**3. Angiotensin II and cardiac remodeling**

Ang II is the primary effector peptide of the RAS. The hypertrophic effects of this peptide on the heart are associated with its vasoconstrictor and hypertensive properties. However, it is currently known that independently of its blood pressure effects, Ang II is a powerful hypertrophic agent. *In vitro* studies show that Ang II activates different hypertrophic signaling pathways in cardiac myocytes [11]. In addition, the crucial components to initiate synthetic route to Ang II production are present in the heart. Thus, Ang II is also locally synthesized at the myocardium, acting as an autocrine factor [11]. Increased cardiac Ang II synthesis is mediated, *in vitro*, by cardiomyocyte under stretch conditions [14]. Similarly, the rise in cardiac Ang II was also observed in hypertrophied heart from animals after overload pressure as well as in patients from end-stage heart failure, which suggests the hypertrophy have resulted from local RAS activation [11, 15].

The Ang II hypertrophic effects are mediated by the activation of specific receptor AT1 that plays a crucial role in heart failure pathophysiology, but both AT1 and AT2 receptor are present in the cardiac tissue [16–19]. The AT1 receptor is 7-transmembrane domain coupled to Gq protein (GPCR). Ang II is able to perform the signaling transduction to adaptive and maladaptive remodeling pathway [2]. AT1 receptor stimulated by Ang II leads to the protein kinase C activation [17], which in turn activates the mitogen-activated protein kinase (MAPK). The intracellular signaling cascade generated from MAPK is constituted by a phosphorylationbased amplification network and results in hypertrophic signals to cardiac adaptive or maladaptive remodeling [2, 10]. Three MAPKs, such as p38 kinases, c-Jun-terminal kinases (JNK) and ERK 1/2 have been described as signaling pathways in cardiac myocytes or extracellular matrix changes along the heart failure progression [10].

In the compensatory response to overload pressure, ERK 1/2 activation has been related to adaptive changes and increased width of the myocytes [20]. Further, some studies have suggested that JNK could contribute to maladaptive remodeling due to its pivotal role in cell death [2, 10, 21]. The MAPK signaling from cardiomyocyte cytoplasm drives to nuclei where transcriptional factors such as factor nuclear kappa B (NF-kB), activating protein-1 (AP-1) and Smad are intracellular proteins to transduce extracellular signals from transforming growth factor beta ligands to the nucleus where they activate downstream gene transcription, rising the transcription of key proteins and developing essential function to the cardiac remodeling progression [22–25].

The NF-kB is an oxidative-sensitive transcriptional factor [26]. Likewise, multiple signal transduction pathways are activated in response to reactive oxygen species (ROS) [27]. In this regard, emergent evidences have shown that Ang II-mediated hypertrophic response may be dependent of increased ROS production, particularly during hypertension [25, 28–30].

Pathogenic cellular and interstitial changes in hypertension-induced cardiac remodeling are orchestrated by several molecular mechanisms that may be transduced from mechanical force into myocardial growth. In this regard, renin-angiotensin system (RAS) is activated in hypertension and may be involved in cardiac hypertrophy and failure. Clinical and experimental studies have shown significant benefit conferred by pharmacological blockade of RAS [7, 11–13] arousing interest by mechanisms underlying the action of angiotensin II (Ang II).

Ang II is the primary effector peptide of the RAS. The hypertrophic effects of this peptide on the heart are associated with its vasoconstrictor and hypertensive properties. However, it is currently known that independently of its blood pressure effects, Ang II is a powerful hypertrophic agent. *In vitro* studies show that Ang II activates different hypertrophic signaling pathways in cardiac myocytes [11]. In addition, the crucial components to initiate synthetic route to Ang II production are present in the heart. Thus, Ang II is also locally synthesized at the myocardium, acting as an autocrine factor [11]. Increased cardiac Ang II synthesis is mediated, *in vitro*, by cardiomyocyte under stretch conditions [14]. Similarly, the rise in cardiac Ang II was also observed in hypertrophied heart from animals after overload pressure as well as in patients from end-stage heart failure, which suggests the hypertrophy have resulted from

The Ang II hypertrophic effects are mediated by the activation of specific receptor AT1 that plays a crucial role in heart failure pathophysiology, but both AT1 and AT2 receptor are present in the cardiac tissue [16–19]. The AT1 receptor is 7-transmembrane domain coupled to Gq protein (GPCR). Ang II is able to perform the signaling transduction to adaptive and maladaptive remodeling pathway [2]. AT1 receptor stimulated by Ang II leads to the protein kinase C activation [17], which in turn activates the mitogen-activated protein kinase (MAPK). The intracellular signaling cascade generated from MAPK is constituted by a phosphorylationbased amplification network and results in hypertrophic signals to cardiac adaptive or maladaptive remodeling [2, 10]. Three MAPKs, such as p38 kinases, c-Jun-terminal kinases (JNK) and ERK 1/2 have been described as signaling pathways in cardiac myocytes or

In the compensatory response to overload pressure, ERK 1/2 activation has been related to adaptive changes and increased width of the myocytes [20]. Further, some studies have suggested that JNK could contribute to maladaptive remodeling due to its pivotal role in cell death [2, 10, 21]. The MAPK signaling from cardiomyocyte cytoplasm drives to nuclei where transcriptional factors such as factor nuclear kappa B (NF-kB), activating protein-1 (AP-1) and Smad are intracellular proteins to transduce extracellular signals from transforming growth factor beta ligands to the nucleus where they activate downstream gene transcription, rising the transcription of key proteins and developing essential function to the cardiac remodeling

extracellular matrix changes along the heart failure progression [10].

**3. Angiotensin II and cardiac remodeling**

54 Renin-Angiotensin System - Past, Present and Future

local RAS activation [11, 15].

progression [22–25].

Several studies have confirmed the key role of ROS in the genesis and progression of cardiac remodeling [28, 31, 32]. Low levels of ROS are important to many downstream regulators in a physiological condition such as ion channel, receptors, kinases, phosphatases and transcriptional factor. However, increased ROS production characterizes oxidative stress, disrupts redox signaling within the cells and the interstice, promoting activation of calcium/calmodulin-dependent protein kinase I (CaMKI), increased NF-kB, AP-1 and other transcriptional factors signaling [33, 34]. Oxidative stress also elicits post-transductional pathways that result in activation of some proteins, e.g., matrix metaloproteinases (MMP) [35]. Consequently, oxidative stress has been associated with cardiac contraction dysfunction, increased collagen deposition and myocytes hypertrophy that contribute to cardiac dysfunction, myocyte hypertrophy and cell death [27, 35].

Considering the relevance of ROS to cardiac diseases, a substantial body of studies has investigated which enzyme could be more important to ROS synthesis. Along the progression of cardiac remodeling, a family of complex enzymes termed nicotinamide adenine dinucleotide phosphate-oxidase (NADPH oxidase) seems to play a central position to ROS production [27, 29]. Increased expression and activity of NADPH oxidase have been persistently observed in both preclinical and clinical studies of heart failure [27, 36]. There are seven Nox family isoforms (Nox1-5 and DUOX1 and 2), and the main cardiac enzymes are Nox2 and Nox4 [37].

Nox4 contribute to myocyte hypertrophy and cardiac fibrosis induced by AngII [38]. However, the role of Nox4 to cardiac hypertrophy is not yet fully comprehended [39].

Amount evidences show Nox2 associated with detrimental effects in the heart [27, 39]. The low-level activity of Nox2 is continuously present in the presence of nanomolar ROS levels but may be increased at the Ang II, endothelin, transforming growth factor (TGF)-β, tumor necrosis factor (TNF)-α presence as well as due to mechanical force [27]. Interestingly, Ang II-induced cardiac hypertrophy and fibrosis were reduced in knockout mice for Nox2 when compared to the wild type [38]. Currently, the contribution of Nox2 to Ang II hypertrophic effects appears to involve ERK1/2, Akt and NF-kB signaling [27, 41, 43]. In addition, increased Ang II-induced MMP activation and expression seem to be dependent of ROS [30, 42, 44] resulting in cardiac adaptive remodeling and fibrosis [43, 44]. **Figure 2** summarizes relationship between Ang IIinduced cardiac remodeling and Nox2.

The important signaling in Ang II-induced fibrosis predominantly requires the differentiation from fibroblast into myofibroblast cells [3]. This phenotypic transformation from fibroblast is characterized by α-smooth muscle actin (α-SMA) expression and increased production of extracellular matrix, which is a key event in connective tissue remodeling involved in the heart failure progression [3, 45]. Rossi [9] has shown an intensive and progressive accumulation of collagen,accompaniedbyincreasedheartweightinhypertensivesubjects.Inaddition,thestudy revealed an association among connective matrix, cardiac systolic and diastolic dysfunction in

**Figure 2.** Main pathways concerning Ang II-induced cardiac remodeling. Nox2 is activated by Ang II via AT1 receptor, triggering ROS formation, which activates intracellular pathways related to the cardiac hypertrophy. Nox2: NADPH oxidase isoform 2; ROS: reactive oxygen species; Akt: serine/threonine-specific protein kinase; ERK1/2: extracellular.

hypertension suggesting that collagen deposition could contribute to decreased myocardial complianceanddisruptedheartelectricalproperties[9].Currently,itiswellknowntherelevance of fibrosis not only to the structure of the cardiac hypertrophy but also to the heart dysfunction [46]. Collagen is the main component of the extracellular matrix in the myocardium, which is synthesized by fibroblasts. However, its deposition in the heart during hypertension also depends of its degradation [3]. Thus, a large body of studies has shown the contribution of the MMP, which is the main proteases to collagen degradation, strongly contributing to the cardiac remodeling after pressure overload or infarct [28, 47]. The imbalances between MMP and endogenous tissue inhibitor (TIMP) are key mechanisms to control the collagen formation and deposition[2,3,48].Indeed, some transcriptionalfactors suchasAP-1 andNF-kBmaymodulate the MMP activity increasing its MMP expression and also TIMP expression [28, 47]. Posttranslationalmechanisms suchasoxidativestress,particularlyperoxynitriteandhydrogenperoxide, may activate MMP and inhibit TIMP activity [35, 49, 50], suggesting a possible mechanism to ROS-induced fibrosis in hypertensive rats [51]. Thus, MMP activity is regulated at three levels: (i) transcriptional level, (ii) endogenous inhibitors and (iii) factor activators (ROS). Interestingly, Ang II may increase MMP activity involving redox-sensitive signaling in fibroblast, thus triggering NF-kB and AP-1 transcriptional factor activation [52]. In fact, antioxidant therapy reverses Ang II-induced cardiac hypertrophy and MMP activity in left ventricle from hypertensive rats [30]. Taken together, Ang II promotes myocyte on the heart and matrix extracellular hypertrophybysimilarmechanismsinvolvingredoxsignaling,whichnotonlyactivatestheRNA expressionofproteinsinthemyocytesorfibroblasts,butalsorisestheactivityofenzymesalready present in the heart, such as MMP.

Furthermore, it must be recognized that Ang II induces inflammation by triggering cardiac remodeling. The proinflammatory effects of Ang II have been described since 1970 by Finn Olsen [53]. Thenceforward, several studies have supported the contribution of the inflammatory processes associated with Ang II to cardiovascular disorders, including hypertension and heart remodeling [23, 25, 54]. Since inflammation contributes to this important clinical condition, numerous evidences have reported the connection between Ang II and two pivotal mediators for heart remodeling, the cytokines transforming growth factor (TGF)-β [23] and the tumoral necrosis factor (TNF)-α [25].
