Cannabinoid CB1/CB2 Receptors in the Heart: Expression, Regulation, and Function

Endocannabinoids exert their actions in the heart and vessels, at least in part, by stimulating the cannabinoid CB1 and the CB2 receptor subtypes which belong to a group of seven transmembrane-spanning receptors and are coupled to Gi/o-proteins. Activation of cardiovascular CB1 receptors leads to depressed cardiac contractility and hypotension. Conversely, in most studies, the CB1 receptor antagonists are cardioprotective against ischemia–reperfusion injury, myocardial ischemia, heart failure, and cardiomyopathies. Evidence to date indicates that CB2 receptor activation is cardioprotective. CB2 receptor-mediated effects such as anti-inflammation and anti-fibrosis may be in part opposite to the actions of the CB1 receptor. The aim of this review is to up-date on recent experimental findings and controversies on the role of endocannabinoid system in the myocardial injury with emphasis on pathophysiological processes such as left ventricular remodeling, cardiac fibrosis, hypertrophy, and endothelial dysfunction. Recent experimental studies employing genetic deficiency of CB1 and CB2 receptors and endocannabinoid anandamide metabolizing enzymes are reviewed. Moreover, the protective mechanisms which are mediated by cannabinoid receptors during ischemic preconditioning as well as in the early and late phase after myocardial infarction are discussed in the context of possible therapeutic implications

The kallikrein-kinin pathway as a mechanism for auto-control of brown adipose tissue activity

Brown adipose tissue (BAT) is known to secrete regulatory factors in response to thermogenic stimuli. Components of the BAT secretome may exert local effects that contribute to BAT recruitment and activation. Here, we found that a thermogenic stimulus leads to enhanced secretion of kininogen (Kng) by BAT, owing to induction of kininogen 2 (Kng2) gene expression. Noradrenergic, cAMP-mediated signals induce KNG2 expression and release in brown adipocytes. Conversely, the expression of kinin receptors, that are activated by the Kng products bradykinin and [Des-Arg9]-bradykinin, are repressed by thermogenic activation of BAT in vivo and of brown adipocytes in vitro. Loss-of-function models for Kng (the circulating-Kng-deficient BN/Ka rat) and bradykinin (pharmacological inhibition of kinin receptors, kinin receptor-null mice) signaling were coincident in showing abnormal overactivation of BAT. Studies in vitro indicated that Kng and bradykinin exert repressive effects on brown adipocyte thermogenic activity by interfering the PKA/p38 MAPK pathway of control of Ucp1 gene transcription, whereas impaired kinin receptor expression enhances it. Our findings identify the kallikrein-kinin system as a relevant component of BAT thermogenic regulation that provides auto-regulatory inhibitory signaling to BAT

Die Rolle des Renin-Angiotensin- und Kallikrein-Kinin-Systems bei der Aneurysmaformation und mögliche therapeutische Interventionen

The overall prevalence of aortic aneurysms has strongly increased in the last 30 years due to an ageing population. Present treatment options such as endovascular stents or open surgery procedures are not appropriate for all patients. The risk of death from aneurysm rupture remains a significant clinical problem. Better understanding of aortic aneurysms is important for development of new pharmacological treatments. In this dissertation, different aspects of AAA were investigated in the context of reninangiotensin- and kallikrein-kinin-systems. In the first study we have found that most RAS components were significantly stronger expressed in human AAA when compared to atherosclerotic lesions (paper 1). Some components which take part in Ang II generation, namely chymase, cathepsin G and cathepsin D, additionally possess features such as chemotaxis or proteolysis which may accessorily contribute to AAA formation. Therefore, in humans increasing of RAS activation may be involved in the transition from atherosclerosis to AAA. The next study (paper 2) was designed to elucidate protective mechanisms of AT1 receptor blockade by AAA formation. We studied telmisartan, the AT1 receptor antagonist with long duration of action and high lipid solubility. Telmisartan prevented aneurysm expansion in an experimental model of AAA independently of blood pressure reduction. Several mechanisms of telmisaratan underlie vascular protection: inhibition of proteolysis, reduced production of proinflammatory cytokines and prevention of apoptotis in the aorta. The AT2 receptors which are known to afford tissue protective actions may be exposed to enhanced Ang II levels after AT1 blockade. The effects of the AT2 receptor stimulation were investigated using a specific AT2 receptor agonist compound 21 (paper 3). In the model of myocardial infarction, compound 21 reduced infarct size and improved heart function. These protective effects were associated with the anti-inflammation and anti-apoptosis. The implication of kallikrein-kinin system in aneurysm formation was discovered in a genetic animal model (paper 4). In this study we reported that Brown Norway Katholiek rats, which feature a deficiency of plasma kininogens, develop severe abdominal aortic aneurysm. A genetically determined kininogen deficiency promoted the formation of AAA but not atherosclerosis and was associated with enhanced elastolysis, FasL- and caspase-3-mediated apoptosis, changes in plasma cytokines and the induction of the MMPs associated proteolytic cascade. Next in vitro study has revealed a molecular basis for these effects (paper 5). HKa affected the regulation of MMP-9 and MMP-2 and their tissue inhibitors TIMPs in VSMCs as demonstrated by a negative regulation of cytokine-induced MMP expression and activity. This study suggested that HKa might contribute to prevent the extracellular matrix from excessive degradation in the context of physiological and pathophysiological vascular remodeling. Thus, pharmacological interference with various components of renin-angiotensin- and kallikrein-kinin-systems is a promising approach for the treatment of AAA.Das Aneurysma der Aorta abdominalis (AAA) ist eine weltverbreitete vaskuläre Erkrankung mit hoher Sterblichkeitsrate. Die steigende Inzidenz des AAA in den letzten Jahrzehnten ist durch die Alterung der Bevölkerung bedingt. Die oft einzige Möglichkeit stellen operative oder endovaskuläre Therapien dar. Vor dem Hintergrund der Rupturgefahr von AAA sind neue therapeutische Strategien wünschenswert. Die genaue Bestimmung von spezifischen Genen und Pathomechanismen, die für die Aneurysma-Entstehung bedeutsam sind, könnte Angriffsmöglichkeiten für eine pharmakologische Therapie dieser Erkrankung identifizieren. In dieser Habilitationsschrift wurden verschiedene Aspekte der AAA im Kontext des Renin-Angiotensin-Systems (RAS) und des Kallikrein-Kinin- Systems (KKS) untersucht. Die Bedeutung des RAS für den Übergang aus der Atherosklerose in die Aneurysmenformation wurde durch eine Untersuchung des humanen Gewebes demonstriert (paper 1). In dieser Studie konnten wir eine Aktivierung/verstärkte Expression von RAS Komponenten (AT1 Rezeptor, Chymase, Cathepsin G und Cathepsin D) in AAA feststellen. Die therapeutischen Effekte der AT1 Rezeptor Blockade bei der AAA Formation wurden am Rattenmodel untersucht (paper 2). Der AT1 Rezeptor Antagonist Telmisartan zeigte eine inhibierende Wirkung auf die Entstehung der Elastase-induzierten AAA, die unabhängig von dem blutdrucksenkenden Effekt dieses Wirkstoffs auftrat. Diesem protektiven Effekt liegen anti-inflammatorische, anti-apoptotische und anti- proteolytische Mechanismen zugrunde. Da der AT2 Rezeptor eine wichtige Rolle bei der Gewebeadaptation und Regeneration spielt, haben wir die Effekte einer direkten AT2 Rezeptor Stimulation mit einem selektiven Agonisten des AT2 Rezeptors Compound 21 im Tiermodell untersucht (paper 3). Compound 21 zeigte einen positiven therapeutischen Effekt auf die Narbenbildung und die Herzfunktion. Dieser therapeutischen Wirkung liegt die Aktivierung von anti- inflammatorischen und anti-apoptotischen Prozessen zugrunde. Die Rolle des Kallikrein-Kinin Systems bei der Aneurysmaformation wurde in einem genetischen Tiermodell demonstriert (paper 4). In dieser Studie konnte eine ausgeprägte Prädisposition von Kininogen-defizienten Ratten für abdominale Aortenaneurysmen aufgezeigt werden, deren Entstehung durch eine Degradation von Elastinfasern, eine FasL- und Caspase-3-vermittelte Apoptoserate, gesteigerte Zytokinproduktion sowie eine erhöhte Expression von MMPs gekennzeichnet war. Hieraus ergaben sich erstmals Hinweise auf eine Beteiligung von Kininogen an der MMP-Regulation. Weiteren Untersuchungen in primären glatten Gefäßmuskelzellen zeigten auf, dass das gespaltene hochmolekulare Kininogen (HKa) die Zytokin-induzierte Sekretion von sowohl MMP-2 als auch MMP-9 konzentrationsabhängig reduzierte (paper 5). Auf der Grundlage dieser Ergebnisse spielen Renin-Angiotensin- und Kallikrein-Kinin- Systeme eine bedeutende Rolle bei der Entstehung von vaskulärem Stress und der dadurch bedingten Aneurysmenformation. Pharmakologische Beeinflussung dieser Systeme bildet einen vielversprechenden Ansatz zur Erweiterung der Behandlungsmöglichkeiten der AAA

Differential Regulation of MMPs, Apoptosis and Cell Proliferation by the Cannabinoid Receptors CB1 and CB2 in Vascular Smooth Muscle Cells and Cardiac Myocytes

Cannabinoids (CB) are implicated in cardiovascular diseases via the two main receptor subtypes CB1R and CB2R. This study investigated whether cannabinoids regulate the activity of matrix metalloproteases (MMP-2, MMP-9) in vascular smooth muscle cells (VSMCs) and in cells of cardiac origin (H9c2 cell line). The influence of CB1- and CB2 receptor stimulation or inhibition on cell proliferation, apoptosis and glucose uptake was also evaluated. We used four compounds that activate or block CB receptors: arachidonyl-2-chloroethylamide (ACEA)-CB1R agonist, rimonabant-CB1R antagonist, John W. Huffman (JWH133)-CB2R agonist and CB2R antagonist-6-Iodopravadoline (AM630). Treatment of cells with the CB2R agonist JWH133 decreased cytokine activated secretion of proMMP-2, MMP-2 and MMP-9, reduced Fas ligand and caspase-3-mediated apoptosis, normalized the expression of TGF-beta1 and prevented cytokine-induced increase in glucose uptake into the cell. CB1R inhibition with rimonabant showed similar protective properties as the CB2R agonist JWH133, but to a lesser extent. In conclusion, CB1R and CB2R exert opposite effects on cell glucose uptake, proteolysis and apoptosis in both VSMCs and H9c2 cells. The CB2R agonist JWH133 demonstrated the highest protective properties. These findings may pave the way to a new treatment of cardiovascular diseases, especially those associated with extracellular matrix degradation

Long-term angiotensin II receptor blockade limits hypertension, aortic dysfunction, and structural remodeling in a rat model of chronic kidney disease

Background/Aims: Chronic kidney disease (CKD) is associated with large artery remodeling, endothelial dysfunction and calcification, with angiotensin II (Ang II) a known driver of these pathologies. We investigated long-term Ang II type 1 receptor inhibition with valsartan on aortic function and structure in the Lewis polycystic kidney (LPK) rat model of CKD. Methods: Mixed sex LPK and Lewis control (total n = 28) treated (valsartan 60 mg/kg/day p.o. from 4 to 18 weeks) and vehicle groups were studied. Functional responses to noradrenaline (NA), potassium chloride and endothelium-dependent and independent relaxations were investigated in vitro using acetylcholine hydrochloride (ACh) and sodium nitroprusside (SNP), respectively. Effects of the nitric oxide synthase (NOS) substrate L-arginine, NOS inhibitor L-NAME and cyclooxygenase inhibitor indomethacin on ACh responses were examined. Results: In the LPK, valsartan reduced systolic blood pressure and urinary protein, ameliorated exaggerated sensitivity to NA, and normalized endothelium-dependent (ACh-Rmax; 91 ± 7 vs. 59 ± 6%, p = 0.0001) and independent dysfunction (SNP-Rmax; 99 ± 1 vs. 82 ± 7%, p = 0.040), as well as improving NO-dependent relaxation (Rmax; -51 ± 6 vs. -26 ± 9%, p = 0.008). Valsartan also reduced aortic wall hypertrophy, elastin disruption/fragmentation, calcification, media cystic degeneration, and levels of matrix metalloproteinase 9. Conclusions: This study highlights the role of Ang II in driving vascular manifestations of CKD and indicates that early treatment can significantly limit pathological changes.14 page(s