Cardiac hypertrophy is characterized by thickening myocardium and decreasing in heart chamber volume in response to mechanical or pathological stress, but the underlying molecular mechanisms remain to be defined. This study investigated altered miRNA expression and autophagic activity in pathogenesis of cardiac hypertrophy. A rat model of myocardial hypertrophy was used and confirmed by heart morphology, induction of cardiomyocyte autophagy, altered expression of autophagy-related ATG9A, LC3 II/I and p62 proteins, and decrease in miR-34a expression. The in vitro data showed that in hypertrophic cardiomyocytes induced by Ang II, miR-34a expression was downregulated, whereas ATG9A expression was up-regulated. Moreover, miR-34a was able to bind to ATG9A 3′-UTR, but not to the mutated 3′-UTR and inhibited ATG9A protein expression and autophagic activity. The latter was evaluated by autophagy-related LC3 II/I and p62 levels, TEM, and flow cytometry in rat cardiomyocytes. In addition, ATG9A expression induced either by treatment of rat cardiomyocytes with Ang II or ATG9A cDNA transfection upregulated autophagic activity and cardiomyocyte hypertrophy in both morphology and expression of hypertrophy-related genes (i.e., ANP and β-MHC), whereas knockdown of ATG9A expression downregulated autophagic activity and cardiomyocyte hypertrophy. However, miR-34a antagonized Ang II-stimulated myocardial hypertrophy, whereas inhibition of miR-34a expression aggravated Ang II-stimulated myocardial hypertrophy (such as cardiomyocyte hypertrophy-related ANP and β-MHC expression and cardiomyocyte morphology). This study indicates that miR-34a plays a role in regulation of Ang II-induced cardiomyocyte hypertrophy by inhibition of ATG9A expression and autophagic activity.
References
[1]
Ahmad F, Seidman JG, Seidman CE (2005) The genetic basis for cardiac remodeling. Annu Rev Genomics Hum Genet 6: 185–216. doi: 10.1146/annurev.genom.6.080604.162132
[2]
Sadoshima J, Izumo S (1997) The cellular and molecular response of cardiac myocytes to mechanical stress. Annu Rev Physiol 59: 551–571. doi: 10.1146/annurev.physiol.59.1.551
[3]
Berk BC, Fujiwara K, Lehoux S (2007) ECM remodeling in hypertensive heart disease. J Clin Invest 117: 568–575. doi: 10.1172/jci31044
[4]
Hill JA, Olson EN (2008) Cardiac plasticity. N Engl J Med 358: 1370–1380. doi: 10.1056/nejmra072139
[5]
Zhu H, Tannous P, Johnstone JL, Kong Y, Shelton JM, et al. (2007) Cardiac autophagy is a maladaptive response to hemodynamic stress. J Clin Invest 117: 1782–1793. doi: 10.1172/jci27523
[6]
Nakai A, Yamaguchi O, Takeda T, Higuchi Y, Hikoso S, et al. (2007) The role of autophagy in cardiomyocytes in the basal state and in response to hemodynamic stress. Nat Med 13: 619–624. doi: 10.1038/nm1574
[7]
Suzuki K, Kirisako T, Kamada Y, Mizushima N, Noda T, et al. (2001) The pre-autophagosomal structure organized by concerted functions of APG genes is essential for autophagosome formation. EMBO J 20: 5971–5981. doi: 10.1093/emboj/20.21.5971
[8]
Yen WL, Klionsky DJ (2007) Atg27 is a second transmembrane cycling protein. Autophagy 3: 254–256.
[9]
Sayed D, Hong C, Chen IY, Lypowy J, Abdellatif M (2007) MicroRNAs play an essential role in the development of cardiac hypertrophy. Circ Res 100: 416–424. doi: 10.1161/01.res.0000257913.42552.23
[10]
Callis TE, Wang DZ (2008) Taking microRNAs to heart. Trends Mol Med 14: 254–260. doi: 10.1016/j.molmed.2008.03.006
[11]
Fujita Y, Kojima K, Hamada N, Ohhashi R, Akao Y, et al. (2008) Effects of miR-34a on cell growth and chemoresistance in prostate cancer PC3 cells. Biochem Biophys Res Commun 377: 114–119. doi: 10.1016/j.bbrc.2008.09.086
[12]
Tazawa H, Tsuchiya N, Izumiya M, Nakagama H (2007) Tumor-suppressive miR-34a induces senescence-like growth arrest through modulation of the E2F pathway in human colon cancer cells. Proc Natl Acad Sci U S A 104: 15472–15477. doi: 10.1073/pnas.0707351104
[13]
He L, He X, Lim LP, de Stanchina E, Xuan Z, et al. (2007) A microRNA component of the p53 tumour suppressor network. Nature 447: 1130–1134. doi: 10.1038/nature05939
[14]
Tivnan A, Tracey L, Buckley PG, Alcock LC, Davidoff AM, et al. (2011) MicroRNA-34a is a potent tumor suppressor molecule in vivo in neuroblastoma. BMC Cancer 11: 33. doi: 10.1186/1471-2407-11-33
[15]
Cheng Y, Ji R, Yue J, Yang J, Liu X, et al. (2007) MicroRNAs are aberrantly expressed in hypertrophic heart: do they play a role in cardiac hypertrophy? Am J Pathol 170: 1831–1840. doi: 10.2353/ajpath.2007.061170
[16]
Yang J, Chen D, He Y, Melendez A, Feng Z, et al. (2013) MiR-34 modulates Caenorhabditis elegans lifespan via repressing the autophagy gene atg9. Age (Dordr) 35: 11–22. doi: 10.1007/s11357-011-9324-3
[17]
Porrello ER, Delbridge LM (2009) Cardiomyocyte autophagy is regulated by angiotensin II type 1 and type 2 receptors. Autophagy 5: 1215–1216. doi: 10.4161/auto.5.8.10153
[18]
Han JJ, Hao J, Kim CH, Hong JS, Ahn HY, et al. (2009) Quercetin prevents cardiac hypertrophy induced by pressure overload in rats. J Vet Med Sci 71: 737–743. doi: 10.1292/jvms.71.737
[19]
Barbosa ME, Alenina N, Bader M (2005) Induction and analysis of cardiac hypertrophy in transgenic animal models. Methods Mol Med 112: 339–352. doi: 10.1385/1-59259-879-x:339
[20]
Zou Y, Hiroi Y, Uozumi H, Takimoto E, Toko H, et al. (2001) Calcineurin plays a critical role in the development of pressure overload-induced cardiac hypertrophy. Circulation 104: 97–101. doi: 10.1161/01.cir.104.1.97
[21]
Zahabi A, Picard S, Fortin N, Reudelhuber TL, Deschepper CF (2003) Expression of constitutively active guanylate cyclase in cardiomyocytes inhibits the hypertrophic effects of isoproterenol and aortic constriction on mouse hearts. J Biol Chem 278: 47694–47699. doi: 10.1074/jbc.m309661200
[22]
Takemoto M, Node K, Nakagami H, Liao Y, Grimm M, et al. (2001) Statins as antioxidant therapy for preventing cardiac myocyte hypertrophy. J Clin Invest 108: 1429–1437. doi: 10.1172/jci200113350
[23]
Perrotta I (2013) The use of electron microscopy for the detection of autophagy in human atherosclerosis. Micron 50: 7–13. doi: 10.1016/j.micron.2013.03.007
[24]
Mizushima N, Yoshimori T, Levine B (2010) Methods in mammalian autophagy research. Cell 140: 313–326. doi: 10.1016/j.cell.2010.01.028
[25]
Boon RA, Iekushi K, Lechner S, Seeger T, Fischer A, et al. (2013) MicroRNA-34a regulates cardiac ageing and function. Nature 495: 107–110. doi: 10.1038/nature11919
[26]
Bernardo BC, Gao XM, Winbanks CE, Boey EJ, Tham YK, et al. (2012) Therapeutic inhibition of the miR-34 family attenuates pathological cardiac remodeling and improves heart function. Proc Natl Acad Sci U S A 109: 17615–17620. doi: 10.1073/pnas.1206432109
[27]
Basak SK, Veena MS, Oh S, Lai C, Vangala S, et al. (2013) Correction: The CD44 Tumorigenic Subsets in Lung Cancer Biospecimens Are Enriched for Low miR-34a Expression. PLoS One 8.
[28]
Wang W, Li T, Han G, Li Y, Shi LH, et al. (2013) Expression and role of miR-34a in bladder cancer. Indian J Biochem Biophys 50: 87–92.
[29]
Ikeda S, He A, Kong SW, Lu J, Bejar R, et al. (2009) MicroRNA-1 negatively regulates expression of the hypertrophy-associated calmodulin and Mef2a genes. Mol Cell Biol 29: 2193–2204. doi: 10.1128/mcb.01222-08
[30]
Lin Z, Murtaza I, Wang K, Jiao J, Gao J, et al. (2009) miR-23a functions downstream of NFATc3 to regulate cardiac hypertrophy. Proc Natl Acad Sci U S A 106: 12103–12108. doi: 10.1073/pnas.0811371106
[31]
Care A, Catalucci D, Felicetti F, Bonci D, Addario A, et al. (2007) MicroRNA-133 controls cardiac hypertrophy. Nat Med 13: 613–618. doi: 10.1038/nm1582
[32]
Huang ZP, Chen J, Seok HY, Zhang Z, Kataoka M, et al. (2013) MicroRNA-22 regulates cardiac hypertrophy and remodeling in response to stress. Circ Res 112: 1234–1243. doi: 10.1161/circresaha.112.300682
[33]
Yin X, Peng C, Ning W, Li C, Ren Z, et al. (2013) miR-30a downregulation aggravates pressure overload-induced cardiomyocyte hypertrophy. Mol Cell Biochem 379: 1–6. doi: 10.1007/s11010-012-1552-z
[34]
Zhang ZH, Li J, Liu BR, Luo CF, Dong Q, et al. (2013) MicroRNA-26 was decreased in rat cardiac hypertrophy model and may be a promising therapeutic target. J Cardiovasc Pharmacol.
[35]
Hu Y, Correa AM, Hoque A, Guan B, Ye F, et al. (2011) Prognostic significance of differentially expressed miRNAs in esophageal cancer. Int J Cancer 128: 132–143. doi: 10.1002/ijc.25330
[36]
Chang TC, Wentzel EA, Kent OA, Ramachandran K, Mullendore M, et al. (2007) Transactivation of miR-34a by p53 broadly influences gene expression and promotes apoptosis. Mol Cell 26: 745–752. doi: 10.1016/j.molcel.2007.05.010
[37]
Reggiori F, Shintani T, Nair U, Klionsky DJ (2005) Atg9 cycles between mitochondria and the pre-autophagosomal structure in yeasts. Autophagy 1: 101–109. doi: 10.4161/auto.1.2.1840
[38]
He C, Klionsky DJ (2009) Regulation mechanisms and signaling pathways of autophagy. Annu Rev Genet 43: 67–93. doi: 10.1146/annurev-genet-102808-114910
[39]
Rifki OF, Bodemann BO, Battiprolu PK, White MA, Hill JA (2013) RalGDS-dependent cardiomyocyte autophagy is required for load-induced ventricular hypertrophy. J Mol Cell Cardiol 59: 128–138. doi: 10.1016/j.yjmcc.2013.02.015
[40]
Chen H, Wang X, Tong M, Wu D, Wu S, et al. (2013) Intermedin suppresses pressure overload cardiac hypertrophy through activation of autophagy. PLoS One 8: e64757. doi: 10.1371/journal.pone.0064757
[41]
Tannous P, Zhu H, Nemchenko A, Berry JM, Johnstone JL, et al. (2008) Intracellular protein aggregation is a proximal trigger of cardiomyocyte autophagy. Circulation 117: 3070–3078. doi: 10.1161/circulationaha.107.763870
[42]
Bjorkoy G, Lamark T, Brech A, Outzen H, Perander M, et al. (2005) p62/SQSTM1 forms protein aggregates degraded by autophagy and has a protective effect on huntingtin-induced cell death. J Cell Biol 171: 603–614. doi: 10.1083/jcb.200507002
[43]
Su H, Li J, Menon S, Liu J, Kumarapeli AR, et al. (2011) Perturbation of cullin deneddylation via conditional Csn8 ablation impairs the ubiquitin-proteasome system and causes cardiomyocyte necrosis and dilated cardiomyopathy in mice. Circ Res 108: 40–50. doi: 10.1161/circresaha.110.230607
[44]
Jian X, Xiao-yan Z, Bin H, Yu-feng Z, Bo K, et al. (2011) MiR-204 regulate cardiomyocyte autophagy induced by hypoxia-reoxygenation through LC3-II. Int J Cardiol 148: 110–112. doi: 10.1016/j.ijcard.2011.01.029
[45]
Ucar A, Gupta SK, Fiedler J, Erikci E, Kardasinski M, et al. (2012) The miRNA-212/132 family regulates both cardiac hypertrophy and cardiomyocyte autophagy. Nat Commun 3: 1078. doi: 10.1038/ncomms2090
[46]
Zhang XD, Wang Y, Wu JC, Lin F, Han R, et al. (2009) Down-regulation of Bcl-2 enhances autophagy activation and cell death induced by mitochondrial dysfunction in rat striatum. J Neurosci Res 87: 3600–3610. doi: 10.1002/jnr.22152
[47]
Zalckvar E, Berissi H, Eisenstein M, Kimchi A (2009) Phosphorylation of Beclin 1 by DAP-kinase promotes autophagy by weakening its interactions with Bcl-2 and Bcl-XL. Autophagy 5: 720–722. doi: 10.4161/auto.5.5.8625
