A polymorphism within the promoter of the dopamine receptor D1 (DRD1 -48A7G) associates with impaired kidney function in white hypertensive patients

Dopamine DRD1 receptor regulates renal function and vascular resistance. It plays a role in the pathogenesis of hypertension in animal models. In humans, the DRD1 gene presents a A-48G polymorphism associated to hypertension in a Japanese population. To explore the role of this polymorphism on blood pressure and renal function in Caucasian hypertensive patients (H), we evaluated the allele frequencies in a populations of 697 H and 100 blood volunteers, and found no difference in the distribution of the alleles between the two groups (AA;AG;GG: 13%;50%;37%; and 12%; 51%;36% respectively). In H, we found a significant difference between AA and GG in serum creatinine (AA: 1.06±.08 mg/dl; GG:0.97±0.02 mg/dl, p<0.03). Treatment restored serum creatinine at levels comparable between genotypes (AA: 0.99±0.03 mg/dl; GG: 0.94±0.02 mg/dl, n.s.). To replicate the finding, in a case control study of 8 AA and 7 GG hypertensive patients matched for age, sex and body mass index, in pharmacological wash out for 30 days, we evaluated serum (Creatinine, Na, Uric Acid, Urea) and urinary (volume/24h, protein/24h, creatinine clearance/24h) biochemistry and renal hemodynamic assessed by ultrasound. Once again, the AA group showed higher serum creatinine, Na, Uric acid and urea, reduced creatinine clearance and a higher level of urinary protein excretion. These changes occurred while no differences were observed in diuresis and renal vascular resistances. In conclusions, the DRD1 A-48G polymorphism identifies a class of H that is prone to hypertension related kidney alterations

Calcium-calmodulin-dependent kinase II (CaMKII) mediates insulin-stimulated proliferation and glucose uptake.

Cellular growth and glucose uptake are regulated by multiple signals generated by the insulin receptor. The mechanisms of individual modulation of these signals remain somewhat elusive. We investigated the role of CaMKII in insulin signalling in a rat skeletal muscle cell line, demonstrating that CaMKII modulates the insulin action on DNA synthesis and the negative feedback that down regulates glucose uptake. Insulin stimulation generated partly independent signals leading to the rapid activation of Akt, Erk-1/2 and CaMKII. Akt activation was followed by Glut-4 translocation to the plasma membrane and increase of glucose uptake. Then, IRS-1 was phosphorylated at S612, the IRS-1/p85PI3K complex was disrupted, Akt was no more phosphorylated and both Glut-4 translocation and glucose uptake were reduced. Inhibition of CaMKII abrogated the insulin-induced Erk-1/2 activation, DNA synthesis and phosphorylation of IRS-1 at S612. Inhibition of CaMKII also abrogated the down-regulation of insulin-stimulated Akt phosphorylation, Glut-4 membrane translocation and glucose uptake. These results demonstrate that: 1 — CaMKII modulates the insulin-induced Erk-1/2 activation and cell proliferation; 2 — after the initial stimulation of the IRS-1/Akt pathway, CaMKII mediates the down- regulation of stimulated glucose uptake. This represents a novel mechanism in the selective control of insulin signals, and a possible site for pharmacological intervention

Integrating GRK2 and NFkappaB in the Pathophysiology of Cardiac Hypertrophy

G protein coupled receptor kinase type 2 (GRK2) plays an important role in the development and maintenance of cardiac hypertrophy and heart failure even if its exact role is still unknown. In this study, we assessed the effect of GRK2 on the regulation of cardiac hypertrophy. In H9C2 cells, GRK2 overexpression increased atrial natriuretic factor (ANF) activity and enhanced phenylephrine-induced ANF response, and this is associated with an increase of NFκB transcriptional activity. The kinase dead mutant and a synthetic inhibitor of GRK2 activity exerted the opposite effect, suggesting that GRK2 regulates hypertrophy through upregulation of NFκB activity in a phosphorylation-dependent manner. In two different in vivo models of left ventricle hypertrophy (LVH), the selective inhibition of GRK2 activity prevented hypertrophy and reduced NFκB transcription activity. Our results suggest a previously undisclosed role for GRK2 in the regulation of hypertrophic responses and propose GRK2 as potential therapeutic target for limiting LVH

Targeting the CaMKII/ERK Interaction in the Heart Prevents Cardiac Hypertrophy

AIMS: Activation of Ca2+/Calmodulin protein kinase II (CaMKII) is an important step in signaling of cardiac hypertrophy. The molecular mechanisms by which CaMKII integrates with other pathways in the heart are incompletely understood. We hypothesize that CaMKII association with extracellular regulated kinase (ERK), promotes cardiac hypertrophy through ERK nuclear localization. METHODS AND RESULTS: In H9C2 cardiomyoblasts, the selective CaMKII peptide inhibitor AntCaNtide, its penetratin conjugated minimal inhibitory sequence analog tat-CN17β, and the MEK/ERK inhibitor UO126 all reduce phenylephrine (PE)-mediated ERK and CaMKII activation and their interaction. Moreover, AntCaNtide or tat-CN17β pretreatment prevented PE induced CaMKII and ERK nuclear accumulation in H9C2s and reduced the hypertrophy responses. To determine the role of CaMKII in cardiac hypertrophy in vivo, spontaneously hypertensive rats were subjected to intramyocardial injections of AntCaNtide or tat-CN17β. Left ventricular hypertrophy was evaluated weekly for 3 weeks by cardiac ultrasounds. We observed that the treatment with CaMKII inhibitors induced similar but significant reduction of cardiac size, left ventricular mass, and thickness of cardiac wall. The treatment with CaMKII inhibitors caused a significant reduction of CaMKII and ERK phosphorylation levels and their nuclear localization in the heart. CONCLUSION: These results indicate that CaMKII and ERK interact to promote activation in hypertrophy; the inhibition of CaMKII-ERK interaction offers a novel therapeutic approach to limit cardiac hypertrophy

Cellular subtype expression and activation of CaMKII regulate the fate of atherosclerotic plaque

Abstract Background and aims Atherosclerosis is a degenerative process of the arterial wall implicating activation of macrophages and proliferation of vascular smooth muscle cells. Calcium-calmodulin dependent kinase type II (CaMKII) in vascular smooth muscle cells (VSMCs) regulates proliferation, while in macrophages, this kinase governs diapedesis, infiltration and release of extracellular matrix enzymes. We aimed at understanding the possible role of CaMKII in atherosclerosis plaques to regulate plaque evolution towards stability or instability. Methods Clinically defined stable and unstable plaques obtained from patients undergoing carotid end arteriectomy were processed for evaluation of CaMKs protein expression, activity and localization. Results The larger content of CaMKII was found in CD14 + myeloid cells that were more abundant in unstable rather than stable plaques. To test the biological effect of activated CD14 + myeloid cells, VSMCs were exposed to the conditioned medium (CM) of macrophages extracted from carotid plaques. CM induced attenuation of CaMKs expression and activity in VSMCs, leading to the reduction of VSMCs proliferation. This appears to be due to the CaMKII dependent release of cytokines. Conclusions These results indicate a pivotal role of CaMKs in atherosclerosis by regulating activated myeloid cells on VSMCs activity. CaMKII could represent a possible target for therapeutic strategies based on macrophages specific inhibition for the stabilization of arteriosclerotic lesions

"Freeze, Don't Move": How to Arrest a Suspect in Heart Failure - A Review on Available GRK2 Inhibitors

Cardiovascular disease and heart failure (HF) still collect the largest toll of death in western societies and all over the world. A growing number of molecular mechanisms represent possible targets for new therapeutic strategies, which can counteract the metabolic and structural changes observed in the failing heart. G protein-coupled receptor kinase 2 (GRK2) is one of such targets for which experimental and clinical evidence are established. Indeed, several strategies have been carried out in place to interface with the known GRK2 mechanisms of action in the failing heart. This review deals with results from basic and preclinical studies. It shows different strategies to inhibit GRK2 in HF in vivo (βARK-ct gene therapy, treatment with gallein, and treatment with paroxetine) and in vitro (RNA aptamer, RKIP, and peptide-based inhibitors). These strategies are based either on the inhibition of the catalytic activity of the kinase ("Freeze!") or the prevention of its shuttling within the cell ("Don't Move!"). Here, we review the peculiarity of each strategy with regard to the ability to interact with the multiple tasks of GRK2 and the perspective development of eventual clinical use

Opposite effects of β2-adrenoceptor gene deletion on insulin signaling in liver and skeletal muscle

Background and aim β2-Adrenoceptors (β2-ARs) are G protein-coupled receptors (GPCRs) expressed in the major insulin target tissues. The interplay between β2-AR and insulin pathways is involved in the maintenance of glucose homeostasis. The aim of this study was to explore the consequences of β2-ARs deletion on insulin sensitivity and insulin signaling cascade in metabolically active tissues. Methods and results We evaluated glucose homeostasis in skeletal muscle and liver of β2-AR-null mice (β2-ARâ\u88\u92/â\u88\u92) by performing in vivo (glucose tolerance test and insulin tolerance test) and ex vivo (glucose uptake and glycogen determination) experiments. β2-AR gene deletion is associated with hepatic insulin resistance and preserved skeletal muscle insulin sensitivity. Importantly, we demonstrate that hepatic β2-AR regulates insulin-induced AKT activation via Grb2-mediated SRC recruitment through a Gi-independent mechanism. Conclusions β-AR stimulation contributes to the development of early stages of insulin resistance progression in the liver. Our findings indicate that the cross-talk between β2-AR and insulin signaling represents a fundamental target towards the development of novel therapeutic approaches to treat type 2 diabetes and metabolic syndrome