| BackgroundAtherosclerosis formation and plaque rupture are the common pathological basis of most of cardio-cerebrovascular diseases. Understanding the molecular and cellular mechanisms that lead to the development of atherosclerosis is critical for identifying strategies to limit disease progression before it leads to clinical consequences. Studies have revealed that many different cell types, including macrophages, lymphocytes, endothelial cells and smooth muscle cells (SMCs), are involved in atherosclerotic lesion formation. Most reviews on atherosclerosis focus on the role of endothelial and inflammatory cells in the initiation of atherosclerosis and discuss SMCs largely in the context of late atherosclerosis when they migrate into the neointima and secrete matrixproteins to stabilize the plaque. Histological studies of autopsy specimens of human coronary arteries ranging from infants to adults provide evidence that regions prone to the development of atherosclerosis contain abundant SMCs whereas regions that are more resistant to atherosclerosis contain few, raising the interesting question: Do SMCs in areas of intimal thickening play a pathogenic role in the increased development of atherosclerosis at these sites? Recent studies, coupled with significant in vitro mechanistic data and in vivo correlative data reviewed herein, provide strong evidence that SMCs proliferation and migration are indeed important in early atherogenesis. Present evidence suggests that intimal SMCs differ significantly from medial SMCs and as such may have unique atherogenic properties that make them fertile ground for the initiation of plaques. Arterial medial SMCs predominantly express proteins involved in the contractile function and quiet, and could swith to the "synthetic" phenotypic states in response to a variety of atherogenic stimuli, while the latter migrate and proliferate more readily than "contractile" SMCs, leading to the atherogenesis.Renin-angiotensin system (RAS) plays a critical role in VSMCs proliferation, migration and atherogenesis and atherosclerosis acceleration. In particular, confirmed data have shown that angiotensinⅡ(AngⅡ) contributed a lot to VSMCs proliferative activity and aterial remodeling. Angiotensin-(1-7) [Ang-(1-7)], a new member of RAS family, was mostly produced by angiotensin converting enzyme-2 (ACE2) activity. Recent studies have found Ang-(1-7) plays an important protective role in various pathophysiological processes, including cardiac remodeling and fibrosis, hypertensive vacular remodeling, inflammation and regulation of endothelial cells function, through antagonizing AngⅡ. Previous studies have shown that Ang-(1-7) inhibited VSMCs proliferation in a balloon-injured model, however, the mechanism of VSMCs proliferation and migration is still to be elucidated.Mitogen-activated protein kinases (MAPKs) play a pivotal role in AngⅡ-mediated VSMCs proliferative activity, and recent studies found that SM22a-knockout promoted VSMCs proliferation and migration, and atherosclerotic process.To sum up, we hypothesize that Ang-(1-7) can effectively inhibit AngⅡ-induced VSMCs proliferation and migration, through blocking the MAPKs and SM22a signaling pathway crosstalk, and to regulate VSMCs bioactivities.According to the questions mentioned above, our present study was designed to test our hypothesis in vitro.Aims1. To test the effect of Ang-(1-7) on AngⅡ-induced VSMCs proliferation and migration. 2. To test the effect of Ang-(1-7) on AngⅡ-induced VSMCs proliferation and migration-related signaling proteins, such as ERK1/2, P38, JNK1/2 and SM22a.3. To clarify the signaling pathway connection and cross-talk in the VSMCs proliferation and migration.Methods1. Cell cultureThe human VSMCs were purchased from Sciencell Company and cultured by the routine methods. When cells confluence reached 80-90%, the cells were removed again and those at passage 4-8 were selected for the study.2. Experimental groupingIn order to inspect the effects of AngⅡand Ang-(1-7) on VSMCs poliferative activity and the related signaling proteins, VSMCs were divided into six groups: control group, AngⅡgroup, Ang-(1-7) group, AngⅡ+Ang-(1-7) group, AngⅡ+Ang-(1-7)+A779 group, A779 group, PD98059 group, SB203580 group, PD98059+SB203580 group and SM22 a knockdown group. In VSMCs culture medium, the concentration of AngⅡ, Ang-(1-7) and A779 were 0.1μM, 1μM and 1μM, respectively.3. MTT and BrdU incorporation assayThese experiments were undertaken to compare VSMCs reproductive activities after AngⅡ, Ang-(1-7), A779, PD98059, SB203580, PD98059+SB203580 and SM22 a knockdown intervention for 24h.4. Transwell plate assayThe transwell plate assay was undertaken to detect VSMCs migration acivity after AngⅡ,Ang-(1-7), A779, PD98059, SB203580, PD98059+SB203580 and SM22 a knockdown intervention for 12h.5. RT-PCRRT-PCR was taken to explore some cytokines mRNA expression in VSMCs after AngⅡand Ang-(1-7) intervention.6. Western blotTo observe the expression of some cytokines in VSMCs after AngⅡ,Ang-(1-7), A779, PD98059, SB203580, PD98059+SB203580 and SM22 a knockdown intervention, Western blot was undertaken. The relative expression of protein was demonstrated by the ratio of integral optical density (IOD) value between target protein andβ-actin.Results1. Ang-(1-7) regulating AngⅡ-mediated VSMCs proliferative activity:Both MTT assay and BrdU incorporation assay confirmed that Ang II could effectively increase the proliferative activity, while Ang-(1-7) could remarkably abolish the efficacy of AngⅡ. Meanwhile, A779 could effectively reverse the effect of Ang-(1-7).2. Effect of Ang-(1-7) on AngⅡ-induced VSMCs migration:Transwell plate assay showed that, compared with the control group, Ang II could effectively increase the migration of VSMCs. Similar with the effect of Ang-(1-7) on VSMCs proliferation, Ang-(1-7) could remarkably reverse the effect of AngⅡ, while A779 effectivley abolish the effect of Ang-(1-7).3. The study of VSMCs proliferation signal transduction pathway:The activities of ERK1/2 and P38 were significantly higher in Ang II group than control group, and Ang-(1-7) treatment effectively decrease the efficacy of AngⅡ, while A779 could effectively abolish the effect of Ang-(1-7). The activity of JNK1/2 significantly increased after AngⅡtreatment, however, no significant change was found with Ang-(1-7) incubation. There were less mRNA and protein expression of SM22a in AngⅡgroup than control group. There were more mRNA and protein expression of SM22a in Ang-(1-7) group than control group, while A779 could effectively abolish the effect of Ang-(1-7).4. Crosstalk between MAPKs and SM22a pathway:Both the proliferative and migrative activities significantly decreased in PD98059 group, SB203580 group and PD98059+SB203580 group, and remarkably increased in SM22a knockdown group. On the other hand, the mRNA and protein expression of SM22a in PD98059 group, SB203580 group and PD98059+SB203580 group were significantly higher than the control group. Conclusion:1. AngⅡcan enhance the VSMCs activities of proliferation and migration, while Ang-(1-7) can effectively inhibit these functions.2.AngⅡcan activate ERK1/2, P38, and JNK1/2 and inhibit SM22a, however, its effects on ERK1/2, P38 and SM22a were significantly abolished by Ang-(1-7).3. The Ang-(1-7)-MAS-ERK/P38-SM22a signaling pathway might play an important role in regulating the VSMCs bioactivities. 1. BackgroundAtherosclerosis formation and progression is the common pathological basis of the most cardio-cerebrovascular diseases. Accumulating evidence indicates that the rennin-angiotensin system (RAS) plays an important role in vascular smooth muscle cell (VSMCs) proliferation, migration and atherosclerosis formation and progression. AngiotensinⅡ(AngⅡ), the main peptide of RAS, is a key cytokine that regulates cell growth, migration, inflammation and oxidative stress and therefore contributed to VSMCs proliferation, migration and atherosclerosis formation and progression.. Angiotensin-(1-7), a new member of RAS, is mainly produced by angiotension converting enzyme (ACE2), through degradating AngⅡ. A large mount of rencent studies demonstrated that Ang-(1-7) was an important endogenous antagonist of AngⅡ, via anti-inflammation and oxidative stress, vascular vasodiation, reduction of fibrosis and phosphatases activation, and thus played an important role in cardiac reconstruction and fibrosis, hypertensive vascular reconstruction, VSMCs proliferation and migration, and endothelial cells function regulation.Previous studies demonstrated that Ang-(1-7) significantly inhibited VSMCs proliferation and migration in a balloon-injured arterty model, and our rencent study showed that overexpression ACE2 inhibted early AS formation. All the above studies indicated that Ang-(1-7) might play a critical role in AS progression. So, chronic injection of Ang-(1-7) in ApoE-/-mice might attenuate over-activited AngⅡeffectively, and thus inhibit AS formation.The effect of Ang-(1-7) on AS formation and the possible mechanism is unclear. Our in vitro study demonstrated that Ang-(1-7) regulated VSMCs ERK1/2, P38 and SM22 a activities and expression, which play a critical role in AS formation and progression. However, whether Ang-(1-7) could regulate the above signaling protein and AS progression in vivo is unknown.So, we raise the following hypothesis:Ang-(1-7) might regulate the MAS-ERK/P38-SM22 a signaling pathway and inhibit AS formation in ApoE-/-mice.2. Objectives2.1 To assess the therapeutic effects of Ang-(1-7) on atherosclerotic lesions formation in ApoE-/-mice.2.2 To elucidate the molecular mechanisms of Ang-(1-7)-mediated therapeutic effects on atherosclerotic lesions.3. Methods3.1 Atherosclerosis animal modelApoE-/-mice aged 8 weeks were fed an atherogenic chow (0.25% cholesterol and 15% cocoa butter). After 8 weeks for atherogenic chow, they were fasted overnight and killed by euthanasia.3.2 Ang-(1-7) treatmentAt the end of week 4 for atherogenic diet, the mice were randomly allocated to phosphate buffered saline (PBS) control group, Ang-(1-7) group and A779 group. All the animals were implanted with a subcutaneous osmotic pump for the duration of 4 weeks. In order to demonstrate the dose-response in mice, the Ang-(1-7) and A779 groups were divided three subgroups(100ng.kg-1.min-1,200ng.kg-1.min-1 and 400ng.kg-1.min-1), respectively.The mice were anesthetized and their aortas were collected for pathological and biochemical analysis at the end of experiment.3.3 Serum lipid measurementSerum samples were collected after 12 hours of fasting and stored at-80℃. Total cholesterol, Triglyceride, high density lipoprotein cholesterol and low density lipoprotein cholesterol were measured.3.4 Histopathological and immunohistochemical measurementThe whole aortas were stained with Oil Red O, and the cardiac muscles were stained by Masson. SM22ain aortas and MOMA-2 in kidney were identified using appropriate primary antibodies.3.5 Western blot analysisThe protein expression of p-ERK1/2 and p-P38 were analyzed by Western blot measurement.4. Results4.1 Ang-(1-7) protects against atherosclerosisAfter 8 weeks of high fat diet, we found that, with the dose of Ang-(1-7) increasing, the inhibitory action of Ang-(1-7) on atherosclerotic lesions formation was increased. The mice treated with high-dose Ang-(1-7) showed significantly smaller atherosclerotic lesions in their aortas, as shown by both en face (2.58% vs.4.14%, P<0.05) and aortic root section analysis (12.08% vs.20.28%, P<0.05), comparing with the control mice.4.2 The effect of A779 on atherosclerosisAfter 8 weeks of high fat diet, we found that high dose A779 significantly increased both the en face (4.79% vs.4.14%, P<0.05) and aortic root section (23.22% vs.20.28%, P<0.05) atherosclerotic lesions formation, comparing with the control mice.4.3 Ang-(1-7) has no influence on the serum lipid profilesSerum analysis demonstrates that no significant differences were found among the various treatment groups, which indicating the protective effect of Ang-(1-7) on atherosclerosis might independent of lipid profiles.4.4 Ang-(1-7) inhibits the activities of ERK1/2 and P38MAPK in vivoWestern blot analysis showed that the phosphorylations of ERK1/2 and P38MAPK were significantly attenuated by Ang-(1-7) in vivo. Conversely, with the administration of A779, the phosphorylations of ERK1/2 and P38 were significantly increased in the A779 treatment group.4.5 Ang-(1-7) increases the protein expression of SM22aImmunohistochemical staining demonstrated that the protein expression of SM22ain aortas of Ang-(1-7)-treated mice were significantly higher than the PBS control mice (59.57% vs.8.80%, P<0.01). However, no significant difference of protein expression of SM22a between PBS and A779 groups (8.80% vs.7.75%).4.6 Ang-(1-7) has no effect on macrophages infiltration in kidneyImmunofluorescence analysis did not found evident macrophages infiltration in kidney of Ang-(1-7)-treated mice, as well as PBS and A779-treated mice.4.7 Ang-(1-7) has no effect on cardiac muscle collagen contentMasson staining demonstrates no remarkable cardiac fibrosis, and no significant differences of collagen contents among various groups.5. Conclusions1. Chronic Ang-(1-7) treatment significantly attenuated atherosclerotic lesions formation in ApoE-/-mice.2. The anti-atherosclerosis effect of Ang-(1-7) were partly mediated by the effect of anti-proliferation and migration activity on VSMCs through cross-talk of MAS-ERK/P38-SM22a signaling pathways 1. BackgroundRecent studies have shown that plaque rupture and subsequent intraluminal thrombosis is the most common cause of acute coronary syndrome (ACS). Early prevention of plaque rupture might be an effective approach to reduce the risk of ACS. Ruptured plaques have a large necrotic core, a thin fibrous cap depleted of extracellular matrix (collagen and proteoglycans) and smooth muscle cells (SMCs), and infiltration by macrophages and T-cells with outward positive remodeling and increased plaque vascularity. In most patients with ACS, more than one vulnerable plaque are present in their coronary arteries, and thus, systemic drug therapy should be the appropriate treatment. Unfortunately, there is still not an ideal drug for stabilizing atherosclerotic plaques. Although clinical and basic trials have demonstrated the capability of angiotensin converting enzyme inhibitors (ACEI) and angiotensin type I receptor blockers (ARB) in stabilizing vulnerable plaques, a large part of patients experienced an acute coronary event despite an intensive ACEI and ARB therapy. Therefore, exploration of new drugs with high efficacy and low side effects for stabilizing vulnerable plaques is clearly warranted.Atherosclerosis is a chronic inflammatory disease. Although the precise mechanisms of plaque rupture are poorly understood, most researchers agree that the disruption of a fibrous cap rich in macrophages and T-lymphocytes in addition to the necrotic core coming in contact with circulating blood are events that lead to the development of thrombi in fatal plaques.Accumulating evidence demonstates that the rennin-angiotensin system (RAS) contributes a lot to atherogenesis and atherosclerotic progress. AngiotensinⅡ(AngⅡ), the main peptide of RAS, is well proved in atherosclerotic formation and progression. Inhibition of the RAS with angiotensin-converting enzyme (ACE) inhibitors or angiotensin receptor blockers (ARB) is one of the current therapeutic strategies with proven beneficial effects in the treatment of atherosclerosis. However, a large part of patients experienced an acute coronary event despite an intensive ACEI and ARB therapy. Recent studies have shown that, angiotensin converting enzyme 2 (ACE2), Ang-(1-7), new components of the RAS, play important roles in atherosclerosis, cardiac and vascular remodeling. Ang-(1-7) may exert actions that oppose the atherogenic effects of AngⅡ, which plays an important role in cardiovascular pathophysiology. Our previous study also demonstrated that overexpression of ACE2 stabilized vulnerable plaques. However, whether Ang-(1-7) has protective effect against vulnerable atherosclerotic plaques has not been elucidated. In this study, we observed the effect of Ang-(1-7) on atherosclerotic plaque stability in ApoE-/-mice by systemic application.2. Objectives2.1 To assess the therapeutic effects of Ang-(1-7) on atherosclerotic lesions stability in ApoE-/-mice.2.2 To explore the mechanisms of Ang-(1-7)-mediated therapeutic effects on atherosclerotic lesions inflammation infiltration.3. Methods3.1 Atherosclerosis animal modelApoE-/-mice (aged 8 weeks), which were fed an atherogenic chow (0.25% cholesterol and 15%cocoa butter) throughout the experimental procedure, were divided into two group:group 1:atherogenic chow; group 2:atherogenic chow+ Carotid collar placement. After 16 (group 1) or 14 (group 2) weeks for atherogenic chow, they were fasted overnight and killed by euthanasia.3.2 Ang-(1-7) treatment Group 1:At the end of 12 weeks for atherogenic diet, the mice were randomly allocated to phosphate buffered saline (PBS) group, Ang-(1-7) (400ng.kg-1.min-1) group and A779 (400ng.kg-1.min-1)group. All the animals were implanted with a subcutaneous osmotic pump for the duration of 4 weeks.Group 2:At the end of 2 weeks for atherogenic diet, a 2-mm long constricting silastic tube was placed on the right common carotid artery. Eight weeks later, the collars were removed and a subcutaneous osmotic pump was implanted for the duration of 4 weeks. The mice were randomly allocated to phosphate buffered saline (PBS) group, Ang-(1-7) (400ng.kg-1.min-1) group and A779 (400ng.kg-1.min-1) group.3.3 Histopathological and immunohistochemical measurementMice were perfused through the left ventricle with PBS, followed by 4% paraformaldehyde under 100 mmHg pressure. The aortic root and right common carotid artery was excised and immersed in 4%formaldehyde overnight (4℃), embedded in OCT, and stored at-80℃until use. In all of the mice, OCT-embedded aortic root and carotid artery were cross-sectioned into pieces 6μm thick at 50-μm intervals. Sections were stained with hematoxylin and eosin (H&E), and further stained for lipids and collagen using Oil-red O and picrosirius red, respectively. Smooth muscle cells (SMCs, anti-a-actin) and macrophages (MOMA-2), tumor necrosis factor-a(TNF-α), Interleukin-6 (IL-6), Interleukin-1β(IL-1β), macrophage chemoattractant protein-1 (MCP-1), matrix matalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase (TIMP-1) were immunostained with corresponding antibodies. The vulnerability index was calculated by the following formula:(macrophage staining %+lipid staining %)/(smooth muscle cell%+fiber%).3.4 RT-PCRThe mRNA expression levels of TNF-α,IL-6,IL-1βand MCP-1 in macrophages were tested by RT-PCR.4. Results4.1 The effect of Ang-(1-7) on atherosclerotic plaque stability Group 1:Immunohistochemistry analysis indicated that plaques from Ang-(1-7)-injected ApoE-knockout mice showed increased percentages of collagen (18.5%) and smooth muscle cells (66.9%), and reduced percentages of macrophages (29.9%) and lipid area (23.7%) compared with those treated with PBS. Accordingly, the histological plaque vulnerability index was also decreased 46.1% by Ang-(1-7) treatment.Group 2:Immunohistochemistry analysis indicated that plaques from carotid arteries in Ang-(1-7)-injected ApoE-knockout mice showed increased percentages of collagen (18.2%) and smooth muscle cells (60.0%), and reduced percentages of macrophages (42.1%) and lipid area (27.8%) compared with those treated with PBS. Accordingly, the histological plaque vulnerability index was also decreased 48.1%by Ang-(1-7) treatment.4.2 The effect of Ang-(1-7) on inflammatory cytokines in plaquesImmunohistochemical analysis indicated the protein expression levels of IL-6, IL-1β, TNF-αand MCP-1 in atherosclerotic plaques of aortic roots were significantly decreased by Ang-(1-7) injection in ApoE-/-mice.4.3 The effect of Ang-(1-7) on MMPs expression in atherosclerotic plaquesImmunohistochemical analysis demonstrated that the differences of protein expression from aortic root plaques between PBS and Ang-(1-7) subgroups were significant for both MMP-2 (25.1%versus 20.2%, p<0.05) and MMP-9 (17.2% versus 11.3%,p<0.05) staining.4.4 The effect of Ang-(1-7) on AngⅡ-mediated proinflammatory cytokines expression in macrophagesThe results in vitro demonstrated that the mRNA expression levels of IL-6, IL-1β, TNF-αand MCP-1 in RAW264.7 mouse macrophage were significantly increased after incubation with AngⅡcompared to control group, and this up-regulation was significantly reversed by Ang-(1-7), while A779 counteracted the effect of Ang-(1-7).5. Conclusions1. Chronic Ang-(1-7) treatment significantly attenuated lipids and macrophages infiltration, and increased the contents of smooth muscle cells and collagen in atherosclerotic plaques in ApoE-/-mice.2. The effect of Ang-(1-7) on plaque stablity was partly mediated by the the inhibitory effect on inflammation reaction and MMPs activities in atherosclerotic plaques. 1. BackgroundA large amount of evidence demonstates that the rennin-angiotensin system (RAS) plays a critical role in the atherogenesis and atherosclerosis progression. AngiotensinⅡ(AngⅡ), the main peptide of RAS, is an important cytokine that regulates cell growth, migration, inflammation and oxidative stress and therefore contributed to atherosclerotic progression. Inhibition of the RAS with angiotensin-converting enzyme (ACE) inhibitors is one of the current therapeutic strategies with proven beneficial effects in the treatment of atherosclerosis. Blockade of RAS with ACE inhibitors significantly attenuate the development of aortic atherosclerosis in the hyperlipidemic rabbit and ApoE-/-mice. The link between RAS and human atherosclerosis has also been proved by the results of the Heart Outcomes Prevention Evaluation (HOPE) trial, which showed a 22% reduction in the incidence of myocardial infarction, stroke, cardiac arrest and revascularization procedures in patients taking the ACE inhibitor ramipril compared with those on placebo. In addition, in the Perindopril pROtection aGainst REcurrent Stroke Study (PROGRESS), treatment with the ACE inhibitor perindopril reduced the incidence of major vascular events by 26% and myocardial infarction by 38%. Thus, ACE inhibitor treatment may have protective effects on vascular cells via the antagonism of AngⅡactions.Recent studies have shown that, angiotensin converting enzyme 2 (ACE2) and angiotensin-(1-7) [Ang-(1-7)], new components of the RAS, play important roles in atherosclerosis, cardiac and vascular remodeling and kidney function. Ang-(1-7) may exert actions that oppose the proliferative, hypertrophic and proinflammatory effects of AngⅡ, which may play an important role in cardiovascular pathophysiology. Our foregoing studies have demonstrated the protective effect of Ang-(1-7) on atherogenesis and plaques stability. This study was carried out to test the hypothesis that Ang-(1-7) enhances stability of atherosclerotic plaque, similar to a high-dose ramipril therapy. Since recent studies revealed a better outcome in patients with vascular disease by ramipril treatment, a high-dose ramipril was chosen as a therapeutic standard to compare with a high dose of Ang-(1-7) in a ApoE-/-mice atherosclerosis model.2. Objectives2.1 To compare the protective effects of ramipril and Ang-(1-7) on atherosclerotic lesions formation in ApoE-/-mice.2.2 To assess the therapeutic effects of ramipril and Ang-(1-7) on atherosclerotic plaque stability in ApoE-/-mice.3. Methods3.1 Atherosclerosis animal modelAll the animals were fed an atherogenic chow(0.25% cholesterol and 15% cocoa butter) during all the experiment period.Group 1:ApoE-/-mice aged 8 weeks (n=36) were fed an athrogenic chow for 4 weeks.Then the mice were divided into three groups randomly:control group (n=12), ramipril group (n=12) and Ang-(1-7) group (n=12).Group 2:ApoE-/-mice aged 8 weeks (n=36) were fed an athro genic chow for 2 weeks. Then, a 2-mm long constricting silastic tube was placed on the right common carotid artery near its bifurcation. Eight weeks later, the collars were removed, and the mice were divided into three groups randomly:control group (n=12), ramipril group (n=12) and Ang-(1-7) group (n=12). After a further 4-week treatment for atherogenic chow, they were fasted overnight and killed by euthanasia.3.2 Serum lipid measurementSerum samples were collected after overnight fasting and stored at-80℃. Total Triglyceride, cholesterol, low density lipoprotein and high density lipoprotein cholesterol cholesterol were measured.3.3 Histopathological and immunohistochemical measurementGroup 1:The whole aortas were stained with Oil Red O, and the aortic root lesions were stained with hematoxylin and eosin (H&E).Group 2:Sections of carotid plaques were stained with H&E for histological analysis. The collagen and lipids in plaques were stained by using picrosirius red and Oil-red O, respectively. Smooth muscle cells (SMCs) and macrophages were immunostained with anti-a-actin antibodies and MOMA-2, respectively.4. Results4.1 The effect of Ramipril and Ang-(1-7) on serum lipid levelsSerum analysis demonstrates that no significant differences were found among the various treatment groups.4.2 Ramipril and Ang-(1-7) protects against atherogenesis in the whole surface of aortasAfter 8 weeks of high fat diet, we found that, both the two groups treated with ramipril and Ang-(1-7) showed significantly smaller atherosclerotic lesions in their aortas, as shown by both en face (ramipril group:2.52%; Ang-(1-7) group:2.58% and PBS group:4.14%). However, the atherosclerotic lesion area in ramipril group was not different from the Ang-(1-7) (P>0.05), while the lesion areas in both ramipril and Ang-(1-7) groups were larger than control group (both.P<0.05).4.3 Ramipril and Ang-(1-7) protects against atherosclerosis in the aortic rootsAfter 8 weeks of high fat diet, we found that, all the three groups treated with ramipril and Ang-(1-7) showed significantly smaller atherosclerotic lesions by aortic root section analysis (ramipril group:15.32%; Ang-(1-7) group:12.08% and PBS group:20.28%). The atherosclerotic lesion area in ramipril group was significantly larger than the Ang-(1-7) group (P<0.05), while the lesion areas in both ramipril and Ang-(1-7) groups were larger than control group (both P<0.05).4.4 The effect of Ramipril and Ang-(1-7) on carotid atherosclerotic plaque stabilityHistopathological and Immunohistochemical analysis indicated that plaques from carotid arteries in ramipril and Ang-(1-7)-treated ApoE-/-mice showed increased percentages of collagen (ramipril group:16.49%; Ang-(1-7) group:17.98% and PBS group:15.21%) and smooth muscle cells (ramipril group:7.72%; Ang-(1-7) group:8.24%and PBS group:5.15%), and reduced percentages of macrophages (ramipril group:6.68%; Ang-(1-7) group:7.21% and PBS group:12.46%) and lipid area (ramipril group:15.48%; Ang-(1-7) group:14.76% and PBS group:20.43%) compared with those treated with PBS. Accordingly, the histological plaque vulnerability index was also decreased (ramipril group:0.91±0.26; Ang-(1-7) group: 0.84±0.24 and PBS group:1.62±0.38).5. Conclusions1. Ramipril and Ang-(1-7) offered similar protection in terms of atherosclerotic lesions formation in ApoE-/-mice.2. Ramipril and Ang-(1-7) showed similar protective effect on the atherosclerotic plaques stability in ApoE-/-mice.
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