Haakon K. Grøgaard

ET-receptor antagonism, myocardial gene expression, and ventricular remodeling during CHF in rats

Both myocardial and plasma endothelin-1 (ET-1) are elevated in congestive heart failure (CHF). However, the role played by endogenous ET-1 in the progression of CHF remains unknown. The aim of the present study was to investigate and correlate myocardial gene expression programs and left ventricular (LV) remodeling during chronic ET-receptor antagonism in CHF rats. After ligation of the left coronary artery, rats were randomized to oral treatment with a nonselective ET-receptor antagonist (bosentan, 100 mg ⋅ kg-1 ⋅ day-1, n = 11) or vehicle (saline, n = 13) for 15 days, starting 24 h after induction of myocardial infarction. Bosentan substantially attenuated LV dilatation during postinfarction failure as evaluated by echocardiography. Furthermore, bosentan decreased LV systolic and end-diastolic pressures and increased fractional shortening. Myocardial expression of preproET-1 mRNA and a fetal gene program characteristic of myocardial hypertrophy were increased in the CHF rats and were not affected by bosentan. Consistently, right ventricular-to-body weight ratios, diameters of cardiomyocytes, and echocardiographic analysis demonstrated a sustained hypertrophic response and a normalized relative wall thickness after intervention with bosentan. Thus the modest reduction of preload and afterload provided by bosentan substantially attenuates LV dilatation, causing improved pressure-volume relationships. However, the compensatory hypertrophic response was not altered by ET-receptor antagonism. Therefore, ET-1 does not appear to play a crucial role in the mechanisms of myocardial hypertrophy during the early phase of postinfarction failure.Both myocardial and plasma endothelin-1 (ET-1) are elevated in congestive heart failure (CHF). However, the role played by endogenous ET-1 in the progression of CHF remains unknown. The aim of the present study was to investigate and correlate myocardial gene expression programs and left ventricular (LV) remodeling during chronic ET-receptor antagonism in CHF rats. After ligation of the left coronary artery, rats were randomized to oral treatment with a nonselective ET-receptor antagonist (bosentan, 100 mg . kg-1 . day-1, n = 11) or vehicle (saline, n = 13) for 15 days, starting 24 h after induction of myocardial infarction. Bosentan substantially attenuated LV dilatation during postinfarction failure as evaluated by echocardiography. Furthermore, bosentan decreased LV systolic and end-diastolic pressures and increased fractional shortening. Myocardial expression of preproET-1 mRNA and a fetal gene program characteristic of myocardial hypertrophy were increased in the CHF rats and were not affected by bosentan. Consistently, right ventricular-to-body weight ratios, diameters of cardiomyocytes, and echocardiographic analysis demonstrated a sustained hypertrophic response and a normalized relative wall thickness after intervention with bosentan. Thus the modest reduction of preload and afterload provided by bosentan substantially attenuates LV dilatation, causing improved pressure-volume relationships. However, the compensatory hypertrophic response was not altered by ET-receptor antagonism. Therefore, ET-1 does not appear to play a crucial role in the mechanisms of myocardial hypertrophy during the early phase of postinfarction failure.

Transient, isopeptide-specific induction of myocardial endothelin-1 mRNA in congestive heart failure in rats

Increased myocardial expression of preproendothelin-1 (ppET-1) mRNA has been associated with congestive heart failure (CHF) in rats. However, the time course and isoform pattern of ppET mRNA induction and the cellular localization of ET in failing hearts are unknown. Thus our aim was to investigate myocardial ppET mRNA expression in CHF rats during the first 6 wk after induction of myocardial infarction. Furthermore, performing immunohistochemical analysis, we also investigated the origin and localization of immunoreactive endothelin (ET) in different regions of the failing heart. Ribonuclease protection assays revealed a marked increase in ppET-1 mRNA levels in rat myocardial tissues during CHF. The induction of ppET-1 mRNA was isopeptide specific and transient. The most substantial upregulation was observed in the infarcted area, where maximal expression of ppET-1 mRNA was observed after 7 days (25-fold increase, P < 0.05). However, a marked and statistically significant induction of ppET-1 mRNA was also observed in the nonischemic myocardium. Immunohistochemical analysis revealed ET-1-like immunoreactivity in cardiomyocytes, vascular endothelial cells, macrophages, and proliferating fibroblasts. Thus immunohistochemistry revealed the structural basis for the dramatic upregulation of the myocardial ET system in the infarcted region, suggesting a role for ET in the healing process after myocardial infarction. However, the global upregulation of ppET-1 mRNA in the heart also suggests an autocrine/paracrine regulatory mechanism in the nonischemic myocardium during CHF.Increased myocardial expression of preproendothelin-1 (ppET-1) mRNA has been associated with congestive heart failure (CHF) in rats. However, the time course and isoform pattern of ppET mRNA induction and the cellular localization of ET in failing hearts are unknown. Thus our aim was to investigate myocardial ppET mRNA expression in CHF rats during the first 6 wk after induction of myocardial infarction. Furthermore, performing immunohistochemical analysis, we also investigated the origin and localization of immunoreactive endothelin (ET) in different regions of the failing heart. Ribonuclease protection assays revealed a marked increase in ppET-1 mRNA levels in rat myocardial tissues during CHF. The induction of ppET-1 mRNA was isopeptide specific and transient. The most substantial upregulation was observed in the infarcted area, where maximal expression of ppET-1 mRNA was observed after 7 days (25-fold increase, P < 0.05). However, a marked and statistically significant induction of ppET-1 mRNA was also observed in the nonischemic myocardium. Immunohistochemical analysis revealed ET-1-like immunoreactivity in cardiomyocytes, vascular endothelial cells, macrophages, and proliferating fibroblasts. Thus immunohistochemistry revealed the structural basis for the dramatic upregulation of the myocardial ET system in the infarcted region, suggesting a role for ET in the healing process after myocardial infarction. However, the global upregulation of ppET-1 mRNA in the heart also suggests an autocrine/paracrine regulatory mechanism in the nonischemic myocardium during CHF.

Age and stress related phenotypical changes in bone marrow CD34+ cells

Objective. Phenotypical changes in the human bone marrow (BM) due to age and stress have not so far been properly addressed in the literature. In the present study, we compared CD34+ BM cells between older and young volunteers. The influence of stress on CD34+ cell phenotype in older patients was investigated in an age‐matched group with acute myocardial infarction (AMI). Cytokines thought to influence BM CD34+ cell homeostasis were also analysed. Material and methods. BM mononuclear cells of 10 older volunteers and of 7 young volunteers (18–25 years), as well as 22 AMI patients, were analysed by flow cytometry for the following markers: CD34, CD38, CD117 (c‐kit) and CD133. Blood samples were analysed for CRP, IL‐6, MCP‐1, IL‐8, MMP‐9, TIMP‐1 and TNFα by ELISA methods. Results. Significantly higher numbers of CD34+ CD38− cells (both absolute and relative) were observed in older volunteers than in young volunteers and AMI patients. Higher numbers of immature progenitors, namely CD34+CD38− cells and CD34+CD38−CD117+CD133+ cells, were observed among older volunteers compared to the other groups. However, the relative number of CD34+ cells lacking CD38 expression or expressing CD133 was higher in the old volunteers and AMI patients. None of the circulating factors investigated correlated with any of the cell population yields. Conclusion. In this study, we found that the absolute and relative numbers of BM CD34+CD38− progenitor cells increase with age. The increment is attenuated in patients with AMI.

Inflammatory responses after percutaneous coronary intervention in patients with acute myocardial infarction or stable angina pectoris

Objective. To investigate the profile of circulating inflammatory markers after percutaneous coronary intervention (PCI) in patients with AMI or stable angina pectoris (AP). Material and methods. Twenty patients with AMI and 10 with stable AP were treated with PCI of a central coronary artery. Blood samples were drawn immediately before PCI, in the AP group and after 3 and 12 h, days 1, 3, 5, 7 and 14 in both groups. Results. Interleukin 6 increased in both groups to time‐point 12 h and day 1 (peak), being significantly higher in the AMI group compared to the AP group at 3 and 12 h, and also at days 1 and 3. A similar profile was demonstrated for CRP with significantly higher levels in the AMI group at days 1, 3 and 5 compared to the AP group. A slightly different pattern was shown for Interleukin 10 (IL‐10) with significantly higher levels in the AMI group at 3 and 12 h, days 1 and 14 compared to the AP group. Conclusion. AMI patients treated with PCI experienced a marked short‐term increase in pro‐inflammatory mediators as well as IL‐10 compared to patients with stable angina pectoris treated with PCI.

The profile of circulating metalloproteinases after PCI in patients with acute myocardial infarction or stable angina

UNLABELLED The objective of this study was to investigate the time profiles of plasma matrix metalloproteinase-9 (MMP-9), tissue inhibitor of metalloproteinases (TIMP-1 and -2), and pregnancy associated plasma protein A (PAPP-A) in patients with acute myocardial infarction (AMI), compared to patients with stable angina pectoris (AP), all treated with percutaneous coronary intervention (PCI) with stent. METHODS Twenty patients with ST-elevation AMI and 10 patients with AP were included. Serum levels of the selected markers were measured before (only in the AP group) PCI, and 3 and 12 hours, 1,3,5,7 and 14 days after PCI in all patients. RESULTS The levels of MMP-9 and MMP-9/TIMP-1 ratio, being higher in the AMI group compared to the AP group at 3 hours, were significantly reduced 1 day after PCI (p<0.01 for both), sustaining during the study period. A similar pattern was observed in PAPP-A levels with significant reduction after 12 hours (p<0.01). In the AP group only smaller changes were observed, except from an increase in PAPP-A levels from before PCI to 3 hours (p<0.001), followed by significant reduction. No significant correlations were found between any of the measured biomarkers and the infarct size, either evaluated in the acute phase or after 6 weeks. CONCLUSION AMI patients treated with PCI had an early significant reduction in MMP-9, MMP-9/TIMP-1 ratio and PAPP-A when compared to patients with stable angina pectoris treated with PCI. This indicates that the metalloproteinases may be involved in the early phase of the plaque rupture process, with limited influence of the PCI procedure.

Gene expression of natriuretic peptides and their receptors type-A and -C after myocardial infarction in rats.

The purpose of this study was to investigate myocardial mRNA expression of brain natriuretic peptide (BNP) and atrial natriuretic peptide (ANP) in different regions of the heart at three different time points after induction of myocardial infarction (MI) in rats. Furthermore, we examined putative changes in mRNA expression of natriuretic peptide receptors (NPRs), NPR-A and NPR-C, in myocardium and peripheral organs. Substantial increase in the mRNA levels of both BNP and ANP in the infarcted as well as non-infarcted regions were observed after induction of MI. These findings were paralleled by elevated circulating concentrations of ANP, BNP and N-terminal proANP (Nt-proANP). In addition, the mRNA levels of the clearance receptor, NPR-C, were augmented in the infarcted and non-infarcted regions of the left ventricular wall (LV), while it was decreased in the kidneys and lungs 28 days post-MI. Based on these data, we propose that, in addition to increased myocardial secretion of BNP and ANP, reduced peripheral clearance by NPR-C may contribute to the observed increase in circulating plasma concentrations of the natriuretic peptides after induction of MI in rats.

Endothelin Receptor Antagonism Attenuates Cardiomyocyte Apoptosis after Induction of Ischemia in Rats

Objective : Apoptosis has recently been implicated as a process that may contribute to loss of cardiomyocytes and to adverse myocardial remodeling in congestive heart failure (CHF). We investigated to what extent myocardial ischemia and CHF lead to induction of apoptotic gene programs and loss of cardiomyocytes through apoptosis, and subsequently to what extent the beneficial effects of endothelin (ET) receptor antagonism after myocardial infarction (MI) could be attributed to reduction of apoptotic cell loss in the myocardium. Design : Northern blot analysis, analysis of DNA fragmentation, and immunohistochemical analysis were performed after induction of MI in rats. Results : After induction of MI, the mRNA levels of the pro-apoptotic genes FAS, BAX, P53, and CASPASE-1 were significantly increased in the non-ischemic region of the left ventricle (LV) with highest levels of expression in the peri-infarct area. High levels of FAS, BAX, P53, and CASPASE-1 mRNA were also observed in the infarcted region. Concomitantly, numerous TUNEL-positive cells and internucleosomal degradation of DNA were found in tissue from the peri-infarct area and in the infarcted zone, indicating apoptotic cell death. Treatment with bosentan, a mixed ET A /ET B receptor antagonist, during the first 24 h after induction of MI significantly reduced the area of TUNEL-positive myocardium in the ischemic region (64 - 2% of LV circumference in the vehicle group vs 55 - 2% of LV circumference in the bosentan group, p < 0.05). Consistently, bosentan also caused a similar reduction of infarct size. Conclusion : Our data demonstrate activation of pro-apoptotic genes and provide evidence of cardiomyocyte apoptosis in the viable peri-infarct area and in the infarcted region after MI in rats. Intervention with bosentan may attenuate cardiomyocyte cell loss through apoptosis in the area at risk after induction of ischemia.

Circulating CD34+ progenitor cells and growth factors in patients treated with PCI for acute myocardial infarction or stable angina pectoris

Abstract Objective. To differentiate the effect of myocardial infarction from the effect of percutaneous coronary intervention (PCI) on the circulatory profiles of CD34+ cells and growth factors in patients with ST-elevation myocardial infarction (STEMI). Methods. Twenty patients with STEMI and 10 with angina pectoris (AP) were included. All were treated with PCI. Blood was drawn before PCI in the AP group, and after 3 and 12 hours, and 1, 3, 5, 7 and 14 days after PCI in both groups. In STEMI patients, correlation analyses between TIMI myocardial perfusion grade (TMP-grade) and circulating CD34+cells were also assessed. Results. Circulating CD34+ cells increased from day 1 to days 5 and 7 after PCI only in STEMI patients (p < 0.05). Between-group analyses revealed a borderline significant difference in change in SDF-1α concentrations from 3 h to 14 days after PCI (p = 0.05), and SDF-1α was significantly higher in STEMI patients 14 days after PCI (p < 0.05). In both groups, peak HGF concentrations were observed 3 h after PCI, whereas IGF-1 increased in AP patients only, 3 h after PCI (p < 0.005). TIMI perfusion grade was negatively correlated to the circulating number of CD34+ cells 5 days after PCI (r =−0.69, p < 0.005). Conclusion. After PCI, STEMI patients have significantly higher numbers of circulating CD34+ progenitor cells compared to patients with AP. STEMI results in a significant increase in SDF-1α after 14 days, and the increase at this time may indicate a favorable environment for progenitor cell therapy.

Cell treatment after acute myocardial infarction prevents early decline in circulating IGF-1

Abstract Objectives. To examine the influence of intracoronary autologous bone marrow cell transplantation after acute myocardial infarction on circulating growth factors and their relationship to left ventricular function. Methods. Circulating insulin-like growth factor-1 (IGF-1), hepatocyte growth factor (HGF), stromal derived factor-1-alpha (SDF-1α), and transforming growth factor beta (TGF-β) were measured in patients randomized to cell treatment or control, in the ASTAMI study. Autologous cells were injected intracoronary on day 6; blood was sampled on days 5, 9, and at three months. Left ventricular ejection fraction was recorded by electrocardiogram-gated single photon emission computed tomography at six months. Results. Only change in IGF-1 from baseline to three months differed between groups (p=0.024). A weak but significant correlation was found between left ventricular ejection fraction and the averaged IGF-1 concentrations of all patients (r=0.24, p=0.02). Patients with IGF-1 above or below median (102 ng/ml) had a left ventricular ejection fraction of 52.3% (±11.4) versus 46.4% (±12.2) respectively (p=0.017). Conclusions. Intracoronary bone marrow cell treatment after myocardial infarction attenuates a reduction in circulating IGF-1. IGF-1 levels over time were weakly, but significantly correlated to left ventricular ejection fraction.