One well-described signalling pathway involved in the regulation of hypertrophic
responses is the 3’, 5-cyclic guanosine monophosphate (cGMP)/ protein kinase G (PKG) signalling pathway. Generation of cGMP by NO activation of soluble guanylyl cyclases (sGC) was shown to reduce the hypertrophic response to norepinephrine (NE) (Calderone et al. 1998) or angiotensin II (Ang II) stimulation (Ritchie et al. 1998). Atrial and brain natriuretic peptides (ANP and BNP) which, by activating the membrane-bound particulate guanylyl cyclases (sGC) stimulate the production of cGMP, have also been found to exert anti-hypertrophic effects in vitro (Calderone et al. 1998; Rosenkranz et al. 2003). ANP and BNP are found at high levels during embryonic development and in early neonates but are absent in healthy adults (Gardner 2003), however in the hypertrophic heart the expression levels of ANP and BNP are found to be significantly increased (Silberbach and Roberts 2001; Tremblay et al. 2002; Molkentin 2003). In fact, expression of the genes encoding ANP and BNP is one of the most reliable markers for activation of the hypertrophic program in clinical states and experimental models associated with hypertrophy. Some recent studies have reported that natriuretic peptides exert a regulatory influence over cardiac hypertrophy. Neonatal rat ventricular myocytes (NRVM) stimulated with an ANP receptor antagonist lead to increased protein syntheses and cell size as well as reactivation of the fetal gene program. Zaprinast, a cGMP analogue and an inhibitor for a cGMP- specific phosphodiesterase, suppressed the basal and PE-stimulated protein syntheses (Horio et al. 2000). Thus, ANP may play a role as an autocrine factor in the regulation of cardiac myocyte growth. Another study reported that in NRVM and cardiac fibroblasts, treatment with ANP or the NO donor S-nitroso-N-acetyl-D, L- penicillamine (SNAP) attenuated the effects of NE-induced hypertrophy. The authors suggest that this effect was most likely by a cGMP-mediated inhibition of NE-stimulated Ca2+ influx (Calderone et al. 1998).
Recent studies using targeted gene deletion in mice have been particularly informative with regard to the role of the NPs in modulating cardiac hypertrophy. Deletion of the Npr1 gene encoding the natriuretic peptide receptor A (NPR-A) in mice resulted in moderate
elevations in blood pressure, with a disproportionate increase in cardiac weight (i.e. hypertrophic growth) and interstitial fibrosis (Lopez et al. 1995; Oliver et al. 1997). Knowles and colleagues used transverse aortic constriction (TAC) to induce pressure overload in the Npr1 -/- mice, which resulted in a 15-fold increase in atrial natriuretic
peptide (ANP) expression, a 55% increase in left ventricular weight/body weight
(LV/BW), dilatation of the LV, and significant decline in cardiac function (Knowles et al. 2001). In TAC treated wild-type mice, the authors described only a three-fold increase in ANP expression, an 11% increase in LV/BW, a 0.2 mm decrease in LV end diastolic dimension, and no change in fractional shortening (Knowles et al. 2001). These results suggest that the NPR-A system has direct anti-hypertrophic actions in the heart.
Failure of the natriuretic peptide system, at the ligand, receptor or post-receptor level, could account for the progression in cardiac dysfunction that accompanies longstanding hypertrophy and heart failure.
The role of cGMP/ PKG effect in cardiac hypertrophy has been described in various knockout mice lacking elements of this signalling pathway. Holtwick and colleagues showed that mice with a cardiomyocyte specific deletion of constituently active guanylyl cyclase-A (GC-A), which is normally activated by natriuretic peptides to produce cGMP, exhibited mild cardiac hypertrophy and an increase in mRNA expression of fetal genes ANP, α-skeletal actin and β-myosin heavy chain. Pressure overload induced by transverse aortic constriction (TAC) in this mouse model lead to an exaggerated hypertrophic
response as well as enhanced cardiac fibrosis and marked cardiac dysfunction (Holtwick et al. 2003). In the same year, Zahabi et al engineered transgenic mice that overexpressed a catalytic fragment of the GC-A domain of the atrial natriuretic peptide receptor in a
cardiomyocyte-specific manner. They found increased GC-A attenuated the effects of both ISO treatment and abdominal aortic constriction on cardiac wall thickness, cardiomyocyte size and prevented the onset of the fetal gene expression program (Zahabi et al. 2003). Mice lacking eNOS and/or nNOS also develop cardiac hypertrophy, dysfunction and increased premature mortality (Barouch et al. 2003; Li et al. 2004; Flaherty et al. 2007). Conventional PKG-I knockout mice die young and perhaps too early to develop any cardiac hypertrophy (Pfeifer et al. 1998). Wollert and colleagues showed that in vitro adenoviral overexpression of PKG-Iβ in neonatal rat cardiac myocytes enhanced the anti- hypertrophic effect of NO in phenylephrine-stimulated cardiac myocytes (Fiedler et al. 2002; Wollert et al. 2002). However, the significance of the PKG pathway in modulating cardiac hypertrophy has been recently questioned. Lukowski and colleagues showed that, when compared to wild type mice, the degree of cardiac hypertrophy induced either by isoproterenol (ISO) infusion or by TAC was not changed in total PKG knock-out mice or in mice lacking PKG specifically in cardiomyocytes, indicating that the development of
cardiac hypertrophy is not amplified by the absence of endogenous PKG (Lukowski et al. 2010).
It has been reported that enhancement of the cGMP/ PKG pathway by inhibiting PDE mediated hydrolysis of cGMP reduces cardiac hypertrophy (Takimoto et al. 2005; Miller et al. 2009). Takimoto and colleagues showed that pharmacological inhibition of PDE5 with Sildenafil reverses hypertrophy, suppresses remodelling and improves cardiac function in mouse hearts subjected to thoracic aortic constriction (TAC). In addition, the authors suggested that the anti-hypertrophic effects of Sildenafil are mediated by the cGMP/PKG signalling pathway (Takimoto et al. 2005). Miller et al. published data which indicated that PDE1 inhibition with IC86340 is able to attenuate the hypertrophic response induced by chronic isoproterenol infusion in vivo (Miller et al. 2009). The authors also showed that expression levels of the dual-specific PDE1 are significantly up-regulated in various in vivo and in vitro models of hypertrophy. Results obtained with selective PKG or PKA inhibitors, indicated that PDE1 seems to regulate cardiac hypertrophy via modulation of cGMP/PKG signalling pathways (Miller et al. 2009). The role of PDE5 in regulating cardiac hypertrophy was also questioned by Lukowski and co-workers, as endogenous PDE5 could not be detected in cardiomyocytes from either wild type mice or mice lacking cardiac-specific PKG (Lukowski et al. 2010). Further studies are required to fully
understand the role of cGMP/ PKG in cardiac hypertrophy.