Interstitial Collagen Content Research Articles

Thalidomide is not able to inhibit radiation-induced heart disease

Purpose: Radiotherapy to the thorax increases the risk of radiation-induced heart disease. We and others have shown that local irradiation to the murine heart results in inflammatory and fibrotic responses and decreased microvascular density. In the present study we tested whether thalidomide is able to inhibit radiation-induced heart disease.Methods: Single doses of 16 Gy or 0 Gy (sham treatment) were delivered to the hearts of mice. At 16 weeks after irradiation the mice were allocated to receive a thalidomide-containing chow (100 mg/kg body weight/day) or control chow until the end of the experiment. At 40 weeks after irradiation, functional imaging was performed and the hearts were examined for histological damage.Results: Irradiation led to an increase in epicardial thickness and infiltrating inflammatory cells in the epicardium as well as an increase in interstitial collagen content. The microvasculature had a decreased alkaline phosphatase activity and reduced pericyte coverage. Thalidomide had no protective role in any of these processes. There were no differences in heart function measured between the treatment groups.Conclusions: Although others have shown protective effects of thalidomide in disease models involving inflammation, fibrosis and blood vessel maturation, thalidomide was not able to reduce radiation-induced heart damage.

Abstract 19325: Genetic Ablation of the P2y2 Nucleotide Receptor Reduces Atherosclerosis in Mice

Objective: We have shown that nucleotide binding to P2Y2 receptor induces endothelial activation via enhanced VCAM-1 expression leading to intimal recruitment of monocyte/macrophage. Here, we sought to determine whether genetic ablation of P2Y2 receptor affects the development of atherosclerosis. Methods and Results: Western blot and immumofluorescence analysis revealed that P2Y2 receptor deficiency in ApoE mice is accompanied by a sharp reduction in basal VCAM-1 expression levels. To analyze the consequence of P2Y2 receptor deficiency in endothelial function, we examined transmigration of monocytes isolated from the peripheral blood of wt or P2Y2R −/− mice across wt or P2Y2R −/− endothelial monolayers in response to monocyte chemoattractant protein-1. Absence of P2Y 2 R in endothelial cells significantly impaired monocyte transmigration. Remarkably, loss of P2Y2 receptor in monocytes did not significantly impair transmigration in the presence (transendothelial) or absence (chemotaxis) of P2Y2R +/+ endothelial monolayers, suggesting that endothelial P2Y2R is relatively more important than monocytic P2Y 2 R in regulating monocyte motility. The absence of P2Y 2 R did not significantly change the plasma cholesterol or triglyceride levels; however, lipid deposition in the lesion, as demonstrated by Oil Red O–positive region, was decreased by 47% in the aortic preparation from ApoE−/−/P2Y2R+/+ mice. Notably, P2Y2R deficiency in ApoE−/− resulted in a 52% reduction in the area of aortic sinus lesion as compared to ApoE −/− mice. Consistent with reduced lesion size, decreased macrophage content was observed in the sinus aortic lesions of ApoE−/−/P2Y 2 R −/− mice. Staining for SMC showed that most of the SMCs were in the intimal area close to the lumen. The staining showed a 36% decrease in ApoE−/−/P2Y 2 R −/− when compared with ApoE −/− /P2Y2R +/+ mice. Staining with Sirius red showed that the interstitial collagen content of the lesions of ApoE −/− /P2Y2R +/+ mice was 2-fold higher than in ApoE−/−/P2Y2R −/−mice. Conclusion: These observations suggest that genetic ablation of P2Y 2 R significantly decreases both macrophage accumulation and interstitial fibrosis but also reduces smooth muscle cell content of sinus aortic lesions.

Abstract 240: Interruption of Ivabradine Treatment in Middle-Aged Post-MI Rats May Lead to Heart Failure

Background: We have previously demonstrated that heart rate reduction (HRR) with the selective cardiac pacemaker I f current inhibitor ivabradine (IVA) improved LV function and coronary perfusion reserve in post-MI middle-aged rats. The beneficial effects of IVA treatment were associated with attenuated collagen accumulation in the surviving LV myocardium and inhibition of the circulatory renin-angiotensin system. However, sustainability of these positive changes after a period of IVA treatment interruption remains to be determined. Methods: MI was induced in 12-month-old male Sprague-Dawley rats by left coronary artery ligation. Twenty four hours later, the rats with a confirmed large transmural MI were randomly assigned in two experimental groups. In a first group, rats were treated with IVA i.p. via osmotic pumps in a dose of 10.5 mg/kg/d for 4 weeks followed by a 4-week period without treatment (MI+IVA). In a second group, rats received placebo for 4 weeks (MI). Sham-operated rats served as a control. Eight weeks after MI, the various LV parameters were examined. Results: During 4 weeks of treatment, heart rate in MI+IVA rats was consistently reduced compared to untreated MI rats (mean of HRR was 27%). After IVA withdrawal, heart rate increased to the levels of the placebo group. Eight weeks after MI, the IVA treated group demonstrated smaller LV free wall infarct size (57.5 ± 2.7% vs. 72.5 ± 2.5%, p<0.01) and greater LV weight/body weight ratio (2.54 ± 0.06 vs. 2.26 ± 0.11 mg/g, p<0.05), indicating greater myocardial compensatory hypertrophy. Furthermore, the interstitial collagen content was significantly lower in the remote LV myocardium of MI+IVA rats compared to MI rats (7.5 ± 0.7% vs. 13.3 ± 1.0% in free wall and 7.3 ± 0.8% vs. 11.2 ± 0.9% in septum, p < 0.01). However, compared to non-infarcted controls, rats of both MI groups had markedly lower LV ejection fraction and coronary perfusion reserve. Conclusions: Our data demonstrate that although beneficial structural changes induced by IVA in post-MI myocardium can be sustainable for a long period of time they cannot prevent deterioration of LV function after withdrawal of treatment. Therefore, we believe that continuous treatment with IVA may be essential for attenuation of heart failure after a large MI.

Beta1-adrenergic receptors promote focal adhesion signaling downregulation and myocyte apoptosis in acute volume overload

Numerous studies demonstrated increased expression of extracellular matrix (ECM) proteins and activation of focal adhesion (FA) signaling pathways in models of pressure overload-induced cardiac hypertrophy. However, little is known about FA signaling in response to volume overload where cardiac hypertrophy is associated with ECM loss. This study examines the role of beta1-adrenergic receptors (β1-ARs) in FA signaling changes and myocyte apoptosis induced during acute hemodynamic stress of volume overload. Rats with eccentric cardiac hypertrophy induced after aorto-caval fistula (ACF) develop reduced interstitial collagen content and decreased tyrosine phosphorylation of key FA signaling molecules FAK, Pyk2 and paxillin along with an increase in cardiac myocyte apoptosis. ACF also increased activation of PTEN, a dual lipid and protein phosphatase, and its interaction with FA proteins. β1-AR blockade (extended-release of metoprolol succinate, 100mg QD) markedly attenuated PTEN activation, restored FA signaling and reduced myocyte apoptosis induced by ACF at 2days, but failed to reduce interstitial collagen loss and left ventricular dilatation. Treating cultured myocytes with β1-AR agonists or adenoviral expression of β1-ARs caused PTEN activation and interaction with FA proteins, thus leading to FA signaling downregulation and myocyte apoptosis. Adenoviral-mediated expression of a catalytically inactive PTEN mutant or wild-type FAK restored FA signaling downregulation and attenuated myocyte apoptosis induced by β1-ARs. Collectively, these data show that β1-AR stimulation in response to ACF induces FA signaling downregulation through an ECM-independent mechanism. This effect involves PTEN activation and may contribute to adverse cardiac remodeling and function in the course of volume overload.

Procollagen propeptides: serum markers for atrial fibrosis?

Atrial fibrosis and its spatial heterogeneity are regarded as a substrate for the perpetuation of atrial arrhythmias. During collagen synthesis and degradation, collagen propeptides and telopeptides are released into the blood. This study tested the hypothesis that serum markers of collagen turnover correlate with atrial fibrosis. We prospectively included 28 patients in sinus rhythm undergoing cardiac surgery. Plasma concentrations of the carboxy- and amino-terminal propeptide of procollagen type-I (PICP and PINP) and type-III (PIIINP), and the C-terminal telopeptide of type-I collagen (ICTP) were determined. Interstitial fibrosis of left (n = 10) and right atrial appendages (n = 28) was analyzed histologically. We found a correlation between left and right atrial fibrosis (r (s) = 0.79, p < 0.01). Interestingly, the higher the interstitial collagen content, the higher was the spatial heterogeneity of fibrosis (r (s) = 0.90, p < 0.001). However, PICP, PIIINP, and ICTP were not correlated to left or right atrial collagen content, or to the spatial heterogeneity of atrial fibrosis. There was a weak and even negative correlation between the serum PINP concentration and the degree of fibrosis in both the left and the right atrium (r (s) = -0.65 (p = 0.04) and r (s) = -0.42 (p = 0.03), respectively). A high degree of interstitial atrial fibrosis indicates a high degree of spatial heterogeneity of interstitial collagen. Although serum PICP is known to be correlated with ventricular fibrosis, this and other serum markers of collagen turnover (PINP, PIIINP, and ICTP) do not directly reflect atrial fibrosis in patients with severe cardiac disease.

Angiotensin Type 2 Receptor Agonist Compound 21 Reduces Vascular Injury and Myocardial Fibrosis in Stroke-Prone Spontaneously Hypertensive Rats

-Angiotensin type 2 receptor-mediated effects of angiotensin II appear to counteract many of the effects mediated via the angiotensin type 1 receptor. Compound 21 (C21), a selective angiotensin type 2 receptor agonist, has demonstrated beneficial effects on cardiac function after myocardial infarction in rodents. We hypothesized that C21 alone or in combination with an angiotensin type 1 receptor antagonist would blunt the development of hypertension and vascular damage in stroke-prone spontaneously hypertensive rats. Six-week-old stroke-prone spontaneously hypertensive rats received C21 (1 mg/kg per day), the angiotensin type 1 receptor antagonist losartan (10 mg/kg per day), C21 plus losartan, or vehicle PO for 6 weeks. Systolic blood pressure was lower in losartan and C21-losartan combination groups (P<0.001). Endothelium-dependent relaxation was enhanced (P<0.001) in the C21-losartan combination group at lower acetylcholine concentrations. C21 or C21-losartan combination reduced vascular stiffness, aortic medial and myocardial interstitial collagen content, and aortic fibronectin (P<0.05). C21 and losartan decreased the expression of 2 genes associated with cardiac hypertrophy, myosin heavy chain-β (myh7) by 30 to 50%, and α-skeletal muscle actin by 30% to 35% (P<0.05). C21-losartan combination caused an additional 40% reduction in myh7 compared with C21 (P<0.01). Aortic superoxide generation was reduced equally by the 3 treatments (P<0.001). Monocyte/macrophage infiltration in the aorta and kidney (P<0.001) and T-lymphocyte infiltration in the renal cortex (P<0.05) were lowered similarly by the 3 treatments. These data suggest that C21 alone or in combination with losartan may improve endothelial function and vascular composition and mechanics by reducing oxidative stress, collagen content, fibronectin, and inflammatory cell infiltration in stroke-prone spontaneously hypertensive rats.

Selective Inhibition of Matrix Metalloproteinase-13 Increases Collagen Content of Established Mouse Atherosclerosis

Evidence has linked collagen loss with the onset of acute coronary events. This study tested the hypothesis that selective matrix metalloproteinase-13 (MMP-13) collagenase inhibition increases collagen content in already established and nascent mouse atheromas. In vitro and in situ experiments documented the selectivity and efficacy of an orally available MMP-13 inhibitor (MMP13i-A). In vivo observations monitored macrophage accumulation and MMP-13 activity using molecular imaging. After 10 weeks of MMP13i-A treatment, apolipoprotein E-deficient mice with evolving or established lesions exhibited reduced MMP-13 activity without affecting macrophage content, measured either by intravital microscopy or fluorescence reflectance imaging. Histological analysis indicated that MMP13-iA did not affect plaque size or macrophage or smooth muscle cell accumulation. Administration of MMP13i-A to mice with evolving or established atheromas substantially increased plaque interstitial collagen content in the intima and locally in the fibrous cap, compared with vehicle-treated controls. Analysis of collagen revealed thicker collagen fibers within the plaques of treated groups. Pharmacological MMP-13 inhibition yields collagen accumulation in plaques (a feature associated in humans with resistance to rupture), even in established plaques. This study, of considerable clinical relevance, furnishes new mechanistic insight into regulation of the plaque's extracellular matrix and validates molecular imaging for studying plaque biology.

Resveratrol Improves Cardiovascular Function in DOCA-Salt Hypertensive Rats

The phytoalexin resveratrol (3,4',5-trihydroxy-trans-stilbene) may attenuate cardiovascular disease in man. This study has determined whether treatment with resveratrol (1 mg/kg/day orally) prevented cardiac fibrosis and the decreased cardiovascular function in the DOCA-salt hypertensive rat as a model of human hypertension. Uninephrectomised rats (UNX) administered DOCA (25mg every 4th day sc) and 1% NaCl in drinking water for 28 days developed cardiac and vascular remodelling. In these DOCA-salt rats, resveratrol decreased inflammatory cell infiltration, decreased cardiac fibrosis (left ventricular interstitial and perivascular collagen content) and improved cardiac and vascular function. Resveratrol attenuated other features of cardiovascular remodelling such as increases in systolic blood pressure, left ventricular wet weight, left ventricular wall thickness, diastolic stiffness constant, as well as decreased cardiac contractility and prolonged action potential duration characteristic of DOCA-salt rats. In summary, resveratrol, at a nutritionally relevant dose, prevents or attenuates the adverse changes in the cardiovascular system. We propose that the anti-inflammatory and anti-fibrotic effects of resveratrol are responsible, at least in part, for its amelioration in cardiovascular remodelling in DOCA-salt rats. These actions of resveratrol could play an important role in the protective effects on the human cardiovascular system reported for this constituent of red wine.

Editorial [Hot topic: Crucial Role of Redox Signaling in the Regulation of Heart Health (Guest Editor: Dipak K. Das)

Editorial [Hot topic: Crucial Role of Redox Signaling in the Regulation of Heart Health (Guest Editor: Dipak K. Das)

Impact of Mechanical Unloading on Microvasculature and Associated Central Remodeling Features of the Failing Human Heart

This study investigates alterations in myocardial microvasculature, fibrosis, and hypertrophy before and after mechanical unloading of the failing human heart. Recent studies demonstrated the pathophysiologic importance and significant mechanistic links among microvasculature, fibrosis, and hypertrophy during the cardiac remodeling process. The effect of left ventricular assist device (LVAD) unloading on cardiac endothelium and microvasculature is unknown, and its influence on fibrosis and hypertrophy regression to the point of atrophy is controversial. Hemodynamic data and left ventricular tissue were collected from patients with chronic heart failure at LVAD implant and explant (n = 15) and from normal donors (n = 8). New advances in digital microscopy provided a unique opportunity for comprehensive whole-field, endocardium-to-epicardium evaluation for microvascular density, fibrosis, cardiomyocyte size, and glycogen content. Ultrastructural assessment was done with electron microscopy. Hemodynamic data revealed significant pressure unloading with LVAD. This was accompanied by a 33% increase in microvascular density (p = 0.001) and a 36% decrease in microvascular lumen area (p = 0.028). We also identified, in agreement with these findings, ultrastructural and immunohistochemical evidence of endothelial cell activation. In addition, LVAD unloading significantly increased interstitial and total collagen content without any associated structural, ultrastructural, or metabolic cardiomyocyte changes suggestive of hypertrophy regression to the point of atrophy and degeneration. The LVAD unloading resulted in increased microvascular density accompanied by increased fibrosis and no evidence of cardiomyocyte atrophy. These new insights into the effects of LVAD unloading on microvasculature and associated key remodeling features might guide future studies of unloading-induced reverse remodeling of the failing human heart.

Hyperoxia increases the uptake of 5-fluorouracil in mammary tumors independently of changes in interstitial fluid pressure and tumor stroma

BackgroundHypoxia is associated with increased resistance to chemo- and radiation-therapy. Hyperoxic treatment (hyperbaric oxygen) has previously been shown to potentiate the effect of some forms of chemotherapy, and this has been ascribed to enhanced cytotoxicity or neovascularisation. The aim of this study was to elucidate whether hyperoxia also enhances any actual uptake of 5FU (5-fluorouracil) into the tumor tissue and if this can be explained by changes in the interstitium and extracellular matrix.MethodsOne group of tumor bearing rats was exposed to repeated hyperbaric oxygen (HBO) treatment (2 bar, pO2 = 2 bar, 4 exposures à 90 min), whereas one group was exposed to one single identical HBO treatment. Animals housed under normal atmosphere (1 bar, pO2 = 0.2 bar) served as controls. Three doses of 5FU were tested for dose response. Uptake of [3H]-5FU in the tumor was assessed, with special reference to factors that might have contributed, such as interstitial fluid pressure (Pif), collagen content, oxygen stress (measured as malondialdehyd levels), lymphatics and transcapillary transport in the tumors.ResultsThe uptake of the cytostatic agent increases immediately after a single HBO treatment (more than 50%), but not 24 hours after the last repeated HBO treatment. Thus, the uptake is most likely related to the transient increase in oxygenation in the tumor tissue. Factors like tumor Pif and collagen content, which decreased significantly in the tumor interstitium after repeated HBO treatment, was without effect on the drug uptake.ConclusionWe showed that hyperoxia increases the uptake of [3H]-5FU in DMBA-induced mammary tumors per se, independently of changes in Pif, oxygen stress, collagen fibril density, or transendothelial transport alone. The mechanism by which such an uptake occur is still not elucidated, but it is clearly stimulated by elevated pO2.

Effects of AT1‐ and β‐adrenergic receptor antagonists on TGF‐β1‐induced fibrosis in transgenic mice

Transforming growth factor-beta1 (TGF-beta1) is involved in interstitial remodelling promoting collagen synthesis and suppressing collagen degradation by inhibition of collagenases. TGF-beta1 mediates angiotensin II-dependent effects and modulates beta1-adrenergic signalling. To study the effect of neuroendocrine antagonism on TGF-beta-induced hypertrophic and fibrotic phenotype, we treated TGF-beta1 (Cys223,225Ser) transgenic mice (TGF-beta1-TG) with either the beta1-receptor blocker metoprolol (MET), the angiotensin II type I (AT1)-receptor antagonist telmisartan (TEL) or an antibody blocking TGF-beta1 signalling (TGFbeta1-sR-Ab). Transforming growth factor-beta1-TG mice (8 weeks) overexpressing TGF-beta1 were treated with either TEL (10 mg kg(-1)), MET (350 mg kg(-1)) or a soluble TGF-beta1 receptor antibody (1 mg kg(-1)) for 6 weeks. Morphological analyses of interstitium and cardiomyocytes were related to expression of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) by immunoblotting and zymography. In TGF-beta1-TG mice, myocardial interstitial total collagen content was fourfold elevated compared to that of controls (P < 0.05) and was lowered under the treatment with TEL (P < 0.05). Protein expression of TIMP-1 and -4 was increased in TGF-beta1-TG but inhibited by TEL (TIMP-1 and TIMP-4) and MET (TIMP-1), while collagenase activity was decreased in TGF-beta1-TG and normalized by treatment with TEL (MMP-1 and MMP-13) and MET (MMP-1) (P < 0.05). Morphometric measurements of cardiomyocyte diameter and area demonstrated similar antihypertrophic effects for all treatment groups. The AT1-antagonist TEL reduced myocardial hypertrophy and interstitial fibrosis in TGF-beta1-TG mice by normalizing MMP/TIMP ratio. beta1-Adrenergic inhibition by MET as well as TGF-beta1 antagonism induced antihypertrophic rather than antifibrotic effects. Inhibition of both renin-angiotensin system and beta1-adrenergic system may exert different but synergistic effects to reduce myocardial remodelling.

Time course and mechanisms of left ventricular systolic and diastolic dysfunction in monocrotaline-induced pulmonary hypertension

Although pulmonary hypertension (PH) selectively overloads the right ventricle (RV), neuroendocrine activation and intrinsic myocardial dysfunction have been described in the left ventricle (LV). In order to establish the timing of LV dysfunction development in PH and to clarify underlying molecular changes, Wistar rats were studied 4 and 6 weeks after subcutaneous injection of monocrotaline (MCT) 60 mg/kg (MCT-4, n = 11; MCT-6, n = 11) or vehicle (Ctrl-4, n = 11; Ctrl-6, n = 11). Acute single beat stepwise increases of systolic pressure were performed from baseline to isovolumetric (LVPiso). This hemodynamic stress was used to detect early changes in LV performance. Neurohumoral activation was evaluated by measuring angiotensin-converting enzyme (ACE) and endothelin-1 (ET-1) LV mRNA levels. Cardiomyocyte apoptosis was evaluated by TUNEL assay. Extracellular matrix composition was evaluated by tenascin-C mRNA levels and interstitial collagen content. Myosin heavy chain (MHC) composition of the LV was studied by protein quantification. MCT treatment increased RV pressures and RV/LV weight ratio, without changing LV end-diastolic pressures or dimensions. Baseline LV dysfunction were present only in MCT-6 rats. Afterload elevations prolonged tau and upward-shifted end-diastolic pressure dimension relations in MCT-4 and even more in MCT-6. MHC-isoform switch, ACE upregulation and cardiomyocyte apoptosis were present in both MCT groups. Rats with severe PH develop LV dysfunction associated with ET-1 and tenascin-C overexpression. Diastolic dysfunction, however, could be elicited at earlier stages in response to hemodynamic stress, when only LV molecular changes, such as MHC isoform switch, ACE upregulation, and myocardial apoptosis were present.

Correlação do colágeno intersticial miocárdico do septo do ventrículo direito com a função ventricular em pacientes com cardiomiopatia isquêmica

FUNDAMENTO: O conteudo de colageno intersticial (CI) no miocardio exerce influencia no relaxamento e na contracao ventricular. A sua relacao com a funcao ventricular em pacientes (pcts) com cardiomiopatia isquemica (CMPI) nao esta plenamente estudada em humanos. OBJETIVO: Avaliar a relacao da quantidade de CI nas areas nao-infartadas no septo do ventriculo direito com a funcao ventricular na CMPI. METODOS: 31pcts com doenca arterial coronariana foram classificados em quatro grupos: Grupo C (Controle): 7pcts com as fracoes de ejecao dos ventriculos esquerdo (FEVE) e direito (FEVD) normais; Grupo 1: 5 pcts com FEVD < 40%;Grupo 2: 9 pcts com FEVE < 40%; Grupo 3: 10 pcts com disfuncao de ambos os ventriculos. A FEVD e a FEVE foram calculadas por meio da angiocardiografia radionuclidica. As amostras para analise do %CI foram obtidas por meio de biopsia endomiocardica do ventriculo direito e coradas pela tecnica do picrosirius red. RESULTADOS: A media do %CI foi significativamente maior no grupo 3 quando comparada com o grupo-c e com os grupos 1 e 2 (30,2 ± 7,9% vs. 6,8 ± 3,3% vs. 15,8 ± 4,1% vs. 17,5 ± 7,7%, respectivamente; p = 0,0001). O %CI foi tambem significativamente maior nos pacientes do grupo 2 quando comparado com o controle(17,5 ± 7,7% vs. 6,8 ± 3,3%, p = 0.0001). O %CI apresentou correlacao inversa com a FEVD (r = -0,50, p = 0.003) e FEVE (r = -0,70, p = 0,0001). CONCLUSAO: Na CMPI, o %CI encontra-se elevado nas areas nao-infartadas no septo do ventriculo direito e apresenta correlacao inversa com o a funcao ventricular direita e esquerda.

Effect of clenbuterol administration on the healthy murine heart

Clenbuterol has recently been shown to reverse pathologic cardiac remodeling during left ventricular assist device (LVAD) support, leading to restored ventricular function and explantation of LVAD devices. However, others have not been able to support these observations. Our hypothesis is that the beta(2)-adrenergic activity of clenbuterol induces cardiac extracellular matrix (ECM) remodeling, resulting in increased interstitial fibrillar collagen content and altered diastolic function that may account for these conflicting reports. The intent of this study is to characterize the effect of clenbuterol on healthy murine hearts with transthoracic echo and histology. C57BL/6 female mice were administered 2.4 microg/kg/day of clenbuterol in the drinking water for 7 days and analysis conducted on day 8-24 hours after the last dose of clenbuterol. Histological analysis demonstrated an increase in left ventricular ECM collagen content in a control group compared with the clenbuterol group (density 0.32 +/- 0.16 compared to 2.01 +/- 0.30 RD/mm(2)). The ventricular fibrosis was supported by altered diastolic function measured by transthoracic echo where there was a significant increase in isovolumic relaxation time, and left atrial dimension and a decrease in left ventricular free wall tissue Doppler ratios. Our study showed no significant differences in left ventricular ejection fraction, cardiac output, or heart rate between the clenbuterol and control groups. These data suggest that the beta-2 adrenergic activity of clenbuterol increases ECM fibrillar collagen concentrations in normal hearts, resulting in altered diastolic function.

The effect of daily administration of IL-18 on cardiac structure and function

Recently, the cytokine Interleukin-18 (IL-18) has been shown to be increased as a result of cardiac surgery. Elevated IL-18 has been associated with neurological dysfunction, systemic inflammatory response syndrome (SIRS), and multiple organ dysfunction syndrome (MODS) post open-heart surgery. The intent of the study contained herein was to determine the effect of IL-18 administration on cardiac function and structure. Eight C57BL/6 female mice were treated daily with 0.5microg/mouse of recombinant IL-18 for 7 days. Long axis echocardiography (ECHO) measurements of the anatomical and hemodynamic function of the heart for all mice were studied 24h after the last dose. The left ventricular wet weights increased from 84 +/- 1 to 93 +/- 3 mg when comparing the placebo (n = 8) with the IL-18 groups, respectively (p = 0.01). With ECHO analysis, IL-18 significantly increased left ventricular (LV) mass, the left atrium dimensions (LA), and the left ventricular posterior wall thickness (LVPW) over the 8-day time period (p < 0.01). There was a 5-fold increase in interstitial cardiac collagen content and a 30% increase in myocyte size in the IL-18 compared with the control groups (p < 0.01). Administration of IL-18 appears to induce interstitial fibrosis and myocyte hypertrophy, resulting in increased ventricular stiffness. Thus, increased IL-18 during and post open-heart surgical procedures may induce left ventricular diastolic dysfunction and affect post-operative outcomes.

Abstract 1346: Rac1 GTPase Activtiy is Associated with Atrial Fibrosis and Fibrillation

Patients with atrial fibrillation (AF) are characterized by increased atrial fibrosis. The underlying signal transduction is incompletely understood. We hypothesized that activation of Rac1 GTPase contributes to increased fibrosis and atrial fibrillation via activation of NADPH oxidase and production of reactive oxygen species. Methods and Results: Samples of the left atrial appendage were analyzed in 8 patients with AF undergoing mitral valve surgery and 7 patients with sinus rhythm (SR) matched for atrial diameter. Despite same size of the atria, collagen content was significantly higher in AF compared to SR (14.9±2.1% vs. 8.5±1.3%). Expression analysis showed that AF was associated with upregulation of connective tissue growth factor (CTGF). Atria of patients with AF were characterized by significant upregulation of Rac1 total protein expression (Western blot), Rac1 activity (PAK pull-down assays) and a 20-fold upregulation of NADPH oxidase activity compared to SR (2225±500%). In order to test whether Rac1 plays a causal role in the pathogenesis in AF, transgenic mice with cardiac overexpres-sion (αMHC promoter) of Rac1 (RacET) were compared to wildtype (WT) and WT undergoing transaortic constriction (TAC, 360 μm). After 16 months, echocardiography showed similar left ventricular hypertrophy in RacET and TAC. RacET but not TAC exhibited atrial enlargement; 75% of RacET but no WT or TAC showed AF. Interstitial collagen content was significantly increased in the atria of RacET (44±1% of area vs 19±5% in WT). In contrast, interstitial fibrosis in TAC atria did not significantly differ from WT (31±6%). In the left ventricle, both RacET and TAC mice showed an elevated collagen content compared to the control group (WT 9±2%, RacET 29±3%; TAC 24±4%). In all mice, atrial collagen content exceeded ventricular collagen. All effects are significant with p&lt;0.05. Conclusion: Left atria of patients with AF exhibit upregulation of Rac1, increased NADPH oxidase activity and interstitial fibrosis. Cardiac-specific overexpression of Rac1 in mice results atrial fibrosis and fibrillation independent of left ventricular hypertrophy. Rac1-GTPase mediated activation of left atrial NADPH-oxidase may represent a novel target for the prevention of atrial fibrosis.

Chronic sodium hydrosulfide treatment decreases medial thickening of intramyocardial coronary arterioles, interstitial fibrosis, and ROS production in spontaneously hypertensive rats

Hydrogen sulfide (H(2)S) is a gasotransmitter that regulates cardiovascular functions. The present study aimed to examine the hypothesis that chronic treatment with sodium hydrosulfide (NaHS, an H(2)S donor) is able to prevent left-ventricular remodeling in spontaneously hypertensive rats (SHR). Four-week-old SHR were treated with NaHS (10, 30, and 90 micromol x kg(-1) x day(-1)), a combination of NaHS (30 micromol x kg(-1) x day(-1)) and glibenclamide (5 mg x kg(-1) x day(-1)), glibenclamide alone (5 mg x kg(-1) x day(-1)), hydralazine alone (10 mg x kg(-1) x day(-1)), and placebo for 3 mo. At the end of the treatment period, variables such as cardiac geometry and function, intramyocardial arterioles ranging in diameter from 25 to 100 microm, perivascular and interstitial collagen content, reactive oxygen species (ROS), thiol groups, conjugated dienes, and DNA base modification were examined. The novel finding of the present study is that chronic NaHS treatment prevented the hypertrophy of intramyocardial arterioles and ventricular fibrosis, as well as decreased myocardial ROS and conjugated diene levels. The cardioprotective effects were blunted by coadministration of glibenclamide, suggesting a role of ATP-sensitive potassium channels in mediating the action of NaHS. Hydralazine caused a comparable reduction of blood pressure compared with NaHS treatment; however, it exerted no effect on the remodeling process or on ROS and conjugated diene levels. Moreover, NaHS treatment caused an increase in myocardial thiol group levels, whereas DNA base modification was not altered by NaHS treatment. In conclusion, the superior cardioprotective effects of NaHS treatment are worthy to be further explored to develop novel therapeutic approaches for the treatment of cardiac remodeling in hypertension.

Effects of endothelin ETAreceptor blocker LU 135252 on cardiac remodeling and survival in a hypertensive rat model of chronic heart failure

To investigate whether the endothelin ETA receptor blocker provides similar benefit on cardiac remodeling and survival in a hypertensive rat model of chronic heart failure (CHF). Male stroke-prone spontaneously hypertensive (SHR-SP) rats were subjected to permanent ligation of the left coronary artery and were treated for 6 weeks with the endothelin ETA receptor blocker LU 135252 (30 mg.kg(-1).d(-1)) starting 24 h after ligation or untreatment. Sham-operated rats served as normal controls. The mean arterial blood pressure (MAP), heart rate (HR), left ventricular end-diastolic pressure (LVEDP), left ventricular contractility (LV dp/dt(max)), left ventricular inner diameter (LVD) and circumference (LVC), septal thickness, left ventricular interstitial collagen content (ICC) and heart weight (HW) were measured at the end of the treatment. Compared with the untreated group, LU 135252 tended to increase HW (1.43 +/-0.03 vs 1.38 +/-0.04 g; P> 0.05), increased LVD (7.65+/-0.24 mm vs 6.58+/-0.14 mm; P<0.05), markedly increased LVC (30.11+/-0.83 mm vs 24.82+/-0.85 mm; P< 0.01) and reduced left ventricular ICC (3.79%+/-0.09% vs 6.71%+/-0.11%; P< 0.01), slightly lowered MAP (132+/-6 mmHg vs 142+/-4 mmHg; P>0.05), reduced LVEDP (14 4 mmHg vs 27+/-4 mmHg; P<0.05) and improved LV dp/dtmax (4230+/-450 mmHg/s vs 1950+/-400 mmHg/s; P<0.05); survival was not prolonged significantly (13% vs 11%; P=NS). In this hypertensive rat model of CHF, chronic endothelin ETA receptor blockade with LU 135252 improves cardiac hemodynamics, however, it does not affect long-term survival and worsens cardiac remodeling. Thus, endothelin ETA receptor antagonists are unlikely to have an important role in the management of patients with CHF.

Morphological and molecular analysis of idiopathic gingival fibromatosis: a case report

We analyse a case of idiopathic gingival overgrowth using morphological and molecular methods. As this overgrowth involves collagen accumulation in the gingival connective tissue, we measured the collagen turnover to clarify the pathogenic mechanisms potentially involved. The patient was a 29-year-old Italian woman with enlargement of the gingivae throughout the entire mandible and maxilla. Morphological analyses were carried out on haematoxylin-eosin and Sirius red-stained paraffin-embedded gingival sections. mRNA levels of collagen type I and III, matrix metalloproteinase (MMP)-1, transforming growth factor-beta1 and lysyl hydroxylase (LH)2b were determined by RT-PCR on cultured gingival fibroblasts and compared with healthy control fibroblasts. Interstitial collagen and MMP-1 content in the supernatants were assessed, respectively, by dot blot and SDS zymography. In Sirius red-stained sections of the patient's overgrown gingivae, interstitial collagen content was 29% higher than controls. Her gingival fibroblasts had higher collagen type I, MMP-1 and LH2b gene expression and unmodified interstitial collagen, type I protein levels in the supernatants. These findings would seem to suggest that in this case collagen accumulation in the gingival connective tissue was not associated with increased synthesis and decreased degradation.