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HTR2A promotes the development of cardiac hypertrophy by activating PI3K-PDK1-AKT-mTOR signaling - PubMed

HTR2A promotes the development of cardiac hypertrophy by activating PI3K-PDK1-AKT-mTOR signaling

Weinian Gao et al. Cell Stress Chaperones. 2020 Nov.

Abstract

5-Hydroxytryptamine receptor 2A (HTR2A) is a central regulator of fetal brain development and cognitive function in adults. However, the roles of HTR2A in the cardiovascular system are not fully understood. Here in this study, we explored the function of HTR2A in cardiac hypertrophy. Significantly, the expression levels of HTR2A mRNA and protein levels were upregulated in hypertrophic hearts of human patients. Besides, the expression of HTR2A was also upregulated in isoproterenol (ISO)-induced cardiac hypertrophy in the mouse. Next, the expression of HTR2A was knocked down with shRNA or overexpressed with adenovirus in neonatal rat cardiomyocytes, and ISO was used to induce cardiomyocyte hypertrophy. We showed that HTR2A knockdown repressed ISO-induced cardiomyocyte hypertrophy, which was demonstrated by decreased cardiomyocyte size and repressed expression of hypertrophic fetal genes (e.g., myosin heavy chain beta (β-Mhc), atrial natriuretic peptide (Anp), and brain natriuretic peptide (Bnp)). By contrast, HTR2A overexpression promoted cardiomyocyte hypertrophy. Of note, we observed that HTR2A promoted the activation (phosphorylation) of AKT-mTOR (mammalian target of rapamycin) signaling in cardiomyocytes, and repression of AKT-mTOR with perifosine or rapamycin blocked the effects of HTR2A on cardiomyocyte hypertrophy. Finally, we showed that HTR2A regulated AKT-mTOR signaling through activating the PI3K-PDK1 pathway, and inhibition of either PI3K or PDK1 blocked the roles of HTR2A in regulating AKT-mTOR signaling and cardiomyocyte hypertrophy. Altogether, these findings demonstrated that HTR2A activated PI3K-PDK1-AKT-mTOR signaling and promoted cardiac hypertrophy.

Keywords: Akt; Cardiac hypertrophy; HTR2A; PDK1; PI3K; mTOR.

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Conflict of interest statement

The authors declare that they have no conflict of interest.

Figures

Fig. 1
Fig. 1

HTR2A is overexpressed in human and mouse hypertrophic hearts. a mRNA level of marker genes of hypertrophy in human hearts. The RNAs from control and hypertrophic heart tissues of human patients were used for mRNA analysis with qRT-PCR (n = 6 in each group). ANP, atrial natriuretic peptide; BNP, brain natriuretic peptide; β-MHC, myosin heavy chain beta. b mRNA level of HTR2A in human hearts from control donors and patients with cardiac hypertrophy (n = 6 in each group). HTR2A, 5-hydroxytryptamine receptor 2A. c The protein level of HTR2A in human hearts from control donors and patients with cardiac hypertrophy (n = 4 in each group). d mRNA level of marker genes of hypertrophy in mouse hearts. Cardiac hypertrophy was induced in male C57BL/6J mice by chronic treatment of isoproterenol (ISO, 50 mg/kg/day) for 28 days. mRNA levels of hypertrophic marker genes were analyzed with qRT-PCR (n = 5 in each group). e mRNA level of HTR2A in mouse control and hypertrophic hearts (n = 5 in each group). f The protein level of HTR2A in mouse control and hypertrophic hearts (n = 4 in each group). **P < 0.01 analyzed by using the Student’s t test

Fig. 2
Fig. 2

HTR2A promotes ISO-induced cardiomyocyte hypertrophy. a, b Short-hairpin RNA (shRNA) knockdown of HTR2A in cardiomyocytes. Rat cardiomyocytes were infected with adenovirus expressing shHTR2A (Ad-shHTR2A) or the control shRNA (Ad-shCtrl) for 48 h. Then, the mRNA (a) and protein (b) levels were analyzed with qRT-PCR and western blot respectively (n = 3 in each group). c, d The knockdown of HTR2A inhibits cardiomyocyte hypertrophy induced by ISO. Rat cardiomyocytes with/without HTR2A knockdown were subjected to hypertrophy induction with ISO (50 μM) treatment for 48 h. Cardiomyocyte size was quantified with the ImageJ software (c), and the expression of hypertrophy-associated fetal genes was analyzed with qRT-PCR (d, n = 3 in each group). e, f Overexpression of rat HTR2A in cardiomyocytes. Rat cardiomyocytes were infected with adenovirus overexpressing rat HTR2A gene (Ad-HTR2A) or control adenovirus (Ad-Ctrl) for 48 h. Then, the mRNA (e) and protein (f) levels were analyzed with qRT-PCR and western blot respectively (n = 3 in each group). g, h Overexpression of HTR2A promotes cardiomyocyte hypertrophy induced by ISO. Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 48 h. Cardiomyocyte size was quantified with the ImageJ software (g), and the expression of hypertrophy-associated fetal genes was analyzed with qRT-PCR (h, n = 3 in each group). **P < 0.01 analyzed by using the Student’s t test (a, e) or two-way ANOVA followed by the Tukey post hoc test (c, d, g, h)

Fig. 3
Fig. 3

HTR2A promotes AKT-mTOR-S6K1 signaling activation. a HTR2A knockdown represses AKT-mTOR-S6K1 signaling activated by ISO in rat cardiomyocytes. Cardiomyocytes with/without HTR2A knockdown were subjected to ISO treatment (50 μM) for 24 h. mTOR, mammalian target of rapamycin; S6K1, p70 ribosomal protein S6 kinase 1. Representative western blot and quantitative results are shown (n = 3). b HTR2A overexpression promotes AKT-mTOR-S6K1 signaling activated by ISO in rat cardiomyocytes. Cardiomyocytes with/without HTR2A overexpression were subjected to ISO treatment (50 μM) for 24 h. Representative western blot and quantitative results are shown (n = 3). c, d Inhibition of AKT represses HTR2A function in cardiomyocyte hypertrophy. Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 48 h. The cardiomyocytes were also treated with/without AKT inhibitor perifosine (1 μM). Cardiomyocyte size was quantified with the ImageJ software (c), and the expression of hypertrophy-associated fetal genes was analyzed with qRT-PCR (d, n = 3 in each group). e, f Inhibition of mTOR represses HTR2A function in cardiomyocyte hypertrophy. Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 48 h. The cardiomyocytes were also treated with/without AKT inhibitor rapamycin (100 nM). Cardiomyocyte size was quantified with the ImageJ software (e), and the expression of hypertrophy-associated fetal genes was analyzed with qRT-PCR (f, n = 3 in each group). **P < 0.01 analyzed by two-way ANOVA followed by the Tukey post hoc test. Cardiomyocyte culture sets were performed on three different dates. Western blot was performed for each experiment set, and representative western blot results were shown

Fig. 4
Fig. 4

PI3K-PDK1-mediated HTR2A activation on the AKT-mTOR pathway. a HTR2A knockdown represses PI3K-PDK1 signaling. Cardiomyocytes with/without HTR2A knockdown were subjected to ISO treatment (50 μM) for 24 h. Representative western blot and quantitative results are shown (n = 3). b HTR2A overexpression promotes PI3K-PDK1 signaling. Cardiomyocytes with/without HTR2A overexpression were subjected to ISO treatment (50 μM) for 24 h. Representative western blot and quantitative results are shown (n = 3). c PI3K inhibition blocks HTR2A function in regulating AKT-mTOR signaling in cardiomyocytes. Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 24 h. The cardiomyocytes were also treated with/without PI3K inhibitor LY294002 (500 nM). d, e PI3K inhibition blocks HTR2A function in cardiomyocyte size. Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 48 h. The cardiomyocytes were also treated with/without PI3K inhibitor LY294002 (500 nM). Cardiomyocyte size was quantified with the ImageJ software (d), and the expression of hypertrophy-associated fetal genes was analyzed with qRT-PCR (e, n = 3 in each group). f PDK1 inhibition blocks HTR2A function in AKT-mTOR signaling. Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 24 h. The cardiomyocytes were also treated with/without PDK1 inhibitor OSU-03012 (2 μM). Representative western blot and quantitative results are shown (n = 3). g, h Rat cardiomyocytes with/without HTR2A overexpression were subjected to hypertrophy induction with ISO (50 μM) treatment for 48 h. The cardiomyocytes were also treated with/without PDK1 inhibitor OSU-03012 (2 μM). Cardiomyocyte size was quantified with the ImageJ software (h), and the expression of hypertrophy-associated fetal genes was analyzed with qRT-PCR (h, n = 3 in each group). **P < 0.01 analyzed by two-way ANOVA followed by the Tukey post hoc test. Cardiomyocyte culture sets were performed on three different dates. Western blot was performed for each experiment set, and representative western blot results were shown

Fig. 5
Fig. 5

Illustrator showing HRT2A function during cardiac hypertrophy. Upon hypertrophic stress, HTR2A expression is upregulated, which leads to the activation of PI3K. Activated PI3K promotes the activation of AKT in a PDK1-dependent manner. Finally, mTOR and S6K1 signalings are activated by AKT, which results in increased protein synthesis and hypertrophic growth of cardiomyocytes

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References

    1. Aoyagi T, Matsui T. Phosphoinositide-3 kinase signaling in cardiac hypertrophy and heart failure. Curr Pharm Des. 2011;17:1818–1824. doi: 10.2174/138161211796390976. - DOI - PMC - PubMed
    1. Ayme-Dietrich E, Marzak H, Lawson R, Mokni W, Wendling O, Combe R, Becker J, El Fertak L, Champy MF, Matz R, Andriantsitohaina R, Doly S, Boutourlinsky K, Maroteaux L, Monassier L. Contribution of serotonin to cardiac remodeling associated with hypertensive diastolic ventricular dysfunction in rats. J Hypertens. 2015;33:2310–2321. doi: 10.1097/HJH.0000000000000695. - DOI - PubMed
    1. L. Cai, G. Fan, F. Wang, S. Liu, T. Li, X. Cong, J. Chun, X. Chen,(2017) Protective role for LPA3 in cardiac hypertrophy induced by myocardial infarction but not by isoproterenol, Front. Physiol., 8. - PMC - PubMed
    1. Chen X-F, Chen X, Tang X. Short-chain fatty acid, acylation and cardiovascular diseases. Clin Sci. 2020;134:657–676. doi: 10.1042/CS20200128. - DOI - PubMed
    1. Choi JR, Jeon M, Koh SB. Association between serotonin 2A receptor (HTR2A) genetic variations and risk of hypertension in a community-based cohort study. BMC Med Genet. 2020;21:5. doi: 10.1186/s12881-019-0927-3. - DOI - PMC - PubMed

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