The sevo postC mediated myocardial protection against I/R-injury was not associated with de novo ferrtin synthesis. Furthermore, simvastatin aggravated myocardial injury after sevo postC in STZ-induced DHs, likely due to increasing NO levels. Despite the known mechanistic overlaps between PC and postC stimuli, distinct differences underlie the cardioprotective interventions against myocardial I/R-injury and are impaired in the DH. Sevo postC mediated cardioprotection, unlike IPC, does not involve de novo ferritin accumulation and cannot be rescued by simvastatin in STZ-induced DHs.

Fig 1. Experimental protocols.

(A) Hearts in groups 1–9 were harvested after 120 min (60 min of reperfusion) and stained to allow infarct size determination. Simvastatin (simv) was injected 48h (day-2) and 24h (day-1) before the start of the experiments. (B) Hearts in groups 10–16 were harvested at different time points along the perfusion protocol (*) to allow molecular biological analyses. Ctr = hearts from normoglycemic animals, Diab = hearts from STZ-induced diabetic animals, SHAM = continues perfused normoxia treated hearts, I/R = ischemia/reperfusion, sevo = sefoflurane.

Fig 2. Myocardial infarct size analyses after the different perfusion protocols.

Myocardial infarct sizes of the normoglycemic Control hearts are depicted by the dark grey bars. Myocardial infarct sizes of the DHs are depicted by the light grey bars. The DHs with simvastatin and I/R+3.6% sevo postC are depicted by the striated light grey bar. Data are presented as Mean ± SEM, * denotes p<0.01 vs. I/R in the respective group (Control heart or Diabetic heart). # denotes p<0.05 vs. I/R.

Fig 3. Myocardial ferritin protein levels in normoglycemic control hearts and in STZ-induced diabetic hearts.

(A) Ferritin protein levels in Control hearts and STZ-induced DHs (B) along different time points of the perfusion protocols. Data are presented as Mean ± SEM and indicate the heart ferritin. * denotes p<0.01 vs. I/R. # denotes p<0.05 vs. I/R. ^ denotes p<0.05 vs. baseline.