A common feature of glomerular disease is a protein leak into the urine. Proteinuria occurs in kidney disease and is an important risk factor for cardiovascular disease (CVD). ET‐1 is a potent vasoconstrictor/pressor peptide that is up‐regulated in CVD and many forms of inflammatory renal diseases. The actions of ET‐1 are mediated via two G‐protein coupled receptors, the ETAR which serves primarily in the pro‐hypertensive actions of ET‐1 and is often considered as the main pathological receptor subtype, with the ETBR serving to clear circulating ET‐1. Antagonism of one or both of receptors has been shown to be of clinical benefit in the treatment of hypertension. This research demonstrated a beneficial effect of selective ETAR antagonism using Sitaxsentan in a rat model of GN. ETAR blockade reduced blood pressure and importantly reduced glomerular inflammation as assessed by glomerular macrophage (Mϕ) infiltration. Further, we aimed to demonstrate that Mϕ, key mediators of inflammation are activated by ET‐1 to adopt a pro‐inflammatoy phenotype. However, early studies demonstrated that ET‐1 does not activate Mϕ as hypothesised. Mϕ were more phagocytic, and ET‐1 was chemokinetic for macrophages, an ETBR medicated event. ET‐1 was also removed by Mϕ, suggesting a potential regulatory role of Mϕ in the ET system. This phenomenon led to inclusion of additional in vivo studies to investigate the role of Mϕ in the regulation of ET‐1 and its pressor effects. These effects were investigated in a murine model of Mϕ ablation using CD11b‐DTR mice. These experiments determined in vivo that Mϕ ablation augments pressor responses to ET‐1, suggesting that Mϕ are required to regulate ET‐1. In vitro, Mϕ remove ET‐1 by several mechanisms involving proteolytic degradation of the peptide and ETBR mediated clearance, demonstrating a potential mechanism for the in vivo observation. Furthermore, proteinuria is believed to be due to damage or effacement of specialized visceral glomerular epithelial cells or podocytes. We identified in vitro that the ETAR mediates ET‐1 induced human podocyte cell effacement by actin cytoskeleton aberrations and slit‐diaphragm protein down-regulation, ET‐1 and pro‐inflammatory cytokine production. This thesis provides evidence to support our initial hypotheses that selective ETAR antagonism ameliorates proteinuric renal disease via its effects on podocytes and macrophages. Continued studies both in vitro and in vivo will strengthen the body of evidence to promote the therapeutic use of ETR antagonists in inflammatory renal disease.

- / Background: Endothelin-1 (ET-1), a strong vasoconstrictive substance acting via stimulation of specific receptors (ET-A and ET-B), has been implicated in the development of renal scarring. Activation of endothelin system was observed in experimental models of glomerular diseases and this was attributed to the toxic action of proteinuria to the tubular epithelial cells. However, we have not enough information about the role of endothelin system in human glomerular diseases and in renal diseases without proteinuria like obstructive nephropathy. The aim of this study was to examine the endothelin system in patients with primary glomerular diseases and in experimental animals with unilateral ureteric obstruction. Patients and Methods: Thirty-seven patients with different types of primary glomerulonephritides and 14 controls were included in the study. Patients presented by either nephrotic syndrome (n=25) or mild proteinuria (<1g/24h, n=12). The expression of ET-A and ET-B receptors in the renal tissue was examined immunohistochemically. At the time of biopsy, urinary ET-1 was determined by RIA. Experimental animals and Methods: Twenty –day old opossum pups (n=6) underwent surgical ligation of the left ureter. Sham operated animals, non-operated controls and normal human kidneys were also used. Animals were sacrificed at 2 (n=2), 3 (n=1), 4 (n=1), 5(n=1) and 8 (n=1) weeks post surgery and their kidneys were examined. Sham operation was performed at equivalent times in pups that served as control. The expression of ET-A and ET-B receptors in the renal tissue was examined immunohistochemically. Results: The expression of both receptors was mainly localized within tubular epithelial cells and was significantly higher in patients with glomerulonephritides compared to controls. The expression of ET-B receptors was higher in nephrotic compared to non-nephrotic patients while no difference was observed in the expression of ET-A receptors. Urinary excretion of ET-1 was increased in patients compared to healthy subjects (579±146 ng/24h vs. 410±78 ng/24h, p<0.01) and it was higher in nephrotic compared to non-nephrotic patients (617±167 ng/24h vs. 485±71 ng/24h, p<0.05). A significant positive correlation of the excreted ET-1 with the degree of proteinuria (r= 0.338, p<0.05) and the extent of immunostaining for ET-B receptors (r=0.427, p<0.05) was observed. The expression of ET-B receptors and the excretion of ET-1 were significantly decreased in patients who present remission of the nephrotic syndrome under immunosuppressive therapy. In tubular epithelial cells of the experimental animals there was a temporal increase in the expression of ET-A receptors with duration of obstruction while there was no significant difference between the expression of ET-B receptors in obstructed kidneys and controls. Conclusions: this study provides evidence that the endothelin system is activated in renal diseases and proteinuria seems to be related only in part to this activation. Further investigation is needed to ascertain if the activation of endothelin system has a causative role in the progression of renal diseases.

no / Prolonged hyperglycaemia impairs vascular reactivity and inhibits voltage-activated K+ (Kv) channels. We examined acute effects of altering glucose concentration on the activity and inhibition by endothelin-1 (ET-1) of Kv currents of freshly isolated rat arterial myocytes. Peak Kv currents recorded in glucose-free solution were reversibly reduced within 200 s by increasing extracellular glucose to 4 mm. This inhibitory effect of glucose was abolished by protein kinase C inhibitor peptide (PKC-IP), and Kv currents were further reduced in 10 mm glucose. In current-clamped cells, membrane potentials were more negative in 4 than in 10 mm glucose. In 4 mmd-glucose, 10 nm ET-1 decreased peak Kv current amplitude at +60 mV from 23.5 ± 3.3 to 12.1 ± 3.1 pA pF−1 (n = 6, P < 0.001) and increased the rate of inactivation, decreasing the time constant around fourfold. Inhibition by ET-1 was prevented by PKC-IP. When d-glucose was increased to 10 mm, ET-1 no longer inhibited Kv current (n = 6). Glucose metabolism was required for prevention of ET-1 inhibition of Kv currents, since fructose mimicked the effects of d-glucose, while l-glucose, sucrose or mannitol were without effect. Endothelin receptors were still functional in 10 mmd-glucose, since pinacidil-activated ATP-dependent K+ (KATP) currents were reduced by 10 nm ET-1. This inhibition was nearly abolished by PKC-IP, indicating that endothelin receptors could still activate PKC in 10 mmd-glucose. These results indicate that changes in extracellular glucose concentration within the physiological range can reduce Kv current amplitude and can have major effects on Kv channel modulation by vasoconstrictors.

Les maladies cardio-vasculaires (MCV), telles que l’hypertension et l’athérosclérose, s’accompagnent de modifications structurales et fonctionnelles au niveau vasculaire. Un fonctionnement aberrant de la migration, l’hypertrophie et la prolifération des cellules musculaires lisses vasculaires (CMLV) sont des évènements cellulaires à l’origine de ces changements. L’endothéline-1 (ET-1) contribue à la pathogénèse des anomalies vasculaires, notamment via l’activation des protéines MAPK et PI3-K/PKB, des composantes clés impliquées dans les voies prolifératives et de croissance cellulaires. Il a été suggéré que le stress oxydant jouerait un rôle intermédiaire dans les effets pathophysiologiques vasculaires de l’ET-1. En conséquence, une modulation de la signalisation induite par l’ET-1 peut servir comme éventuelle stratégie thérapeutique contre le développement des MCV. Il apparaît de nos jours un regain d’intérêt dans l’utilisation des agents phyto-chimiques pour traiter plusieurs maladies. La curcumine, constituant essentiel de l’épice curcuma, est dotée de plusieurs propriétés biologiques parmi lesquelles des propriétés anti-oxydantes, anti-prolifératrices et cardio-protectrices. Cependant, les mécanismes moléculaires de son effet cardio-protecteur demeurent obscurs. Dans cette optique, l’objectif de cette étude a été d’examiner l’efficacité de la curcumine à inhiber la signalisation induite par l’ET-1 dans les CMLV. La curcumine a inhibé la phosphorylation des protéines IGF-1R, PKB, c-Raf et ERK1/2, induite par l’ET-1 et l’IGF-1. De plus, la curcumine a inhibé l’expression du facteur de transcription Egr-1 induite par l’ET-1 et l’IGF-1, dans les CMLV. Ces résultats suggèrent que la capacité de la curcumine à atténuer ces voies de signalisation serait un mécanisme d’action potentiel de ses effets protecteurs au niveau cardiovasculaire. / Cardiovascular diseases (CVDs), including hypertension and atherosclerosis, are associated with vascular functional and structural changes. Some of the cellular events underlying these processes include aberrant vascular smooth muscle cell (VSMC) proliferation, hypertrophy and migration. Endothelin-1 (ET-1) has been implicated in the pathogenesis of vascular abnormalities through the hyperactivation of key components of growth promoting and proliferative signaling pathways, including MAPKs and PI3-K/PKB. Vascular oxidative stress has also been suggested to play an intermediary role in mediating ET-1-induced pathophysiological effects. Interference with ET-1-induced signaling may therefore serve as a potential therapeutic strategy against the progression of cardiovascular disorders. There is presently a surge of interest in the use of plant-derived phytochemicals for the treatment of various diseases. Curcumin, the main constituent of the spice turmeric, exhibits multiple biological properties, amongst them, antioxidant, anti-proliferative and cardioprotective properties. However, the molecular mechanisms of its cardiovascular protective action remain obscure. Therefore, in the present studies, we investigated the effectiveness of curcumin to inhibit ET-1-induced signaling events in VSMC. Curcumin inhibited ET-1-induced as well as IGF-1-induced phosphorylation of IGF-1R, PKB, c-Raf and ERK1/2, in VSMC. Furthermore, curcumin inhibited the expression of transcription factor early growth response-1 (Egr-1) induced by ET-1 and IGF-1, in VSMC. In summary, these results demonstrate that curcumin is a potent inhibitor of ET-1 and IGF-1-induced mitogenic and proliferative signaling events in VSMC, suggesting that the ability of curcumin to attenuate these effects may contribute as potential mechanism for its cardiovascular protective response.

Les maladies cardio-vasculaires (MCV), telles que l’hypertension et l’athérosclérose, s’accompagnent de modifications structurales et fonctionnelles au niveau vasculaire. Un fonctionnement aberrant de la migration, l’hypertrophie et la prolifération des cellules musculaires lisses vasculaires (CMLV) sont des évènements cellulaires à l’origine de ces changements. L’endothéline-1 (ET-1) contribue à la pathogénèse des anomalies vasculaires, notamment via l’activation des protéines MAPK et PI3-K/PKB, des composantes clés impliquées dans les voies prolifératives et de croissance cellulaires. Il a été suggéré que le stress oxydant jouerait un rôle intermédiaire dans les effets pathophysiologiques vasculaires de l’ET-1. En conséquence, une modulation de la signalisation induite par l’ET-1 peut servir comme éventuelle stratégie thérapeutique contre le développement des MCV. Il apparaît de nos jours un regain d’intérêt dans l’utilisation des agents phyto-chimiques pour traiter plusieurs maladies. La curcumine, constituant essentiel de l’épice curcuma, est dotée de plusieurs propriétés biologiques parmi lesquelles des propriétés anti-oxydantes, anti-prolifératrices et cardio-protectrices. Cependant, les mécanismes moléculaires de son effet cardio-protecteur demeurent obscurs. Dans cette optique, l’objectif de cette étude a été d’examiner l’efficacité de la curcumine à inhiber la signalisation induite par l’ET-1 dans les CMLV. La curcumine a inhibé la phosphorylation des protéines IGF-1R, PKB, c-Raf et ERK1/2, induite par l’ET-1 et l’IGF-1. De plus, la curcumine a inhibé l’expression du facteur de transcription Egr-1 induite par l’ET-1 et l’IGF-1, dans les CMLV. Ces résultats suggèrent que la capacité de la curcumine à atténuer ces voies de signalisation serait un mécanisme d’action potentiel de ses effets protecteurs au niveau cardiovasculaire. / Cardiovascular diseases (CVDs), including hypertension and atherosclerosis, are associated with vascular functional and structural changes. Some of the cellular events underlying these processes include aberrant vascular smooth muscle cell (VSMC) proliferation, hypertrophy and migration. Endothelin-1 (ET-1) has been implicated in the pathogenesis of vascular abnormalities through the hyperactivation of key components of growth promoting and proliferative signaling pathways, including MAPKs and PI3-K/PKB. Vascular oxidative stress has also been suggested to play an intermediary role in mediating ET-1-induced pathophysiological effects. Interference with ET-1-induced signaling may therefore serve as a potential therapeutic strategy against the progression of cardiovascular disorders. There is presently a surge of interest in the use of plant-derived phytochemicals for the treatment of various diseases. Curcumin, the main constituent of the spice turmeric, exhibits multiple biological properties, amongst them, antioxidant, anti-proliferative and cardioprotective properties. However, the molecular mechanisms of its cardiovascular protective action remain obscure. Therefore, in the present studies, we investigated the effectiveness of curcumin to inhibit ET-1-induced signaling events in VSMC. Curcumin inhibited ET-1-induced as well as IGF-1-induced phosphorylation of IGF-1R, PKB, c-Raf and ERK1/2, in VSMC. Furthermore, curcumin inhibited the expression of transcription factor early growth response-1 (Egr-1) induced by ET-1 and IGF-1, in VSMC. In summary, these results demonstrate that curcumin is a potent inhibitor of ET-1 and IGF-1-induced mitogenic and proliferative signaling events in VSMC, suggesting that the ability of curcumin to attenuate these effects may contribute as potential mechanism for its cardiovascular protective response.