Background-Recent studies have demonstrated that lesions of aortic sclerosis and stenosis share several similarities with lesions of atherosclerosis. In atherosclerosis, angiotensin-converting enzyme (ACE) is expressed by a subset of macrophages. This study was undertaken to determine whether ACE might be present in aortic sclerosis or stenosis lesions. Methods and Results-Immunohistochemistry was performed on 26 paraffin-embedded human aortic valves. Monospecific antibodies were used to identify ACE, macrophages, angiotensin II type 1 receptor (AT-1 receptor), angiotensin II, and apolipoprotein B. Human low-density lipoprotein (LDL) and high-density lipoprotein (HDL) were isolated from plasma of normal volunteers by sequential density-gradient ultracentrifugation. ACE was not present in normal valves but was present in all valves with aortic sclerosis or stenosis lesions. ACE was detected on a subset of lesion macrophages but was present primarily in an extracellular distribution, where it colocalized with apolipoprotein B. ACE was detected by Western blotting on plasma LDL but not on HDL isolated from normal volunteers. Angiotensin II, the enzymatic product of ACE, was colocalized with ACE in valve lesions. ACE also was colocalized with apolipoprotein B in an adjacent coronary atherosclerotic plaque. Conclusions-ACE is present in aortic sclerosis and stenosis lesions, where it may participate in lesion development, as is evidenced by the presence of its enzymatic product, angiotensin II. The observation of an association of ACE with LDL in both lesions and plasma suggests that LDL may deliver ACE to lesions and has implications for the role of ACE-containing LDL in other diseases, such as atherosclerosis.

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Abstract-Lipoprotein retention in the vascular extracellular matrix (ECM) plays a critical role in atherogenesis. Previous studies demonstrated the presence of apo A-I and E in atherosclerotic lesions, suggesting that HDL may be trapped by the artery wall. We sought to determine mechanisms by which HDL could be bound and retained by the arterial wall, and whether apo E was a principal determinant of this binding. We evaluated in situ accumulation of fluorescently labeled DiI-human HDLϮapo E in perfused carotid arteries from apo E-null mice. Apo E was important in mediating HDL binding to the vascular wall, with a 48Ϯ16% increase in accumulation of DiI-labeled apo E-containing HDL (HDL 3 ϩE) compared with DiI-apo E-free HDL (HDL 3 ϪE) (Pϭ0.003). To investigate possible mechanisms responsible for retention, we assessed binding of unlabeled HDL 3 ϪE and HDL 3 ϩE to ECM generated by cultured arterial smooth muscle cells. Similar to the in situ carotid artery data, HDL 3 ϩE bound better to the ECM than did HDL 3 ϪE (3-fold lower K a and 3.5-fold higher B max for HDL 3 ϩE versus HDL 3 ϪE). These differences were eliminated after either neutralization of arginine residues on apo E or digestion of matrix with chondroitin ABC lyase, suggesting that chondroitin and/or dermatan sulfate proteoglycans were responsible for apo E-mediated increased binding. These findings demonstrate that HDL can bind to both intact murine carotid arteries and smooth muscle cell-derived ECM, and that apo E is a principal determinant in mediating the ability of HDL to be trapped and retained via its interaction with ECM proteoglycans. Key Words: apolipoprotein E Ⅲ high-density lipoproteins Ⅲ extracellular matrix Ⅲ artery wall Ⅲ proteoglycan T he retention of lipoproteins in the arterial extracellular matrix (ECM) is a critical step in atherogenesis. 1-3 Apolipoproteins (apo) B and E, found on triglyceride-rich, atherogenic lipoproteins, have been found extensively in atherosclerotic lesions, often associated with ECM proteoglycans. 4 However, immunohistochemical studies show that apo A-I, which is the primary apolipoprotein on HDL, is present in human atherosclerotic lesions as well. 4 -7 Further, apo A-I and E were found to colocalize with biglycan, 4 a major arterial extracellular proteoglycan, suggesting that HDL can be trapped by ECM proteoglycans. We also have shown that apo E-containing HDL (HDLϩE) binds to purified matrix proteoglycans in vitro, whereas apo E-free HDL (HDLϪE) does not bind. 4,8 Because HDLϪE contains a large amount of apo A-I, this suggests that apo A-I itself does not bind to proteoglycans. This hypothesis is supported by reports that mammalian apo A-I does not contain heparin binding domains. 9 Therefore, these data indicate that apo E may play an important role in the retention of HDL by ECM proteoglycans.Because the artery wall contains ECM components in addition to proteoglycans, we conducted studies to evaluate the hypothesis that the presence of apo E on HDL could affect its retention by the intact vascular ECM. ...