Cardiac Specimens Research Articles

Increased myocardial prevalence of C-reactive protein in human coronary heart disease: direct effects on microvessel density and endothelial cell survival

Elevated plasma C-reactive protein (CRP) is a biomarker of cardiovascular diseases (CVDs), but its potential roles as a participant of the disease process are not well defined. Although early endothelial cell injury and dysfunction are recognized events in CVD, the initiating events are not well established. Here we investigated the local myocardial CRP levels and cardiac microvessel densities in control and CVD tissue samples. Using in vitro methodologies, we investigated the direct effects of CRP on human endothelial cells. Cardiac specimens were collected at autopsy within 4 h of death and were classified as normal controls or documented evidence of CVD. The regional prevalence of CRP and the cardiac microvessels (<40 μm) were investigated using immunohistochemistry. For in vitro experiments, human umbilical vein endothelial cells were incubated with CRP. Intracellular oxidant levels were assessed using 2',7'-dichlorofluorescein diacetate fluorescence microscopy, and cell survival was concurrently determined. Effects of chemical antioxidants on endothelial cell survival were also tested. Myocardial CRP levels were elevated in CVD specimens. This was associated with reduced cardiac microvessels, and this rarefaction was inversely correlated to adjacent myocardial CRP prevalence. CRP caused concentration-dependent increases in oxidant production and cell apoptosis. These findings provide evidence supporting myocardial CRP as a locally produced inflammatory marker and as a potential participant in endothelial toxicity and microvascular rarefaction.

Congestive heart failure in acromegaly: A review of 6 cases.

Background:Though cardiac involvement is common in acromegaly, overt congestive heart failure is uncommon.Materials and Methods:This is retrospective analysis of hospital record between 1996 and 2007. We analyzed records of 150 consecutive patients with acromegaly. We included the patients with acromegaly those who had overt congestive heart failure either at presentation or during the course of illness for the present analysis. The diagnosis of acromegaly and congestive cardiac failure were based on standard criteria.Results:Out of 150 patients with acromegaly, 6 patients had overt CHF (4.0%), of which 4 presented with the features of CHF and 2 developed during the course of illness. Three patients had hypertension and 1 had diabetes. Baseline echocardiography showed severe biventricular dysfunction and global hypokinesia in all. Angiography showed dilated hypokinetic left ventricle with normal coronaries in 3, it was confirmed at autopsy in 1. Three underwent trans-sphenoidal surgery, 1 received somatostatin analogue as primary treatment modality. Normalization of growth hormone and IGF-1 led to improvement in cardiac function in 1, 1 patient lost to follow up, and 4 died during the course of illness. In 1 patient, autopsy was performed and cardiac specimen revealed normal coronaries, concentric ventricular hypertrophy, and dilatation with myofibrolysis and interfascicular fibrosis.Conclusion:Prevalence of overt CHF is 4% in present series. Overt CHF carries poor prognosis and hence, this complication should be recognized at earliest, and medical management to normalized cardiac function should be given utmost priority.

Sarcoidosis does not belong to or overlap with immunoglobulin G4–related diseases based on an assessment of serum immunoglobulin G4 levels in cardiac and noncardiac sarcoidosis

Although sarcoidosis may exhibit histopathologic features similar to those of a newly emerging clinical entity, immunoglobulin G4-related sclerosing disease, sarcoidosis is currently not considered to be associated with immunoglobulin G4-related immunoinflammation. Not many studies on this association have been reported. We investigated serum immunoglobulin G4 levels among patients with sarcoidosis with or without cardiac involvement (cardiac sarcoidosis and non-cardiac sarcoidosis patients). The mean serum immunoglobulin G4 level among the 65 patients with sarcoidosis was 56.8 ± 43.0 mg/dL, which did not significantly differ between patients with cardiac sarcoidosis (54 ± 48 mg/dL, n = 12) and patients without cardiac sarcoidosis (58 ± 42 mg/dL; n = 53). Serum level of soluble interleukin 2 receptor, a potent marker that may reflect sarcoidosis activity, was elevated in cardiac sarcoidosis (910 ± 683 U/L) and noncardiac sarcoidosis (689 ± 399 U/L) but did not significantly differ between the groups. Immunohistochemistry of cardiac or lymph node specimens from patients with cardiac sarcoidosis showed only sparse or no infiltration of immunoglobulin G4-positive lymphocytes, in contrast to the moderate to severe infiltration of CD68-positive macrophages and CD45-positive lymphocytes. Although the number of study subjects was small, these findings collectively suggest that regardless of the presence or absence of cardiac involvement, sarcoidosis does not belong to or overlap with immunoglobulin G4-related sclerosing disease.

Methods to Prepare Perfusion Fixed Cardiac Specimens for Multimodal Imaging: The Use of Formalin and Agar Gels

Recent advances in cardiac imaging have resulted in a growing understanding of both the form and function of the heart in vivo. Currently, the primary modalities for cardiac imaging are (1) two-dimensional cardiac ultrasound or echocardiography, (2) computed tomography (CT), and (3) magnetic resonance imaging (MRI). Yet, high resolution imaging with these modalities can be complicated by motion artifacts and long acquisition times resulting in most of the high resolution anatomical cardiac imaging protocols being reserved for ex vivo studies. Our laboratory has had the privilege to obtain fresh human heart specimens for educational and research purposes. These specimens have been perfusion fixed in 10% buffered formalin, by attaching cannulas to the great vessels, so to create a pressure head of approximately 50 mm Hg. The hearts were then suspended in containers and positioned in anatomically correct orientations before being embedded in 0.7% agar gel, at approximately 45 °C. The cooled specimens were then scanned using the aforementioned clinical imaging modalities (2D and 3D echocardiography, CT, and 3T MRI). The stability of the embedded specimen, the physical properties of the gel, and the lack of motion artifacts allows for the acquisition of extremely high resolution images. These images have subsequently been used in the analysis of cardiac anatomies for a variety of pathologic investigations, not possible with current clinical imaging protocols, and/or for high resolution diffusion tensor MR imaging studies (e.g., of fiber orientations in heart failure in swine ventricles). Future work will include investigations as to whether this gelling approach could be used to prepare other organ specimens for such imaging.

Clinicopathological features of senile systemic amyloidosis: an ante- and post-mortem study

Senile systemic amyloidosis is a common age-related amyloidosis that involves accumulation of wild-type transthyretin, with cardiac dysfunction being a predominant result. The importance of obtaining an accurate diagnosis of senile systemic amyloidosis has been increasingly recognized, so that novel treatments are being developed. However, the clinicopathological features of senile systemic amyloidosis remain to be completely understood. Here, we evaluated cardiac specimens from 181 consecutive post-mortem cases older than 40 years, including 6 cases of senile systemic amyloidosis, and 5 cases of familial amyloidotic polyneuropathy, which is a hereditary systemic amyloidosis caused by mutant forms of transthyretin. Furthermore, we studied ante-mortem clinicopathological findings of 11 senile systemic amyloidosis cases, in which 9 cases underwent gastrointestinal tract biopsy and/or subcutaneous tissue biopsy, at Kumamoto University Hospital. Of the autopsied cases of elderly Japanese (older than 80 years), 12% had senile systemic amyloidosis, with the percentage increasing with age. The occurrence of senile systemic amyloidosis in elderly Japanese patients was lower than that in previous reports, which suggests that a genetic background and/or environmental factor(s) may have important roles in the occurrence of senile systemic amyloidosis. Transthyretin amyloid deposits in familial amyloidotic polyneuropathy cases developed mainly in the pericardium and the surrounding muscle fascicles, whereas in cases with senile systemic amyloidosis the transthyretin amyloid deposits had a patchy plaque-like shape and developed mainly inside the ventricular wall. Biopsies from senile systemic amyloidosis patients evidenced amyloid deposits in 44% (4/9) of gastrointestinal tract and subcutaneous tissue samples combined. As myocardial biopsy may be dangerous for elderly people, the use of a combination of gastrointestinal tract and subcutaneous tissue biopsies may make diagnosis of senile systemic amyloidosis easier.

Pathology of the Aortic Arch in Hypoplastic Left Heart Syndrome: Surgical Implications

Aortic arch reconstruction plays an important role in the success of the Norwood procedure (NP) for hypoplastic left heart syndrome (HLHS). This study investigated the cardiac specimens to determine the etiology of distal aortic arch obstruction after the NP for HLHS and to locate coarctation of the aorta in HLHS untreated by surgery. This study examined 17 cardiac specimens: 9 that had NP and 8 not treated by surgery. The findings after NP showed frequent failure to resect the coarctation segment completely and failure to extend the augmentation patch into the descending aorta. Five (62.5%) of the eight hearts not treated by surgery had significant periductal coarctation of the aorta. After NP for nine patients, three (33%) had residual coarctation of the aorta. To minimize the risk of recurrent or persistent aortic arch obstruction after NP and to improve the long- and short-term outcome, the ductal tissue and the coarctation segment encircling the aortic lumen should be resected. The distal wall incision should be extended at least 5 mm beyond the distal aspect of the ductal tissue. These steps could avoid major aortic arch obstruction, promote growth of the native aortic tissue, and avoid ventricular dysfunction.

Case of Paracoccus yeei infection documented in a transplanted heart

After a successful cardiac transplantation, routine endomyocardial biopsies showed severe infiltrates comparable with myocarditis. Polymerase chain reaction analysis of native myocardial samples revealed infection with Paracoccus yeei, and the clinical condition of the patient deteriorated. After administration of ciprofloxacin, his clinical condition improved, and further biopsies showed no infiltrates in the cardiac specimens. To our knowledge this is the first documented case of P. yeei infection in a heart transplant patient.

Magnetic resonance of hearts in a jar: breathing new life into old pathological specimens

Specimens of the normal and congenitally abnormal heart have been long preserved, collected, and studied. It is increasingly difficult to add to such pathological collections. These museum pieces are often inaccessible for teaching purposes. Magnetic resonance imaging of old pathological specimens could produce high-resolution unalterable datasets that could be processed to create three-dimensional reconstructions using inexpensive systems that could be used by untrained individuals. To our knowledge, the concept of "Virtual Autopsy" has not been applied to cardiac specimens of museum collections. To determine optimal sequences and assure specimen safety, five different pulse sequences designed to create three-dimensional datasets were tried on a uterus specimen suspended in a fluid-filled glass container, using a 1.5 Tesla scanner with an eight-channel phased-array coil. Having found the best sequences and established specimen integrity, we scanned six historical heart specimens in their original fluid-filled glass containers. The datasets were processed on a laptop with a DICOM viewer available as freeware. All specimens were successfully scanned. The best image quality was obtained by using a three-dimensional FSPGR and the BRAVO pulse sequences. High-resolution three-dimensional and multi-planar image processing was possible for all datasets. Detailed examination of the specimens could be easily performed. Pathological specimens can successfully be scanned in minutes resulting in unalterable and portable high-resolution three-dimensional datasets that can be processed by using inexpensive readily available software. The final cardiac reconstructions can be widely shared for educational and scientific purposes and ensure a lasting access to pathological specimens.

The cardiac veins in congenitally corrected transposition of the great arteries: Delivery options for cardiac devices

Device implantation is sometimes required in congenitally corrected transposition of the great arteries (ccTGA) because of morphologic right ventricular (RV) dysfunction and complete heart block. The anatomical course of the veins remains unknown, despite well-described coronary arterial anatomy. Knowledge of the venous anatomy may facilitate planning of percutaneous cardiac procedures in these patients. This study sought to characterize the venous anatomy in ccTGA. Pathologic cardiac specimens from patients with ccTGA were identified from the Mayo Clinic pathology database. Coronary sinus (CS) anatomy and distances from the CS ostium to the major cardiac veins were evaluated. Thebesian veins with ostial openings >1 mm, epicardial veins, and venous collaterals were also quantified. There were 56 hearts with a diagnosis of ccTGA identified. The CS was unidentifiable in 5 hearts due to slicing, and assessment of the Thebesian veins was not possible in 16. Seven hearts had an abnormal CS, 2 of which had atretic ostia and the other 5 of which either had an abnormal ostial location or multiple ostia. There were 28 hearts with at least 1 Thebesian vein with an ostial opening >1 mm. All 12 hearts with unidentifiable Thebesian veins had venous collaterals from the right ventricle (RV) to the major cardiac veins. Epicardial veins extended to the proximal, middle, and distal thirds of the RV in 71%, 23%, and 6%, respectively. In ccTGA, the ventricular venous anatomy is abnormal and follows the morphologic RV. However, large interventricular and Thebesian veins may offer options for percutaneous lead or catheter placement when approaching the systemic RV.

Brugada-like EKG pattern and myocardial effects in a chronic propofol abuser

Introduction. Cases of death are reported due to medical use of propofol, whereas deaths due to recreational purpose are unusual. Case report. A 26-year-old Caucasian man, physician trainee in anesthesiology, was referred to an intensive care unit. The man was found unconscious in his bed with a butterfly-needle canalized into the vein of the left forearm and connected to an empty syringe. Transferred to the local hospital, the patient was monitored, and EKG showed typical Brugada features in V1–V3. Profound hypotension and metabolic acidosis were registered. Half an hour after admission, the patient developed prolonged QT interval, idioventricular rhythm, and ventricular fibrillation. Strong positive reaction for tumor necrosis factor alpha in cardiac myocytes and a diffuse apoptotic process in the heart specimens were observed. The multiple needle marks on the hands and forearms, and the propofol concentration in the hair examined (0.73 µg/g), led us to believe that the young man was a long-term propofol abuser. Discussion. Development of the EKG pattern of ST-segment elevation in leads V1–V3 may be the first indicator of electrical instability and high risk for imminent sudden death. Whether this finding applies to other patients poisoned with propofol is unclear, but the association of sudden death and the acquired EKG pattern has been observed in other disease states. Conclusion. This article describes a fatal propofol-related death case because of recreational purpose; the EKG pattern, the cardiac morphology, and the expression of tumor necrosis factor alpha and apoptosis in cardiac tissue specimens are discussed to elucidate the mechanism of death.

Multidetector-row computed tomographic evaluation of myocardial perfusion in reperfused chronic myocardial infarction: value of color-coded perfusion map in a porcine model

We aimed to develop color-coded CT perfusion maps (CPM) of infarcted myocardium and assess the utility of CPM in evaluating ischemic heart disease on a cardiac multi-detector CT (MDCT) in a porcine reperfused-myocardial-infarction model. Myocardial infarctions were induced by 30 min occlusions of the proximal left anterior descending coronary artery (LAD) in 17 healthy adult female pigs. First-pass and 5 min-delayed cardiac MDCTs were performed after 4 weeks of LAD occlusion. Myocardial CPMs were obtained by using the CPM program. Triphenyltetrazolium chloride (TTC)-staining was performed on the cardiac specimens. We analyzed the intermodality agreement on the size and location of the myocardial infarctions. TTC staining revealed myocardial infarction in 16 of 17 pigs, and 15 of these (94%) showed matched infarcts on the CPM and first-pass images. The areas of perfusion deficit noted in early arterial phase images and CPM coincided exactly with the areas of poor TTC staining in 12 of 15 pigs (80%). In the three remaining pigs, the areas of poor TTC staining were larger than those of a perfusion deficit demonstrated by either early arterial phase images or CPM. The agreement between these tests is calculated to be moderate to good (k = 0.736, P < 0.05). Ten myocardial segments in 4 of the 15 pigs (27%) with hypoattenuated myocardium showed a delayed enhancement on the 5 min-delayed images. Contrast-enhanced MDCT was useful and accurate in detecting chronic myocardial infarction; CPM was helpful in visualizing the infarcted myocardium.

Occlusive Myocardial Infarct in Rabbits as Detected by the Use of Multidetector-row CT: A Radiological-pathological Correlation According to the Infarct Age

Purpose: This study was designed to evaluate the imaging findings and the diagnostic accuracy for determining the transmural extent of a myocardial infarct (MI) in rabbits with the use of multidetector row CT (MDCT) according to the infarct age, and we determined the radiological and pathological correlation. Materials and Methods: MIs were demonstrated in 10 of 15 rabbits in which ligation of the left coronary arteries was performed. The cardiac scans were obtained according to the infarct age (hyperacute (within 24 hours), acute (24 hours to seven days) or chronic (eight weeks)) by using a 16-MDCT scanner in the contiguous transverse plane with 120.75 mm collimation, 420 ms rotation and retrospective ECG-gating. Early and late scans were obtained at one minute and six minutes, respectively, after intravenous contrast injection. The rabbits were sacrificed according to the infarct age. Slices of the cardiac specimens stained with triphenyltetrazolium chloride (TTC) and the MDCT images were evaluated for scoring the transmural extent of the MIs. The agreement of the scores between the MDCT images and TTC-stained specimens was statistically analyzed. The radiological and pathological correlation was determined for the specimens and the MDCT images. Results: The presence of an infarcted myocardium in the hyperacute phase was demonstrated as a mixed low and high-attenuation area on the early and late scans. In the acute phase, a low-attenuation area was seen on the early scan and a low-attenuation area with enhancement along the endocardial and pericardial sides was seen on the late scan. In the chronic phase, a high-attenuation area was seen on the late scan. There was fair agreement for the scores of the MDCT scan in hyperacute MI (kappa value = 0.380) and excellent agreement of the early scan in acute MI and the late scan of chronic MI (kappa value = 0.865 and 0.858, respectively). A pathological examination of the TTC-unstained areas of the MI specimens demonstrated the presence of necrosis of myocardial cells and the increased inflammatory infiltrates and repair process surrounding the damaged myocytes, according to the infarct age. Conclusion: The MDCT findings of MI were various according to the infarct age, which might be caused by the degree of the inflammatory and repair process. For diagnosing the transmural extent of MI, the early MDCT scan was useful for the acute MI and the late scan was useful for the chronic MI. However, MDCT was not useful for diagnosing the transmural extent of the hyperacute MI.

Abstract 5672: Novel SCN5A Mutations and SNP in Patients with Brugada Syndrome

Background: Sodium channel alpha 5 subunit (SCN5A) mutations are important genetic abnormalities of Brugada syndrome (BS). Here we report novel SCN5A mutations in patients with BS. Methods and Results: Genomic mutations in the SCN5A gene were analyzed by PCR and direct sequencing in 121 patients with BS (119 men and 2 women). Thirteen (11%) of the patients had SCN5A mutation and five (4%) of the patients had single nucleotide polymorphisms (SNP) associated with BS. Eleven patients had single point mutations, one patient had a deletion (1380 del N) and one patient had a mutation within the splicing junction (IVS21+1 g&gt;a). The whole-cell patch clamp technique revealed that four novel mutations (F532C, R814Q, G833R, R878C) showed loss of function, and peak INa of a novel SNP (L1988R) transiently expressed in HEK 293 cells was significantly reduced (P&lt;0.05). RT-PCR analysis revealed that the intronic mutation (IVS21+1 g&gt;a) resulted in exon 21 deletion of SCN5A in cardiac biopsy specimens from the patient (Figure ). Conclusions: We detected five novel SCN5A mutations (1380 del N, F532C, R814Q, G833R, R878C) and a novel disease-associated SNP (L1988R) in patients with BS. We showed a novel analysis of splicing mutation (IVS21+1 g&gt;a) that resulted in exon skipping in the heart.

Abstract P184: Novel SCN5A Mutations and SNP in Patients with Brugada Syndrome

Background: Sodium channel alpha 5 subunit (SCN5A) mutations are important genetic abnormalities of Brugada syndrome (BS). Here we report novel SCN5A mutations in patients with BS. Methods and Results: Genomic mutations in the SCN5A gene were analyzed by PCR and direct sequencing in 121 patients with BS (119 men and 2 women). Thirteen (11%) of the patients had SCN5A mutation and five (4%) of the patients had single nucleotide polymorphisms (SNP) associated with BS. Eleven patients had single point mutations, one patient had a deletion (1380 del N) and one patient had a mutation within the splicing junction (IVS21+1 g&gt;a). The whole-cell patch clamp technique revealed that four novel mutations (F532C, R814Q, G833R, R878C) showed loss of function, and peak INa of a novel SNP (L1988R) transiently expressed in HEK 293 cells was significantly reduced (P&lt;0.05). RT-PCR analysis revealed that the intronic mutation (IVS21+1 g&gt;a) resulted in exon 21 deletion of SCN5A in cardiac biopsy specimens from the patient (Figure ). Conclusions: We detected five novel SCN5A mutations (1380 del N, F532C, R814Q, G833R, R878C) and a novel disease-associated SNP (L1988R) in patients with BS. We showed a novel analysis of splicing mutation (IVS21+1 g&gt;a) that resulted in exon skipping in the heart.

Abstract 3726: Right Ventricular Microcirculatory Changes in Pulmonary Hypertension-Associated Right Heart Failure Imaged by Three-Dimensional Confocal Microscopy

Background: Right ventricular (RV) ischemia occurs in pulmonary hypertension (PH) patients despite normal coronary angiograms. O 2 supply-demand balance in the pressure-overloaded RV is determined by the interplay between reduced coronary perfusion (due to systemic hypotension and compression of coronary vessels) and a wall stress-related increase in myocardial O 2 consumption. The role of capillary rarefaction, which is involved in pressure overload induced left heart failure, has not been investigated in PH-associated RV failure. Methods: Rats received a single dose of the VEGF receptor blocker SU5416 and were exposed to 4 wks of normobaric hypoxia and 2 wks of sea level room air. This regimen induces angioproliferative changes in the pulmonary circulation, severe PH and RV failure. At 6 wks, the rats received intravital injections of Texas-Red conjugated tomato lectin, which is an N-acetylglucosamine specific marker of blood vessels in rodents. After circulation of the lectin for 5 min, papaverin was injected to promote maximal vasodilation of all perfused blood vessels. Animals were exsanguinated and perfusion-fixed with paraformaldehyde. After overnight fixation, 300 μm thick transverse sections of the myocardium were bleached in a methanol/DMSO mixture and nuclei were stained using DAPI. Prior to 3-dimensional imaging under a confocal microscope, further tissue clearing was achieved using organic solvents. Results: Z-stacks of 2-dimensional images were made in whole-mount cardiac specimens, using a step size of 0.32 μm. 3-dimensional reconstruction allowed quantification of the total capillary volume, which was reduced in the PH model compared to controls (2.9±1% vs. 5.7±1% of tissue volume, p&lt;0.05). The reconstructions showed a remarkable increased morphological heterogeneity of the capillaries in the failing RV. In contrast, images of the LV microcirculation in the SU5416 treated animals were not different from those of controls. Conclusions: RV failure in experimental PH is associated with capillary rarefaction. 3-dimensional imaging of the myocardial microcirculation can be achieved using intravital injections of tomato lectin and subsequent confocal imaging.

Classification of amyloid deposits in diagnostic cardiac specimens by immunofluorescence

At least 12 distinct forms of amyloidosis are known to involve the heart or great vessels. Patient treatment regimens require proper subtyping of amyloid deposits in small diagnostic cardiac specimens. A growing lack of confidence in immunohistochemical staining for subtyping amyloid has arisen primarily as a result of studies utilizing immunoperoxidase staining of formalin-fixed paraffin-embedded tissue. Immunofluorescence staining on fresh frozen tissue is generally considered superior to immunoperoxidase staining for subtyping amyloid; however, this technique has not previously been reported in a series of cardiac specimens. Amyloid deposits were subtyped in 17 cardiac specimens and 23 renal specimens using an immunofluorescence panel. Amyloid deposits were successfully subtyped as AL, AH, or AA amyloid by immunofluorescence in 82% of cardiac specimens and 87% of renal specimens. In all cases, the amyloid classification was in good agreement with available clinical and laboratory assessments. A cross-study analysis of 163 cases of AL amyloidosis reveals probable systemic misdiagnosis of cardiac AL amyloidosis by the immunoperoxidase technique, but not by the immunofluorescence technique. Amyloid deposits can be reliably subtyped in small diagnostic cardiac specimens using immunofluorescence. The practical aspects of implementing an immunofluorescence approach are compared with those of other approaches for subtyping amyloid in the clinical setting.

Morphologic Validation of Reperfused Hemorrhagic Myocardial Infarction by Cardiovascular Magnetic Resonance

The purposes of this study were to assess the ex vivo cardiovascular magnetic resonance (CMR) signals of pathologically proved hemorrhagic myocardial infarction (MI) and to correlate these with in vivo CMR findings. Late gadolinium hypoenhancement within a hyperenhanced area in reperfused acute MI is ascribed to severe microvascular obstruction. The hearts of 2 patients, who died from cardiogenic shock after acute MIs and who had undergone coronary recanalization and in vivo CMR, were examined by T(2) and T(1) late enhancement sequences as well as by gross and histologic investigation. Four corresponding short-axis slices of each cardiac specimen from the base to the left ventricular apex were selected to assess the extent of MI and hemorrhage and were compared with the in vivo T(2) and late enhancement CMR scans. On pathologic examination, the extent of MI was 57 +/- 30% and 44 +/- 24%, and the extent of hemorrhage was 23 +/- 13% and 19 +/- 8% of the left ventricular area, respectively, showing progressive increases from the base to the apex. The low-signal intensity areas observed by ex vivo T(2) CMR strongly correlated with the hemorrhage quantified on histology (R = 0.93, p = 0.0007). Using ex vivo late gadolinium sequences, bright areas surrounded by thin dark rims, consistent with magnetic susceptibility effects, were detected, corresponding with hemorrhage. On in vivo CMR images, low-signal intensity and hyperintense areas with peripheral susceptibility artifacts were observed within the MI core on T(2) and late gadolinium sequences, respectively. In conclusion, in reperfused MI, CMR hypointense T(2) signal and susceptibility effects within the late gadolinium hypoenhanced areas are consistent with interstitial hemorrhage due to irreversible vascular injury, as proved by pathologic study.

Post-transcriptional modifications of VEGF-A mRNA in non-ischemic dilated cardiomyopathy

Vascular endothelial growth factor (VEGF-A) is one of the most important proangiogenic factors. It has many isoforms encoded by one gene. The occurrence of these isoforms is associated with the process of alternative splicing of mRNA. Some of the splice forms are perceived as tissue specific. The aim of this study was to determine the alternative splicing of VEGF-A mRNA in dilated cardiomyopathy, especially at the level of particular myocardial layers. The assessment of post-transcriptional modifications of VEGF-A mRNA was made on specimens taken from the explanted hearts of patients undergoing cardiac transplantation. Molecular and histopathological studies were perfomed on particular layers of the myocardial muscle (endocardium, myocardium, epicardium). A molecular analysis of cardiac samples was performed by quantitative analysis of the mRNA of the studied VEGF-A isoforms (VEGF121, -145, -165, -183, -189, and -206) using QRTPCR with an ABI-PRISM 7700-TaqMan sequence detector. 72 cardiac specimens taken from the explanted hearts were analyzed. Each of the studied VEGF-A splice forms was present in the evaluated hearts, but the types of alternative splicing of mRNA were different in particular layers. Quantitative analysis revealed different amounts of the studied isoforms. Generally, significantly increased expression of the VEGF-A isoforms was observed in samples taken from hearts with post-inflammatory etiology of cardiomyopathy. Our conclusions are: 1. All the studied VEGF-A isoforms were found in the human hearts, including those thusfar considered characteristic for other tissues. 2. Significant differences were observed in the expression of the VEGF-A splice forms with respect to the myocardial layers and the location of the cardiac biopsy. 3. Repetitive and comparable results for samples with post-inflammatory etiology were obtained, and they revealed considerably higher amounts of VEGF-A isoforms compared to specimens with idiopathic etiology.

Increased myocardial prevalence of C‐reactive protein in human coronary heart disease: relation to local endothelial injury and microvessel rarefaction

Increased plasma C-reactive protein (CRP) is a biomarker of coronary heart disease (CHD) but its potential roles as a disease participant are not well defined. A key component of CHD is endothelial injury and dysfunction. Here we tested the hypotheses that CRP is detectable in human cardiac specimens and is elevated in patients with CHD, and these changes are related to local endothelial stress and/or microvessel rarefaction. Cardiac LV specimens were collected at autopsy within 4h of death and were classified as normal controls (CTL) or evidence of CHD (w/o MI) (n=12/group). Immunohistochemistry and digital image analyses were used to define discrete regional distributions of CRP in myocyte, coronary artery and microvessel regions. Prevalence of subendocardial microvessels was determined using an anti-CD31 antibody and automated macro. Increased interstitial fibrosis & hypertrophy was seen in CHD. CTL had detectable levels of CRP, which was increased in CHD group (p<0.05). Microvessel prevalence was significantly reduced in CHD myocardium vs. CTL, and rarefaction was inversely correlated to myocardial CRP prevalence (p<0.05). These findings provide evidence that locally produced myocardial CRP is a potential participant of myocardial toxicity and microvascular rarefaction. These observations may also help to explain the strong predictive value of increased blood CRP to CHD progression in humans

Robust gene expression with amplified RNA from biopsy-sized human heart tissue

The need to assess heart failure at an early stage highlights the importance of accurate microarray analysis using small tissue samples. To test our ability to obtain high quality RNA from biopsy-sized cardiac specimens, amplification was performed on RNA from biopsy-sized samples of left ventricle (LV) tissue from one explanted failing human heart and one non-failing heart. Two methods were used: one-cycle (1C) amplification of 1.6 μg of RNA, and two-cycle (2C) amplification of 50 ng of RNA. The resulting cRNA was hybridized to Affymetrix GeneChip® arrays. Over 65% of all differentially expressed genes for failing vs non-failing hearts were concordant between 1C and 2C RNA amplification. Differentially expressed genes between 1C and 2C RNA amplification in our study were highly correlated (R2 = 0.957 and changes in gene expression agreed with prior studies on genes and heart failure; e.g., decreased α-myosin heavy chain and α-tropomyosin, as well as increased expression of insulin-like growth factor). Two cycles of amplification from cardiac biopsies will permit accurate transcription profiling of heart failure at pre-symptomatic stages. Ability to measure gene expression from nanogram amounts of RNA will provide new opportunities to predict progression to symptomatic heart failure, and to identify potential targets for therapy.