Left Ventricular Endomyocardial Biopsy Research Articles

Circulation : Clinical Summaries

<i>Circulation</i> : Clinical Summaries

Toll-like receptor 4 is a marker of immune-mediated damage in patients with inflammatory cardiomyopathy

Purpose: Toll-like receptor 4 has been involved in the development of experimental autoimmune myocarditis. The aim of the study is to determine whether Toll-like receptor 4 plays a role in human myocarditis responsive to immunosuppression. Methods: Left ventricular endomyocardial biopsies from a subgroup of patients from the TIMIC trial were analyzed for the immunohistochemical expression of Toll-like receptor 4. In particular 18 patients (8 males,16 females, age 48.6±12.1 ys) with chronic (>6 months), virus negative (negative PCR for EV, AV, EBV, PVB19, HCV, Inf A/B, HSV1/2) inflammatory cardiomyopathy (LVEF 29.4±7.3%), responders to a 6 months treatment with prednisone and azathioprine were studied (group A). Controls were 10 left ventricular biopsies of patients with virus-negative inflammatory cardiomyopathy not responders to immunosuppression (group B) and 10 normal left ventricular surgical samples (group C). A semiquantitative evaluation of the immunoreactivity (grading from 0 to 4) was applied. Results: A focal intense positive cytoplasmic immunoreactivity was observed in cardiac myocytes of all patients in group A (grading 3.5±0.5) (panel A), with a statistically significant difference (p<0.001) compared with group B (grading 0.8±0.4) (panel B) and controls (0.2±0.4) (panel C). ![Figure][1] Conclusion: Toll-like receptor 4 is highly expressed in human immune-mediated myocarditis. It can be considered as a new marker in patient selection for immunosuppressive/immunomodulatory therapy. [1]: pending:yes

Blood myeloid dendritic cell reduction and differences between myocarditis and dilated cardiomyopathy

Aims: The role of dendritic cells (DCs) in cardiomyopathies is unknown yet. Dilated cardiomyopathy (DCM) is often preceded by myocarditis, hence the two conditions often clinically overlap. In order to investigate the role of DCs in diagnosing myocarditis/DCM, we prospectively investigated the circulating DC precursors (DCPs) as well as mature DCs in endomyocardial biopsies (EMBs) from patients with acute myocarditis and newly diagnosed DCM. Methods: Blood myeloid (mDCPs) and plasmacytoid (pDCPs) DC precursors were flow cytometrically analyzed in patients with newly onset heart failure (HF) and biopsy proven DCM (n=43, mean EF 29±10%) and patients with MRI confirmed myocarditis (n=23). Individuals with excluded heart disease were used as controls (n=52). Left-ventricular EMBs were performed when clinically indicated (n=45) and were immunohistologically analyzed for the presence of mature myeloid DCs (CD83+) and tissue fibrosis (Sirius red). Analyses of circulating DCPs were performed initially and at repeated follow-up intervals of 6 months. Correlation analysis was performed between circulating DCPs and mature DCs in EMBs, as well as clinical, echocardiographic and invasive left-ventricular parameters. Results: Relative and absolute numbers of circulating mDCPs were reduced in patients with myocarditis compared to controls (0.065% vs 0.2%,p<0.001). Following clinical recovery of myocarditis, circulating mDCPs tended to normalise (0.015% vs 0.2%,p=0.008). Circulating mDCPs of DCM patients were at initial presentation unchanged compared to controls (0.18% vs 0.2%,p=0.258), but were significantly reduced at 6- and 12-month-follow-up (0.16% vs 0.2%,p=0.047 and 0.14% vs 0.2%,p=0.021). No statistical significance was observed for pDCPs. Correlation analysis revealed inverse correlation of blood mDCPs with myocardial mature DCs in EMBs (r=-0.522,p=0.026). We also found an inverse correlation of blood mDCPs with cTNI (r=-0.826,p<0.001), CRP (r =-0.388,p=0.014), invasively measured EF (r=-0.383,p=0.026), but not BNP (p=0.921). Blood mDCPs positively correlated with tissue fibrosis (r=0.395,p=0.05), mitral and tricuspid regurgitation (r=0.395,p=0.012 and r=0.47,p=0.002). Conclusions: Circulatory mDCs migrated into the myocardium of these patients. While their peripheral reduction was acute in myocarditis, it was delayed and chronic/progressive in DCM. Thus, inflammatory processes in DCM were not causative, but rather secondary to structural myocardial changes. DCs could prove to be discriminatory between myocarditis and DCM in early diagnosis.

High prevalence of myocarditis in patients with hypertensive heart disease and cardiac deterioration

Structural abnormalities causing cardiac deterioration in hypertensive heart disease (HHD) are poorly understood. The aim of the study was to evaluate left ventricular (LV) myocardial changes in patients with HHD and cardiac failure. Among 1229 patients undergoing an LV or biventricular endomyocardial biopsy because of unexplained LV dysfunction from 2000 to 2010, 45 had HHD. The HHD population had non-invasive assessment of cardiac wall thickness, diameters, and function; endomyocardial samples were processed for histology, immunohistochemistry, and polymerase chain reaction (PCR) for cardiotropic viruses. Mean LV end-diastolic diameter was 63.3 ± 5.9 mm, LV ejection fraction 29.7 ± 7.6%, and maximal wall thickness 12.8 ± 0.9 mm. At histology, hypertrophy with degeneration of cardiomyocytes, increased external/lumen ratio of intramural arterioles, and myocardial fibrosis were observed in 17 patients (38%). In the remaining 28 patients (62%), these histological changes were associated with myocarditis. Myocarditis was present in 40.4% of the cohort without HHD. PCR was negative in HHD without inflammation while it was positive in 10 subjects with myocarditis (enterovirus in 3 cases, adenovirus 4, influenza virus 3). Addition of immunosuppression to supportive therapy in the 18 virus-negative myocarditis subjects was followed at 6 months by significant recovery of LV function in 15 (83%; ejection fraction from 25.3 ± 9.3% to 49.5 ± 9.8%) while LV function improved to a minor extent ( ejection fraction from 25 ± 5.1% to 36 ± 4.4%) in 12 of 27 patients (44%) on supportive treatment alone. Myocarditis is a major cause of cardiac deterioration in patients with HHD; its recognition may improve patient treatment and outcome.

The Results and Technique of Left Ventricular Endomyocardial Biopsy

The Results and Technique of Left Ventricular Endomyocardial Biopsy

Coronary telangiectasia associated with hypertrophic cardiomyopathy

Coronary telangiectasia (CT) is a rare congenital anomaly causing ventricular shunt and myocardial ischaemia. Its prevalence, genetic background, and impact in human hypertrophic cardiomyopathy (HCM) are unknown and were therefore investigated in this study. Among 445 patients with HCM, 195 had a coronary angiography and 124 a left ventricular endomyocardial biopsy. CT draining into the ventricular cavities was observed in 5 of 195 HCM patients (2.5%), whereas it was detected in 0.1% of 1000 consecutive subjects without congenital anomalies undergoing coronary angiography. Patients with CT-HCM underwent a total body computed tomography scan to investigate the presence of systemic vascular malformations. HCM-related MYH7, MYBPC3, TNNT2, and TPM1 genes and hereditary haemorragic telangiectasia-related endoglin and activin receptor-like kinase 1 genes were analysed. Histology, clinical profile, and outcome of CT-HCM patients were correlated with those of 22 control HCM patients. No mucocutaneous or systemic vascular malformations were detected. Gene analysis showed a MYH7 mutation in two patients, with an associated endoglin point mutation. Histology showed in the CT-HCM cohort a more pronounced myocardial fibrosis (29.8 ± 3.8%) compared with HCM controls (13 ± 2.6%), and disorganized cardiomyocytes separated by thin-walled large vessels adherent to the endocardium. Clinically, the CT-HCM cohort had a higher arrhythmic profile at diagnosis and increased incidence of implantable cardioverter defibrillator (ICD) implantations and arrhythmic deaths during a long-term follow-up. CT is detectable in 2.5% of HCM patients vs. 0.1% of the general population; it may derive from a co-existing endoglin gene mutation and cause a prominent, potentially arrhythmogenic myocardial fibrosis.

Diagnostic contribution of left ventricular endomyocardial biopsy in patients with clinical phenotype of hypertrophic cardiomyopathy

Hypertrophic cardiomyopathy phenotype is shared by heterogeneous entities. The purpose of the study was to evaluate the diagnostic role of left ventricular endomyocardial biopsy. One hundred fifty-one consecutive patients with unexplained left ventricular hypertrophy and normal/elevated QRS voltages or left bundle-branch block underwent left ventricular endomyocardial biopsy because of associated left ventricular dysfunction (37%), presence of sporadic form of left ventricular hypertrophy (32%), or patient desire for a definite diagnosis (31%). Biopsy samples were processed for histology and electron microscopy. Blood samples were collected for histologically oriented gene analysis of major sarcomeric (MYH7, MYBPC3, TNNT2, TPM1) and lysosomal (LAMP2, PRKAG2, α-galactosidase A) proteins. Histology showed changes consistent/compatible with hypertrophic cardiomyopathy in 124 patients: myocardial storage disease in 18 due to Fabry disease in 12 and glycogen-storage disease in 6 and myocardial infiltrative disease in 9 because of amyloidosis in 7 and sarcoidosis in 2. Gene analysis was positive in 67% of patients with hypertrophic cardiomyopathy (MYH7 mutation in 36, MYBP in 29, TNNT2 in 14, and TPM1 in 5) and in 83% of patients with lysosomal storage disease (α-galactosidase A mutation in 12, PRKAG2 in 2, and LAMP2 in 1). In patients with hypertrophic cardiomyopathy phenotype, left ventricular endomyocardial biopsy is safe and may recognize infiltrative/storage diseases in up to 18% of evolving and sporadic cases.

Circulation : Clinical Summaries

<i>Circulation</i> : Clinical Summaries

Diagnostic Performance of CMR Imaging Compared With EMB in Patients With Suspected Myocarditis

The goal of this study was to assess the diagnostic performance of cardiac magnetic resonance (CMR) compared with endomyocardial biopsy in patients with suspected acute myocarditis (AMC) and chronic myocarditis (CMC). Several studies have reported an encouraging diagnostic performance of CMR in myocarditis. However, the comparison of CMR with clinical data only and the use of preselected patient populations are important limitations of the majority of these reports. One hundred thirty-two consecutive patients with suspected AMC (defined by symptoms ≤ 14 days; n = 70) and CMC (defined by symptoms >14 days; n = 62) were included. Patients underwent cardiac catheterization with left ventricular endomyocardial biopsy and CMR, including T(2)-weighted imaging for assessment of edema, T(1)-weighted imaging before and after contrast administration for evaluation of hyperemia, and assessment of late gadolinium enhancement. CMR results were considered to be consistent with the diagnosis of myocarditis if 2 of 3 CMR techniques were positive. Within the total population, myocarditis was the most common diagnosis on endomyocardial biopsy analysis (62.9%). Viral genomes were detected in 30.3% (40 of 132) of patients within the total patient population and significantly more often in patients with AMC than CMC (40.0% vs. 19.4%; p = 0.013). For the overall cohort of patients with either suspected AMC or CMC, the diagnostic sensitivity, specificity, and accuracy of CMR were 76%, 54%, and 68%, respectively. The best diagnostic performance was observed in patients with suspected AMC (sensitivity, 81%; specificity, 71%; and accuracy, 79%). In contrast, diagnostic performance of CMR in suspected CMC was found to be unsatisfactory (sensitivity, 63%; specificity, 40%; and accuracy, 52%). The results of this study underline the usefulness of CMR in patients with suspected AMC. In contrast, the diagnostic performance of CMR in patients with suspected CMC might not be sufficient to guide clinical management.

Peripartum Cardiomyopathy Presenting with Syncope due to Torsades de Pointes: a Case of Long QT Syndrome with a Novel KCNH2 Mutation

Peripartum cardiomyopathy (PPCM) is a cardiomyopathy of unknown cause that occurs in the peripartum period. We report a case of PPCM presenting with syncope 1 month after an uncomplicated delivery. Electrocardiography showed Torsades de pointes (TdP) and QT interval prolongation. Echocardiography showed left ventricular systolic dysfunction and endomyocardial biopsy showed myocyte degeneration and fibrosis. Administration of magnesium sulfate and temporary pacing eliminated recurrent TdP. Genetic analyses revealed that recurrent TdP occurred via electrolyte disturbance and cardiac failure due to PPCM on the basis of a novel mutation in KCNH2, a gene responsible for inherited type 2 long QT syndrome.

High prevalence of intramural coronary infection in patients with drug-resistant cardiac syndrome X: comparison with chronic stable angina and normal controls

BackgroundCoronary microvascular dysfunction has been reported along with myocardial viral infection. Whether intramural coronary vessels infection plays a role in patients with cardiac syndrome X (CSX) is unknown.MethodsThirteen consecutive patients...

Comparative Evaluation of Left and Right Ventricular Endomyocardial Biopsy

Endomyocardial biopsy (EMB) represents the gold standard for diagnosing myocarditis and nonischemic cardiomyopathies. This study focuses on the risk of complications and the respective diagnostic performance of left ventricular (LV), right ventricular (RV), or biventricular EMB in patients with suspected myocarditis and/or cardiomyopathy of unknown origin. In this 2-center study, 755 patients with clinically suspected myocarditis (n=481) and/or cardiomyopathy of nonischemic origin including those with infiltrative or connective tissue disease (n=274) underwent either selective LV-EMB (n=265; 35.1%), selective RV-EMB (n=133; 17.6%), or biventricular EMB (n=357; 47.3%) after coronary angiography and exclusion of significant coronary artery disease. Cardiovascular magnetic resonance, including late gadolinium enhancement, imaging was performed in 540 patients (71.5%). The major complication rate for LV-EMB was 0.64% and for RV-EMB, 0.82%. Considering postprocedural pericardial effusion that occurred after biventricular EMB, the minor complication rate for LV-EMB varied between 0.64% to 2.89% and for RV-EMB, between 2.24% and 5.10%. Diagnostic EMB results were achieved significantly more often in those patients who underwent biventricular EMBs (79.3%) compared to those who underwent either selective LV-EMB or selective RV-EMB (67.3%; P<0.001). In patients with biventricular EMB, myocarditis was diagnosed in LV-EMB samples in 18.7% and in RV-EMB samples in 7.9% (P=0.002) , and it was diagnosed in both ventricles in 73.4%. There were no differences in the number of positive LV-EMB, RV-EMB, or LV- and RV-EMB findings when related to the site of cardiovascular magnetic resonance-based late gadolinium enhancement. Both LV-EMB and RV-EMB are safe procedures if performed by experienced interventionalists. The diagnostic yield of EMB may be optimized when samples from both ventricles are available. Preferential biopsy in regions showing late gadolinium enhancement on cardiovascular magnetic resonance does not increase the number of positive diagnoses of myocarditis.

Myocardial Fiber Diameter as a Good Indicator of Outcome in Batista's Operation

Despite the initial promissory results of partial left ventriculectomy, or Batista's operation, the postoperative mortality associated with the procedure has been too high. We described a histopathologic study performed to identify histological parameters that could help to determine outcomes of patients undergoing this procedure. Myocardial fiber diameter, myocardial fibrosis, thickness of the compact wall, and number of cells presenting from the endocardium to epicardium were analyzed in 32 patients with idiopathic dilated cardiomyopathy who underwent Batista's operation. Data were grouped by patients who died < or = 6 months and patients who survived for > 6 months after the surgical procedure. Additional analyses were performed to compare results according the causes of death and to test the application of these results to biopsy. Myocardial fiber diameter was the only index that could distinguish the two groups. Myocardial fiber diameter < 22 microm distinguished the group of patients who survived the 6-month postoperative period from patients who died during that time with sensitivity of 85.7 and specificity of 72.2. The subendocardial region of the compact wall and the trabecular portion of the wall exhibited comparable results. Our results indicate that the myocardial fiber diameter of samples from the trabecular or subendocardial compact wall regions may help predict the outcome of left ventriculectomy. Samples from the trabecular or subendocardial compact wall regions were used for analysis. Further prospective studies involving left ventricular endomyocardial biopsies are necessary to confirm if the use of myocardial fiber diameter in the selection of patients for surgery improves the index of success of Batista's operation. Other factors that are involved remain unclear.

Low level myocardial parvovirus B19 persistence is a frequent finding in patients with heart disease but unrelated to ongoing myocardial injury

While myocardial parvovirus B19 (B19V), aside from enteroviruses (EV) and adenoviruses (ADV), has recently been found often in patients with myocarditis and idiopathic dilated cardiomyopathy (IDC), the pathogenetic significance of B19V genomes in those patients has not yet been sufficiently elucidated. In the present study, left ventricular endomyocardial biopsies from 24 patients with left ventricular ejection fraction (LVEF) below 55% due to IDC, and tissue from the right atrial appendage of 10 control patients undergoing bypass surgery with normal LVEF (>55%) were investigated for B19V, ADV, and EV genomes by specific nested polymerase chain reaction (PCR), by real time PCR or by reverse-transcription PCR, respectively. The myocardial tissue samples from the 10 controls were analyzed each in three different virological laboratories for B19V. In the IDC group, the frequency of the myocardial virus genomes found in 54% (13/24) of the patients was as follows: B19V: 50% (12/24), EV: 8% (2/24), including one patient with B19V and EV, and ADV: 0% (0/24). For comparison, the prevalence of B19V genomes was between 30% and 60% in the control group as detected in three different laboratories, but all these control subjects were EV- and ADV-negative. The number of B19V gene copies, however, was very low and similar both in the IDC and control group. In the majority of patients myocardial B19V persistence was associated with a low virus load irrespective of the underlying heart disease so that it may be of no importance in the pathogenesis of IDC.

Cardiac Titin

Titin constitutes the third myofilament of cardiac muscle, with a single giant polypeptide spanning from Z-disk to the M-band region of the sarcomere1 (Fig. 1). The ∼1.0 MDa region in the I-band is extensible and consists of tandemly arranged immunoglobulin (Ig)-like domains that make up proximal (near Z-disk) and distal (near A-I junction) segments, interspersed by the PEVK sequence (rich in proline, glutamate, valine, and lysine residues) and the N2B element2. Each functions as a distinct spring element3. The C-terminal ∼2 MDa of titin is located in the A-band and is inextensible. It is composed of regular arrays of Ig and fibronectin type 3 (Fn3) modules forming so-called super-repeats2. A-band titin may function as a molecular ruler, regulating assembly of the thick filament2,4,5. Titin's ∼250-kDa COOH-terminal region is an integral part of the M-band and contains a kinase domain6,7. As in the Z-disk, where titin filaments from opposite sarcomeres overlap, titin filaments from opposite half-sarcomeres overlap within the M-band, where they are interconnected by M-band proteins8. Thus, titin filaments with opposite polarity overlap in both Z-disk and M-band, forming a contiguous filament along the myofibril. In this review, we discuss titin's functions in the heart, with an emphasis on its role in diastolic function and the various mechanisms whereby passive stiffness can be tuned. Due to space constraints, it has not been possible to provide inclusive references to all original articles in the field. Figure 1 Schematic of titin in sarcomere. Differential splicing Titin is encoded by a single gene containing 368 exons. Multiple splice pathways in the I-band encoding region (∼230 exons) give rise to isoforms with different spring composition9. The three cardiac isoform classes are shown in Fig. 1. The relatively small ∼3.0 MDa isoform is known as N2B titin (it contains the N2B element)9. A second class also contains the N2A element, and is termed N2BA titin. N2BA titins have a longer PEVK segment and a variable number of additional Ig domains resulting in a ∼3.3-3.5 MDa size9. The third class includes isoforms that predominate in fetal-neonatal life which contain additional spring elements in both tandem Ig and PEVK regions, resulting in a ∼3.6-3.8 MDa size protein10-12. These isoforms gradually disappear during postnatal development. Regulation of the spring composition of titin in fetal-neonatal myocardium allows adjustment of diastolic filling properties during development.

Persistent Diastolic Dysfunction Late After Valve Replacement in Severe Aortic Regurgitation

Regression of left ventricular (LV) hypertrophy with normalization of diastolic function has been reported in patients with aortic stenosis late after aortic valve replacement (AVR). The purpose of the present study was to evaluate the effect of AVR on LV function and structure in chronic aortic regurgitation early and late after AVR. Twenty-six patients were included in the present analysis. Eleven patients with severe aortic regurgitation were studied before, early (21 months) and late (89 months) after AVR through the use of LV biplane angiograms, high-fidelity pressure measurements, and LV endomyocardial biopsies. Fifteen healthy subjects were used as controls. LV systolic function was determined from biplane ejection fraction and midwall fractional shortening. LV diastolic function was calculated from the time constant of LV relaxation, peak filling rates, and myocardial stiffness constant. LV structure was assessed from muscle fiber diameter, interstitial fibrosis, and fibrous content. LV muscle mass decreased significantly by 38% early and 55% late after surgery. Ejection fraction was significantly reduced preoperatively and did not change after AVR (P=NS). LV relaxation was significantly prolonged before surgery (89+/-28 ms) but was normalized late after AVR (42+/-14 ms). Early and late peak filling rates were increased preoperatively but normalized postoperatively. Diastolic stiffness constant was increased before surgery (22+/-6 versus 9+/-3 in control subjects; P=0.0003) and remained elevated early and late after AVR (23+/-4; P=0.002). Muscle fiber diameter decreased significantly after AVR but remained increased at late follow-up. Interstitial fibrosis was increased preoperatively and increased even further early but decreased late after AVR. Fibrosis was positively linearly correlated to myocardial stiffness and inversely correlated to LV ejection fraction. Patients with aortic regurgitation show normalization of macroscopic LV hypertrophy late after AVR, although fiber hypertrophy persists. These changes in LV myocardial structure late after AVR are accompanied by a change in passive elastic properties with persistent diastolic dysfunction.

Distinct myocardial effects of beta-blocker therapy in heart failure with normal and reduced left ventricular ejection fraction

Left ventricular (LV) myocardial structure and function differ in heart failure (HF) with normal (N) and reduced (R) LV ejection fraction (EF). This difference could underlie an unequal outcome of trials with beta-blockers in heart failure with normal LVEF (HFNEF) and heart failure with reduced LVEF (HFREF) with mixed results observed in HFNEF and positive results in HFREF. To investigate whether beta-blockers have distinct myocardial effects in HFNEF and HFREF, myocardial structure, cardiomyocyte function, and myocardial protein composition were compared in HFNEF and HFREF patients without or with beta-blockers. Patients, free of coronary artery disease, were divided into beta-(HFNEF) (n = 16), beta+(HFNEF) (n = 16), beta-(HFREF) (n = 17), and beta+(HFREF) (n = 22) groups. Using LV endomyocardial biopsies, we assessed collagen volume fraction (CVF) and cardiomyocyte diameter (MyD) by histomorphometry, phosphorylation of myofilamentary proteins by ProQ-Diamond phosphostained 1D-gels, and expression of beta-adrenergic signalling and calcium handling proteins by western immunoblotting. Cardiomyocytes were also isolated from the biopsies to measure active force (F(active)), resting force (F(passive)), and calcium sensitivity (pCa(50)). Myocardial effects of beta-blocker therapy were either shared by HFNEF and HFREF, unique to HFNEF or unique to HFREF. Higher F(active), higher pCa(50), lower phosphorylation of troponin I and myosin-binding protein C, and lower beta(2) adrenergic receptor expression were shared. Higher F(passive), lower CVF, lower MyD, and lower expression of stimulatory G protein were unique to HFNEF and lower expression of inhibitory G protein was unique to HFREF. Myocardial effects unique to either HFNEF or HFREF could contribute to the dissimilar outcome of beta-blocker therapy in both HF phenotypes.

Risikostratifizierung und Therapie kardialer Amyloidosen

Cardiac amyloidoses are a heterogeneous group of cardiomyopathies that are resistant to treatment and are associated with a poor outcome. Standard heart failure treatment is usually not well tolerated and the underlying disease remains unaffected. The clinical picture is uncharacteristic. Cardiac amyloidosis is often associated with dysfunction of additional organs. Early cardiac amyloid involvement usually reveals left ventricular hypertrophy, impairment of longitudinal shortening and diastolic ventricular function. Without adequate therapy (bi-)ventricular hypertrophy will progress to severe systolic ventricular function decrease. The combination of low voltage pattern, left ventricular hypertrophy and granular sparkling is characteristic for advanced cardiac amyloid involvement. Cardiac magnetic resonance imaging and scintigraphy yield further information on the pattern and severity of cardiac involvement. In unclear cases (left ventricular) endomyocardial biopsy is necessary. Detection of early cardiac involvement and proper identification of patients at high risk for sudden cardiac death due to rapid progressive amyloidosis is still incompletely defined. Referral to specialized centers is strongly recommended.

Morphological and Quantitative Changes of the Initial Myocardial Lymphatics in Terminal Heart Failure

Terminal heart failure is associated with chronic myocardial edema, which in part is compensated by increased myocardial lymph flow. However, little is known about the impact of terminal heart failure on lymphangiogenesis. The purpose of the study was to investigate the morphological and quantitative changes of the initial myocardial lymphatics in terminal heart failure. Paraffin-embedded left ventricular endomyocardial biopsies, taken during heart transplantation from 7 heart transplant recipients (failing heart) and 8 heart transplant donors (control), were investigated by immunohistostaining and triple immunofluorescence for lymphatic endothelial markers LYVE-1, PROX-1, and VEGFR-3. The vessel density was calculated and the ratio of open versus collapsed vessels was estimated by analyzing randomly selected marked vessels. The absolute densities of lymph vessels in failing and control myocardium were not significantly different for all investigated markers. The ratio of open LYVE-1 positive lymph vessels in failing heart was significantly higher than in control (64+/-12.5 vs. 44.3+/-9.3, p<0.008). There was no difference for the ratio of open VEGFR-3 vessels between groups (69.0+/-17.5 vs. 70.7+/-17.2). Triple fluorescent immunohistostaining revealed in failing hearts LYVE-1 and PROX-1 positive open vessels, which were VEGFR-3 negative. VEGFR-3 positive, but LYVE-1 and PROX-1 negative vessels could also be seen. Myocardial initial lymphatics in patients with terminal heart failure undergo significant morphological changes in comparison to normal hearts. The ratio of open LYVE-1 vessels was higher in failing hearts by no difference in absolute densities for all markers. These findings suggest that appositional growth of initial lymphatics, rather than "de novo" genesis from pluripotent stem cells or sprouting from preexisting venous vessels, may be the predominant mechanism of lymphangiogenesis in terminal heart failure.

Abstract 4329: Myocardial Protein Kinase G Activity Differs in Heart Failure with Normal and Reduced Ejection Fraction

Concentric left ventricular (LV) remodeling and cardiomyocyte (CM) hypertrophy characterize heart failure with normal ejection fraction (HFNEF) whereas eccentric LV remodeling and low myofilamentary density characterize heart failure with reduced ejection fraction (HFREF). In rodents, low myocardial protein kinase G (PKG) activity has been implicated in the development of pathological CM hypertrophy induced by LV pressure-overload. The present study therefore compared LV remodeling, CM hypertrophy and myocardial PKG activity in patients (pts) with HFNEF (n=36) and HFREF (n=43). All pts had been admitted to hospital for worsening heart failure (NYHA 3–4) and were free of coronary artery disease. HFNEF pts had a LVEF &gt; 50% and a LV end-diastolic pressure &gt; 16 mmHg. HFREF pts had a LVEF &lt;45%. LV remodeling was measured by LV mass/volume ratio derived from biplane LV angiograms and 2D-echocardiograms. CM hypertrophy and myocardial PKG activity were assessed in LV endomyocardial biopsies by histomorphometry of CM diameter and by immunohistochemistry using antibodies against the specific PKG substrate vasodilator stimulated phosphoprotein (VASP) and against phosphorylated VASP ((P)-VASP). Myocardial PKG activity corresponded to the ratio of (P)-VASP/VASP. LV mass/volume ratio and CM diameter were significantly larger in HFNEF than in HFREF (Table ). As evident from (P)-VASP/VASP ratio, PKG activity was lower in HFNEF than in HFREF (Table ). Conclusion: HFNEF patients have lower myocardial PKG activity than HFREF patients. This low myocardial PKG activity could contribute to CM hypertrophy and concentric LV remodeling observed in HFNEF. Raising myocardial PKG activity by phosphodiesterase 5A inhibition could be useful to limit CM hypertrophy and concentric LV remodeling in HFNEF.