Robert E. Stroud

Specific Temporal Profile of Matrix Metalloproteinase Release Occurs in Patients After Myocardial Infarction Relation to Left Ventricular Remodeling

Background— Changes in matrix metalloproteinase (MMP) and tissue inhibitors of MMPs (TIMPs) contribute to left ventricular (LV) remodeling after myocardial infarction (MI). We tested the hypothesis that a specific plasma MMP/TIMP profile would emerge after MI and be associated with the degree of LV dilation. Methods and Results— LV end-diastolic volume and MMP/TIMP plasma profiles were determined in 53 age-matched control subjects and 32 post-MI patients from day 1 through 180 after MI. LV end-diastolic volume increased by >38% at day 90 after MI (P<0.05). MMP-9 increased by >150% from control at day 1 after MI (P<0.05) and remained elevated. MMP-8 rose to >120% at day 3 after MI (P<0.05) and fell to control values by day 5. TIMP-1 increased by >60% from control at day 1 after MI (P<0.05), whereas TIMP-2 increased only at later time points. Cardiac-specific TIMP-4 fell by 40% at day 5 after MI and remained reduced. A persistent or elevated MMP-9 at day 5 was accompanied by a 3-fold end-diastolic volume increase at day 28 (P<0.05). Conclusions— A specific temporal pattern of MMP/TIMPs occurred in post-MI patients that included an early and robust rise in MMP-9 and MMP-8 and a uniform fall in TIMP-4. These findings suggest that a specific MMP/TIMP plasma profile occurs after MI and holds both prognostic and diagnostic significance.

Myocardial Stiffness in Patients With Heart Failure and a Preserved Ejection Fraction Contributions of Collagen and Titin

Background— The purpose of this study was to determine whether patients with heart failure and a preserved ejection fraction (HFpEF) have an increase in passive myocardial stiffness and the extent to which discovered changes depend on changes in extracellular matrix fibrillar collagen and cardiomyocyte titin. Methods and Results— Seventy patients undergoing coronary artery bypass grafting underwent an echocardiogram, plasma biomarker determination, and intraoperative left ventricular epicardial anterior wall biopsy. Patients were divided into 3 groups: referent control (n=17, no hypertension or diabetes mellitus), hypertension (HTN) without (–) HFpEF (n=31), and HTN with (+) HFpEF (n=22). One or more of the following studies were performed on the biopsies: passive stiffness measurements to determine total, collagen-dependent and titin-dependent stiffness (differential extraction assay), collagen assays (biochemistry or histology), or titin isoform and phosphorylation assays. In comparison with controls, patients with HTN(–)HFpEF had no change in left ventricular end-diastolic pressure, myocardial passive stiffness, collagen, or titin phosphorylation but had an increase in biomarkers of inflammation (C-reactive protein, soluble ST2, tissue inhibitor of metalloproteinase 1). In comparison with both control and HTN(–)HFpEF, patients with HTN(+)HFpEF had increased left ventricular end-diastolic pressure, left atrial volume, N-terminal propeptide of brain natriuretic peptide, total, collagen-dependent, and titin-dependent stiffness, insoluble collagen, increased titin phosphorylation on PEVK S11878(S26), reduced phosphorylation on N2B S4185(S469), and increased biomarkers of inflammation. Conclusions— Hypertension in the absence of HFpEF did not alter passive myocardial stiffness. Patients with HTN(+)HFpEF had a significant increase in passive myocardial stiffness; collagen-dependent and titin-dependent stiffness were increased. These data suggest that the development of HFpEF depends on changes in both collagen and titin homeostasis.

Plasma Biomarkers That Reflect Determinants of Matrix Composition Identify the Presence of Left Ventricular Hypertrophy and Diastolic Heart Failure

Background— Chronic pressure overload (such as arterial hypertension) may cause left ventricular (LV) remodeling, alterations in cardiac function, and the development of diastolic heart failure. Changes in the composition of the myocardial extracellular matrix may contribute to the development of pressure-overload–induced LV remodeling. We hypothesized that a specific pattern of plasma biomarker expression that reflected changes in these pathophysiological mechanisms would have diagnostic application to identify (1) patients who have development of LV hypertrophy (LVH) and (2) patients with LVH who have development of diastolic heart failure. Methods and Results— Plasma concentration of 17 biomarkers (matrix metalloproteinase [MMP]-1, -2, -3, -7, -8, and -9; tissue inhibitors -1, -2, -3, and -4; N-terminal propeptide of brain natriuretic peptide (NT-proBNP); cardiotrophin; osteopontin; soluble receptor for advanced glycation end products; collagen I teleopeptide; collagen I NT-proBNP; and collagen III N-terminal propetide [PIIINP]), an echocardiogram, and 6-minute hall walk were performed on 241 referent control subjects, 144 patients with LVH but no evidence of heart failure, and 61 patients with LVH and diastolic heart failure (DHF). A plasma multibiomarker panel consisting of increased MMP-7, MMP-9, TIMP-1, PIIINP, and NT-proBNP predicted the presence of LVH with an area under the curve of 0.80. A plasma multibiomarker panel consisting of increased MMP-2, TIMP-4, PIIINP, and decreased MMP-8 predicted the presence of DHF with an area under the curve of 0.79. These multibiomarker panels performed better than any single biomarker including NT-proBNP and better than using clinical covariates alone (area under the curve, 0.73 for LVH and 0.68 for DHF). Conclusions— Plasma biomarkers reflecting changes in extracellular matrix fibrillar collagen homeostasis, combined into a multibiomarker panel, have discriminative value in identifying the presence of structural remodeling (LVH) and clinical disease (DHF).

Expression of Matrix Metalloproteinases and Endogenous Inhibitors Within Ascending Aortic Aneurysms of Patients With Marfan Syndrome

Background— Marfan syndrome (MFS) is known to cause ascending thoracic aortic aneurysms (ATAAs). Transforming growth factor beta (TGF-&bgr;) has recently been implicated in this process. Imbalances between the matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) have also been shown to contribute to aneurysm formation. Whether and to what degree MMP, TIMP, and TGF-&bgr; signaling profiles are altered in ATAAs in MFS compared with non-MFS patients remains unknown. Methods and Results— ATAA samples taken during aortic replacement from age-matched MFS (n=9) and non-MFS (n=18) patients were assessed for representative subtypes of all MMP classes, all 4 known TIMPs, and type 2 TGF-&bgr; receptors (TGFBR2). Results were expressed as a percentage (mean±SEM) of reference control samples (100%; n=18) obtained from patients without ATAA. In MFS, decreased MMP-2 (76±7; P<0.05 versus control), increased MMP-12 (161±27% versus control; P<0.05), and increased MT1-MMP (248±64% versus 91±21 non-MFS and control; P<0.05) were observed. TIMP-3 (74±23%) was reduced compared with control values (P<0.05) and TIMP-2 was elevated (128±31%) compared with non-MFS (73±19%; P<0.05). In non-MFS samples, MMP-1 (70±16%), MMP-3 (77±18%), MMP-8 (75±11%), MMP-9 (69±14%), and MMP-12 (85±15%) were decreased compared with control (P<0.05). TIMPs 1 to 3 were reduced in non-MFS compared with control values (P<0.05). TGFBR2 were increased in MFS (193±32%) compared with non-MFS (95±16%) and controls (P<0.05). Conclusions— A unique MMP and TIMP portfolio was observed in ATAAs from MFS compared with non-MFS patients. In addition, MFS samples showed evidence of increased TGF-&bgr; signaling. These differences suggest disparate mechanisms of extracellular matrix remodeling between these 2 groups of patients.

Cardiac Support Device Modifies Left Ventricular Geometry and Myocardial Structure After Myocardial Infarction

Background—Whether mechanical restraint of the left ventricle (LV) can influence remodeling after myocardial infarction (MI) remains poorly understood. This study surgically placed a cardiac support device (CSD) over the entire LV and examined LV and myocyte geometry and function after MI. Methods and Results—Post-MI sheep (35 to 45 kg; MI size, 23±2%) were randomized to placement of the CorCap CSD (Acorn Cardiovascular, Inc) (MI+CSD; n=6) or remained untreated (MI only; n=5). Uninstrumented sheep (n=10) served as controls. At 3 months after MI, LV end-diastolic volume (by MRI) was increased in the MI only group compared with controls (98±8 versus 43±4 mL; P<0.05). In the MI+CSD group, LV end-diastolic volume was lower than MI only values (56±7 mL; P<0.05) but remained higher than controls (P<0.05). Isolated LV myocyte shortening velocity was reduced by 35% from control values (P<0.05) in both MI groups. LV myocyte &bgr;-adrenergic response was reduced with MI but normalized in the MI+CSD group. LV myocyte length increased in the MI group and was reduced in the MI+CSD group. Relative collagen content was increased and matrix metalloproteinase-9 was decreased within the MI border region of the CSD group. Conclusions—A CSD beneficially modified LV and myocyte remodeling after MI through both cellular and extracellular mechanisms. These findings provide evidence that nonpharmacological strategies can interrupt adverse LV remodeling after MI.

Mesenchymal Cell Transplantation and Myocardial Remodeling After Myocardial Infarction

Background— Targeted delivery of mesenchymal precursor cells (MPCs) can modify left ventricular (LV) cellular and extracellular remodeling after myocardial infarction (MI). However, whether and to what degree LV remodeling may be affected by MPC injection post-MI, and whether these effects are concentration-dependent, remain unknown. Methods and Results— Allogeneic MPCs were expanded from sheep bone marrow, and direct intramyocardial injection was performed within the borderzone region 1 hour after MI induction (coronary ligation) in sheep at the following concentrations: 25×106 (25 M, n=7), 75×106 (75 M, n=7), 225×106 (225 M, n=10), 450×106 (450 M, n=8), and MPC free media only (MI Only, n=14). LV end diastolic volume increased in all groups but was attenuated in the 25 and 75 M groups. Collagen content within the borderzone region was increased in the MI Only, 225, and 450 M groups, whereas plasma ICTP, an index of collagen degradation, was highest in the 25 M group. Within the borderzone region matrix metalloproteinases (MMPs) and MMP tissue inhibitors (TIMPs) also changed in a MPC concentration–dependent manner. For example, borderzone levels of MMP-9 were highest in the 25 M group when compared to the MI Only and other MPC treatment group values. Conclusions— MPC injection altered collagen dynamics, MMP, and TIMP levels in a concentration-dependent manner, and thereby influenced indices of post-MI LV remodeling. However, the greatest effects with respect to post-MI remodeling were identified at lower MPC concentrations, thus suggesting a therapeutic threshold exists for this particular cell therapy.

Targeted Myocardial Microinjections of a Biocomposite Material Reduces Infarct Expansion in Pigs

BACKGROUND Left ventricular (LV) remodeling after myocardial infarction (MI) commonly causes infarct expansion (IE). This study sought to interrupt IE through microinjections of a biocompatible composite material into the post-MI myocardium. METHODS MI was created in 21 pigs (coronary ligation). Radiopaque markers (2-mm diameter) were placed for IE (fluoroscopy). Pigs were randomized for microinjections (25 injections; 2- x 2-cm array; 200 microL/injection) at 7 days post-MI of a fibrin-alginate composite (Fib-Alg; fibrinogen, fibronectin, factor XIII, gelatin-grafted alginate, thrombin; n = 11) or saline (n = 10). RESULTS At 7 days after injection (14 days post-MI), LV posterior wall thickness was higher in the Fib-Alg group than in the saline group (1.07 +/- 0.11 vs 0.69 +/- 0.07 cm, respectively, p = 0.002). At 28 days post-MI, the area within the markers (IE) increased from baseline (1 cm2) in the saline (1.71 +/- 0.13 cm2, p = 0.010) and Fib-Alg groups (1.44 +/- 0.23 cm2, p < 0.001). However, the change in IE at 21 and 28 days post-MI was reduced in the Fib-Alg group (p=0.043 and p=0.019). Total collagen content within the MI region was similar in the saline and Fib-Alg groups (12.8 +/- 1.7 and 11.6 +/- 1.5 microg/mg, respectively, p = NS). However, extractable collagen, indicative of solubility, was lower in the Fib-Alg group than the saline group (59.1 +/- 3.5 vs 71.0 +/- 6.1 microg/mL, p = 0.020). CONCLUSIONS Targeted myocardial microinjection of the biocomposite attenuated the post-MI decrease in LV wall thickness and infarct expansion. Thus, intraoperative microinjections of biocompatible material may provide a novel approach for interrupting post-MI LV remodeling.

Relationship between the temporal profile of plasma microRNA and left ventricular remodeling in patients after myocardial infarction.

Background— microRNAs (miRs) are small noncoding RNAs that recognize and bind to mRNAs and inhibit protein translation or degrade mRNA. Studies in animal models have suggested that miRs play a translational or posttranslational regulatory role in myocardial growth, fibrosis, viability, and remodeling. However, whether specific temporal changes in miRs occur in patients during the left ventricular (LV) remodeling process that follows a myocardial infarction (post-MI) remains unknown. The current pilot study tested the hypotheses that plasma miRs could be reliably measured in post-MI patients and that there is a relationship between temporal changes in specific miRs and post-MI LV structural remodeling. Methods and Results— LV end-diastolic volume (echocardiography) and plasma miR were measured in age-matched referent controls (CTLs, n=12) and post-MI patients (n=12) from day 2 through day 90 post-MI. Selected miRs (miR-1, miR-21, miR-29a, miR-133a, and miR-208) were measured using quantitative reverse transcription–polymerase chain reaction and normalized for endogenous small nuclear RNA U6. After MI, LV end-diastolic volume increased progressively compared with CTL; this was accompanied by time-dependent changes in specific miRs. For example, miR-21 initially decreased 2 days post-MI (0.3±0.1-fold versus CTL; P<0.05), increased 5 days post-MI (2±1-fold versus CTL; P<0.05), and returned to CTL values at later post-MI time points. In contrast, miR-29a increased 5 days post-MI (4±1-fold versus CTL; P<0.05) and then decreased to CTL at later time points. miR-208 increased 5 days post-MI (3±1-fold versus CTL; P<0.05) and remained elevated up to 90 days post-MI. Conclusions— A time-dependent change in miRs occurred in post-MI patients, including an early and robust increase in miRs that has affected myocardial growth, fibrosis, and viability. Thus, serially profiling miRs in the plasma of post-MI patients may hold both mechanistic and prognostic significance.

Mitral Regurgitation Augments Post-Myocardial Infarction Remodeling: Failure of Hypertrophic Compensation

OBJECTIVES We examined whether mitral regurgitation (MR) augments post-myocardial infarction (MI) remodeling. BACKGROUND MR doubles mortality after MI, but its additive contribution to left ventricular (LV) remodeling is debated and has not been addressed in a controlled fashion. METHODS Apical MIs were created in 12 sheep, and 6 had an LV-to-left atrial shunt implanted, consistently producing regurgitant fractions of approximately 30%. The groups were compared at baseline, 1, and 3 months. RESULTS Left ventricular end-systolic volume progressively increased by 190% with MR versus 90% without MR (p < 0.02). Pre-load-recruitable stroke work declined by 82 +/- 13% versus 25 +/- 16% (p < 0.01) with MR, with decreased remote-zone sarcoplasmic reticulum Ca(2+)-ATPase levels (0.56 +/- 0.03 vs. 0.76 +/- 0.02, p < 0.001), and decreased isolated myocyte contractility. In remote zones, pro-hypertrophic Akt and gp130 were upregulated in both groups at 1 month, but significantly lower and below baseline in the MR group at 3 months. Pro-apoptotic caspase 3 remained high in both groups. Matrix metalloproteinase (MMP)-13 and membrane-type MMP-1 were increased in remote zones of MR versus infarct-only animals at 1 month, then fell below baseline. The MMP tissue inhibitors rose from baseline to 3 months in all animals, rising higher in the MI + MR-group border zone. CONCLUSIONS In this controlled model, moderate MR worsens post-MI remodeling, with reduced contractility. Pro-hypertrophic pathways are initially upregulated but subsequently fall below infarct-only levels and baseline; with sustained caspase 3 elevation, transformation to a failure phenotype occurs. Extracellular matrix turnover increases in MR animals. Therefore, MR can precipitate an earlier onset of dilated heart failure.

Selective MicroRNA Suppression in Human Thoracic Aneurysms Relationship of miR-29a to Aortic Size and Proteolytic Induction

Background— Increasing evidence points to a direct role for altered microRNA (miRNA or miR) expression levels in cardiovascular remodeling and disease progression. Although alterations in miR expression levels have been directly linked to cardiac hypertrophy, fibrosis, and remodeling, their role in regulating gene expression during thoracic aortic aneurysm (TAA) development has yet to be explored. Methods and Results— The present study examined miR expression levels in aortic tissue specimens collected from patients with ascending TAAs by quantitative real-time PCR, and observed decreased miR expression (miRs -1, -21, -29a, -133a, and -486) as compared with normal aortic specimens. A significant relationship between miR expression levels (miRs -1, -21, -29a, and -133a) and aortic diameter was identified; as aortic diameter increased, miR expression decreased. Through the use of a bioinformatics approach, members of the matrix metalloproteinase (MMP) family, proteins involved in TAA development, were examined for putative miR binding sites. MMP-2 and MMP-9 were identified as potential targets for miR-29a and miR-133a, respectively, and MMP-2 was subsequently verified as a miR-29a target in vitro. A significant inverse relationship between miR-29a and total MMP-2 was then identified in the clinical TAA specimens. Conclusions— These findings demonstrate altered miR expression patterns in clinical TAA specimens, suggesting that the loss of specific miR expression may allow for the elaboration of specific MMPs capable of driving aortic remodeling during TAA development. Importantly, these data suggest that these miRs have biological and clinical relevance to the behavior of TAAs and may provide significant targets for therapeutic and diagnostic applications.Background— Increasing evidence points to a direct role for altered microRNA (miRNA or miR) expression levels in cardiovascular remodeling and disease progression. Although alterations in miR expression levels have been directly linked to cardiac hypertrophy, fibrosis, and remodeling, their role in regulating gene expression during thoracic aortic aneurysm (TAA) development has yet to be explored. Methods and Results— The present study examined miR expression levels in aortic tissue specimens collected from patients with ascending TAAs by quantitative real-time PCR, and observed decreased miR expression (miRs -1, -21, -29a, -133a, and -486) as compared with normal aortic specimens. A significant relationship between miR expression levels (miRs -1, -21, -29a, and -133a) and aortic diameter was identified; as aortic diameter increased, miR expression decreased. Through the use of a bioinformatics approach, members of the matrix metalloproteinase (MMP) family, proteins involved in TAA development, were examined for putative miR binding sites. MMP-2 and MMP-9 were identified as potential targets for miR-29a and miR-133a, respectively, and MMP-2 was subsequently verified as a miR-29a target in vitro. A significant inverse relationship between miR-29a and total MMP-2 was then identified in the clinical TAA specimens. Conclusions— These findings demonstrate altered miR expression patterns in clinical TAA specimens, suggesting that the loss of specific miR expression may allow for the elaboration of specific MMPs capable of driving aortic remodeling during TAA development. Importantly, these data suggest that these miRs have biological and clinical relevance to the behavior of TAAs and may provide significant targets for therapeutic and diagnostic applications.