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Diabetic Nephropathy (DN) is the leading cause of end-stage renal disease. Preclinical and experimental studies show that PDE5 inhibitors (PDE5is) exert protective effects in DN improving perivascular inflammation. Using a mouse model of diabetic kidney injury we investigated the protective proprieties of PDE5is on renal hemodynamics and the molecular mechanisms involved. PDE5i treatment prevented the development of DN-related hypertension (P

RationaleIn heart failure, myofilament proteins display abnormal phosphorylation, which contributes to contractile dysfunction. The mechanisms underlying the dysregulation of protein phosphorylation on myofilaments is not clear.ObjectiveThis study aims to understand the mechanisms underlying altered phosphorylation of myofilament proteins in heart failure.Methods and resultsWe generate a novel genetically encoded protein kinase A (PKA) biosensor anchored onto the myofilaments in rabbit cardiac myocytes to examine PKA activity at the myofilaments in responses to adrenergic stimulation. We show that PKA activity is shifted from the sarcolemma to the myofilaments in hypertrophic failing rabbit myocytes. In particular, the increased PKA activity on the myofilaments is because of an enhanced β2 adrenergic receptor signal selectively directed to the myofilaments together with a reduced phosphodiesterase activity associated with the myofibrils. Mechanistically, the enhanced PKA activity on the myofilaments is associated with downregulation of caveolin-3 in the hypertrophic failing rabbit myocytes. Reintroduction of caveolin-3 in the failing myocytes is able to normalize the distribution of β2 adrenergic receptor signal by preventing PKA signal access to the myofilaments and to restore contractile response to adrenergic stimulation.ConclusionsIn hypertrophic rabbit myocytes, selectively enhanced β2 adrenergic receptor signaling toward the myofilaments contributes to elevated PKA activity and PKA phosphorylation of myofilament proteins. Reintroduction of caveolin-3 is able to confine β2 adrenergic receptor signaling and restore myocyte contractility in response to β adrenergic stimulation.

Phosphodiesterase 5A (PDE5A) is involved in cGMP hydrolysis, regulating many physiological processes. Increased activity of PDE5A has been found in several pathological conditions, and the pharmacological inhibition of PDE5 has been demonstrated to have several therapeutic applications. We have identified the presence of three different Pde5a isoforms in cardiomyocytes, and we have found that the expression of specific Pde5a isoforms may have a causal role in the onset of pathological responses in these cells. In our previous study, we demonstrated that PDE5A inhibition could ameliorate muscular dystrophy by acting at different levels, as assessed by the altered genomic response of muscular cells following treatment with the PDE5A inhibitor tadalafil. Thus, considering the importance of PDE5A in various pathophysiological conditions, we further investigated the regulation of this enzyme. Here, we analysed the expression of Pde5a isoforms in the pathophysiology of skeletal muscle. We found that skeletal muscle tissues and myogenic cells express Pde5a1 and Pde5a2 isoforms, and we observed an increased expression of Pde5a1 in damaged skeletal muscles, while Pde5a2 levels remained unchanged. We also cloned and characterized the promoters that control the transcription of Pde5a isoforms, investigating which of the transcription factors predicted by bioinformatics analysis could be involved in their modulation. In conclusion, we found an overexpression of Pde5a1 in compromised muscle and identified an involvement of MyoD and Runx1 in Pde5a1 transcriptional activity. [ABSTRACT FROM AUTHOR]

[beta] adrenoceptor ([beta]AR) signaling is finely regulated to mediate the sympathetic nervous system control of cardiovascular function. In neonatal cardiac myocytes, [[beta].sub.1]AR activates the conventional [G.sub.s]/cAMP pathway, whereas [[beta].sub.2]AR sequentially activates both the [G.sub.s] and [G.sub.i] pathways to regulate the myocyte contraction rate. Here, we show that phosphodiesterase 4D (PDE4D) selectively impacts signaling by [[beta].sub.2]AR in neonatal cardiac myocytes, while having little or no effect on [[beta].sub.1]AR signaling. Although [[beta].sub.2]AR activation leads to an increase in cAMP production, the cAMP generated does not have access to the protein kinase A-dependent signaling pathways by which the [[beta].sub.1]AR regulates the contraction rate. However, this restricted access is lost in the presence of PDE4 inhibitors or after ablation of PDE4D. These results not only suggest that PDE4D is an integral component of the [[beta].sub.2]AR signaling complex, but also underscore the critical role of subcellular cAMP regulation in the complex control of receptor signaling. They also illustrate a mechanism for fine-tuned [beta]AR subtype signaling specificity and intensity in the cardiac system. camp | heart | knockout

Cytoplasmic [Ca.sup.2+] concentration ([[Ca.sup.2+]].sub.i]) variation is a key event in myoblast differentiation, but the mechanism by which it occurs is still debated. Here we show that increases of extracellular [Ca.sup.2+] concentration ([[[Ca.sup.2+]].sub.o]) produced membrane hyperpolarization and a concentration-dependent increase of [[[Ca.sup.2+]].sub.i] due to [Ca.sup.2+] influx across the plasma membrane. Responses were not related to inositol phosphate turnover and [Ca.sup.2+]-sensing receptor. [[[Ca.sup.2+]].sub.o]-induced [[[Ca.sup.2+]].sub.i] increase was inhibited by [Ca.sup.2+] channel inhibitors and appeared to be modulated by several kinase activities. [[[Ca.sup.2+]].sub.i] increase was potentiated by depletion of intracellular [Ca.sup.2+] stores and depressed by inactivation of the [Na.sup.+]/[Ca.sup.2+] exchanger. The response to arginine vasopressin (AVP), which induces inositol 1,4,5-trisphosphate-dependent [[[Ca.sup.2+]].sub.i] increase in L6-C5 cells, was not modified by high [[[Ca.sup.2+]].sub.o]. On the contrary, AVP potentiated the [[[Ca.sup.2+]].sub.i] increase in the presence of elevated [[[Ca.sup.2+]].sub.o]. Other clones of the L6 line as well as the rhabdomyosarcoma RD cell line and the satellite cell-derived C2-C12 line expressed similar responses to high [[[Ca.sup.2+]].sub.o], and the amplitude of the responses was correlated with the myogenic potential of the cells. calcium; myogenesis; calcium channels; sodium-calcium exchanger