Left ventricular pressure-volume relationship in a rat model of advanced aging-associated heart failure.

Aging is associated with profound changes in the structure and function of the heart. A fundamental understanding of these processes, using relevant animal models, is required for effective prevention and treatment of cardiovascular disease in the elderly. Here, we studied cardiac performance in 4- to 5-mo-old (young) and 24- to 26-mo-old (old) Fischer 344 male rats using the Millar pressure-volume (P-V) conductance catheter system. We evaluated systolic and diastolic function in vivo at different preloads, including preload recruitable stroke work (PRSW), maximal slope of the systolic pressure increment (+dP/dt), and its relation to end-diastolic volume (+dP/dt-EDV) as well as the time constant of left ventricular pressure decay, as an index of relaxation. The slope of the end-diastolic P-V relation (EDPVR), an index of left ventricular stiffness, was also calculated. Aging was associated with decrease in left ventricular systolic pressure, +dP/dt, maximal slope of the diastolic pressure decrement, +dP/dt-EDV, PRSW, ejection fraction, stroke volume, cardiac and stroke work indexes, and efficiency. In contrast, total peripheral resistance, left ventricular end-diastolic volume, left ventricular end-diastolic pressure, and EDPVR were greater in aging than in young animals. Taken together, these data suggest that advanced aging is characterized by decreased systolic performance accompanied by delayed relaxation and increased diastolic stiffness of the heart in male Fischer 344 rats. P-V analysis is a sensitive method to determine cardiac function in rats

Cannabidiol attenuates high glucose-induced endothelial cell inflammatory response and barrier disruption.

A nonpsychoactive cannabinoid cannabidiol (CBD) has been shown to exert potent anti-inflammatory and antioxidant effects and has recently been reported to lower the incidence of diabetes in nonobese diabetic mice and to preserve the blood-retinal barrier in experimental diabetes. In this study we have investigated the effects of CBD on high glucose (HG)-induced, mitochondrial superoxide generation, NF-kappaB activation, nitrotyrosine formation, inducible nitric oxide synthase (iNOS) and adhesion molecules ICAM-1 and VCAM-1 expression, monocyte-endothelial adhesion, transendothelial migration of monocytes, and disruption of endothelial barrier function in human coronary artery endothelial cells (HCAECs). HG markedly increased mitochondrial superoxide generation (measured by flow cytometry using MitoSOX), NF-kappaB activation, nitrotyrosine formation, upregulation of iNOS and adhesion molecules ICAM-1 and VCAM-1, transendothelial migration of monocytes, and monocyte-endothelial adhesion in HCAECs. HG also decreased endothelial barrier function measured by increased permeability and diminished expression of vascular endothelial cadherin in HCAECs. Remarkably, all the above mentioned effects of HG were attenuated by CBD pretreatment. Since a disruption of the endothelial function and integrity by HG is a crucial early event underlying the development of various diabetic complications, our results suggest that CBD, which has recently been approved for the treatment of inflammation, pain, and spasticity associated with multiple sclerosis in humans, may have significant therapeutic benefits against diabetic complications and atherosclerosis

Decreased age-related cardiac dysfunction, myocardial nitrative stress, inflammatory gene expression, and apoptosis in mice lacking fatty acid amide hydrolase.

Recent studies have uncovered important cross talk between inflammation, generation of reactive oxygen and nitrogen species, and lipid metabolism in the pathogenesis of cardiovascular aging. Inhibition of the endocannabinoid anandamide metabolizing enzyme, the fatty acid amide hydrolase (FAAH), is emerging as a promising novel approach for the treatment of various inflammatory disorders. In this study, we have investigated the age-associated decline of cardiac function and changes in inflammatory gene expression, nitrative stress, and apoptosis in FAAH knockout (FAAH(-/-)) mice and their wild-type (FAAH(+/+)) littermates. Additionally, we have explored the effects of anandamide on TNF-alpha-induced ICAM-1 and VCAM-1 expression and monocyte-endothelial adhesion in human coronary artery endothelial cells (HCAECs). There was no difference in the cardiac function (measured by the pressure-volume conductance catheter system) between 2- to 3-mo-old (young) FAAH(-/-) and FAAH(+/+) mice. In contrast, the aging-associated decline in cardiac function and increased myocardial gene expression of TNF-alpha, gp91phox, matrix metalloproteinase (MMP)-2, MMP-9, caspase-3 and caspase-9, myocardial inducible nitric oxide synthase protein expression, nitrotyrosine formation, poly (ADP-ribose)polymerase cleavage and caspase-3/9 activity, observed in 28- to 31-mo-old (aging) FAAH(+/+) mice, were largely attenuated in knockouts. There was no difference in the myocardial cannabinoid CB(1) and CB(2) receptor gene expression between young and aging FAAH(-/-) and FAAH(+/+) mice. Anandamide dose dependently attenuated the TNF-alpha-induced ICAM-1 and VCAM-1 expression, NF-kappaB activation in HCAECs, and the adhesion of monocytes to HCAECs in a CB(1)- and CB(2)-dependent manner. These findings suggest that pharmacological inhibition of FAAH may represent a novel protective strategy against chronic inflammation, oxidative/nitrative stress, and apoptosis associated with cardiovascular aging and atherosclerosis

Effects of inosine on reperfusion injury after cardiopulmonary bypass

Objective: Inosine, a break-down product of adenosine has been recently shown to exert inodilatory and anti-inflammatory properties. Furthermore inosine might be a key substrate of pharmacological post-conditioning. In the present pre-clinical study, we investigated the effects of inosine on cardiac function during reperfusion in an experimental model of cardioplegic arrest and extracorporal circulation. Methods: Twelve anesthetized dogs underwent hypothermic cardiopulmonary bypass. After 60 minutes of hypothermic cardiac arrest, reperfusion was started after application of either saline vehicle (control, n = 6), or inosine (100 mg/kg, n = 6). Left ventricular end-systolic pressure volume relationship (ESPVR) was measured by a combined pressure-volume-conductance catheter at baseline and after 60 minutes of reperfusion. Left anterior descendent coronary blood flow (CBF), endothelium-dependent vasodilatation to acetylcholine (ACh) and endothelium-independent vasodilatation to sodium nitroprusside (SNP) were also determined. Results: The administration of inosine led to a significantly better recovery (given as percent of baseline) of ESPVR 90 ± 9% vs. 46 ± 6%, p &lt; 0.05. CBF and was also significantly higher in the inosine group (56 ± 8 vs. 23 ± 4, ml/min, p < 0.05). While the vasodilatatory response to SNP was similar in both groups, ACh resulted in a significantly higher increase in CBF (58 ± 6% vs. 25 ± 5%, p < 0.05) in the inosine group. Conclusions: Application of inosine improves myocardial and endothelial function after cardiopulmonary bypass with hypothermic cardiac arrest

CB2-receptor stimulation attenuates TNF-alpha-induced human endothelial cell activation, transendothelial migration of monocytes, and monocyte-endothelial adhesion.

Targeting cannabinoid-2 (CB(2)) receptors with selective agonists may represent a novel therapeutic avenue in various inflammatory diseases, but the mechanisms by which CB(2) activation exerts its anti-inflammatory effects and the cellular targets are elusive. Here, we investigated the effects of CB(2)-receptor activation on TNF-alpha-induced signal transduction in human coronary artery endothelial cells in vitro and on endotoxin-induced vascular inflammatory response in vivo. TNF-alpha induced NF-kappaB and RhoA activation and upregulation of adhesion molecules ICAM-1 and VCAM-1, increased expression of monocyte chemoattractant protein, enhanced transendothelial migration of monocytes, and augmented monocyte-endothelial adhesion. Remarkably, all of the above-mentioned effects of TNF-alpha were attenuated by CB(2) agonists. CB(2) agonists also decreased the TNF-alpha- and/or endotoxin-induced ICAM-1 and VCAM-1 expression in isolated aortas and the adhesion of monocytes to aortic vascular endothelium. CB(1) and CB(2) receptors were detectable in human coronary artery endothelial cells by Western blotting, RT-PCR, real-time PCR, and immunofluorescence staining. Because the above-mentioned TNF-alpha-induced phenotypic changes are critical in the initiation and progression of atherosclerosis and restenosis, our findings suggest that targeting CB(2) receptors on endothelial cells may offer a novel approach in the treatment of these pathologies

Poly (ADP-ribose) polymerase-1 is a key mediator of liver inflammation and fibrosis.

Poly (ADP-ribose) polymerase 1 (PARP-1) is a constitutive enzyme, the major isoform of the PARP family, which is involved in the regulation of DNA repair, cell death, metabolism, and inflammatory responses. Pharmacological inhibitors of PARP provide significant therapeutic benefits in various preclinical disease models associated with tissue injury and inflammation. However, our understanding the role of PARP activation in the pathophysiology of liver inflammation and fibrosis is limited. In this study we investigated the role of PARP-1 in liver inflammation and fibrosis using acute and chronic models of carbon tetrachloride (CCl4 )-induced liver injury and fibrosis, a model of bile duct ligation (BDL)-induced hepatic fibrosis in vivo, and isolated liver-derived cells ex vivo. Pharmacological inhibition of PARP with structurally distinct inhibitors or genetic deletion of PARP-1 markedly attenuated CCl4 -induced hepatocyte death, inflammation, and fibrosis. Interestingly, the chronic CCl4 -induced liver injury was also characterized by mitochondrial dysfunction and dysregulation of numerous genes involved in metabolism. Most of these pathological changes were attenuated by PARP inhibitors. PARP inhibition not only prevented CCl4 -induced chronic liver inflammation and fibrosis, but was also able to reverse these pathological processes. PARP inhibitors also attenuated the development of BDL-induced hepatic fibrosis in mice. In liver biopsies of subjects with alcoholic or hepatitis B-induced cirrhosis, increased nitrative stress and PARP activation was noted. CONCLUSION: The reactive oxygen/nitrogen species-PARP pathway plays a pathogenetic role in the development of liver inflammation, metabolism, and fibrosis. PARP inhibitors are currently in clinical trials for oncological indications, and the current results indicate that liver inflammation and liver fibrosis may be additional clinical indications where PARP inhibition may be of translational potential

Ecto-5′-Nucleotidase: A Candidate Virulence Factor in Streptococcus sanguinis Experimental Endocarditis

Streptococcus sanguinis is the most common cause of infective endocarditis (IE). Since the molecular basis of virulence of this oral commensal bacterium remains unclear, we searched the genome of S. sanguinis for previously unidentified virulence factors. We identified a cell surface ecto-5′-nucleotidase (Nt5e), as a candidate virulence factor. By colorimetric phosphate assay, we showed that S. sanguinis Nt5e can hydrolyze extracellular adenosine triphosphate to generate adenosine. Moreover, a nt5e deletion mutant showed significantly shorter lag time (P<0.05) to onset of platelet aggregation than the wild-type strain, without affecting platelet-bacterial adhesion in vitro (P = 0.98). In the absence of nt5e, S. sanguinis caused IE (4 d) in a rabbit model with significantly decreased mass of vegetations (P<0.01) and recovered bacterial loads (log10CFU, P = 0.01), suggesting that Nt5e contributes to the virulence of S. sanguinis in vivo. As a virulence factor, Nt5e may function by (i) hydrolyzing ATP, a pro-inflammatory molecule, and generating adenosine, an immunosuppressive molecule to inhibit phagocytic monocytes/macrophages associated with valvular vegetations. (ii) Nt5e-mediated inhibition of platelet aggregation could also delay presentation of platelet microbicidal proteins to infecting bacteria on heart valves. Both plausible Nt5e-dependent mechanisms would promote survival of infecting S. sanguinis. In conclusion, we now show for the first time that streptococcal Nt5e modulates S. sanguinis-induced platelet aggregation and may contribute to the virulence of streptococci in experimental IE

Differentiation and Glucocorticoid Regulated Apopto-Phagocytic Gene Expression Patterns in Human Macrophages. Role of Mertk in Enhanced Phagocytosis

The daily clearance of physiologically dying cells is performed safely mainly by cells in the mononuclear phagocyte system. They can recognize and engulf dying cells utilizing several cooperative mechanisms. In our study we show that the expression of a broad range of apopto-phagocytic genes is strongly up-regulated during differentiation of human monocytes to macrophages with different donor variability. The glucocorticoid dexamethasone has a profound effect on this process by selectively up-regulating six genes and down-regulating several others. The key role of the up-regulated mer tyrosine kinase (Mertk) in dexamethasone induced enhancement of phagocytosis could be demonstrated in human monocyte derived macrophages by gene silencing as well as blocking antibodies, and also in a monocyte-macrophage like cell line. However, the additional role of other glucocorticoid induced elements must be also considered since the presence of autologous serum during phagocytosis could almost completely compensate for the blocked function of Mertk