Automated Quantification of Myocardial Salvage in a Rat Model of Ischemia–Reperfusion Injury Using 3D High‐Resolution Magnetic Resonance Imaging (MRI)

Abstract

BackgroundQuantification of myocardial “area at risk” (AAR) and myocardial infarction (MI) zone is critical for assessing novel therapies targeting myocardial ischemia–reperfusion (IR) injury. Current “gold‐standard” methods perfuse the heart with Evan's Blue and stain with triphenyl tetrazolium chloride (TTC), requiring manual slicing and analysis. We aimed to develop and validate a high‐resolution 3‐dimensional (3D) magnetic resonance imaging (MRI) method for quantifying MI and AAR.Methods and ResultsForty‐eight hours after IR was induced, rats were anesthetized and gadopentetate dimeglumine was administered intravenously. After 10 minutes, the coronary artery was re‐ligated and a solution containing iron oxide microparticles and Evan's Blue was infused (for comparison). Hearts were harvested and transversally sectioned for TTC staining. Ex vivo MR images of slices were acquired on a 9.4‐T magnet. T2* data allowed visualization of AAR, with microparticle‐associated signal loss in perfused regions. T1 data demonstrated gadolinium retention in infarcted zones. Close correlation (r=0.92 to 0.94; P<0.05) of MRI and Evan's Blue/TTC measures for both AAR and MI was observed when the combined techniques were applied to the same heart slice. However, 3D MRI acquisition and analysis of whole heart reduced intra‐observer variability compared to assessment of isolated slices, and allowed automated segmentation and analysis, thus reducing interobserver variation. Anatomical resolution of 81 μm3 was achieved (versus ≈2 mm with manual slicing).ConclusionsThis novel, yet simple, MRI technique allows precise assessment of infarct and AAR zones. It removes the need for tissue slicing and provides opportunity for 3D digital analysis at high anatomical resolution in a streamlined manner accessible for all laboratories already performing IR experiments.

Adenosine 2B Receptor Activation Reduces Myocardial Reperfusion Injury by Promoting Anti-Inflammatory Macrophages Differentiation via PI3K/Akt Pathway

“…In present study, we did not analyze the cardiac function after myocardial reperfusion injury. However, it is well known that histological determination of myocardial infarction has been widely used to estimate the cardiac injury in animal models [ 38 ] and utilized as the golden standard to validate novel imaging methods of infarction quantification, including cardiac MRI [ 39 ] and contrast echocardiography [ 40 ]. Moreover, using a cardiac MRI imaging technique, we have defined in our previous publication that left ventricular function is depressed proportionally to the size of infarction, which was determined by the same TTC staining technique in the present study [ 41 ].…”

Practical guidelines for rigor and reproducibility in preclinical and clinical studies on cardioprotection

“…TTC staining is the widely accepted method to measure infarct size [ 214 ]. Transthoracic echocardiography [ 71 , 436 ], CMR [ 124 , 166 , 288 ] and positron emission tomography [ 175 , 177 ] have also been used in studies of myocardial infarction in rats.…”

Imaging the Area at Risk in Myocardial Infarction With Cardiovascular Magnetic Resonance

“…In this issue of JAHA: Journal of the American Heart Association , Grieve et al publish a novel and elegant “whole heart” 3‐dimensional method to measure MI size and AAR ex vivo in rodents. 9 Analogous to microspheres, they use iron oxide microparticles to mark perfused myocardium that excludes the AAR. The signal loss from T2* decay introduced by the iron oxide microparticles causes black dots in perfused myocardium evident on CMR gradient echo images.…”