Maija Kaartinen

Expression of Angiotensin II and Interleukin 6 in Human Coronary Atherosclerotic Plaques Potential Implications for Inflammation and Plaque Instability

BACKGROUND Patients with an activated renin-angiotensin system (RAS) or genetic alterations of the RAS are at increased risk of myocardial infarction (MI). Administration of ACE inhibitors reduces the risk of MI, and acute coronary syndromes are associated with increased interleukin 6 (IL-6) serum levels. Accordingly, the present study evaluated the expression of angiotensin II (Ang II) in human coronary atherosclerotic plaques and its influence on IL-6 expression in patients with coronary artery disease. METHODS AND RESULTS Immunohistochemical colocalization of Ang II, ACE, Ang II type 1 (AT(1)) receptor, and IL-6 was examined in coronary arteries from patients with ischemic or dilated cardiomyopathy undergoing heart transplantation (n=12), in atherectomy samples from patients with unstable angina (culprit lesion; n=8), and in ruptured coronary arteries from patients who died of MI (n=13). Synthesis and release of IL-6 was investigated in smooth muscle cells and macrophages after Ang II stimulation. Colocalization of ACE, Ang II, AT(1) receptor, and IL-6 with CD68-positive macrophages was observed at the shoulder region of coronary atherosclerotic plaques and in atherectomy tissue of patients with unstable angina. Ang II was identified in close proximity to the presumed rupture site of human coronary arteries in acute MI. Ang II induced synthesis and release of IL-6 shortly after stimulation in vitro in macrophages and rat smooth muscle cells. CONCLUSIONS Ang II, AT(1) receptor, and ACE are expressed at strategic sites of human atherosclerotic coronary arteries, suggesting that Ang II is produced primarily by ACE within coronary plaques. The observation that Ang II induces IL-6 and their colocalization with the AT(1) receptor and ACE is consistent with the notion that the RAS may contribute to inflammatory processes within the vascular wall and to the development of acute coronary syndromes.

Infiltrates of Activated Mast Cells at the Site of Coronary Atheromatous Erosion or Rupture in Myocardial Infarction

BACKGROUND Erosion and rupture of coronary atheromas are the events preceding the vast majority of acute coronary syndromes. The shoulder regions of atheromas, the sites at which erosion or rupture is most likely to occur, are the sites at which mast cells accumulate. These cells are filled with neutral proteases capable of triggering extracellular matrix degradation via activation of matrix metalloproteinases. To obtain more direct evidence for the participation of mast cells in the acute coronary syndromes, we quantified the numbers of mast cells at eroded or ruptured sites of coronary atheromas in patients who died of myocardial infarction. METHODS AND RESULTS In specimens of coronary arteries from 20 patients who had died of acute myocardial infarction, the site of atheromatous erosion or rupture was identified. The specimens were stained with monoclonal antibodies against the two major proteases of mast cells, tryptase and chymase, and against macrophages, T lymphocytes, and smooth muscle cells. At the immediate site of erosion or rupture, mast cells amounted to 6% of all nucleated cells, in the adjacent atheromatous area to 1%, and in the unaffected intimal area to 0.1%. The proportions of these mast cells that were activated, ie, had been stimulated to degranulate and release some of their tryptase and chymase contents, were 86% at the site of erosion or rupture, 63% in the adjacent atheromatous area, and 27% in the unaffected intima. At the site of erosion or rupture, the numbers of macrophages and T lymphocytes were also increased, but the number of smooth muscle cells was decreased. CONCLUSIONS The accumulation of activated mast cells (200-fold more than in the unaffected coronary intima) at the site of atheromatous erosion or rupture suggests that in thrombotic coronary occlusion the role played by mast cells is significant.

Accumulation of activated mast cells in the shoulder region of human coronary atheroma, the predilection site of atheromatous rupture.

BACKGROUND Rupture in the shoulder region of a coronary atheroma is considered to be a sequel to local extracellular matrix degradation in this highly vulnerable site. Such degradation could be triggered by mast cells, which are filled with neutral proteases and are present in coronary atheromas. However, the distribution and phenotype of mast cells within coronary atheromas have not been studied. METHODS AND RESULTS Specimens of normal and atherosclerotic human coronary intima from 32 autopsy cases with ages ranging from 13 to 67 years were stained with monoclonal antibodies against the two major proteases of mast cells, tryptase and chymase. Of the tryptase-containing mast cells, a variable proportion (average, 40%; range, 0% to 100%) also contained chymase. In the normal coronary intimas, mast cells amounted to 0.1% of all nucleated cells. In the fatty streaks, this proportion was higher by 9-fold, and in the cap, core, and shoulder regions of atheromas by 5-, 5-, and 10-fold, respectively. Electron and light microscopic studies of mast cells in the shoulder region of atheromas revealed degranulation of mast cells, a sign of their activation, and moreover, that the proportion of activated mast cells was much higher (85%) in this region than in the normal intima (18%). CONCLUSIONS The far higher proportion (50-fold) of activated mast cells in the shoulder region of atheromas supports the hypothesis that mast cells, a cell type capable of triggering matrix degradation, actively participate in the destabilization and ensuing rupture of coronary atheromas and thus may trigger an acute coronary event.

Association Between Myocardial Infarction and the Mast Cells in the Adventitia of the Infarct-Related Coronary Artery

BACKGROUND Histamine, a product of mast cells, is an effective vasoconstrictor of atherosclerotic coronary arteries. Because it has been suggested that coronary spasm plays a role in acute coronary syndromes such as myocardial infarction (MI), we quantified and characterized the mast cells in the adventitia of infarct-related coronary arteries. METHODS AND RESULTS In a series of 17 autopsied MI patients, we identified the segment of the left coronary artery with ruptured plaque responsible for the infarction. More distal segments from the infarct-related coronary artery, either with nonruptured plaques or with normal intima, were also studied. Corresponding segments taken from left coronary arteries obtained from 17 patients who had died of noncardiac causes served as controls. Adventitial mast cells in the infarct-related and the control coronary arteries were identified immunohistochemically by staining for tryptase. In the infarct-related coronary arteries, we also stained for chymase and histamine. Moreover, T lymphocytes and macrophages were identified immunohistochemically and counted. In the infarct-related coronary arteries, significantly larger numbers of mast cells were present in the adventitia backing ruptured plaques (98+/-40 mast cells/mm2, mean+/-SD) than in the adventitia backing nonruptured plaques (41+/-12 mast cells/mm2; P<0.001) or backing normal intima (19+/-8 mast cells/mm2; P<0.001). No such difference was found among the 3 different segments in the control coronary arteries. The majority of mast cells contained not only tryptase but also chymase. Mast cells were the only cells in the coronary adventitia that contained histamine. The proportion of adventitial mast cells that were degranulated was highest in the segments with ruptured plaques. The numbers of adventitial macrophages and T lymphocytes were also increased in the segments with plaque rupture. CONCLUSIONS In infarct-related coronary arteries, the number of degranulated mast cells in the adventitia backing ruptured plaques is increased. Histamine released from the degranulated mast cells may reach the media, where it may locally provoke coronary spasm and thus contribute to the onset of MI.

Mast Cells in Rupture-Prone Areas of Human Coronary Atheromas Produce and Store TNF-α

BACKGROUND Mast cells, a cell type involved in inflammatory reactions, are present in coronary atheromas and localize to the erosion or rupture site of atheromas in myocardial infarction. Here we report the presence of TNF-alpha, a proinflammatory cytokine, in mast cells of human coronary atheromas. METHODS AND RESULTS From samples of 37 coronary arteries from subjects autopsied for medicolegal reasons, sections of the bifurcation area of the left coronary artery were stained immunohistochemically for mast cells and TNF-alpha. In addition, macrophages, T lymphocytes, smooth muscle cells, and endothelial cells were investigated for their content of TNF-alpha. In normal intimas and fatty streaks, none of the cell types studied were TNF-alpha-positive. In 14 of the 24 atheromas found, TNF-alpha-positive cells were present. Of the total number of mast cells, 23% stained for TNF-alpha; of the macrophages, 1.3%; and of the smooth muscle cells, 0.4%. The majority (55%) of TNF-alpha-positive mast cells in the atheromas were located in the shoulder region and the remaining 35% in the cap and 10% in the core regions. Immunoelectron microscopy showed that the TNF-alpha in mast cells resided within their cytoplasmic secretory granules, demonstrating that these cells contain stores of TNF-alpha that will be released on degranulation. CONCLUSIONS This study demonstrates the presence of mast cells with TNF-alpha-containing secretory granules, particularly in the shoulder region of human coronary atheromas. By releasing their TNF-alpha, mast cells may play an active role in the inflammatory reactions of these rupture-prone areas of atheromas.

Mast cells of two types differing in neutral protease composition in the human aortic intima. Demonstration of tryptase- and tryptase/chymase-containing mast cells in normal intimas, fatty streaks, and the shoulder region of atheromas.

Biochemical studies in vitro have demonstrated that stimulated mast cells induce macrophage foam cell formation through the synergistic action of mast cell granule neutral proteases and proteoglycans. To determine the presence and number of mast cells in human arterial intima, the site of atherogenesis, specimens of normal and atherosclerotic human aortic intima from 35 autopsies of persons ranging from 13 to 67 years old were stained with monoclonal antibodies against the two major proteases of mast cells, tryptase and chymase. All mast cells present were found to contain tryptase, and an average of 40% contained chymase as well. In sections of normal intimas, fatty streaks, and atheromas, the mast cells had average densities of 15/mm2, 15/mm2, and 3/mm2, respectively. In contrast to the normal intimas and fatty streaks, however, the atheromas had mast cells distributed unevenly in a typical pattern: 8/mm2 in the shoulder region, 1/mm2 in the fibrous cap, and none in the core region. In normal intimas, fatty streaks, and the shoulder region of atheromas, the mast cells amounted to 3% of all nucleated cells. The ratios of mast cells to T lymphocytes and to macrophages, respectively, were 2:1 and 1:4 in normal intimas, 1:3 and 1:10 in fatty streaks, and 1:5 and 1:20 in the shoulder region of atheromas. Thus, among the blood-borne cells in the human aortic intima, mast cells compose a significant cell population, and in terms of their protease content, these intimal mast cells are heterogeneous.(ABSTRACT TRUNCATED AT 250 WORDS)

Mast cells accompany microvessels in human coronary atheromas: implications for intimal neovascularization and hemorrhage

Mast cells have been assigned a role in neovascularization. Therefore, we examined the deep regions of human coronary atheromas, the areas known to be prone to neovascularization, for the presence of mast cells. Specimens of atherosclerotic human coronary intima from 37 autopsy cases with ages of 24-84 years were stained with elastica-van Gieson to detect atheroma formation and with monoclonal antibody against von Willebrand factor to detect neovascularization. Mast cells were detected by staining the atheromas with monoclonal antibodies against the two major proteases of mast cells, tryptase and chymase. Of the 24 coronary atheromas found, 13 contained mast cells in the deep regions. All these 13 deep regions also displayed neovascularization, and the number of microvessels and the number of mast cells around the microvessels correlated strongly with the size of the atheroma. On the other hand, of the 11 deep regions lacking mast cells, only one displayed neovascularization. In the neovascularized areas of the coronary atheromas, the mast cells were in close proximity to the microvessels. All the mast cells contained tryptase, and some of them chymase, both known for their angiogenic and matrix-degrading potential. In light microscopic studies, degranulated mast cells were observed indicating activation of these cells, with release of tryptase and chymase. The selective localization of activated mast cells containing angiogenic factors around newly formed microvessels in human coronary atheromas suggests that mast cells play a role in the neovascularization of these lesions. Moreover, mast cells may also, by virtue of their neutral proteases, injure the microvessels, and thereby produce intraplaque hemorrhages and, ultimately, unstable lesions.

Genetics and genotype-phenotype correlations in Finnish patients with dilated cardiomyopathy

Genetic analysis among patients with dilated cardiomyopathy (DCM) is becoming an important part of clinical assessment, as it is in hypertrophic cardiomyopathy (HCM). The genetics of DCM is complex and therefore next-generation sequencing strategies are essential when providing genetic diagnostics. To achieve maximum yield, the diagnostic approach should include comprehensive clinical phenotyping combined with high-quality, high-coverage deep sequencing of DCM-associated genes and clinical variant classification as a basis for defining true yield in genetic testing. Our study has combined a novel sequencing strategy and clinical interpretation to analyse the yield and genotype–phenotype correlations among well-phenotyped Finnish DCM patients.

Late gadolinium enhanced cardiovascular magnetic resonance of lamin A/C gene mutation related dilated cardiomyopathy

BackgroundThe purpose of this study was to identify early features of lamin A/C gene mutation related dilated cardiomyopathy (DCM) with cardiovascular magnetic resonance (CMR). We characterise myocardial and functional findings in carriers of lamin A/C mutation to facilitate the recognition of these patients using this method. We also investigated the connection between myocardial fibrosis and conduction abnormalities.MethodsSeventeen lamin A/C mutation carriers underwent CMR. Late gadolinium enhancement (LGE) and cine images were performed to evaluate myocardial fibrosis, regional wall motion, longitudinal myocardial function, global function and volumetry of both ventricles. The location, pattern and extent of enhancement in the left ventricle (LV) myocardium were visually estimated.ResultsPatients had LV myocardial fibrosis in 88% of cases. Segmental wall motion abnormalities correlated strongly with the degree of enhancement. Myocardial enhancement was associated with conduction abnormalities. Sixty-nine percent of our asymptomatic or mildly symptomatic patients showed mild ventricular dilatation, systolic failure or both in global ventricular analysis. Decreased longitudinal systolic LV function was observed in 53% of patients.ConclusionsCardiac conduction abnormalities, mildly dilated LV and depressed systolic dysfunction are common in DCM caused by a lamin A/C gene mutation. However, other cardiac diseases may produce similar symptoms. CMR is an accurate tool to determine the typical cardiac involvement in lamin A/C cardiomyopathy and may help to initiate early treatment in this malignant familiar form of DCM.

A novel mutation, Arg71Thr, in the δ-sarcoglycan gene is associated with dilated cardiomyopathy

Approximately 20–35% of cases of idiopathic dilated cardiomyopathy are familial. DCM-associated mutations have been reported in 13 genes including the desmin, δ-sarcoglycan, and metavinculin genes. This study screened for variants in these genes in Finnish patients with DCM. All coding regions of the desmin and δ-sarcoglycan genes and the metavinculin-specific exon of the vinculin gene were screened in 52 DCM patients from eastern Finland by PCR-SSCP. We detected a novel mutation, Arg71Thr, in the δ-sarcoglycan gene in two members of a small DCM family. One of the mutation carriers fulfills diagnostic criteria for DCM and is also symptomatic. The other mutation carrier has slightly dilated left ventricle and well preserved systolic function. Therefore carriers of the Arg71Thr mutation had a relatively mild phenotype and a late onset of the disease. Disease-associated mutations were not found in the desmin gene or the metavinculin-specific exon of the vinculin gene. We conclude that the desmin and δ-sarcoglycan genes are not predominant disease-causing genes in patients with DCM in eastern Finland.