NIMA Related Kinase6Attenuates Cardiac Remodeling Via AKT Signaling Pathway

BackgroundHeart failure has been seen as a common endpoint of cardiovascular diseases such as coronary heart diseases, hypertension, valvular heart diseases, cardiomyopathis, congenital heart diseases and so on. However, the treatment of heart failure has been a difficult issue in the treatment of cardiovascular diseases. The main cause lies in the lack of basic understanding of the pathogenesis. Cardiac remodeling is a compensatory response to the increased ventricular pressure overload. The main pathological changes are myocardial hypertrophy, cardiomyocyte hypertrophy, increased myocardial fibrosis. Recent studies have shown that myocardial remodeling play an important role in the initial process and development of heart failure. In order to improve the treatment of heart failure, further studies are needed in the field of myocardial remodeling. Because he pathogenesis of cardiac remodeling is not entirely clear, looking for the new targets in the prevention and treatment of cardiac remodeling is meaningful.Never in mitosis gene A (NIMA) related expressed kinase6(Nek6), one of the members of Neks family, was widely expressed in plants and animals. In mammal, it could be expressed in multiple organs, especially in skeletal muscle and heart. Recent studies have shown that Nek6is closely associated with cell cycle and tumor, but the roles in cardiovascular disease is unclear.ObjectivesTo determine the relationship of Nek6and cardiac remodeling and clarify the possible mechanism.MethodsPart one:The Nek6protein expression in the left ventricular samples of patients with dilated cardiomyopathy (DCM) and volunteer were detect in this part. Besides, aortic banding (AB) was used to build hypertrophic models in mice. We also detect the protein expression of Nek6in mice。Immunohistochemical (IHC) was used to find the location and expression of Nek6in cardiomyocytes.Part two:Nek6knockout (KO) mice and wide type (WT) mice were use in this part. The approach for mice modeling is the same as part one. Echocardiography and PV were used in the assessment of cardiac function. Heart weight/body weight (HW/BW), lung weight/body weight (LW/BW) and heart weight/tibial length (HW/TL) were used to assess weight change of heart and lungs of mice. The picture of whole heart was used to assess the volume of heart, and HE staining was used to evaluate the cross-sectional area (CSA) of cardiomyocytes. PSR staining was used to evaluate the collagen volume of LV. RT-PCR was used in the detection of mRNA expression levels of hypertrophic and fibrotic markers. To find a possible mechanism, the expressions of some proteins in the possible signaling pathway were detected by Western Blot.Part three:H9c2cardiomyocytes with and without pCMV-Nek6transfection was used in this part. Cardiomyocytes were stimulated by AngⅡ or PBS. Immunofluore scence (IF) was use to assess CSA of cardiomyocytes. To find a possible mechanism, the expressions of some proteins in the possible signaling pathway were detected by Western Blot.ResultsPart one:The Nek6level was higher in the DCM, compared with volunteers. In mice, the protein expression of Nek6is tripled after AB1w, and doubled in AB8w。In IHC, Nek6was expressed in the cytoplasm, and was heightened after AB.Part two:As a result of echocardiography, LVEDD and LVESD have been raised after AB, and elevated more remarkably in KO mice. EF and FS have been declined after AB, and descended more obviously in KO mice. HW/BW and HW/TL have been raised after AB, and elevated more obviously in KO mice. LW/BW has no significant change after AB in WT mice, but ascended obviously in KO mice. The sizes of whole heart have been raised after AB, and elevated more remarkably in KO mice. As a result of HE staining, the CSA of cardiomyocytes have been raised after AB, and ascended more remarkably in KO mice. As a result of PSR staining, the collagen volume of LV has been elevated after AB, and raised more remarkably in KO mice. As results of RT-PCR, the mRNA levels of ANP, BNP and β-MHC have been raised after AB, and elevated more remarkably in KO mice. The levels of α-MHC and SERC2a have been declined after AB, and descended more obviously in KO mice. The mRNA levels of some fibrotic markers, such as CTGF, TGF-β2, Collagen la and Collagen III, have been raised after AB, and elevated more remarkably in KO mice. As a result of Western Blot, the phos-protein expressions of some markers in AKT signaling pathway, such as AKT, GSK3β, mTOR,4EBP1, eIF4e and p70, have been raised after AB, and elevated more remarkably in KO mice. But there are no significant changes in the expressions of total protein.Part three:As a result of IF, the CSA of cardiomyocytes have been enlarged after stimulations of AngⅡ, and elevated more remarkably in H9c2group. As a result of Western Blot, the phos-protein expressions of some markers in AKT signaling pathway, such as AKT, GSK3β, mTOR,4EBP1and eIF4e, have been raised after AB, and elevated more remarkably H9c2group. But there are no significant changes in the expressions of total protein.Conclusions1. Cardiac remodeling increase the expression of Nek6;2. Nek6knockout aggravates cardiac hypertrophy and fibrosis through the activation of AKT signaling pathway;3. Nek6overpress suppresses cardiomyocyte hypertrophy via the inhibition of AKT signaling pathway;...