The Function Studies Of MR-1 In Cardiac Hypertrophy Induced By Angiotensin Ⅱ

Myofibrillogenesis regulator-1 (MR-1) was cloned from a human skeletal muscle cDNA library. The MR-1 gene (NCBI GeneBank accession no. AF417001) is located on human chromosome 2q35 (AC021016) and spans about 2887 bp of contiguous DNA. The MR-1 gene is composed of three distinct exons and encodes a protein of 142 amino acids. Myofibrillogenesis regulator-1 (MR-1) as a novel homo gene augments cardiomyocytes hypertrophy induced by AngII in vivo and in vitro. Overexpression of MR1 aggravates cardiac hypertrophy induced by AngII in mice. In this study the effect of silencing MR-1 with RNAi on cardiac hypertrophy induced by AngII in mice was investigated. Cardiac hypertrophy mice was induced by chronic infusion of AngII through implanted pumps. Recombinant adenoviral vector (pAdxsi) expressing MR-1 SiRNA was constructed and used to treat cardiac hypertrophy in mice. AdSiR-MR-1 was administered to the mice via the jugular or tail vein at first in order to define the route of delivery and the effective silencing dose. Significant silencing was observed with 2×10~9 pfu of AdSiR-MR-1 via jugular vein. Western blot analysis showed that AdSiR-MR-1 effectively silenced the expression of MR-1 in cardiomyocytes as well as in other tissues except the skeletal muscle. AdSiR-MR-1 suppressed dose-dependent myocardial MR-1 protein expression in AngII-infused mice. Administration of AdSiR-MR-1 via the jugular vein almost abolished the overexpression of MR-1 induced by AngII resulting in decreased left-ventricular hypertrophy assessed by echocardiography. Moreover, cardiac hypertrophy-related protein levels, such as ANF, andβ-myosin were attenuated significantly in the heart of AngII-infused mice by knockdown of MR-1. AdSiR-MR-1 treatment also significantly diminishes the cardiac fibrosis accompanied with the decrease of collagen contents as observed morphologically and by immunohistochemical determination. Our results identify, for the first time, that MR-1 serves as a new target in protecting and suppressing cardiac hypertrophy induced by AngII in vivo.Genechip microarray expression analysis ( Affymetrix, over 18, 000cDNA sequences) was applied to evaluate the changes in the genes expression in AngII-infused AdSiR-MR-1 treated mice verse to AngII-infused mice without treatment(control). About 1453 genes expression were increased in more than two fold, among them 6 genes were expressed in ten fold higher than control. In the same time 1081 genes were expressed lower in two fold more and 43 genes expression were decreased in ten fold comparing with control. The altered expression genes mostly involved in the pathways related to muscle relaxation and contraction, immune reaction, G-protein related signaling , calcium ion binding or regulation as well as amino-acid metabolisms. Differentially expressed genes in AdSiR-MR-1 treated mice could be grouped into the functions of muscle contraction and relaxation, signaling transduction, calcium ion biding and signaling etc. Nine genes encoding for heat shock proteins (HSP) and eight genes encoding for thioredoxins (TRX) were up-regulated and eighty four genes related to calcium functions were down-regulated in AdSiR-MR-1 treated mice vs control.. The up-regulated gene expression of HSP72 and TRX as well as down-regulated calcineurin gene expression were confirmed by RT-PCR and Western blot analyses. These findings indicate the possible involvement of HSPs, TRXs and calcineurin in the MR1 regulatory pathways governing the development of cardiac hypertrophy induced by AngII. This work paves the way for better understanding of MR1 regulation mechanism in cardiac hypertrophy induced by AngII and suggests that MR1 would be considered as a new target for the treatment or prevention of cardiac hypertrophy and heart failure in future.