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Microvascular Remodeling on Endomyocardial Biopsy and Coronary Flow Reserve by Transthoracic Doppler Echocardiography in Heart Transplant Patients: A Clinical-Pathological Study
S242
The Journal of Heart and Lung Transplantation, Vol 32, No 4S, April 2013
Purpose: Hemodynamic assessment by catheterization (CATH) remains the cornerstone of cardiac performance evaluation. However, it is expensive and invasive. At the opposite end of the spectrum is BNP followed by echocardiography (ECHO). We sought to examine the correlation of BNP with ECHO and CATH parameters. Methods and Materials: Single-center, retrospective review of recipients transplanted at o21yrs of age. Joint data from BNP, ECHO, CATH, and clinical were collected at each annual surveillance biopsy/coronary angiography. Study spanned from 2007-2011 and included 71 patients. Results: The cohort median age at transplant was 3.4 yrs (IQR 0.38-10) with 55% male and 45% cardiomyopathy. There were 225 annual datasets (3.2/patient) with a median followup of 3.4 years (IQR 2.06.1). The median BNP was 54 pg/mL (IQR 26-96). BNP was correlated with HCT (R¼-0.29, po0.001) and weight (R¼-.225, p¼0.001). BNP correlated with ECHO E/E’ (R¼0.166, p¼0.028), PW systolic thickening fraction, and LV mass gm/M2.7 (R¼0.265, p 40.001). Evaluating the hemodynamic data, BNP correlated with right sided filling pressures: RAP (R¼0.341, p¼0.001), RVEDP (R¼.344, po0.001). BNP also correlated with pulmonary artery hemodynamics: diastolic (R¼0.280, po 0.001), mean (R ¼ 0.289, po0.001), PVRI (R¼0.179, p¼0.012). It also correlated with PCWP (R¼0.289, po0.001) and LVEDP (R¼0.299, p¼0.001). Patients with allograft vasculopathy had higher BNP vs. those without, 77 (IQR 36-197) vs. 53 pg/mL (IQR 25-92, p¼0.019). Using logistic regression, HCT (p¼0.006), vasculopathy (p¼0.006), RAP (p¼0.024), and LV mass (p¼0.085) were independently associated with a BNP4100 pg/mL. Conclusions: BNP as measured at annual catheterization was correlated with left and right side filling pressures and some ECHO parameters. In multivariate analysis, a BNP cutoff of 4100 pg/mL was associated with RAP, vasculopathy, HCT, and possibly with LV mass. This simple test may have utility in the monitoring of cardiac performance with the caveat that a low HCT can also affect its levels. 665 Long-Term Survival after Heart Transplantation Depends on Proper Outward Remodeling of Epicardial Arteries with Cardiac Allograft Vasculopathy M.M.H. Huibers,1 J. Kaldeway,1 A. Huisman,1 K. Timmermans,2 M.B. Leenders,3 M.E.I. Schippers,1 A. Vink,1 J.R. Lahpor,4 5 H.J.H. Kirkels,5 C. Klopping, N. de Jonge,5 R.A. de Weger.1 ¨ 1 Pathology, University Medical Center Utrecht, Utrecht, Netherlands; 2 Anesthesiology & Intensive Care Medicine, Radboud University Nijmegen Medical Center, Nijmegen, Gelderland, Netherlands; 3 Gastroenterology & Hepatology, University Medical Center Utrecht, Utrecht, Netherlands; 4Cardio Thoracic Surgery, University Medical Center Utrecht, Utrecht, Netherlands; 5Cardiology, University Medical Center Utrecht, Utrecht, Netherlands. Purpose: Long term survival after heart transplantation (HTx) is hampered by chronic rejection. The immune reaction in the coronary arteries leads to Cardiac Allograft Vasculopathy (CAV). This is the first study to describe histological changes in the artery wall in a quantitative way in relation to time post HTx. Methods and Materials: Coronary arterial cross sections from 51 HTx patients, made available after autopsy, were stained (immuno) histochemically. Histological CAV stages (H-CAV 0-3) were identified and all arteries classified accordingly. The relative surface area of the media (M), neo-intima (NI), NI-luminal layer (NI-LL), NI-smooth muscle cell layer (NI-SMC), and the lumen was measured digitally (n¼38).
Results: The H-CAV stage of the patients was higher with increasing post HTx time. Stenosis of the lumen was seen in 11 patients, predominantly scored as H-CAV 1&2. Patients that died within 2 years post HTx (n¼25) showed a significant correlation between time post HTx and all vessel layer areas. This correlation was negative for luminal area and M, and positive for the NI, NI-LL, and NI-SMC (all po0.05). A statistically significant positive correlation between luminal area and a negative correlation between NI-SMC area and time post HTx was observed (both po0.05) for patients with a longer survival time. Conclusions: Distinct patterns of CAV and arterial remodeling can be observed between patients that die within 2 years after HTx and long term HTx survivors. In the first group, CAV induces luminal stenosis without compensatory remodeling. In long term HTx survivors compensatory outward remodeling of the vessel wall seems to prevent luminal narrowing due to CAV. 666 Changes of Plasma microRNAs in Heart Transplantation Patients Do Not Reflect microRNA Changes in the Cardiac Allograft Vasculopathy Vessel Wall M.M.H. Huibers,1 H. Vroman,1 J. van Kuik,1 E. Siera-de Koning,1 N. de Jonge,2 R.A. de Weger.1 1Pathology, University Medical Center Utrecht, Utrecht, Netherlands; 2Cardiology, University Medical Center Utrecht, Utrecht, Netherlands. Purpose: A limiting factor for long term survival after heart transplantation (HTx) is cardiac allograft vasculopathy (CAV). CAV is characterized by hyperplasia of the intima in coronary arteries. Recently the importance of microRNAs (miRs) was described in multiple vascular diseases. The aim of our study was to determine whether changes in miR expression in plasma post HTx reflect changes in the coronary vessel wall of patients with CAV. Methods and Materials: Pooled plasma of HTx patients (two pools, each n¼3) was analyzed at 6 and 52 weeks after HTx and compared to healthy control plasma. CAV vessels (n¼6) were obtained after autopsy and the intimal layer was isolated by tissue laser microdissection. MiR expression in plasma and vessel wall was determined using Q-PCR arrays. Eight miRs were selected from both array analyses (plasma & vessel wall). Criteria for selection were: fold change 42x, Cq-values o32, and the miR should be detectable in all samples. These 8 miRs were validated with single Q-PCR assays in a larger cohort of patients (both plasma and CAV intima: n¼15). Results: Array analyses showed miR expression changes over time after HTx and in intimal tissue. Within the Q-PCR validation of plasma 1 miR (1/8) showed a significant up-regulation over time (miR132). The intimal tissue analysis showed 5 miRs (5/8) to have a significant up- (miR-21, -223, and -146b-5p) or down-regulation (miR886-5p and -214). Within the top 8 of plasma and intima array analysis no overlap was found in the detected miRs. Conclusions: MiRs in CAV patients do alter both in plasma and vessel wall. However, the changes in plasma do not reflect changes in the CAV vessel wall. Q-PCR validation on tissue samples confirmed more miRs than validation in plasma, indicating that miRs in tissue show a more representative pattern in array analysis. The miRs selected from the neo-intimal layer might have a role in CAV development and could be considered potential therapeutic targets in the future. 667 Microvascular Remodeling on Endomyocardial Biopsy and Coronary Flow Reserve by Transthoracic Doppler Echocardiography in Heart Transplant Patients: A Clinical-Pathological Study M. Fedrigo, F. Tona, B. Schiavon, G. Feltrin, G. Toscano, C. Castellani, S. Iliceto, G. Gerosa, G. Thiene, M. Valente, A. Angelini. Department of Cardiac, Thoracic and Vascular Science, University of Padua, Padua, Italy. Purpose: Cardiac allograft vasculopathy (CAV) is the main limiting factor of long-term survival after heart transplantation (HT). CAV
Abstracts affects both epicardial coronary vessels and microvasculature. Coronary flow reserve (CFR) by transthoracic Doppler echocardiography (TDE) is an independent predictor of death in HT patients. The aim of the study was to correlate the microvascular remodeling, as assessed by endomyocardial biopsy (EMB), with CFR by TDE in HT patients with normal coronary angiograms. Methods and Materials: We studied 28 consecutive HT patients without angiographic CAV (25 male, aged 54⫾10 years at HT, time from HT 7.5⫾5 years). Coronary flow velocity in the left anterior descending coronary artery was detected by TDE at rest and during adenosine infusion. CFR was the ratio of hyperaemic diastolic flow velocity (DFV) to resting DFV. A CFRr2.5 was considered abnormal. In the 1st year post-HT EMBs, myocytes diameter, fibrosis percentage, capillary density and microvascular remodeling (vessel media area/total vessel area ratio (%)) were evaluated by digital morphometry. Results: Microvascular remodeling was higher in patients with CFRr2.5 (39.2% 11/28 pts) compared with patients with CFR42.5 (72.3⫾8 vs 65.2⫾4.7 %, po0.007). Myocytes diameter, fibrosis percentage and capillary density were similar in the two groups. Male gender prevalence, gender donor-recipient mismatch, age at HT, time from HT, hypertension and diabetes prevalence, and other CAV risk factors were comparable in the two groups. Conclusions: Our results suggest that coronary microvascular remodeling at EMBs may be the main predictor of abnormal CFR in HT patients. CFR by TDE may be a useful noninvasive tool in the evaluation of microvascular dysfunction/damage in HT. Further studies are warranted. 668 Fibroblast Growth Factor 23 Is a Promissing Marker of Myocardial Pathology V. Polyakova,1 M. Richter,1 T. Kubin,2 T. Walther.1 1Heart Surgery, Kerckhoff-Clinic, Bad Nauheim, Germany; 2Cardiac Development and Remodelling, Max-Planck-Institute, Bad Nauheim, Germany. Purpose: Fibroblast growth factor 23 (FGF23) is a main bone hormone responsible for the phosphate homeostasis. Moreover it is a good biomarker of chronic kidney disease. The only experimental report concerning FGF23 in the heart displayed its toxic effect on mice myocardium resulted in hypertrophy after intravenous injection of FGF23. However, there is no notification of FGF23 heart expression. Currently we observed that isolated neonatal as well as adult cardiomyocytes (CM) are able to produce FGF23 in stress conditions, but data in vivo are missing. Methods and Materials: For this purpose we studied heart explants of 10 patients suffering from ischemic cardiomyopathy (ICM), 14 patients with dilated cardiomyopathy (DCM) and 12 patients with myocarditis (Myo). 2 healthy donor hearts (which were not used for transplantation) served as controls. In addition we used animal models: wild-type mice with myocardial infarction (WT-MI) induced by LAD ligation and transgenic mice with a cardiac restricted overexpression of monocyte chemotactic protein (MCP1) as a model of chronic inflammation. Results: We firstly detected FGF23 positive CM in vivo. They were localized in diseased myocardium as isolated cells or in groups showing week or marked cross-striation. The number of FGF23 expressing CM was increased 14.8 fold in patients with ICM, 12.3 fold in DCM patients and 13.5 fold in patients with Myo in comparison with controls (po0.001, po0.01, po0.001 correspondingly). The ischemic stress in WT-MI mice induced 4.7 fold upregulation of FGF23 positive CM (po0.01). MCP1 mice demonstrated 6.5 fold higher amount of FGF23 than in control group (po0.001). In addition we detected an increased expression of FGFR1c (the main receptor of FGF23) in interstitial cells surrounding FGF23 positive CM. Conclusions: For the first time we demonstrated FGF 23 expression in myocardium of patients with heart failure and in mice models. An increased activity of FGF23 and its receptor FGFR1c in stress conditions characterizes the FGF23 as a promising diagnostic and prognostic biomarker of cardiac pathology.
S243 669 A Specific Mechanism for Late Loss of Cardiac Allograft: The Antibody Mediated Rejection (AMR) as a Major Factor of Cardiac Allograft Vasculopathy (CAV) C. Toquet,1 A. Loupy,2 P. Rouvier,3 S. Varnous,3 A. Cazes,2 M. Tible,2 T. Beuscart,2 X. Jouven,2 P. Bruneval,2 J.-P. Duong Van Huyen.2 1CHU Nantes, Nantes, France; 2University Paris Descartes, Paris, France; 3 Hopital La Pitie, Paris, France. Purpose: In heart transplantation CAV is the main cause of late cardiac graft loss. Its pathogenesis is unclear. The rational of this study was to investigate explanted failing grafts in the light of new concepts in rejection mechanisms, namely AMR. Methods and Materials: This retrospective multicenter study collected 40 explanted cardiac grafts failing more than 1 year post-transplantation. The vasculature of the grafts was assessed from epicardial coronary arteries to myocardial microcirculation. The lesions were classified as follows: Classical coronary atherosclerosis; CAV with concentric intimal hyperplasia and media atrophy (arteriosclerosis); Vascular inflammation in the arteries (endothelitis) ⫾ media infiltration and in the microcirculation intracapillary leukocyte stasis. The lesions were scored and submitted to clustering analysis. Immunohistochemistry was performed for C4d and CD68-macrophages. The protocol EMB performed before the graft removal were assessed again by histology and immunohistochemistry. DSA were assessed in stored sera using Luminex SA technique. Results: A pure classical coronary atherosclerosis pattern was observed in 15.5%. A pure pattern of CAV was present in 40% and a mixed pattern associating CAV and atherosclerosis features in 32.5%. The coronary arteries were devoid of significant lesions in 10%. Interestingly the CAV pure and mixed patterns were significantly associated with vascular inflammation within the arteries and/or the microcirculation with C4d deposits and macrophages in 14 out of 29 (48%) explanted grafts versus nil (0/7) in pure atherosclerosis pattern (po0.002). Furthermore they were associated with previous AMR episodes on EMB (52%) as shown by intravascular macrophages and/ or C4d deposits and by positive DSA (66%) versus nil in pure atherosclerosis pattern (po0.05 and p ¼ 0.05, respectively). Conclusions: In failing cardiac grafts CAV lesions are associated with makers of AMR. CAV should be the consequence in coronary arteries of an ongoing AMR process. 670 C4d Immunostaining Is an Independent Predictor of Graft Dysfunction, Cardiac Allograft Vasculopathy and Death in Heart Transplant Recipients A. Luk,1 C. Alba,1 J. Butany,2 K. Tinckam,2 D.H. Delgado,1 H.J. Ross.1 1 Division of Cardiology, Department of Medicine, University Health Network, Toronto, ON, Canada; 2Departments of Laboratory Medicine and Pathobiology and Medicine, University of Toronto, Toronto, ON, Canada. Purpose: Antibody-mediated rejection (AMR) occurs in 10-20% of patients post heart transplant (HT), and is associated with hemodynamic instability, rejection, coronary allograft vasculopathy (CAV) and death. C4d immunostaining, histology, allograft dysfunction and presence of donor specific antibodies are used in the diagnosis of AMR. This study aimed to determine if C4d staining has prognostic significance in patients post-HT. Methods and Materials: Consecutive patients receiving an endomyocardial biopsy between July 2007-June 2008 were selected. Left ventricular function, angiography, episodes of antibody and cellular rejection (CMR) and death were noted. Subsequent biopsies after enrollment were reviewed. Immunostaining for C4d deposition was performed on paraffin-embedded tissue using an anti human antibody (rabbit polyclonal, [ALPCO, Salem, NH]), and were graded from 0-3. We used Cox proportional models and recurrent events analysis to evaluate the association between C4d staining (time-varying covariate) and mortality, graft dysfunction, CAV and episodes of Z2R-CMR. Results: We analyzed 2525 biopsy specimens from 217 patients. The average age of patients were 45 ⫾ 13 years at transplant (70% were males). During a mean follow up of 4.5 ⫾ 2 years, 35 died, 49 patients
The Journal of Heart and Lung Transplantation, Vol 32, No 4S, April 2013
Purpose: Hemodynamic assessment by catheterization (CATH) remains the cornerstone of cardiac performance evaluation. However, it is expensive and invasive. At the opposite end of the spectrum is BNP followed by echocardiography (ECHO). We sought to examine the correlation of BNP with ECHO and CATH parameters. Methods and Materials: Single-center, retrospective review of recipients transplanted at o21yrs of age. Joint data from BNP, ECHO, CATH, and clinical were collected at each annual surveillance biopsy/coronary angiography. Study spanned from 2007-2011 and included 71 patients. Results: The cohort median age at transplant was 3.4 yrs (IQR 0.38-10) with 55% male and 45% cardiomyopathy. There were 225 annual datasets (3.2/patient) with a median followup of 3.4 years (IQR 2.06.1). The median BNP was 54 pg/mL (IQR 26-96). BNP was correlated with HCT (R¼-0.29, po0.001) and weight (R¼-.225, p¼0.001). BNP correlated with ECHO E/E’ (R¼0.166, p¼0.028), PW systolic thickening fraction, and LV mass gm/M2.7 (R¼0.265, p 40.001). Evaluating the hemodynamic data, BNP correlated with right sided filling pressures: RAP (R¼0.341, p¼0.001), RVEDP (R¼.344, po0.001). BNP also correlated with pulmonary artery hemodynamics: diastolic (R¼0.280, po 0.001), mean (R ¼ 0.289, po0.001), PVRI (R¼0.179, p¼0.012). It also correlated with PCWP (R¼0.289, po0.001) and LVEDP (R¼0.299, p¼0.001). Patients with allograft vasculopathy had higher BNP vs. those without, 77 (IQR 36-197) vs. 53 pg/mL (IQR 25-92, p¼0.019). Using logistic regression, HCT (p¼0.006), vasculopathy (p¼0.006), RAP (p¼0.024), and LV mass (p¼0.085) were independently associated with a BNP4100 pg/mL. Conclusions: BNP as measured at annual catheterization was correlated with left and right side filling pressures and some ECHO parameters. In multivariate analysis, a BNP cutoff of 4100 pg/mL was associated with RAP, vasculopathy, HCT, and possibly with LV mass. This simple test may have utility in the monitoring of cardiac performance with the caveat that a low HCT can also affect its levels. 665 Long-Term Survival after Heart Transplantation Depends on Proper Outward Remodeling of Epicardial Arteries with Cardiac Allograft Vasculopathy M.M.H. Huibers,1 J. Kaldeway,1 A. Huisman,1 K. Timmermans,2 M.B. Leenders,3 M.E.I. Schippers,1 A. Vink,1 J.R. Lahpor,4 5 H.J.H. Kirkels,5 C. Klopping, N. de Jonge,5 R.A. de Weger.1 ¨ 1 Pathology, University Medical Center Utrecht, Utrecht, Netherlands; 2 Anesthesiology & Intensive Care Medicine, Radboud University Nijmegen Medical Center, Nijmegen, Gelderland, Netherlands; 3 Gastroenterology & Hepatology, University Medical Center Utrecht, Utrecht, Netherlands; 4Cardio Thoracic Surgery, University Medical Center Utrecht, Utrecht, Netherlands; 5Cardiology, University Medical Center Utrecht, Utrecht, Netherlands. Purpose: Long term survival after heart transplantation (HTx) is hampered by chronic rejection. The immune reaction in the coronary arteries leads to Cardiac Allograft Vasculopathy (CAV). This is the first study to describe histological changes in the artery wall in a quantitative way in relation to time post HTx. Methods and Materials: Coronary arterial cross sections from 51 HTx patients, made available after autopsy, were stained (immuno) histochemically. Histological CAV stages (H-CAV 0-3) were identified and all arteries classified accordingly. The relative surface area of the media (M), neo-intima (NI), NI-luminal layer (NI-LL), NI-smooth muscle cell layer (NI-SMC), and the lumen was measured digitally (n¼38).
Results: The H-CAV stage of the patients was higher with increasing post HTx time. Stenosis of the lumen was seen in 11 patients, predominantly scored as H-CAV 1&2. Patients that died within 2 years post HTx (n¼25) showed a significant correlation between time post HTx and all vessel layer areas. This correlation was negative for luminal area and M, and positive for the NI, NI-LL, and NI-SMC (all po0.05). A statistically significant positive correlation between luminal area and a negative correlation between NI-SMC area and time post HTx was observed (both po0.05) for patients with a longer survival time. Conclusions: Distinct patterns of CAV and arterial remodeling can be observed between patients that die within 2 years after HTx and long term HTx survivors. In the first group, CAV induces luminal stenosis without compensatory remodeling. In long term HTx survivors compensatory outward remodeling of the vessel wall seems to prevent luminal narrowing due to CAV. 666 Changes of Plasma microRNAs in Heart Transplantation Patients Do Not Reflect microRNA Changes in the Cardiac Allograft Vasculopathy Vessel Wall M.M.H. Huibers,1 H. Vroman,1 J. van Kuik,1 E. Siera-de Koning,1 N. de Jonge,2 R.A. de Weger.1 1Pathology, University Medical Center Utrecht, Utrecht, Netherlands; 2Cardiology, University Medical Center Utrecht, Utrecht, Netherlands. Purpose: A limiting factor for long term survival after heart transplantation (HTx) is cardiac allograft vasculopathy (CAV). CAV is characterized by hyperplasia of the intima in coronary arteries. Recently the importance of microRNAs (miRs) was described in multiple vascular diseases. The aim of our study was to determine whether changes in miR expression in plasma post HTx reflect changes in the coronary vessel wall of patients with CAV. Methods and Materials: Pooled plasma of HTx patients (two pools, each n¼3) was analyzed at 6 and 52 weeks after HTx and compared to healthy control plasma. CAV vessels (n¼6) were obtained after autopsy and the intimal layer was isolated by tissue laser microdissection. MiR expression in plasma and vessel wall was determined using Q-PCR arrays. Eight miRs were selected from both array analyses (plasma & vessel wall). Criteria for selection were: fold change 42x, Cq-values o32, and the miR should be detectable in all samples. These 8 miRs were validated with single Q-PCR assays in a larger cohort of patients (both plasma and CAV intima: n¼15). Results: Array analyses showed miR expression changes over time after HTx and in intimal tissue. Within the Q-PCR validation of plasma 1 miR (1/8) showed a significant up-regulation over time (miR132). The intimal tissue analysis showed 5 miRs (5/8) to have a significant up- (miR-21, -223, and -146b-5p) or down-regulation (miR886-5p and -214). Within the top 8 of plasma and intima array analysis no overlap was found in the detected miRs. Conclusions: MiRs in CAV patients do alter both in plasma and vessel wall. However, the changes in plasma do not reflect changes in the CAV vessel wall. Q-PCR validation on tissue samples confirmed more miRs than validation in plasma, indicating that miRs in tissue show a more representative pattern in array analysis. The miRs selected from the neo-intimal layer might have a role in CAV development and could be considered potential therapeutic targets in the future. 667 Microvascular Remodeling on Endomyocardial Biopsy and Coronary Flow Reserve by Transthoracic Doppler Echocardiography in Heart Transplant Patients: A Clinical-Pathological Study M. Fedrigo, F. Tona, B. Schiavon, G. Feltrin, G. Toscano, C. Castellani, S. Iliceto, G. Gerosa, G. Thiene, M. Valente, A. Angelini. Department of Cardiac, Thoracic and Vascular Science, University of Padua, Padua, Italy. Purpose: Cardiac allograft vasculopathy (CAV) is the main limiting factor of long-term survival after heart transplantation (HT). CAV
Abstracts affects both epicardial coronary vessels and microvasculature. Coronary flow reserve (CFR) by transthoracic Doppler echocardiography (TDE) is an independent predictor of death in HT patients. The aim of the study was to correlate the microvascular remodeling, as assessed by endomyocardial biopsy (EMB), with CFR by TDE in HT patients with normal coronary angiograms. Methods and Materials: We studied 28 consecutive HT patients without angiographic CAV (25 male, aged 54⫾10 years at HT, time from HT 7.5⫾5 years). Coronary flow velocity in the left anterior descending coronary artery was detected by TDE at rest and during adenosine infusion. CFR was the ratio of hyperaemic diastolic flow velocity (DFV) to resting DFV. A CFRr2.5 was considered abnormal. In the 1st year post-HT EMBs, myocytes diameter, fibrosis percentage, capillary density and microvascular remodeling (vessel media area/total vessel area ratio (%)) were evaluated by digital morphometry. Results: Microvascular remodeling was higher in patients with CFRr2.5 (39.2% 11/28 pts) compared with patients with CFR42.5 (72.3⫾8 vs 65.2⫾4.7 %, po0.007). Myocytes diameter, fibrosis percentage and capillary density were similar in the two groups. Male gender prevalence, gender donor-recipient mismatch, age at HT, time from HT, hypertension and diabetes prevalence, and other CAV risk factors were comparable in the two groups. Conclusions: Our results suggest that coronary microvascular remodeling at EMBs may be the main predictor of abnormal CFR in HT patients. CFR by TDE may be a useful noninvasive tool in the evaluation of microvascular dysfunction/damage in HT. Further studies are warranted. 668 Fibroblast Growth Factor 23 Is a Promissing Marker of Myocardial Pathology V. Polyakova,1 M. Richter,1 T. Kubin,2 T. Walther.1 1Heart Surgery, Kerckhoff-Clinic, Bad Nauheim, Germany; 2Cardiac Development and Remodelling, Max-Planck-Institute, Bad Nauheim, Germany. Purpose: Fibroblast growth factor 23 (FGF23) is a main bone hormone responsible for the phosphate homeostasis. Moreover it is a good biomarker of chronic kidney disease. The only experimental report concerning FGF23 in the heart displayed its toxic effect on mice myocardium resulted in hypertrophy after intravenous injection of FGF23. However, there is no notification of FGF23 heart expression. Currently we observed that isolated neonatal as well as adult cardiomyocytes (CM) are able to produce FGF23 in stress conditions, but data in vivo are missing. Methods and Materials: For this purpose we studied heart explants of 10 patients suffering from ischemic cardiomyopathy (ICM), 14 patients with dilated cardiomyopathy (DCM) and 12 patients with myocarditis (Myo). 2 healthy donor hearts (which were not used for transplantation) served as controls. In addition we used animal models: wild-type mice with myocardial infarction (WT-MI) induced by LAD ligation and transgenic mice with a cardiac restricted overexpression of monocyte chemotactic protein (MCP1) as a model of chronic inflammation. Results: We firstly detected FGF23 positive CM in vivo. They were localized in diseased myocardium as isolated cells or in groups showing week or marked cross-striation. The number of FGF23 expressing CM was increased 14.8 fold in patients with ICM, 12.3 fold in DCM patients and 13.5 fold in patients with Myo in comparison with controls (po0.001, po0.01, po0.001 correspondingly). The ischemic stress in WT-MI mice induced 4.7 fold upregulation of FGF23 positive CM (po0.01). MCP1 mice demonstrated 6.5 fold higher amount of FGF23 than in control group (po0.001). In addition we detected an increased expression of FGFR1c (the main receptor of FGF23) in interstitial cells surrounding FGF23 positive CM. Conclusions: For the first time we demonstrated FGF 23 expression in myocardium of patients with heart failure and in mice models. An increased activity of FGF23 and its receptor FGFR1c in stress conditions characterizes the FGF23 as a promising diagnostic and prognostic biomarker of cardiac pathology.
S243 669 A Specific Mechanism for Late Loss of Cardiac Allograft: The Antibody Mediated Rejection (AMR) as a Major Factor of Cardiac Allograft Vasculopathy (CAV) C. Toquet,1 A. Loupy,2 P. Rouvier,3 S. Varnous,3 A. Cazes,2 M. Tible,2 T. Beuscart,2 X. Jouven,2 P. Bruneval,2 J.-P. Duong Van Huyen.2 1CHU Nantes, Nantes, France; 2University Paris Descartes, Paris, France; 3 Hopital La Pitie, Paris, France. Purpose: In heart transplantation CAV is the main cause of late cardiac graft loss. Its pathogenesis is unclear. The rational of this study was to investigate explanted failing grafts in the light of new concepts in rejection mechanisms, namely AMR. Methods and Materials: This retrospective multicenter study collected 40 explanted cardiac grafts failing more than 1 year post-transplantation. The vasculature of the grafts was assessed from epicardial coronary arteries to myocardial microcirculation. The lesions were classified as follows: Classical coronary atherosclerosis; CAV with concentric intimal hyperplasia and media atrophy (arteriosclerosis); Vascular inflammation in the arteries (endothelitis) ⫾ media infiltration and in the microcirculation intracapillary leukocyte stasis. The lesions were scored and submitted to clustering analysis. Immunohistochemistry was performed for C4d and CD68-macrophages. The protocol EMB performed before the graft removal were assessed again by histology and immunohistochemistry. DSA were assessed in stored sera using Luminex SA technique. Results: A pure classical coronary atherosclerosis pattern was observed in 15.5%. A pure pattern of CAV was present in 40% and a mixed pattern associating CAV and atherosclerosis features in 32.5%. The coronary arteries were devoid of significant lesions in 10%. Interestingly the CAV pure and mixed patterns were significantly associated with vascular inflammation within the arteries and/or the microcirculation with C4d deposits and macrophages in 14 out of 29 (48%) explanted grafts versus nil (0/7) in pure atherosclerosis pattern (po0.002). Furthermore they were associated with previous AMR episodes on EMB (52%) as shown by intravascular macrophages and/ or C4d deposits and by positive DSA (66%) versus nil in pure atherosclerosis pattern (po0.05 and p ¼ 0.05, respectively). Conclusions: In failing cardiac grafts CAV lesions are associated with makers of AMR. CAV should be the consequence in coronary arteries of an ongoing AMR process. 670 C4d Immunostaining Is an Independent Predictor of Graft Dysfunction, Cardiac Allograft Vasculopathy and Death in Heart Transplant Recipients A. Luk,1 C. Alba,1 J. Butany,2 K. Tinckam,2 D.H. Delgado,1 H.J. Ross.1 1 Division of Cardiology, Department of Medicine, University Health Network, Toronto, ON, Canada; 2Departments of Laboratory Medicine and Pathobiology and Medicine, University of Toronto, Toronto, ON, Canada. Purpose: Antibody-mediated rejection (AMR) occurs in 10-20% of patients post heart transplant (HT), and is associated with hemodynamic instability, rejection, coronary allograft vasculopathy (CAV) and death. C4d immunostaining, histology, allograft dysfunction and presence of donor specific antibodies are used in the diagnosis of AMR. This study aimed to determine if C4d staining has prognostic significance in patients post-HT. Methods and Materials: Consecutive patients receiving an endomyocardial biopsy between July 2007-June 2008 were selected. Left ventricular function, angiography, episodes of antibody and cellular rejection (CMR) and death were noted. Subsequent biopsies after enrollment were reviewed. Immunostaining for C4d deposition was performed on paraffin-embedded tissue using an anti human antibody (rabbit polyclonal, [ALPCO, Salem, NH]), and were graded from 0-3. We used Cox proportional models and recurrent events analysis to evaluate the association between C4d staining (time-varying covariate) and mortality, graft dysfunction, CAV and episodes of Z2R-CMR. Results: We analyzed 2525 biopsy specimens from 217 patients. The average age of patients were 45 ⫾ 13 years at transplant (70% were males). During a mean follow up of 4.5 ⫾ 2 years, 35 died, 49 patients