Study Of Effect Of Sca1on Cardiac Hypertrophy And The Mechanisms Involved In

BackgroundCardiac hypertrophy, a critical clinical course of heart failure, could be occurred by a variety of diseases including hypertension, valvular stenosis and myocardial infarction. Although cardiac hypertrophy is a compensatory mechanism in the initial phase to reduce the wall stress and maintain cardiac output, sustained cardiac hypertrophy will eventually lead to ventricular dysfunction and the development of heart failure. Therefore, we can curb the early progression of heart failure and prevent deterioration of cardiac function if cardiac hypertrophy could be inhibited or even reversed. Cardiac hypertrophy is induced by mechanical and neurohormonal stimulation, which affect a variety of signaling pathways and cause the phenotypic changes of related genes in nucleus. However, the mechanisms of cardiac hypertrophy have not yet been fully elucidated, and it is still lack of effective interventions.Stem cell antigen-1(Seal) is a glycosyl phosphatidylinositol-anchored protein (GPI-AP), originally found upregulated in activated lymphocytes, also known as lymphocyte activation protein-6A (Ly-6A), which is a member of Ly6gene family. The GPI-APs are often located in lipid rafts which are sphingolipid and cholesterol-rich-domains of the plasma membrane, containing a variety of signaling and transport proteins, and are able to recruit a variety of intracellular kinases, including Src family kinases, mitogen activated protein kinases (MAPKs) and phosphatidylinositol3-kinase (PI3K)/Akt. Previous studies suggested that Seal is a negative regulator of the Src family kinase molecules, and may have effect on PI3K/Akt and the MAPK family member ERK. However, the signaling pathways mediated by Seal remain unclear.The appropriate expression of Seal is very important to maintain homeostasis. It has been confirmed that Seal is involved in lymphocyte activation, tumorigenesis and muscle remodeling. In addition, the cardiac expression of Seal protein increased significantly after myocardial infarction in mice, and the Seal signaling may play an important role in regulation of cardiac stem cell survival and proliferation after transplantation, which promotes the repair response after myocardial infarction and improves cardiac function. However, the role of Seal in cardiac hypertrophy is still unclear. This study used aortic banding to induce cardiac hypertrophy in mice, examined the effect of Seal on cardiac hypertrophy, and explored the underlying mechanisms.MethodsPart one:The male wild-type C57BL/6mice aged8to10weeks and body weight ranged from23.5-27.5g were included. Aortic Banding (AB) was used to establish pressure overload-induced cardiac hypertrophy in mice. The mice were randomly divided into7groups, respectively, for the sham operation group (sham), AB-1day,3days,1week,2weeks,4weeks and8weeks groups (n=15in each group).Part two:The male Scal knockout mice (sham, n=15; AB, n=30), Scal cardiac-specific transgenic mice (sham, n=15; AB, n=20), and their respective wild-type littermates (sham, n=15; AB, n=20), aged8to10weeks and body weight ranged from23.5-27.5g were included. The mice were randomly divided into AB group and sham group.4weeks after AB, cardiac function of mice was examined by echocardiography and hemodynamics, and the HW/BW, LW/BW, and HW/TL were compared in mice of each group. HE, WGA, and PSR staining were used for morphology analysis. Molecular markers of cardiac hypertrophy and fibrosis were determined by real-time quantitative RT-PCR.Part three:The male Scal knockout mice (sham, n=15; AB, n=30), Scal cardiac-specific transgenic mice (sham, n=15; AB, n=20), and their respective wild-type littermates (sham, n=15; AB, n=10), aged8to10weeks and body weight ranged from23.5～27.5g were included. The mice were randomly divided into AB group and sham group. The levels of phosphorylated and total protein of Src, MAPKs, and Akt in hearts from each group were determined by Western blot analysis after4weeks of AB.ResultsWestern blot analysis showed that the expression of cardiac Scal protein increased strikingly1day after AB surgery compared to sham group, and was maintained at a high level until8weeks after AB (P<0.05vs. sham). Immunohistochemistry and immunofluorescence analysis showed that Scal protein was detected little on the surface of myocytes in sham mice but was obvious after AB.After4weeks of AB surgery, the HW/BW, LW/BW, HW/TL, cardiac myocyte cross-sectional area, and left ventricular collagen volume fraction of Scal knockout mice were significantly higher than those of wild-type mice (P<0.05). The echocardiographic and hemodynamics results showed that Scal knockout mice exhibited higher left ventricular wall thickness and ventricular diameter, and lower left ventricular systolic and diastolic function than wild-type mice (P<0.05). The mRNA levels of cardiac hypertrophic and fibrotic markers including ANP, BNP,β-MHC, TGF-β, CTGF, collagen la and collagen Ⅲα in Scal knockout mice were higher than those in wild-type mice (P<0.05), and the mRNA levels of a-MHC and SERCA2a were lower (P<0.05). Conversely, Scal cardiac-specific transgenic mice exhibited the opposite phenotypes to Scal knockout mice after4weeks of AB surgery.After4weeks of AB, we detected that Scal deficiency enhanced the phosphorylation of Src, MAPKs, and Akt induced by pressure overload using western blot analysis (P<0.05), while the cardiac-specific overexpression of Scal was able to inhibit the activation of Src, MAPKs and Akt (P<0.05).Conclusions1. Cardiac Scal expression is significantly up-regulated in response to pressure overload induced by aortic banding, and could be expressed on cardiomyocyte.2.Scal deficiency promotes pressure overload-induced cardiac hypertrophy and fibrosis, while cardiac-specific overexpression of Scal attenuates cardiac hypertrophy and fibrosis.3. The protective effect of Scal on cardiac hypertrophy is mediated by inhibition of Src, MAPKs, and Akt pathways.