Cardiac contractility is regulated by several neural, hormonal, paracrine, and autocrine factors. Amongst these, signaling through β-adrenergic and serotonin receptors generates the second messenger cyclic AMP (cAMP), whereas activation of natriuretic peptide receptors and soluble guanylyl cyclases generates cyclic GMP (cGMP). Both cyclic nucleotides regulate cardiac contractility through several mechanisms. Phosphodiesterases (PDEs) are enzymes that degrade cAMP and cGMP and therefore determine the dynamics of their downstream effects. In addition, the intracellular localization of the different PDEs may contribute to regulation of compartmented signaling of cAMP and cGMP. In this review, we will focus on the role of PDEs in regulating contractility and evaluate changes in heart failure.

Autoimmune diseases are highly prevalent and affect people at all ages, women more often than men. The most prominent immunological manifestation is the production of antibodies directed against self-antigens. In many cases, these antibodies (Abs) drive the pathogenesis by attacking the body’s own healthy cells, causing serious health problems that may be life threatening. Most autoantibodies are of the immunoglobulin G (IgG) isotype, which has a long plasma half-life and potent effector functions. Thus, there is a need for specific treatment options that rapidly eliminate these pathogenic IgG auto-Abs. In this review, we discuss how the neonatal Fc receptor (FcRn) acts as a regulator of the high levels of not only IgG Abs, but also albumin, by rescuing both these soluble proteins from cellular catabolism, and how a molecular and cellular understanding of this complex biology has spurred an intense interest in the development of FcRn-targeting strategies for the treatment of IgG-driven autoimmune diseases. We find that this emerging therapeutic class demonstrates efficacy within several autoimmune diseases with distinct pathophysiology. This offers hope for both new therapeutic avenues for highly prevalent diseases currently treated by other means, and rare diseases with no approved therapies to date. In addition, we elaborate on studies that have led to approval of the first FcRn antagonists, the clinical progress and structural design of molecules in the pipeline, their position in the overall therapeutic landscape of autoimmunity, the design of next-generation antagonists as well as the use of this receptor-targeting principle for other therapeutic applications.

Aims: Guanylyl cyclase-B (GC-B; natriuretic peptide receptor-B, NPR-B) stimulation by C-type natriuretic peptide (CNP) increases cGMP and causes a lusitropic and negative inotropic response in adult myocardium. These effects are not mimicked by NPR-A (GC-A) stimulation by brain natriuretic peptide (BNP), despite similar cGMP increase. More refined methods are needed to better understand the mechanisms of the differential cGMP signalling and compartmentation. The aim of this work was to measure cGMP near proteins involved in regulating contractility to understand compartmentation of cGMP signalling in adult cardiomyocytes. Methods and results: We constructed several fluorescence resonance energy transfer (FRET)-based biosensors for cGMP subcellularly targeted to phospholamban (PLB) and troponin I (TnI). CNP stimulation of adult rat cardiomyocytes increased cGMP near PLB and TnI, whereas BNP stimulation increased cGMP near PLB, but not TnI. The phosphodiesterases PDE2 and PDE3 constrained cGMP in both compartments. Local receptor stimulation aided by scanning ion conductance microscopy (SICM) combined with FRET revealed that CNP stimulation both in the t-tubules and on the cell crest increases cGMP similarly near both TnI and PLB. In ventricular strips, CNP stimulation, but not BNP, induced a lusitropic response, enhanced by inhibition of either PDE2 or PDE3, and a negative inotropic response. In cardiomyocytes from heart failure rats, CNP increased cGMP near PLB and TnI more pronounced than in cells from sham-operated animals. Conclusion: These targeted biosensors demonstrate that CNP, but not BNP, increases cGMP near TnI in addition to PLB, explaining how CNP, but not BNP, is able to induce lusitropic and negative inotropic responses.

The therapeutic benefit of stimulating the cGMP pathway as a form of treatment to combat heart failure, as well as other fibrotic pathologies, has become well established. However, the development and signal compartmentation of this crucial pathway has so far been overlooked. We studied how the three main cGMP pathways, namely, nitric oxide (NO)-cGMP, natriuretic peptide (NP)-cGMP, and β₃-adrenoreceptor (AR)-cGMP, mature over time in culture during cardiomyocyte differentiation from human pluripotent stem cells (hPSC-CMs). After introducing a cGMP sensor for Förster Resonance Energy Transfer (FRET) microscopy, we used selective phosphodiesterase (PDE) inhibition to reveal cGMP signal compartmentation in hPSC-CMs at various times of culture. Methyl-β-cyclodextrin was employed to remove cholesterol and thus to destroy caveolae in these cells, where physical cGMP signaling compartmentalization is known to occur in adult cardiomyocytes. We identified PDE3 as regulator of both the NO-cGMP and NP-cGMP pathway in the early stages of culture. At the late stage, the role of the NO-cGMP pathway diminished, and it was predominantly regulated by PDE1, PDE2, and PDE5. The NP-cGMP pathway shows unrestricted locally and unregulated cGMP signaling. Lastly, we observed that maturation of the β₃-AR-cGMP pathway in prolonged cultures of hPSC-CMs depends on the accumulation of caveolae. Overall, this study highlighted the importance of structural development for the necessary compartmentation of the cGMP pathway in maturing hPSC-CMs.

Natriuretic peptides (NPs) increase cGMP, show beneficial cardiovascular effects and regulate energy metabolism in other tissues. However, little is known about their direct effect on cardiac mitochondria and cardiomyocyte apoptosis. Here, we examined whether NPs increase cGMP around mitochondria and alter apoptosis in cardiomyocytes. We constructed a novel FRET-based biosensor with high selectivity towards cGMP and found that ANP and CNP increase cGMP at the outer mitochondrial membrane. Moreover, ANP and CNP increased phosphorylation of the pro-apoptotic protein Drp1 and CNP prevented fragmentation of mitochondria. Stimulating cardiomyocytes with ANP or CNP reduced apoptosis, caspase 9 activation and cytochrome c release, suggesting that NPs decrease apoptosis through the intrinsic pathway that involves mitochondria. We suggest that cGMP increase in the outer mitochondrial membrane microdomain that inhibits the pro-apoptotic protein Drp1, leading to reduced mitochondrial fragmentation and thereby reduced apoptosis.

The natriuretic peptides ANP, BNP and CNP activate transmembrane guanylyl cyclases (GC) that producecyclic GMP (cGMP). We have previously employed targeted FRET-based biosensors to demonstrate that thenatriuretic peptides have differential effects in cardiomyocytes and intact heart, where CNP activates GC-B thatincreases cGMP near troponin I and phospholamban (PLB), enhancing relaxation, while activation of GC-A withANP/BNP modestly increases cGMP only near PLB and do not enhance relaxation, suggesting spatiallyrestricted cGMP signaling from GC-A and GC-B. Using a biosensor targeted to the outer mitochondrialmembrane (OMM), we have found that GC-A and GC-B increase cGMP at the OMM and reduce cardiomyocyteapoptosis. To understand this spatial cGMP signaling from GC-A and GC-B, we combined targeted variants ofthe cGMP scavenger SponGee with our targeted FRET-based cGMP biosensors in cardiac H9c2 cells.Activation of GC-B increased cytosolic cGMP, which was modestly reduced by the lipid raft (Lyn-SponGee),non-raft (SponGee-Kras) or OMM-targeted (OMM-SponGee) cGMP scavengers compared to the untargetedSponGee. At the OMM, cGMP increase from GC-B was reduced in content and kinetics only by the OMMSponGee,while cGMP increase from GC-A was reduced by SponGee-Kras and the untargeted SponGee.Our results indicate that GC-A and GC-B are differentially organized on the plasma membrane and that cGMPreaching the OMM could have different origin. Using cGMP scavengers can therefore be used to deciphersignaling from different areas of the plasma membrane to various subcellular locations in cardiac cells.