Faculty Bibliography

Primary vasculitis is the inflammation and necrosis of vessel walls not associated with infections, drugs, and autoimmune and lymphoproliferative disorders. It is important to make the correct diagnosis of different types of vasculitis, as their prognosis may be significantly different. Classification of vasculitis based on the size of the vessel is helpful, but there is often an overlap. Whereas the criteria proposed by the American College of Rheumatology are primarily clinical, the definitions set forth by the Chapel Hill Consensus Conference are based only on histologic observations. Correct diagnosis requires appropriate incorporation of the clinical history, laboratory parameters, and the histologic data. Incorporation of antineutrophil cytoplasmic antibodies in defining the pathogenesis of vasculitis has been particularly useful in diagnosing those small vessel vasculitides that are life threatening and need immediate intervention.

UNLABELLED:We used a model of porcine coronary atherosclerosis characterized by smooth muscle cell apoptosis to test the hypothesis that apoptosis of cells in the vascular wall of coronary arteries can be detected on SPECT images using a technetium-labeled radiotracer that targets apoptosis. METHODS:Eleven juvenile male swine received a high-fat diet combined with injury to 22 coronary vessels. After 51 +/- 9 d (mean +/- SD), the animals underwent coronary angiography, were injected with 403.3 +/- 48.1 MBq of 99mTc-annexin V, underwent SPECT, and were sacrificed. The coronary arteries underwent autoradiography and well counting, and immunostaining was performed for alpha-actin, caspase, and macrophages. RESULTS:Atherosclerotic lesions were predominantly of American Heart Association class II. Thirteen of the 22 injured vessels showed focal uptake of 99mTc-annexin V in vivo (scan positive), and 9 injured vessels and all control vessels showed no focal uptake (scan negative). The count ratios of the injured vessels to the control vessels were 2.38 +/- 0.61 for scan-positive vessels and 1.27 +/- 0.23 for scan-negative vessels (P < 0.001). The percentages of injected dose for the scan-positive and scan-negative vessels were 1.73 +/- 0.83 x 10(-3) and 0.68 +/- 0.20 x 10(-3), respectively (P < 0.001). Immunohistopathologic examination found that the cells undergoing apoptosis were smooth muscle cells. The apoptotic index (caspase-positive cells to total cells) was 63% +/- 7% for scan-positive vessels and 16% +/- 10% for scan-negative vessels (P < 0.001). Both the count ratio of injured vessels to control vessels and the percentage injected dose correlated significantly with death rate by regression analysis. CONCLUSION/CONCLUSIONS:Annexin is a noninvasive method to identify plaque apoptosis in the coronary vessels.

Endomyocardial biopsy (EMB) remains the "gold standard" for diagnosing rejection after cardiac transplantation. In addition, it has value in monitoring patients during treatment with doxorubicin. It also is important in the setting of acute-onset heart failure for the diagnosis of myocarditis, particularly giant cell myocarditis because earlier transplantation usually is undertaken in patients with giant cell morphologic features. EMB has a role in the unexplained cardiomyoapthy for excluding specific disease processes that might lead to similar morphofunctional changes but might be reversible or a contraindication for transplantation. This review focuses on the growing number of diseases that can be diagnosed by EMB in adult and pediatric age groups.

BACKGROUND:Myofibrillarlytic (MFL) cells are commonly observed in subendocardial myocardium in myocardial infarction. Because ischemic damage to myocytes is also known to induce apoptosis, we evaluated the prevalence of apoptosis in MFL cells in nine ischemic cardiomyopathic hearts explanted during transplantation. METHODS:Myocytes with partial or complete clearing of cytoplasm, observed commonly in the subendocardium, were recognized as MFL cells. Prevalence of apoptosis was defined by TUNEL and ISOL staining and further characterized by immunohistochemical staining for caspase-3, Bcl2, BCL-X(L), Bax, proliferating cell nuclear antigen (PCNA), and Ki67. RESULTS:Of 4131 MFL cells examined, 1305 (32%) possessed nuclei in a given histologic section; 1140 (88%) of the nucleated myocardial cells were TUNEL positive. Of 842 cells with normal appearance, 257 (31%) cells demonstrated nuclei in the given histologic section. TUNEL staining was observed in 5 (1.9%) in these control areas. All MFL cells stained positive for caspase 3. The antiapoptotic proteins, Bcl2 and BCL-X(L), demonstrated intense upregulation within and surrounding MFL cells, whereas pro-apoptotic protein Bax expression was only seen at control level. The MFL cells had Ki67 negative and PCNA positive nuclei. CONCLUSIONS:The present study demonstrates that the majority of MFL cells are apoptotic and are associated with upregulation of caspase 3. Simultaneous upregulation of Bcl2 represents a survival effort in these myocytes. This is consistent with the review of the literature that MFL cells are viable, persist in myocardium for long time and may be functionally reversible. Evidence for concurrent apoptosis and survival instinct represent a conceptual paradox and suggests that myocytes undergoing apoptosis should be amenable to reconstitution of function.

We investigated a novel polyepoxide crosslinker that was hypothesized to confer both material stabilization and calcification resistance when used to prepare bioprosthetic heart valves. Triglycidylamine (TGA) was synthesized via reacting epichlorhydrin and NH(3). TGA was used to crosslink porcine aortic cusps, bovine pericardium, and type I collagen. Control materials were crosslinked with glutaraldehyde (Glut). TGA-pretreated materials had shrink temperatures comparable to Glut fixation. However, TGA crosslinking conferred significantly greater collagenase resistance than Glut pretreatment, and significantly improved biomechanical compliance. Sheep aortic valve interstitial cells grown on TGA-pretreated collagen did not calcify, whereas sheep aortic valve interstitial cells grown on control substrates calcified extensively. Rat subdermal implants (porcine aortic cusps/bovine pericardium) pretreated with TGA demonstrated significantly less calcification than Glut pretreated implants. Investigations of extracellular matrix proteins associated with calcification, matrix metalloproteinases (MMPs) 2 and 9, tenascin-C, and osteopontin, revealed that MMP-9 and tenascin-C demonstrated reduced expression both in vitro and in vivo with TGA crosslinking compared to controls, whereas osteopontin and MMP-2 expression were not affected. TGA pretreatment of heterograft biomaterials results in improved stability compared to Glut, confers biomechanical properties superior to Glut crosslinking, and demonstrates significant calcification resistance.

BACKGROUND AND AIM OF THE STUDY/OBJECTIVE:Previous immunohistochemistry studies have shown that the transcription factor, Egr-1, is increased in human atherosclerotic lesions but is absent from the normal adjacent aortic wall. The hypothesis was investigated that Egr-1 is also increased in calcified heart valve cusps because of the unique presence in these tissues of proteins known to be regulated by Egr-1, such as tenascin C (TN-C). METHODS:Non-calcified and calcified human aortic valves were obtained at autopsy or from cardiac surgery. Egr-1 immunohistochemical studies were performed. The effects of Egr-1 on cellular proliferation and on mechanisms of calcification were also investigated using sheep aortic valve interstitial cell (SAVIC) cultures. Signal transduction pathways involving Egr-1 were studied with specific inhibitors. RESULTS:Immunohistochemical studies revealed that calcific aortic stenosis cusps contained a significantly higher level of Egr-1 in the spindle-shaped interstitial cells of calcified human aortic valves, but not white blood cells. By comparison, Egr-1 was detected at very low levels in the interstitial cells of non-calcified human aortic valve cusps. SAVIC cultivated on denatured versus native collagen substrates demonstrated a marked increase in Egr-1 levels (by Western blotting), and an absence of calcification in these cultures, compared to SAVIC grown on native collagen which calcified severely with little Egr-1 expression. Parallel increases in TN-C and osteopontin (OPN), both of which are proteins associated with heart valve calcification, were observed (by Western blotting) in SAVIC grown on denatured collagen. Furthermore, a protein kinase-C (PKC) inhibitor blocked the up-regulation of Egr-1 and TN-C, implicating PKC-dependent signaling control of Egr-1 and TN-C up-regulation. CONCLUSION/CONCLUSIONS:Egr-1 is up-regulated in human calcific aortic stenosis cusps compared to non-calcified normal cusps. Egr-1 up-regulation involves a PKC-dependent signaling pathway. TN-C and OPN appear to be co-regulated with Egr-1. Furthermore, in SAVIC cultures on denatured collagen, Egr-1 up-regulation was associated with inhibition of calcification. Taken together, these results suggest that complex Egr-1 mechanisms may be operative in calcific aortic stenosis.