The IMproving Preclinical Assessment of Cardioprotective Therapies (IMPACT): multicenter pig study on the effect of ischemic preconditioning

Abstract

Numerous cardioprotective interventions have been reported to reduce myocardial infarct size (IS) in pre-clinical studies. However, their translation for the benefit of patients with acute myocardial infarction (AMI) has been largely disappointing. One reason for the lack of translation is the lack of rigor and reproducibility in pre-clinical studies. To address this, we have established the European IMproving Preclinical Assessment of Cardioprotective Therapies (IMPACT) pig AMI network with centralized randomization and blinded core laboratory IS analysis and validated the network with ischemic preconditioning (IPC) as a positive control. Ten sites in the COST Innovators Grant (IG16225) network participated in the IMPACT network. Three sites were excluded from the final analysis through quality control of infarct images and use of pre-defined exclusion criteria. Using a centrally generated randomization list, pigs were allocated to myocardial ischemia/reperfusion (I/R, N = 5/site) or IPC + I/R (N = 5/site). The primary endpoint was IS [% area-at-risk (AAR)], as quantified by triphenyl-tetrazolium-chloride (TTC) staining in a centralized, blinded core laboratory (5 sites), or IS [% left-ventricular mass (LV)], as quantified by a centralized, blinded cardiac magnetic resonance (CMR) core laboratory (2 sites). In pooled analyses, IPC significantly reduced IS when compared to I/R (57 ± 14 versus 32 ± 19 [%AAR] N = 25 pigs/group; p < 0.001; 25 ± 13 versus 14 ± 8 [%LV]; N = 10 pigs/group; p = 0.021). In site-specific analyses, in 4 of the 5 sites, IS was significantly reduced by IPC when compared to I/R when quantified by TTC and in 1 of 2 sites when quantified by CMR. A pig AMI multicenter European network with centralized randomization and core blinded IS analysis was established and validated with the aim to improve the reproducibility of cardioprotective interventions in pre-clinical studies and the translation of cardioprotection for patient benefit.

Keywords: Acute myocardial infarction; Ischemia/reperfusion injury; Ischemic preconditioning; Multicenter network; Pig; Randomized-controlled trial.

Fig. 1

Study flow of IMPACT pig acute myocardial infarction multicenter network. Ten sites agreed to participate in the IMproving Preclinical Assessment of Cardioprotective Therapies (IMPACT) pig acute myocardial infarction network. One site was excluded as it did not pass the QC. Central randomization lists were provided to 9 sites to undertake I/R and IPC + I/R studies. Six sites used TTC images for central blinded core laboratory quantification of IS [%AAR] and 3 sites used CMR for central blinded core laboratory quantification of infarct size [% LV]. One TTC site was excluded as the AAR was smaller than the pre-defined criterion and one CMR site was excluded as it did not follow the central randomization list. Five TTC sites and 2 CMR sites underwent final analysis of infarct size data. AAR area-at-risk, CMR cardiac magnetic resonance imaging, IPC ischemic preconditioning, I/R ischemia/reperfusion, IS infarct size, LV left ventricle, QC quality control

Fig. 2

Pooled and site-specific analysis of area-at-risk from the 5 sites providing triphenyl-tetrazolium-chloride images. A Pooled analysis of AAR quantified by histochemistry (fluorescein or blue dye) was comparable between I/R (open squares) and IPC + I/R (filled squares). B Site-specific analysis of AAR was comparable between the 5 sites. Data are presented as minimum and maximum (whiskers), interquartile range from 25 to 75% (box), mean (square), median (line), and outlier (x) in a box plot and as intra-individual single data points. AAR area-at-risk, ANOVA analysis of variance, IPC ischemic preconditioning, I/R ischemia/reperfusion, LV left ventricle

Fig. 3

Pooled and site-specific analysis of infarct size from the 5 sites providing triphenyl-tetrazolium-chloride images. A Pooled analysis of IS quantified by TTC revealed a significant reduction in IS with IPC (closed squares) when compared to I/R (open squares). B Site-specific analysis of IS revealed significant reduction in IS with IPC in 4 sites and no reduction in IS with IPC at one site. Data are presented as minimum and maximum (whiskers), interquartile range from 25 to 75% (box), mean (square), median (line), and outlier (x) in a box plot and as intra-individual single data points. AAR area-at-risk, ANOVA analysis of variance, IPC ischemic preconditioning, I/R ischemia/reperfusion, IS infarct size, TTC triphenyl-tetrazolium-chloride

Fig. 4

Pooled and site-specific analysis of infarct size from the 2 sites providing cardiac magnetic resonance images. A Pooled analysis of IS quantified by CMR revealed a significant reduction in IS with IPC (closed squares) when compared to I/R (open squares). B Site-specific analysis of IS quantified by CMR revealed significant reduction in IS with IPC at one site but no reduction in IS at the other site. Data are presented as minimum and maximum (whiskers), interquartile range from 25 to 75% (box), mean (square), median (line), and outlier (x) in a box plot and as intra-individual single data points. ANOVA analysis of variance, CMR cardiac magnetic resonance imaging, IPC ischemic preconditioning, I/R ischemia/reperfusion, IS infarct size, LV left ventricle

Fig. 5

Total number of animals required for testing a novel cardioprotective strategy when using our IMPACT network with infarct size quantified with TTC (A) or CMR (B). The Y-axis depicts the total number of animals and the X-axis depicts the relative reduction in infarct size. Calculations are based on the observed IMPACT network IS data with I/R and IPC + I/R and α ≤ 0.05 with statistical power 1-β of ≥ 0.9. AAR area-at-risk, CMR cardiac magnetic resonance imaging, ICC intraclass correlation coefficient, IPC ischemic preconditioning, I/R ischemia/reperfusion, IS infarct size, LV left ventricle, TTC triphenyl-tetrazolium-chloride

Fig. 6

The multicenter network evaluated the cardioprotective effect of IPC as the decrease in infarct size measured ex vivo by TTC and in vivo by CMR. CMR cardiac magnetic resonance imaging, IPC ischemic preconditioning, I/R ischemia/reperfusion, LAD left anterior descending coronary artery, TTC triphenyl-tetrazolium-chloride. Created with BioRender.com