No abstract

One of the major conceptual advances in our understanding of the pathogenesis of age-associated cardiovascular diseases has been the insight that age-related oxidative stress may promote vascular inflammation even in the absence of traditional risk factors associated with atherogenesis (e.g., hypertension or metabolic diseases). In the present review we summarize recent experimental data suggesting that mitochondrial production of reactive oxygen species, innate immunity, the local TNF-alpha-converting enzyme (TACE)-TNF-alpha, and the renin-angiotensin system may underlie NF-kappaB induction and endothelial activation in aged arteries. The theme that emerges from this review is that multiple proinflammatory pathways converge on NF-kappaB in the aged arterial wall, and that the transcriptional activity of NF-kappaB is regulated by multiple nuclear factors during aging, including nuclear enzymes poly(ADP-ribose) polymerase (PARP-1) and SIRT-1. We also discuss the possibility that nucleophosmin (NPM or nuclear phosphoprotein B23), a known modulator of the cellular oxidative stress response, may also regulate NF-kappaB activity in endothelial cells.

Objective-To study matrix metalloproteinase 2 (MMP-2) effects on transforming growth factor-␤1 (TGF-␤1) activation status and downstream signaling during arterial aging. Methods and Results-Western blotting and immunostaining showed that latent and activated TGF-␤1 are markedly increased within the aorta of aged Fisher 344 cross-bred Brown Norway (30 months of age) rats compared with adult (8 months of age) rats. Aortic TGF-␤1-type II receptor (T␤RII), its downstream molecules p-similar to mad-mother against decapentaplegic (SMAD)2/3 and SMAD4, fibronectin, and collagen also increased with age. Moreover, TGF-␤1 staining is colocalized with that of activated MMP-2 within the aged arterial wall and vascular smooth muscle cell (VSMC) in vitro, and this physical association was confirmed by coimmunoprecipitation. Incubation of young aortic rings ex vivo or VSMCs in vitro with activated MMP-2 enhanced active TGF-␤1, collagen, and fibronectin expression to the level of untreated old counterparts, and this effect was abolished via inhibitors of MMP-2. Interestingly, in old untreated rings or VSMCs, the increased TGF-␤1, fibronectin, and collagen were also substantially reduced by inhibition of MMP-2. Conclusions-Active TGF-␤1, its receptor, and receptor-mediated signaling increase within the aortic wall with aging.TGF-␤1 activation is dependent, in part at least, by a concomitant age-associated increase in MMP-2 activity. Thus, MMP-2-activated TGF-␤1, and subsequently T␤RII signaling, is a novel molecular mechanism for arterial aging.

Increased angiotensin II (Ang II), matrix metalloproteinase type II (MMP2), and sympathetic activity accompany age-associated arterial remodeling. To analyze this relationship, we infused a low subpressor dose of Ang II into young (8 months old) rats. This increased carotid arterial MMP2 transcription, translation, and activation, as well as transforming growth factor-␤1 activity and collagen deposition. A higher Ang II concentration, which increased arterial pressure to that of old (30 months old) untreated rats, produced carotid media thickening and intima infiltration by vascular smooth muscle cells (VSMCs). Ex vivo, Ang II increased MMP2 activity in carotid rings from young rats to that of untreated old rats. Ang II also increased the ability of early passage VSMCs from young rats to invade a synthetic basement membrane, similar to that of untreated VSMCs from old rats. The MMP inhibitor GM6001 and the AT 1 receptor antagonist Losartan inhibited these effects. The ␣-adrenoreceptor agonist phenylephrine increased arterial Ang II protein, causing MMP2 activation and intima and media thickening. Exposure of young VSMCs to phenylephrine in vitro increased Ang II protein and MMP2 activity to the levels of old VSMCs; Losartan abolished these effects. Thus, Ang II-induced effects on MMP2, transforming growth factor-␤1, collagen, and VSMCs are central to the arterial remodeling that accompanies advancing age. (Am J Pathol 2005,

Abstract-Studies in animal models demonstrate that angiotensin II and its downstream signaling molecules, that is, matrix metalloproteinases and monocyte chemoattractant protein-1, increase within the diffusely thickened intima of central arteries with aging. Whether such age-related changes occur within the human arterial wall is unknown. We harvested "grossly normal thoracic aortas" from 5 young (20Ϯ3 years) and 5 old white males (65Ϯ6 years) at necropsy, after death from traumatic causes. The intimae of older samples were markedly and diffusely thickened compared with younger intimae and contained increased levels of angiotensin-converting enzyme, angiotensin II, angiotensin II receptor type 1, matrix metalloproteinases 2/9, monocyte chemoattractant protein-1, and collagen I and III proteins. In situ activities of metalloproteinases 2/9 were also significantly enhanced within old, normal aortas. The thickened intima of older aortas also contained a 5-fold increase in the embryonic form of smooth muscle myosin heavy chain-labeled cells than that of younger aortas, and these fetal-type cells were colocalized with angiotensin II protein staining. The ability of isolated smooth muscle cells to invade an artificial basement membrane in response to a monocyte chemoattractant protein-1 gradient increased with age. Furthermore, angiotensin II increased the invasive capacity of young smooth muscle cells, and this effect was reduced by a metalloproteinase inhibitor or an angiotensin II receptor blocker. Thus, in the absence of lipid infiltration, the aged human aortic wall exhibits a proinflammatory profile that renders it a fertile substrate for the development of arterial disease, for example, atherosclerosis and hypertension. (Hypertension. 2007;50:219-227.)

Abstract-To seek evidence that the nonhuman primate arterial wall, as it ages in the absence of atherosclerosis, exhibits alterations in pathways that are involved in the pathogenesis of experimental atherosclerosis, we assessed aortic matrix metalloproteinase-2 (MMP-2) and its regulators, ie, membrane type-1 of matrix metalloproteinase (MT1-MMP) and tissue inhibitor of matrix metalloproteinase-2 (TIMP-2), and the expression of angiotensin II (Ang II), angiotensinconverting enzyme (ACE), and chymase in young (6.4Ϯ0.7 years) and old (20.0Ϯ1.9 years) male monkeys. With advancing age, (1) the intimal thickness increased 3-fold and contained numerous vascular smooth muscle cells and matrix, but no inflammatory cells; (2) the intimal MMP-2 antibody-staining fraction increased by 80% (PϽ0.01); (3) in situ zymography showed that MMP-2 activity, mainly confined to the intima, increased 3-fold (PϽ0.01); (4) the MT1-MMP antibody-staining fraction increased by 150% (PϽ0.001), but the TIMP-2 antibody-staining fraction did not significantly change; (5) steady levels of the mRNA-staining fraction (via in situ hybridization) for MMP-2 increased 7-fold, for MT1-MMP increased 9-fold, and for TIMP-2 increased 2-fold (all PϽ0.001); and (6) intimal Ang II and ACE immunofluorescence were increased 5-fold and 5.6-fold, respectively, and colocalized with MMP-2. Thus, age-associated arterial remodeling and the development and progression of experimental atherosclerosis in young animals share common mechanisms, ie, MMP-2 activation and increased Ang II signaling. This might explain, in part, the dramatically exaggerated prevalence and severity of vascular diseases with aging.

Arterial aging is the major contributing factor to increases in the incidence and prevalence of cardiovascular disease, due mainly to the presence of chronic, low-grade, “sterile” arterial inflammation. Inflammatory signaling driven by the angiotensin II cascade perpetrates adverse age-associated arterial structural and functional remodeling. The aged artery is characterized by endothelial disruption, enhanced vascular smooth muscle cell migration and proliferation, extracellular matrix deposition, elastin fracture, and matrix calcification/amyloidosis/glycation. Importantly, the molecular mechanisms of arterial aging are also relevant to the pathogenesis of hypertension, and atherosclerosis. Age-associated arterial proinflammation is, to some extent, mutable, and interventions to suppress or delay it may have the potential to ameliorate or retard age-associated arterial diseases.

Objective-With age, rat arterial walls thicken and vascular smooth muscle cells (VSMCs) exhibit enhanced migration and proliferation. Monocyte chemotactic protein-1 (MCP-1) affects these VSMC properties in vitro. Because arterial angiotensin II, which induces MCP-1 expression, increases with age, we hypothesized that aortic MCP-1 and its receptor CCR2 are also upregulated and affect VSMC properties. Methods and Results-Both MCP-1 and CCR2 mRNAs and proteins increased in old (30-month) versus young (8-month) F344ϫBN rat aortas in vivo. Cellular MCP-1 and CCR2 staining colocalized with that of ␣-smooth muscle actin in the thickened aortas of old rats and were expressed by early-passage VSMCs isolated from old aortas, which, relative to young VSMCs, exhibited increased invasion, and the age difference was abolished by vCCI, an inhibitor of CCR2 signaling. MCP-1 treatment of young VSMCs induced migration and increased their ability to invade a synthetic basement membrane. The MCP-1-dependent VSMC invasiveness was blocked by vCCI. After MCP-1 treatment, migration and invasion capacities of VSMCs from young aortas no longer differed from those of VSMCs isolated from older rats. Key Words: chemokines Ⅲ aging Ⅲ aorta Ⅲ vascular smooth muscle cells Ⅲ invasion A ging is the major risk factor for the development of atherosclerosis and hypertension, which have an important impact on Western society because they lead to myocardial infarction, stroke, and heart failure. 1-3 Arterial remodeling accompanies advancing age in all species, from rodents to nonhuman primates to humans, and the mechanisms involved in this remodeling likely confer on aging the status of the major risk factor for vascular diseases. 1,2 Specific facets of age-associated remodeling include luminal dilation, thickening of the intimal and medial layers with cellular and extracellular matrix reorganization, increased stiffness, and endothelial dysfunction. 4 -6 Studies from our laboratory and others have shown that diffuse intimal thickening with aging is characterized by accumulation of fibronectin, collagen, and vascular smooth muscle cells (VSMCs), with an increase in matrix metalloproteinase (MMP)-2 and angiotensin II (Ang II) expression. [7][8][9][10][11] In addition, the expression of aortic intracellular adhesion molecule and transforming growth factor-␤1 markedly increases with age, and these molecules localize to MMP-2-staining positive areas. 12 Arterial aging is also characterized by an increase in NAD(P)H oxidase activity and reactive oxygen species production and a reduction in nitric oxide (NO) bioavailability. [13][14][15][16] In addition, increased arterial levels of proinflammatory cytokines, such as tumor necrosis factor-␣ and interleukin-6, among others, accompany aging. [17][18][19][20] Structural and biochemical changes that occur within large arteries with aging are accompanied by a shift of the VSMC phenotype from the "contractile" to the "synthetic" state, characterized by an increased proliferative and migratory responsiveness to...