There are some reports which indicate that congestive heart failure has increased in recent years worldwide [1Yusuf S, Hawken S, Ounpuu S. Effect of potentially modifiable risk factors associated with myocardial infarction in 52 countries (the INTERHEART study): Case-control study. Lancet 2004; 364(9438): 937-52.
[http://dx.doi.org/10.1016/S0140-6736(05)67663-5] [PMID: 16271645] , 2Thygesen K, Alpert JS, White HD. Universal definition of myocardial infarction. J Am Coll Cardiol 2007; 50(22): 2173-95.
[http://dx.doi.org/10.1016/j.jacc.2007.09.011] [PMID: 18036459] ]. It is important to mention that congestive heart failure (translated as myocardial infarction) can induce cardiomyocyte cell death and result in a decrease of cardiac work [3Pfeffer MA. Left ventricular remodeling after acute myocardial infarction. Annu Rev Med 1995; 46: 455-66.
[http://dx.doi.org/10.1146/annurev.med.46.1.455] [PMID: 7598478] -5Figueroa-Valverde L, Díaz-Cedillo F, Rosas-Nexticapa M, García-Cervera E, Pool-Gómez E. Evaluation of activity exerted by a steroid derivative on injury by ischaemia/reperfusion. Afr J Pharm Pharmacol 2014; 8(5): 157-67.
[http://dx.doi.org/10.5897/AJPP2013.3908] ]. Some drugs are used for the treatment of this clinical pathology, such as angiotensin-converting-enzyme inhibitors [6Gilewski W, Błażejewski J, Karasek D, et al. Are changes in heart rate, observed during dobutamine stress echocardiography, associated with a response to cardiac resynchronisation therapy in patients with severe heart failure? Results of a multicentre ViaCRT study. Kardiol Pol 2018; 76(3): 611-7.
[PMID: 29297189] ], β₁-adrenergic agonist or β₁-adrenergic blockers [7Liggett SB, Mialet-Perez J, Thaneemit-Chen S, et al. A polymorphism within a conserved β(1)-adrenergic receptor motif alters cardiac function and β-blocker response in human heart failure. Proc Natl Acad Sci USA 2006; 103(30): 11288-93.
[http://dx.doi.org/10.1073/pnas.0509937103] [PMID: 16844790] ], diuretics [8Domanski M, Norman J, Pitt B, Haigney M, Hanlon S, Peyster E. Diuretic use, progressive heart failure, and death in patients in the Studies of Left Ventricular Dysfunction (SOLVD). J Am Coll Cardiol 2003; 42(4): 705-8.
[http://dx.doi.org/10.1016/S0735-1097(03)00765-4] [PMID: 12932605] ], angiotensin-receptor inhibitors [9Corvalán H, Casanegra P, Chamorro S, Jalil M, Valenzuela P. Comparación de los efectos clínicos y neurohumorales de milrinona y captopril en pacientes con insuficiencia cardíaca crónica. Bol Cardiol Chile 1988; 7(4): 289-300.], calcium sensitizer [10Follath F, Cleland J, Just H, Papp J, Scholz H, Peuhkurinen K. Steering committee and investigators of the Levosimendan Infusion versus Dobutamine (LIDO) study. Efficacy and safety of intravenous levosimendan compared with dobutamine in severe low-output heart failure (the LIDO study): A randomized double-blind trial. Lancet 2002; 360(9328): 196-202.
[http://dx.doi.org/10.1016/S0140-6736(02)09455-2] [PMID: 12133653] ], phosphodiesterase III inhibitors [11Bregagnollo E, Fortes A, Cicogna A. Avaliação dos efeitos inotrópicos e vasodilatadores do lactato de milrinona em pacientes com cardiomiopatia dilatada e insuficiência cardíaca grave. Arq Bras Cardiol 1999; 72(2): 149-54.
[http://dx.doi.org/10.1590/S0066-782X1999000200003] [PMID: 10488574] ], ATP-ase inhibitors [12Tischler M, Smith T. Digitalis: Its current place in the treatment of heart failure. Mod Concepts Cardiovasc Dis 1990; 59: 67-72.] and others. However, some of these drugs can produce adverse effects [13Secknus MA, Marwick TH. Evolution of dobutamine echocardiography protocols and indications: Safety and side effects in 3,011 studies over 5 years. J Am Coll Cardiol 1997; 29(6): 1234-40.
[http://dx.doi.org/10.1016/S0735-1097(97)00039-9] [PMID: 9137218] -18Pitt B, Zannad F, Remme WJ, et al. The effect of spironolactone on morbidity and mortality in patients with severe heart failure. Randomized Aldactone Evaluation Study Investigators. N Engl J Med 1999; 341(10): 709-17.
[http://dx.doi.org/10.1056/NEJM199909023411001] [PMID: 10471456] ]; therefore, in the search for a new drug for the treatment of heart failure and ischemia-reperfusion injury, several drugs have been developed such as MK-7145 (potassium channel inhibitor) [19Tang H, Zhu Y, Teumelsan N, et al. Discovery of MK-7145, an oral small molecule ROMK inhibitor for the treatment of hypertension and heart failure. ACS Med Chem Lett 2016; 7(7): 697-701.
[http://dx.doi.org/10.1021/acsmedchemlett.6b00122] [PMID: 27437080] ], some heteroaryl-substituted naphthalenes (CYP11B2 inhibitors) [20Voets M, Antes I, Scherer C, et al. Heteroaryl-substituted naphthalenes and structurally modified derivatives: Selective inhibitors of CYP11B2 for the treatment of congestive heart failure and myocardial fibrosis. J Med Chem 2005; 48(21): 6632-42.
[http://dx.doi.org/10.1021/jm0503704] [PMID: 16220979] , 21Voets M, Antes I, Scherer C, et al. Synthesis and evaluation of heteroaryl-substituted dihydronaphthalenes and indenes: Potent and selective inhibitors of aldosterone synthase (CYP11B2) for the treatment of congestive heart failure and myocardial fibrosis. J Med Chem 2006; 49(7): 2222-31.
[http://dx.doi.org/10.1021/jm060055x] [PMID: 16570918] ], BAY-1021189 (guanylate cyclase stimulator) [22Follmann M, Ackerstaff J, Redlich G, et al. Discovery of the soluble guanylate cyclase stimulator vericiguat (BAY 1021189) for the treatment of chronic heart failure. J Med Chem 2017; 60(12): 5146-61.
[http://dx.doi.org/10.1021/acs.jmedchem.7b00449] [PMID: 28557445] ], N-benzylcarboxamide (G-protein-coupled receptor kinase 2 inhibitor) [23Okawa T, Aramaki Y, Yamamoto M, et al. Design, synthesis, and evaluation of the highly selective and potent G-protein-coupled receptor kinase 2 (GRK2) inhibitor for the potential treatment of heart failure. J Med Chem 2017; 60(16): 6942-90.
[http://dx.doi.org/10.1021/acs.jmedchem.7b00443] [PMID: 28699740] ], a naphthalene-prazosin derivative (calcium channels activation) [24Sarabia-Alcocer B, Figueroa-Valverde L, Díaz-Cedillo F, Hau-Heredia L, Rosas-Nexticapa M. García-Cervera. Activity induced by a naphthalene-prazosin derivative on ischemia/reperfusion injury in rats. Pharmacol Pharm 2014; 5: 1130-42.
[http://dx.doi.org/10.4236/pp.2014.512123] ], a guanidine analog (sodium-hydrogen exchanger isoform inhibitor) and others. All these data indicate that several drugs may exert effects on heart failure and ischemia-reperfusion injury; however, the molecular mechanism is very confusing, perhaps, this phenomenon can be due to; 1) differences in the chemical structure of each drug; or 2) to different protocols used in each experiment. Therefore, in this study, the biological activity of an Amino-Steroid-Anthracenone (ASA) (Fig. 1) was evaluated using an ischemia-reperfusion injury model.

Fig. (1) Chemical structure of ASA (8-Amino-10-(3-hydroxy-pentyl)-13a-methyl-3,3a,3b,4,5,10,11b,12,13,13a-decahydro-2H-7-oxa-indeno[5,4-a]anthracen-1-one).

In this study, the biological activity induced by an Amino-Steroid-Anthracenone derivative (ASA) with biological activity against ischemia-reperfusion injury was evaluated.

3.1. First Stage

3.1.1. Effect Exerted of ASA on Ischemia/Reperfusion Injury

The results (Fig. 2) showed that the ASA reduces infarct size significantly (p = 0.05) expressed as a percentage of the area at a risk compared with vehicle-treated hearts (control).

Fig. (2) Biological activity exerted by an amino-steroid-anthracenone derivative (ASA; 0.001-100 nM) against ischemia-reperfusion injury (translated as infarct area). The experimental data showed that ASA decreased infarct size (expressed as a percentage of the area at risk) in a dose-dependent manner compared with the vehicle-treated hearts (control). Each bar represents the mean ± S.E. of 9 experiments.

3.1.2. Biological Activity Induced by Both Estrone and ASA on Ischemia/Reperfusion Injury

The experimental data (Fig. 3) showed that the infarct area was significantly reduced (p = 0.05) in the presence of ASA compared with estrone in a dose-dependent manner (0.001 to 100 nM).

Fig. (3) Effect exerted by both estrone and an amino-steroid-anthracenone derivative (ASA; 0.001-100 nM) against ischemia-reperfusion injury (translated as infarct area). The results showed that ASA significantly (p = 0.05) reduced infarct size expressed as a percentage of the area at risk compared with the estrone. Each data points represents the mean ± S.E. of 9 experiments.

3.2. Second Stage

3.2.1. Effects Induced by ASA on Left Ventricular Pressure Through Estrogen Receptor Activation

Fig. 4 shows that ASA (0.001 to 100 nM) decreased the left ventricular pressure in a dose-dependent manner, both in the absence or the presence of tamoxifen (1 nM).

Fig. (4) Biological activity induced by an Amino-Steroid-Anthracenone derivative (ASA; 0.001-100 nM) on Left Ventricular Pressure (LVP) in absence or presence of tamoxifen. The experimental results showed that ASA significantly decreased LVP (p = 0.05) in a dose-dependent manner and this effect was not inhibited in presence of tamoxifen. Each data points represents the mean ± S.E. of 9 experiments.

3.2.2. Effects Induced by the ASA on Left Ventricular Pressure Through Β₂-Adrenergic Receptor Stimulation

The experimental data (Fig. 5) indicated that PDC (0.001 to 100 nM) decreased the left ventricular pressure in a dose-dependent manner in the absence or the presence of butoxamine [1 nM].

Fig. (5) Effect exerted by an Amino-Steroid-Anthracenone (ASA; 0.001-100 nM) on Left Ventricular Pressure (LVP) in absence or presence of butoxamine. The results showed that ASA significantly decreased LVP (p = 0.05) in a dose-dependent manner and this effect was not inhibited in presence of butoxamine. Each data points represents the mean ± S.E. of 9 experiments.

3.2.3. Effect Exerted by ASA on Left Ventricular Pressure Through A₂-Purinergic Receptor Activation

The results shown in Fig. 6 showed that ASA (0.001 to 100 nM) decreased the left ventricular pressure in a dose-dependent manner and this effect was inhibited in the presence of ZM-241,385 [1 nM].

Fig. (6) Biological activity exerted by an Amino-Steroid-Anthracenone derivative (ASA) on Left Ventricular Pressure (LVP) in absence or presence of the compound ZM241385. The experimental results showed that ASA significantly decreased LVP (p = 0.05) in a dose-dependent manner and this effect was inhibited by ZM241385. Each data points represents the mean ± S.E. of 9 experiments.

3.2.4. Effect Exerted of ASA on cAMP Concentration

The results (Fig. 7) showed that ASA (1 nM/min) increased the cAMP levels in a time-dependent manner (3-18 min).

Fig. (7) Effect exerted by an Amino-Steroid-Anthracenone derivative (ASA) on cAMP levels in a time dependent manner (3 - 18 min). The experimental data showed that ASA increases cAMP levels (p = 0.05) with increase in time (3-18 min) compared to the control. Each bar represents the mean ± S.E. of 9 experiments.

There are several drugs that have been used to treat heart failure resulting from myocardial ischemia [30Kersten JR, Montgomery MW, Pagel PS, Warltier DC. Levosimendan, a new positive inotropic drug, decreases myocardial infarct size via activation of K(ATP) channels. Anesth Analg 2000; 90(1): 5-11.
[http://dx.doi.org/10.1097/00000539-200001000-00003] [PMID: 10624967] , 31Node K, Kitakaze M, Kosaka H, Minamino T, Funaya H, Hori M. Amelioration of ischemia- and reperfusion-induced myocardial injury by 17β-estradiol: Role of nitric oxide and calcium-activated potassium channels. Circulation 1997; 96(6): 1953-63.
[http://dx.doi.org/10.1161/01.CIR.96.6.1953] [PMID: 9323086] ]. Nevertheless, there is scarce information about the effects of the Amino-Steroid-Anthracenone (ASA) on this clinic pathology. Therefore, in this study, the biological activity of an ASA against myocardial ischemia translated as infarct area was evaluated using an ischemia-reperfusion injury model [32Lauro FV, Francisco DC, Elodia GC, et al. Evaluation of activity of an estrogen-derivative as cardioprotector drug using an ischemia-reperfusion injury model. Int J Clin Exp Med 2015; 8(8): 12041-55.
[PMID: 26550116] ]. The results showed that ASA reduces infarct size (expressed as a percentage of the area at risk) to different doses compared with conditions control. In order to evaluate whether the biological activity exerted by ASA could be related to the steroid nucleus involved in their chemical structure, in this study, estrone was used as a pharmacological tool. The experimental data showed that ASA reduced infarct size, in a dose-dependent manner, compared with estrone; these data suggest that possibly the pyran ring or the functional groups bound to pyran ring of ASA could be responsible for its biological activity. This finding is supported by some reports which indicate that several pyran derivatives can exert an effect on the cardiovascular system [33Ashwood VA, Cassidy F, Evans JM, Gagliardi S, Stemp G. Synthesis and antihypertensive activity of pyran oxygen and amide nitrogen replacement analogues of the potassium channel activator cromakalim. J Med Chem 1991; 34(11): 3261-7.
[http://dx.doi.org/10.1021/jm00115a015] [PMID: 1659637] -35Auchampach JA, Maruyama M, Cavero I, Gross GJ. The new K+ channel opener Aprikalim (RP 52891) reduces experimental infarct size in dogs in the absence of hemodynamic changes. J Pharmacol Exp Ther 1991; 259(3): 961-7.
[PMID: 1762089] ] and can reduce the area infarction by inducing changes in left ventricular pressure [36Lee B, Seo H, Yoo S, Kim S, Lim H, Shin H. Differential action of KR 31378, a novel potassium channel activator, on cardioprotective and hemodynamic effects. Drug Dev Res 2001; 54(4): 182-90.
[http://dx.doi.org/10.1002/ddr.10028] , 37Lee BH, Seo HW, Yoo SE. Cardioprotective effects of (2S,3R,4S)-N′-benzyl-N"-cyano-N-(3,4-dihydro-2-dimethoxymethyl-3-hydroxy-2-methyl-6-nitro-2H-benzopyran-4-yl)-guanidine (KR-31372) in rats and dogs. Pharmacology 2004; 70(2): 74-82.
[http://dx.doi.org/10.1159/000074671] [PMID: 14685010] ]. Analyzing these data, in this study, the effect exerted by ASA on left ventricular pressure was evaluated; the results showed that ASA decreases the left ventricular pressure in a dose-dependent manner compared with estrone and conditions control.

On the other hand, in the search for a molecular mechanism involved in biological activity induced by ASA on left ventricular pressure, earlier reports indicated that several compounds can induce changes in the left ventricular pressure through estrogen receptor activation [38Skavdahl M, Steenbergen C, Clark J, et al. Estrogen receptor-beta mediates male-female differences in the development of pressure overload hypertrophy. Am J Physiol Heart Circ Physiol 2005; 288(2): H469-76.
[http://dx.doi.org/10.1152/ajpheart.00723.2004] [PMID: 15374829] , 39Donaldson C, Eder S, Baker C, et al. Estrogen attenuates left ventricular and cardiomyocyte hypertrophy by an estrogen receptor-dependent pathway that increases calcineurin degradation. Circ Res 2009; 104(2): 265-75.
[http://dx.doi.org/10.1161/CIRCRESAHA.108.190397] [PMID: 19074476] ]. To evaluate this hypothesis, in this study, the biological activity exerted by ASA on left ventricular pressure in the absence or presence of tamoxifen was evaluated. The results showed that ASA decreases left ventricular pressure and this effect was not inhibited by tamoxifen; this phenomenon indicated that molecular mechanism involved in the biological activity of ASA on left ventricular pressure was not via estrogen-receptor activation.

Since earlier reports indicated that some compounds can exert changes on left ventricular pressure through β₂-adrenergic receptor activation [40Levy B. The adrenergic blocking activity of N-tert.-butylmethoxamine (butoxamine). J Pharmacol Exp Ther 1966; 151(3): 413-22.
[PMID: 4380047] ], in this study, the biological activity of ASA on left ventricular pressure was evaluated in the absence or presence of butoxamine (β₂-adrenergic receptor inhibitor) [41Akatsuka Y, Egashira K, Katsuda Y, et al. ATP sensitive potassium channels are involved in adenosine A2 receptor mediated coronary vasodilatation in the dog. Cardiovasc Res 1994; 28(6): 906-11.
[http://dx.doi.org/10.1093/cvr/28.6.906] [PMID: 7923297] ]. The results showed that ASA decreased left ventricular pressure and this effect was not inhibited by butoxamine; this phenomenon indicated that molecular mechanism involved in the biological activity of ASA on left ventricular pressure was not via β₂-adrenergic receptor activation.

On the other hand, other reports indicated that some substances can exert changes in the pressure of the left ventricle through A₂-adenosine receptor and this phenomenon may result in a decrease in the infarct area [41Akatsuka Y, Egashira K, Katsuda Y, et al. ATP sensitive potassium channels are involved in adenosine A2 receptor mediated coronary vasodilatation in the dog. Cardiovasc Res 1994; 28(6): 906-11.
[http://dx.doi.org/10.1093/cvr/28.6.906] [PMID: 7923297] , 42Jordan JE, Zhao ZQ, Sato H, Taft S, Vinten-Johansen J. Adenosine A2 receptor activation attenuates reperfusion injury by inhibiting neutrophil accumulation, superoxide generation and coronary endothelial adherence. J Pharmacol Exp Ther 1997; 280(1): 301-9.
[PMID: 8996210] ]. Therefore, in this study, the effect exerted by ASA on left ventricular pressure was determined, both in the absence or presence of ZM241385 (A₂-receptor adenosine antagonist) [43Keddie JR, Poucher SM, Shaw GR, Brooks R, Collis MG. In vivo characterisation of ZM 241385, a selective adenosine A2A receptor antagonist. Eur J Pharmacol 1996; 301(1-3): 107-13.
[http://dx.doi.org/10.1016/0014-2999(96)00020-9] [PMID: 8773453] ] The results showed that ASA decreases left ventricular pressure and this effect was inhibited by ZM241385. These data indicated that the effect exerted by ASA on left ventricular pressure was via A₂-adenosine receptor activation.

On the other hand, it is important to mention that the interaction of some substances with A₂-adenosine receptor could involve the synthesis and release of cAMP [44Smits GJ, McVey M, Cox BF, Perrone MH, Clark KL. Cardioprotective effects of the novel adenosine A1/A2 receptor agonist AMP 579 in a porcine model of myocardial infarction. J Pharmacol Exp Ther 1998; 286(2): 611-8.
[PMID: 9694911] ]. Therefore, in this study, an experimental alternative was carried out to evaluate the possibility that ASA could exert changes on cAMP levels; the results showed that ASA significantly increases the cAMP concentration compared with the conditions control.

The Amino-Steroid-Anthracenone derivative (ASA) is a particularly interesting compound because the biological activity exerted against ischemia-reperfusion injury involves a molecular mechanism different in comparison with other drugs; this phenomenon may constitute a novel therapy for ischemia-reperfusion injury translated as a decrease in the heart failure; however, it is important to carry out toxicity studies of this compound to rule out any adverse effect.