Cardiac interventions were performed 1) under X-ray for coronary artery occlusion/reperfusion and 2) MR-guidance for transendocardial delivery of gene. Animals (n=16) showed myocardial infarct 5 weeks prior to delivery of gene and (n=12) 5 weeks after delivery of gene on DE-MR imaging. The pCK-HGF and pCK-LacZ animals showed no significant difference in body weight, heart rate, and mean arterial blood pressure at any time point (). The advancement of endovascular active catheter from the aorta into the LV chamber then to the target, is shown in . The brightness of endovascular catheter in the dark background LV chamber facilitates the navigation of the catheter to the target. The injection was performed after turning off the active coils to eliminate the heating produced by coil activation.
Body weight, heart rate and mean blood pressure measurements during the 10 weeks observation period.
LV mass and volumes
There was no significant difference in LV mass or wall structure (diastolic and systolic LV wall thickness) of remote myocardial segments at any time point between the groups. The segments with infarct scar were significantly thinner compared to remote myocardial segments in both groups at 5 weeks after infarction (). Five weeks after delivery of pCK-HGF gene, there was a significant increase in wall thickness of segments with infarct scar at both diastole (from 5.7±0.5mm at 5 weeks after infarction to 6.6±0.3mm at 5 weeks after therapy, P<0.05) and systole (5.7±0.4mm to 8.1±0.5, P<0.05) compared with pCK-LacZ (from 5.7±0.4mm to 5.4±0.6mm at diastole and from 5.5±0.5mm to 5.4±0.5mm at systole).
Figure 2 Long and short axis views of DE-MR images illustrate the hyperenhanced infarct at 5 weeks after infarction of two representative animals (top block). The bottom block shows the effects of pCK-LacZ (top row) and pCK-HGF genes (bottom row) on infarct transmurality (more ...)
Furthermore, end-diastolic, end-systolic, stroke volumes, cardiac output and ejection fraction were also not significantly different between the groups 5 weeks after infarction (). Animals treated with pCK-HGF gene also showed significant decrease in end systolic volume (from 1.27±0.05 at 5 weeks after infarction to 1.10±0.05ml/kg body weight at 5 weeks after delivery of gene, P<0.05) and an average increase of 5%, in ejection fraction (from 38±1 at 5 weeks after infarction to 43±1% at 5 weeks after delivery of gene, P<0.05). PCK-LacZ animals showed no significant change in end systolic volume (1.31±0.06 at 5 weeks after infarction to 1.28±0.05 ml/kg body weight at 5 weeks after delivery of gene, P=ns) or ejection fraction (38±2 at 5 weeks after infarction to 37±1% at 5 weeks after delivery of gene, P=ns), respectively. Furthermore, both end systolic volume and ejection fraction in pCK-HGF treated animals were significantly different from control pCK-LacZ at 5 weeks after gene delivery ().
Serial MR measurements of global LV function and mass before and after delivery of the genes. Note that the genes were derived 5 weeks after infarction.
At 5 weeks after infarction, quantitative analysis of cine MR images revealed no significant difference between the groups in regional % systolic wall thickening (radial strain) (). The segments with infarct scar were dyskinetic in pCK-LacZ control and akinetic in pCK-HGF animals at 5 weeks after delivery of the gene. However, there was significant improvement in systolic wall thickening of remote myocardial segments in pCK-HGF treated animals. In contrast, control animals showed a trend of further dysfunction in radial strain of remote myocardium at 5 weeks after pCK-LacZ delivery.
Figure 3 Percent systolic wall thickening (radial strain) from 8 circumferential segments (average of 3 slices) of LV from pCK-LacZ control (black bars) and pCK-HGF treated (white bars) animals. The data were obtained at 5 weeks after infarction (top) and 5 weeks (more ...)
Data obtained from tagged and phase-contrast velocity-encoded MR images showed severe dysfunction in segments with infarct scar at 5 weeks after infarction in both groups (). PCK-HGF, but not pCK-LacZ, treated animals showed significant improvement in both circumferential and longitudinal strain suggesting that these imaging sequences were sensitive enough to demonstrate the improvements at 5 weeks after delivery of gene. shows a pixel-by-pixel illustration of the difference in LV longitudinal strain between treated and control animals.
Figure 4 Percent peak circumferential (top) and longitudinal (bottom) strain measurements obtained from remote myocardium and chronic infarct scar containing the infarct shown on DE-MR images for guidance. The circumferential and longitudinal strain were markedly (more ...)
Figure 5 Longitudinal strain in pixel-by-pixel obtained from phase contrast velocity encoded MR images in animals treated with pCK-LacZ and pCK-HGF gene. Longitudinal strain at diastole is shown in green (left images) and at systole in blue. Top block demonstrates (more ...)
At 5 weeks after infarction, first pass perfusion MR imaging showed the infarcted segments as hypoenhanced, due to the delayed arrival of the contrast medium, compared to remote myocardium. The perfusion parameters measured in the infarcted and remote segments were not significantly different between the groups (). At 5 weeks after delivery of HGF gene, however, maximum upslope, P<0.05, peak signal intensity, P<0.05 and time to peak, P<0.05 were improved compared to 5 weeks after infarction in the same animals. Furthermore, treated animals at 5 weeks after delivery of pCK-HGF gene showed better perfusion compared with pCK-LacZ control animals ().
Regional perfusion parameters obtained from first pass MR imaging.
Infarct size and infarct transmurality
DE-MR imaging highlighted infarct scar (the target) in short and long axis views allowing precise delivery of the genes (). At 5 weeks after infarction the infarct size was not significantly different between the groups (). PCK-HGF treated animals showed significantly smaller infarct size (6.7±1.3%) compared to pCK-LacZ control (11.3±2.0%) at 5 weeks after delivery (P<0.01). These changes are associated with a significant decrease in infarct transmurality in 4 of 5 segments with infarct scar (P<0.05) on DE-MR imaging (). TTC infarct size at autopsy was not significantly different from MR infarct size (). MR images and TTC histochemical staining shows the non-transmural infarct at 5 weeks in pCK-HGF, but not in pCK-LacZ, group (). There was no significant change in infarct transmurality in all segments with infarct scar in pCK-LacZ control animals.
Figure 6 Percent infarct transmurality before (upper block) and after (lower block) delivery of pCK-LacZ (black bars) and pCK-HGF (white bars). Prior to therapy, there was no significant difference in infarct transmurality between the groups. pCK-HGF significantly (more ...)
Masson trichrome stain showed homogeneous chronic infarct scar at 5 weeks after pCK-LacZ delivery. However, pCK-HGF treated animals showed different infarct architecture compared to controls. For example, Masson trichrome stain showed few viable islands/peninsulas at the border of segments with infarct scar, while isolectin B5 showed homogeneous increase in vascularity of infarct scar in pCK-HGF treated animals but disarray of vessels in control animals (). The number of vessels in border and core of chronic infarct was greater in pCK-HGF treated (206±10 capillaries/mm2) compared with pCK-LacZ control animals (78±25 capillaries/mm2, P<0.05). A significant difference was also observed in the number of arterioles within the infarct scar in pCK-HGF (11.7±1.9 arterioles/mm2) and pCK-LacZ control (3.4±0.5 arterioles/mm2, P<0.05) animals. On the other hand, there was no significant difference in capillary and arterial densities in remote myocardium between treated (269±30 capillaries/mm2, 10.9±2.1 arterioles/mm2, respectively) and control animals (267±54 capillaries/mm2 and 10.7±1.1 arterioles/mm2, respectively). Myocyte diameters of remote myocardium in pCK-HGF (16.4±0.8μm) and pCK-LacZ control animals (18.3±0.2μm) were not significantly different, respectively. There was no evidence of inflammation in the segments with infarct scar in pCK-LacZ control and pCK-HGF treated animals.
Figure 7 Histopathological heart sections stained with isolectin B5 obtained from two animals 5 weeks after delivery of pCK-LacZ (left) and pCK-HGF (right). Remote myocardium obtained from both animals (top row) showed no difference in vascular density or myocyte (more ...)