Objective-Whereas growth factors, via their ability to stimulate vascular smooth muscle cell (VSMC) proliferation and migration, have been thought to play a permissive role in atherosclerosis initiation and progression, the role of insulin-like growth factor-1 (IGF-1) is unknown. Here we report for the first time that IGF-1 infusion decreased atherosclerotic plaque progression in ApoE-deficient mice on a Western diet. Methods and Results-ApoE-null mice (8 weeks) were infused with vehicle or recombinant human IGF-1 and fed a high-fat diet for 12 weeks. Analysis of aortic sinuses revealed that IGF-1 infusion decreased atherosclerotic plaque progression and macrophage infiltration into lesions. Furthermore, IGF-1 decreased vascular expression of the proinflammatory cytokines interleukin-6 and tumor necrosis factor-␣, reduced aortic superoxide formation and urinary 8-isoprostane levels, and increased aortic pAkt and eNOS expression and circulating endothelial progenitor cells, consistent with an antiinflammatory, antioxidant, and prorepair effect on the vasculature. Conclusions-Our data indicate that an increase in circulating IGF-1 reduces vascular inflammatory responses, systemic and vascular oxidant stress and decreases atherosclerotic plaque progression. These findings have major implications for the treatment of atherosclerosis. Key words: insulin-like growth factor Ⅲ atherosclerosis Ⅲ apolipoprotein E Ⅲ inflammatory response Ⅲ oxidative stress A therosclerosis is the principal underlying cause of most cardiovascular disease-related deaths, the leading cause of mortality in the USA. 1 Long considered to result from progressive vascular lipid accumulation, atherosclerosis is now recognized as a chronic inflammatory disease, 2-5 in which oxidative stress plays a key initiating role. Thus, multiple oxidative stimuli including oxidation of low-density lipoprotein (LDL) in the subendothelial space can result in endothelial cell adhesion molecule expression, monocyte and T cell recruitment, and macrophage lipid accumulation and foam cell formation. 6 Growth factors, cytokines, chemokines, and proteases modulate many steps in the atherosclerotic process including the migration and proliferation of smooth muscle cells. Furthermore, vascular and extravascular progenitor cells contribute to atherogenesis, and these include circulating endothelial progenitor cells (EPCs) that are likely part of a vascular repair system. 7 Insulin-like growth factor-1 (IGF-1) is an endocrine and autocrine/paracrine growth factor that is the primary mediator of the effect of growth hormone (GH) on developmental growth. 8 IGF-1 is expressed in vascular cells 9 and in monocyte/macrophages, 10,11 but its role in atherogenesis is unknown. IGF-1 is a mitogen for endothelial cells 12 and vascular smooth muscle cells, 13 and can potentiate endothelial cell TNF-␣-induced c-Jun and NF-B activation. 14 Furthermore, IGF-1 has potent survival effects on vascular cells and prevents oxidized LDL-induced apoptosis of vascular smooth muscle cells. 15 We ha...

Insulin-like growth factor 1 (IGF-1) is an endocrine and autocrine/paracrine growth factor that circulates at high levels in the plasma and is expressed in most cell types. IGF-1 has major effects on development, cell growth and differentiation, and tissue repair. Recent evidence indicates that IGF-1 reduces atherosclerosis burden and improves features of atherosclerotic plaque stability in animal models. Potential mechanisms for this atheroprotective effect include IGF-1-induced reduction in oxidative stress, cell apoptosis, proinflammatory signaling, and endothelial dysfunction. Aging is associated with increased vascular oxidative stress and vascular disease, suggesting that IGF-1 may exert salutary effects on vascular aging processes. In this review, we will provide a comprehensive update on IGF-1's ability to modulate vascular oxidative stress and to limit atherogenesis and the vascular complications of aging.

Background We have previously shown that systemic infusion of insulin-like growth factor-1 (IGF-1) exerts anti-inflammatory and anti-oxidant effects and reduces atherosclerotic burden in apolipoprotein E (Apoe) deficient mice. Monocytes/macrophages express high levels of IGF-1 receptor (IGF1R) and play a pivotal role in atherogenesis but the potential effects of IGF-1 on their function are unknown. Methods and Results To determine mechanisms whereby IGF-1 reduces atherosclerosis and to explore the potential involvement of monocytes/macrophages, we created monocyte/ macrophage specific IGF1R knockout (MΦ-IGF1R-KO) mice on Apoe−/− background. We assessed atherosclerotic burden, plaque features of stability, and monocyte recruitment to atherosclerotic lesions. Phenotypic changes of IGF1R-deficient macrophages were investigated in culture. MΦ-IGF1R-KO significantly increased atherosclerotic lesion formation, as assessed by Oil-red-O staining of en face aortae and aortic root cross-sections, and changed plaque composition to a less stable phenotype, characterized by increased macrophage and decreased α-smooth muscle actin-positive cell population, fibrous cap thinning, and decreased collagen content. Brachiocephalic artery lesions of MΦ-IGF1R-KO mice had histological features implying plaque vulnerability. Macrophages isolated from MΦ-IGF1R-KO mice showed enhanced proinflammatory responses upon stimulation by IFNγ and oxidized LDL and elevated antioxidant gene expression levels. Moreover, IGF1R deficient macrophages had decreased expression of ABCA1 and ABCG1 and reduced lipid efflux. Conclusions Our data indicate that macrophage IGF1R signaling suppresses macrophage and foam cell accumulation in lesions and reduces plaque vulnerability, providing a novel mechanism whereby IGF-1 exerts anti-atherogenic effects.

The largest subgroup of integrins is that containing the ␤1 subunit. ␤1 integrins have been implicated in a wide array of biological processes ranging from adhesion to cell growth, organogenesis, and mechanotransduction. Global deletion of ␤1 integrin expression results in embryonic death at ca. embryonic day 5 (E5), a developmental time point too early to determine the effects of this integrin on vascular development. To elucidate the specific role of ␤1 integrin in the vasculature, we conditionally deleted the ␤1 gene in the endothelium. Homozygous deletion of ␤1 integrins in the endothelium resulted in failure of normal vascular patterning, severe fetal growth retardation, and embryonic death at E9.5 to 10, although there were no overt effects on vasculogenesis. Heterozygous endothelial ␤1 gene deletion did not diminish fetal or postnatal survival, but it reduced ␤1 subunit expression in endothelial cells from adult mice by approximately 40%. These mice demonstrated abnormal vascular remodeling in response to experimentally altered in vivo blood flow and diminished vascularization in healing wounds. These data demonstrate that endothelial expression of ␤1 integrin is required for developmental vascular patterning and that endothelial ␤1 gene dosing has significant functional effects on vascular remodeling in the adult. Understanding how ␤1 integrin expression is modulated may have significant clinical importance.The largest subgroup of integrins is that containing the ␤1 subunit (5, 6). ␤1 integrins have been implicated in a wide array of biological processes, including cell migration, adhesion, formation of basement membrane, and control of the cell cycle (5,6,11,13,20,25,31). They have also been purported to play an essential role in angiogenesis, although the data supporting this role have been obtained in an indirect manner using techniques such as blocking antibodies or RNA interference (15,22,30). The role of ␤1 integrin in angiogenesis has also been inferred by the finding that tumor formation by ␤1 null cells is defective, with absence of tumor vascularization (4). Further, ␤1 integrin expression is highly upregulated in the central nervous system vasculature during maturation, and it has thus been proposed that this integrin plays an important role in maintaining the integrity of the mature central nervous system vasculature (26). Interestingly, cerebral cavernous malformations (CCMs) have been linked to defects in the KRIT-1 gene, the protein product of which interacts with the integrin cytoplasmic domain-associated protein 1 alpha (ICAP-1␣) (24,(36)(37)(38). ICAP-1␣ directly binds the cytoplasmic tail of ␤1 integrin. Further, the ICAP-1␣ binding sites for both KRIT-1 and ␤1 integrin are similar, and both contain a crucial NPXY motif (36). It has thus been postulated that KRIT-1 and ␤1 integrin compete for binding to ICAP-1␣ and that this competition modulates ␤1 integrin signaling and function. Therefore, the vascular defects observed in patients with KRIT-1 gene mutations may involve abnormal ␤1 i...

Angiotensin-converting enzyme (ACE) is a zinc-containing dipeptidyl carboxypeptidase that catalyzes the conversion of angiotensin I to the potent vasoconstrictor angiotensin II. By analyzing cDNA and genomic DNA, we have constructed a consensus sequence encoding the testis isozyme of mouse ACE. Testis ACE cDNA contains 2,435 base pairs and encodes a protein of 732 amino acids. The N-terminal 66 amino acids are unique to the testis isozyme, while the remaining 666 are identical to the carboxyl half of mouse somatic ACE. The overall conservation of amino acid sequence between the testis isozymes of the mouse, rabbit, and human is 78 to 84%. The conservation of amino acids for the N-terminal domain uniquely expressed within the testis is 63 to 67% between these species. Primer extension and RNase protection experiments show that RNA transcription of the testis ACE isozyme begins 16 or 17 bases upstream from the translation start site. A sequence element resembling a TATA box is found 25 bases 5' of the transcription start site. To create its unique isozyme of ACE, the testis begins mRNA transcription in the middle of the exonic-intronic structure of somatic ACE, within a sequence treated as an intron by somatic tissues. Testis ACE is not the result of alternative RNA splicing but seems due to the start of transcription at a unique site within the ACE gene.Angiotensin-converting enzyme (ACE) (EC 3.4.15.1) is a single-chain peptidase (a dipeptidyl carboxypeptidase) containing 1 molar equivalent of Zn (29). It functions to convert angiotensin I to the potent vasoconstrictor angiotensin II (13). However, ACE is rather nonspecific, and other peptides such as bradykinin have been implicated as potential substrates (35). There are two isozymes of ACE, one produced within the testis by developing sperm and the other produced by many somatic tissues, including vascular endothelium, renal tubular epithelium, ciliated gut epithelium, stimulated macrophages, and areas of the brain (12,25). Recently, the complete amino acid sequence of the somatic form of mouse and human ACE has been determined from cloned cDNA (3,4,30). These studies show that ACE is encoded by a single gene and is a protein containing two highly homologous internal domains, each roughly half the size of the parent molecule.The production of ACE by the testis is unique in that this organ expresses two isozymes that differ in their molecular masses, N-terminal amino acids, and patterns of expression (10, 11). The larger protein (150 kilodaltons [kDa] in the mouse) is identical to the enzyme found in somatic tissues and is thought to be the product of vascular endothelium and perhaps other nonspermatogenic tissues within the testis (10, 11). The smaller protein (92 kDa in the mouse) appears unique to the testis. For convenience, we will refer to the testis-specific isozyme of ACE as testis ACE. Prepubertal animals do not express testis ACE, and immunohistochemical staining has localized the enzyme to spermatid heads and the lumen of seminiferous tubules in stag...

Background:The la ribonucleoprotein domain family member 6, LARP6, regulates collagen type 1 mRNA translation. Results: IGF-1 increases LARP6 expression, resulting in increased LARP6-collagen type 1 mRNA complex and collagen synthesis in smooth muscle. Conclusion: IGF-1 enhances collagen fibrillogenesis via induction of LARP6. Significance: This report uncovers a critical mechanism whereby IGF-1 induces a more stable plaque phenotype in atherosclerosis.

Atherosclerosis is a chronic inflammatory disease in which early endothelial dysfunction and subintimal modified lipoprotein deposition progress to complex, advanced lesions that are predisposed to erosion, rupture and thrombosis. Oxidative stress plays a critical role not only in initial lesion formation but also in lesion progression and destabilization. While growth factors are thought to promote vascular smooth muscle cell proliferation and migration, thereby increasing neointima, recent animal studies indicate that IGF-1 exerts pleiotropic anti-oxidant effects along with anti-inflammatory effects that together reduce atherosclerotic burden. This review discusses the effects of IGF-1 in vascular injury and atherosclerosis models, emphasizing the relationship between oxidative stress and potential atheroprotective actions of IGF-1.

Objective-Growth factors may play a permissive role in atherosclerosis initiation and progression, in part via their promotion of vascular smooth muscle cell (VSMC) accumulation in plaques. However, unstable human plaques often have a relative paucity of VSMC, which has been suggested to contribute to plaque rupture and erosion and to clinical events. Insulin-like growth factor-1 (IGF-1) is an endocrine and autocrine/paracrine growth factor that is a mitogen for VSMC, but when infused into Apoe Ϫ/Ϫ mice it paradoxically reduces atherosclerosis burden. Methods and Results-To determine the effect of stimulation of VSMC growth on atherosclerotic plaque development and to understand mechanisms of IGF-1's atheroprotective effect, we assessed atherosclerotic plaques in mice overexpressing IGF-1 in smooth muscle cells (SMC) under the control of the ␣-smooth muscle actin promoter, after backcrossing to the Apoe Ϫ/Ϫ background (SMP8/Apoe Ϫ/Ϫ ). Compared with Apoe Ϫ/Ϫ mice, these SMP8/Apoe Ϫ/Ϫ mice developed a comparable plaque burden after 12 weeks on a Western diet, suggesting that the ability of increased circulating IGF-1 to reduce plaque burden was mediated in large part via non-SMC target cells. However, advanced plaques in SMP8/Apoe Ϫ/Ϫ mice displayed several features of plaque stability, including increased fibrous cap area, ␣-smooth muscle actin-positive SMC and collagen content, and reduced necrotic cores. Traditionally, the role of growth factors in atherosclerosis has been thought to be permissive and to promote neointima formation. 6 -8 Insulin-like growth factor-1 (IGF-1) is an endocrine and autocrine/paracrine growth factor that exerts pleiotropic effects on cells involved in atherogenesis. 9 Notably, it has mitogenic and antiapoptotic actions on endothelial cells and VSMC and stimulates VSMC migration. 10 Consistent with the role of IGF-1 as a VSMC mitogen, most [11][12][13][14] but not all 15 studies using inhibitors have demonstrated that reduced VSMC IGF-1 signaling correlates with decreased neointimal responses to mechanical arterial injury. Furthermore, targeted overexpression of IGF-1 in SMC increases neointimal formation. 16 However, although mechanical arterial injury models provide much information about the restenotic process, they have significant limitations when addressing mechanisms of atherogenesis, and the role of VSMC IGF-1 in the latter process remains unclear. Conclusion-TheseWe have previously shown that oxidized low-density lipoprotein downregulates IGF-1 and IGF-1 receptor expression in VSMC 17,18 and that expression of IGF-1 and IGF-1 receptor is reduced in areas of advanced human plaque staining positive for oxidized low-density lipoprotein. 19 These findings suggest that decreased IGF-1 activity could contribute to the atherosclerotic process and notably to depletion of VSMC in advanced unstable plaque. Similar findings have been reported in cultured plaque-derived VSMC. 20 We recently reported that infusion of recombinant human IGF-1 into Apoe-deficient mice on a Western diet for ...