Shinichiro Fujimoto

Molecular imaging of matrix metalloproteinase in atherosclerotic lesions : resolution with dietary modification and statin therapy

OBJECTIVES This study sought to evaluate the feasibility of noninvasive detection of matrix metalloproteinase (MMP) activity in experimental atherosclerosis using technetium-99m-labeled broad matrix metalloproteinase inhibitor (MPI) and to determine the effect of dietary modification and statin treatment on MMP activity. BACKGROUND The MMP activity in atherosclerotic lesions contributes to the vulnerability of atherosclerotic plaques to rupture. METHODS Atherosclerosis was produced in 34 New Zealand White rabbits by balloon de-endotheliazation of the abdominal aorta and a high-cholesterol diet. In addition, 12 unmanipulated rabbits were used as controls and 3 for blood clearance characteristics. In vivo micro-single-photon emission computed tomography (SPECT) imaging was performed after radiolabeled MPI administration. Subsequently, aortas were explanted to quantitatively measure percent injected dose per gram (%ID/g) MPI uptake. Histological and immunohistochemical characterization was performed and the extent of MMP activity was determined by gel zymography or enzyme-linked immunosorbent assays. RESULTS The MPI uptake in atherosclerotic lesions (n = 18) was clearly visualized by micro-SPECT imaging; MPI uptake was markedly reduced by administration of unlabeled MPI before the radiotracer (n = 4). The MPI uptake was also significantly reduced after diet withdrawal (n = 6) and fluvastatin treatment (n = 6); no uptake was observed in normal control rabbits (n = 12). The %ID/g MPI uptake (0.10 +/- 0.03%) in the atherosclerotic lesions was significantly higher than the uptake in control aorta (0.016 +/- 0.004%, p < 0.0001). Uptake in fluvastatin (0.056 +/- 0.011%, p < 0.0005) and diet withdrawal groups (0.043 +/- 0.011%, p < 0.0001) was lower than the untreated group. The MPI uptake correlated with immunohistochemically verified macrophage infiltration (r = 0.643, p < 0.0001), and MMP-2 (r = 0.542, p < 0.0001) or MMP-9 (r = 0.578, p < 0.0001) expression in plaques. CONCLUSIONS The present data show the feasibility of noninvasive detection of MMP activity in atherosclerotic plaques, and confirm that dietary modification and statin therapy reduce MMP activity.

Molecular Imaging of Matrix Metalloproteinase Expression in Atherosclerotic Plaques of Mice Deficient in Apolipoprotein E or Low-Density-Lipoprotein Receptor

Matrix metalloproteinases (MMPs) are expressed in atherosclerotic plaques and play an important role in plaque instability. Methods: Using 99mTc-labeled broad-spectrum MMP inhibitor (MPI), we performed noninvasive imaging of MMP expression with micro-SPECT/micro-CT in mice deficient in apolipoprotein E (ApoE−/−, n = 14), mice deficient in low-density-lipoprotein receptor (LDLR−/−, n = 14), and C57/BL6 mice as controls (n = 7). Seven ApoE−/− and 7 LDLR−/− received a high-cholesterol diet. After in vivo imaging, aortas were explanted, ex vivo images acquired, and the percent injected dose of MPI per gram (%ID/g) determined, followed by histologic characterization of atherosclerotic lesions. Results: MPI uptake was noninvasively visualized in atherosclerotic lesions by micro-SPECT, with confirmation by micro-CT of anatomic location and aortic calcification. %ID/g in each part of the aorta was highest in ApoE−/− that were fed a high-cholesterol diet, followed by LDLR−/− that were fed a high-cholesterol diet, ApoE−/− that were fed normal chow, and LDLR−/− that were fed normal chow. The control mice had minimal MPI uptake. A significant correlation was noted between %ID/g and % area positive for macrophages (r = 0.81, P = 0.009), MMP-2 (r = 0.65, P = 0.013), and MMP-9 (r = 0.62, P = 0.008). Conclusion: This study demonstrates the usefulness of molecular imaging for noninvasive assessment of the extent of MMP expression in various transgenic mouse models of atherosclerosis receiving a normal or hyperlipidemic diet. It is conceivable that such a strategy may be translationally developed for identification of unstable atherosclerotic plaques.

Molecular Imaging for Efficacy of Pharmacologic Intervention in Myocardial Remodeling

OBJECTIVES Using molecular imaging techniques, we examined interstitial alterations during postmyocardial infarction (MI) remodeling and assessed the efficacy of antiangiotensin and antimineralocorticoid intervention, alone and in combination. BACKGROUND The antagonists of the renin-angiotensin-aldosterone axis restrict myocardial fibrosis and cardiac remodeling after MI and contribute to improved survival. Radionuclide imaging with technetium-99m-labeled Cy5.5 RGD imaging peptide (CRIP) targets myofibroblasts and indirectly allows monitoring of the extent of collagen deposition post-MI. METHODS CRIP was intravenously administered for gamma imaging after 4 weeks of MI in 63 Swiss-Webster mice and in 6 unmanipulated mice. Of 63 animals, 50 were treated with captopril (C), losartan (L), spironolactone (S) alone, or in combination (CL, SC, SL, and SCL), 8 mice received no treatment. Echocardiography was performed for assessment of cardiac remodeling. Hearts were characterized histopathologically for the presence of myofibroblasts and thick and thin collagen fiber deposition. RESULTS Acute MI size was similar in all groups. The quantitative CRIP percent injected dose per gram uptake was greatest in the infarct area of untreated control mice (2.30 +/- 0.14%) and decreased significantly in animals treated with 1 agent (C, L, or S; 1.71 +/- 0.35%; p = 0.0002). The addition of 2 (CL, SC, or SL 1.31 +/- 0.40%; p < 0.0001) or 3 agents (SCL; 1.16 +/- 0.26%; p < 0.0001) demonstrated further reduction in tracer uptake. The decrease in echocardiographic left ventricular function, strain and rotation parameters, as well as histologically verified deposition of thin collagen fibers, was significantly reduced in treatment groups and correlated with CRIP uptake. CONCLUSIONS Radiolabeled CRIP allows for the evaluation of the efficacy of neurohumoral antagonists after MI and reconfirms superiority of combination therapy. If proven clinically, molecular imaging of the myocardial healing process may help plan an optimal treatment for patients susceptible to heart failure.

Radiolabeled Monocyte Chemotactic Protein 1 for the Detection of Inflammation in Experimental Atherosclerosis

Chemotactic peptides, such as Monocyte Chemotactic Protein 1 (MCP-1), play a key role in transendothelial migration of mononuclear cells during the development and progression of atherosclerotic disease. Because atherosclerotic plaques that are precursors of acute coronary events harbor abundant macrophage infiltration, we hypothesized that the detection of a high concentration of MCP-1 receptors on inflammatory cells should noninvasively identify vulnerable plaques. Methods: Atherosclerotic lesions were induced by balloon deendothelialization of the abdominal aorta, which was followed by a 0.5% cholesterol diet for 16 wk in 7 New Zealand White rabbits; 5 unmanipulated rabbits, fed normal chow for 16 wk, were used as controls. Radionuclide imaging was performed immediately after intravenous 99mTc-labeled MCP-1 administration and 3 h later. At the end of imaging session, aortas were explanted and submitted for estimation of quantitative MCP-1 uptake (in percentage injected dose per gram, %ID/g) and pathologic characterization. Results: Atherosclerotic lesions were clearly visible in all hyperlipidemic animal γ-imaging. No tracer uptake was seen in the control rabbits. The mean quantitative MCP-1 uptake in atherosclerotic lesions was 4-fold higher than that of the aortic specimens from the control rabbits (0.065 ± 0.005 vs. 0.016 ± 0.006; P < 0.0001). Histology confirmed a strong correlation between MCP-1 uptake and the number of macrophages in American Heart Association type II−IV lesions (r = 0.87, P < 0.0001). Conclusion: Noninvasive radionuclide imaging of inflammation is feasible by MCP-1 in experimentally induced atherosclerosis. It is proposed that detection of the extent of inflammation in advanced atherosclerotic plaques may allow identification of unstable plaques.

Computed Tomographic Angiography–Verified Plaque Characteristics and Slow-Flow Phenomenon During Percutaneous Coronary Intervention

OBJECTIVES This study sought to identify whether computed tomographic angiographic (CTA) plaque characteristics are associated with slow-flow phenomenon (SF) during percutaneous coronary intervention (PCI). BACKGROUND SF during PCI is associated with myocardial damage and prolonged hospitalization. Intracoronary ultrasound-verified large echolucent lesions have been reported to predict SF. METHODS The authors evaluated pre-PCI CTA plaque characteristics in 40 consecutive patients (male/female, 31/9; age, 69 ± 10 years) with stable angina pectoris who developed SF during PCI; patients with ≥ 600 Agatston coronary artery calcium score were not included. They were compared with 40 age-, sex-, and culprit coronary artery-matched patients (male/female, 31/9; age, 69 ± 9 years) who underwent PCI during the same period and did not develop SF. Plaque characteristics, including vascular remodeling, plaque consistency, including low-attenuation plaques representing lipid-rich lesions and high-attenuation plaque patterns of calcium deposition, were analyzed. RESULTS Calcium deposition in the perimeter of a plaque, or circumferential plaque calcification (CPC), was significantly more frequent in the SF group (25 of 40, 63%) than the no-SF group (2 of 40, 5.0%) (p < 0.001). Presence of CPC on CTA was confirmed at the same location in the nonenhanced CT during Agatston coronary artery calcium score calculation. The positive remodeling index was significantly higher (1.5 [1.3 to 1.8] vs. 1.2 [1.0 to 1.5]; p < 0.001) and plaque density significantly lower (23.5 [9.5 to 40] HU vs. 45 [29 to 86] HU; p = 0.001) in the SF group. The conditional logistic regression analysis revealed that CPC, plaque density, and dyslipidemia were the predictors of SF, with CPC being the strongest (odds ratio: 79; 95% confidence interval: 8 to 783, p < 0.0001). CONCLUSIONS CTA-verified CPC with low-attenuation plaque and positive remodeling were determinants of SF during PCI. If CTA findings are available in patients undergoing PCI, the interventionists should be aware of the likelihood of SF.

Prevalence of computed tomographic angiography-verified high-risk plaques and significant luminal stenosis in patients with zero coronary calcium score.

BACKGROUND Some patients were detected with coronary artery disease even if the coronary artery calcium score was (CACS)=0. We evaluated the prevalence and predictor of significant stenosis and computed tomography (CT) based vulnerable plaque (CTVP) for patients with CACS=0. METHODS Subjects were 2160 patients (M/F=1110/1050, 64.7 ± 11.6 years) who underwent measurement of calcium score and CT coronary angiography. As for CACS=0 group, age, gender, coronary risk factor (family history (FH), hypertension (HT), hyperlipidemia (HL), diabetes (DM), and smoking), body mass index, history of cerebral infarction, the presence of chest symptom, and abnormal rest ECG findings were investigated as predictors for significant stenosis and CTVP by multivariate analysis using logistic regression analysis. RESULTS Out of 2160 patients, 1141 (52.8%, M/F=655/486, 68.4 ± 9.8 years) were of CACS>0 and 1019 (47.2%, M/F=455/564, 60.5 ± 12.0 years) were of CACS=0. In the CACS=0 group, 24 patients (2.4%) were found with significant stenosis and 47 (4.6%) with 2FPP. In 104 patients with spotty calcification (10.2%), 10 (9.6%) out of these 104 had significant stenosis and also had CTVP. Multivariate analysis using logistic regression analysis revealed significant predictor for significant stenosis to be only male (Odds ratio (OR): 3.075, 95%CI 1.166-8.109, p=0.0232) and significant predictor for CTVP to be age (OR: 1.032, 95%CI 1.001-1.063, p=0.0437) and male (OR: 2.386, 95%CI 1.193-4.775, p=0.0140). CONCLUSIONS The present study suggests that the presence of CTVP must be noted, when patients are male and elderly even if CACS=0 and the presence of spotty calcification increases the prevalence of significant stenosis and CTVP in patients with CACS=0.

Effect of an antimicrobial agent on atherosclerotic plaques: assessment of metalloproteinase activity by molecular imaging.

OBJECTIVES Technetium-99m-labeled matrix metalloproteinase inhibitor (MPI) was used for the noninvasive assessment of matrix metalloproteinase (MMP) activity in atherosclerotic plaques after minocycline (MC) intervention. BACKGROUND MMP activity in atherosclerosis contributes to plaque instability. Some antimicrobial agents may attenuate MMP activity. METHODS Atherosclerotic lesions were produced in 38 rabbits with a high cholesterol diet for 4 months; 5 groups of rabbits, in the fourth month, received fluvastatin (FS) (n = 6), low-dose MC (n = 7), high-dose MC (n = 7), a combination of low-dose MC and FS (n = 6), or no intervention (n = 12); 8 unmanipulated rabbits were used as disease controls. Micro-single-photon emission computed tomography imaging was performed in all animals after intravenous MPI administration, followed by pathologic characterization of the aorta. A cell culture study evaluated the effect of MC on MMP production by activated human monocytes. RESULTS MPI uptake was visualized best in untreated atherosclerotic animals (percent injected dose per gram MPI uptake, 0.11 +/- 0.04%). MPI uptake was reduced in the FS (0.06 +/- 0.01%; p < 0.0001), high-dose MC (0.05 +/- 0.01%; p < 0.0001), and MC-FS (0.05 +/- 0.005%; p < 0.0001) groups. Low-dose MC did not resolve MPI uptake significantly (0.08 +/- 0.02; p = 0.167). There was no incremental benefit of the combination of MC and FS. MPI uptake showed a significant correlation with plaque MMP-2, and MMP-9 activity. MMP-9 release from tumor necrosis factor-alpha-activated macrophages was abrogated by incubation with MC. CONCLUSIONS Molecular imaging of MMP activity in atherosclerotic plaque allows for the study of the efficacy of therapeutic interventions. MC administration resulted in substantial reduction in plaque MMP activity and histologically verified plaque stabilization. MC was found to be equally effective as FS.

Targeting of matrix metalloproteinase activation for noninvasive detection of vulnerable atherosclerotic lesions

IntroductionInflammation plays an important role in vulnerability of atherosclerotic plaques to rupture and hence acute coronary events. The monocyte–macrophage infiltration in plaques leads to upregulation of cytokines and metalloproteinase enzymes.Matrix metalloproteinases result in matrix dissolution and consequently expansive remodeling of the vessel. They also contribute to attenuation of fibrous cap and hence susceptibility to rupture. Assessment of metalloproteinase expression and activity should provide information about plaque instability.

Image Quality and Radiation Dose Stratified by Patient Heart Rate for Coronary 64- and 320-MDCT Angiography

OBJECTIVE. The purpose of this study is to retrospectively measure and compare estimated radiation doses between consecutive patient cohorts who underwent coronary imaging CT with 64- and 320-MDCT scanners. MATERIALS AND METHODS. Subjects without arrhythmia (n = 4475) underwent imaging with 64-MDCT (n = 770) and 320-MDCT (n = 3705) scanners and were classified into one of five subgroups according to the patient heart rate and the image acquisition strategy. For all patients, image quality was subjectively evaluated using a 3-point scale. Estimated radiation dose and image quality were compared between subjects stratified by CT scanner and by subgroups imaged with each technology. RESULTS. For patients with a heart rate of 60 beats/min or less, the estimated radiation dose was halved (3.8 ± 2.0 vs 7.6 ± 2.6 mSv) when the 320-MDCT scanner (n = 2787) replaced the 64-MDCT scanner (n = 511). For the entire cohort, image quality score was significantly better (2.9 ± 0.4 vs 2.8 ± 0.5; p < 0.0001) and the effective dose was significantly lower (4.9 ± 3.3 vs 9.9 ± 5.4 mSv; p < 0.0001) for 320-MDCT scanners, compared with 64-MDCT scanners. CONCLUSION. Wide area-detector coronary CT angiography protocols have reduced radiation dose, with image quality maintained at the same level, compared with 64-MDCT technologies.

Prospective ECG-Gated Coronary 320-MDCT Angiography With Absolute Acquisition Delay Strategy for Patients With Persistent Atrial Fibrillation

OBJECTIVE The purpose of this study was to evaluate image quality and radiation dose when patients with atrial fibrillation undergo coronary CT angiography (CTA) using prospectively ECG-gated 320-MDCT technology with an absolute-delay strategy. MATERIALS AND METHODS A cohort of 75 consecutive patients (60 men and 15 women; age (± SD), 71 ± 10 years) who underwent prospectively ECG-gated coronary CTA using a 320-MDCT scanner during atrial fibrillation was matched with 75 control patients imaged in sinus rhythm. All coronary CTA for the atrial fibrillation cohort used absolute-delay strategy. Subjective image quality score and the dose-length product (DLP) were compared between the two cohorts and, for the atrial fibrillation cohort, among those patients imaged over a different number of heartbeats. The accuracy of stenosis detection was evaluated in 17 studies of the atrial fibrillation cohort using catheter angiography as a reference standard. RESULTS For those patients imaged in atrial fibrillation, one- and two-beat acquisitions were performed in 26.7% (n = 20) and 40% (n = 30) of patients, respectively. There was no significant difference in image quality between the atrial fibrillation (2.9 ± 0.4) and sinus rhythm (2.9 ± 0.3) cohorts, nor was there a difference in image quality with respect to the number of heartbeats used in the acquisition. The atrial fibrillation cohort had an 80% higher DLP (680 ± 470 vs 372 ± 236 mGy × cm, p < 0.0001). The patient-based sensitivity and negative predictive value for stenosis detection were both 100%. CONCLUSION Using an absolute-delay strategy, two thirds of patients who underwent prospectively ECG-gated coronary CTA using a 320-MDCT scanner were imaged within two heartbeats or fewer. Compared with patients imaged in sinus rhythm, the image quality was comparative and the radiation dose was 1.8-fold higher.