Endotoxin causes impaired vascular contractility proposed to be mediated mainly by induction of inducible nitric oxide synthase (iNOS). Evidence suggests that calcium/calmodulin dependent protein kinase II (CaMKII) may lead to activation of cytosolic phospholipase A(2alpha) (cPLA(2alpha))/inducible cyclooxygenase (COX-2) pathway in response to endotoxin in vascular smooth muscle cells. This study was conducted to determine if CaMKII is involved in the endotoxin-induced vascular hyporeactivity by activating of iNOS and/or cPLA(2alpha)/COX-2 enzymes in rat isolated superior mesenteric artery with endothelium. Incubation with endotoxin (100 microg ml(-1)) for 4h caused vascular hyporeactivity to norepinephrine which was completely abolished by phenylene-1,3-bis[ethane-2-isothiourea] dihydrobromide (1,3-PBIT), a selective iNOS inhibitor, methyl arachidonyl fluorophosphonate (MAFP), a selective 85kDa cPLA(2alpha) inhibitor, DFU, a selective COX-2 inhibitor, and KN-93, a selective CaMKII inhibitor. Endotoxin-induced increase in tissue nitrite production was decreased by 1,3-PBIT and DFU, and further increased by MAFP. MAFP, DFU and KN-93 reversed the endotoxin-induced decrease in tissue 6-keto-PGF(1alpha). These data suggest that reversal of the endotoxin-induced vascular hyporeactivity by inhibition of CaMKII in rat superior mesenteric artery may be related to increased production of vasodilator arachidonic acid products by cPLA(2alpha)/COX-2 pathway rather than prostacyclin and nitric oxide.
"release and PGE 2 production by the up-regulation of cPLA 2 and COX-2 via Ca 2+ /PKC/MAPKs (Lee et al., 2009). Ozveren et al. demonstrated that involvement of calcium/CaMKII in endotoxin-induced COX-2 expression (Ozveren et al., 2006). Indeed, we found that in A549 cells, ATP␥S-induced COX-2 expression and PGE 2 generation were reduced via inhibition of PI-PLC, PC-PLC, and Ca 2+ /CaMKII. "
[Show abstract][Hide abstract] ABSTRACT: Endotoxic shock is a systemic inflammatory response that is associated with an increase in nitric oxide production and a decrease in the formation of 20-hydroxyeicosatetraenoic acid (20-HETE), which may contribute to the fall in blood pressure and vascular reactivity. The present study examined the effects of a synthetic analogue of 20-HETE, N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-HEDGE), on the fall in blood pressure and vascular responsiveness to vasoscontrictors and acetylcholine in rats treated with endotoxin. The MAP fell by 31 mmHg, and the heart rate rose by 90 beats/min in male Wistar rats treated with endotoxin (10 mg/kg, intraperitoneally). The fall in MAP was associated with a decrease in the vasoconstrictor response to norepinephrine in isolated aorta and superior mesenteric artery and increased levels of nitrite in the serum, kidney, heart, and vascular tissues. The effects of endotoxin were prevented by 5,14-HEDGE (30 mg/kg, s.c.) given 1 h after injection of endotoxin. Furthermore, a competitive antagonist of vasoconstrictor effects of 20-HETE, 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (30 mg/kg, s.c.), prevented the beneficial effects of 5,14-HEDGE on MAP and vascular tone in rats treated with endotoxin. These data are consistent with the view that a fall in the production of 20-HETE contributes to the fall in MAP and vascular reactivity in rats treated with endotoxin, and that 5,14-HEDGE has a beneficial effect.
[Show abstract][Hide abstract] ABSTRACT: Muscular lymphatics use both phasic and tonic contractions to transport lymph for conducting their vital functions. The molecular mechanisms regulating lymphatic muscle contractions are not well understood. Based on the well-established finding that the phosphorylation of myosin light chain 20 (MLC(20)) plays an essential role in blood vessel smooth muscle contraction, we investigated if phosphorylated MLC(20) (pMLC(20)) would modulate the tonic and/or phasic contractions of lymphatic muscle. The effects of ML-7, a MLC kinase inhibitor (1-10 microM), were tested on the contractile parameters of isolated and cannulated rat mesenteric lymphatics during their responses to the known modulators, pressure (1-5 cm H(2)O) and substance P (SP; 10(-7) M). Immunohistochemical and Western blot analyses of pMLC(20) were also performed on isolated lymphatics. The results showed that 1) increasing pressure decreased both the lymphatic tonic contraction strength and pMLC(20)-to-MLC(20) ratio; 2) SP increased both the tonic contraction strength and phosphorylation of MLC(20); 3) ML-7 decreased both the lymphatic tonic contraction strength and pMLC(20)-to-MLC(20) ratio; and 4) the increase in lymphatic phasic contraction frequency in response to increasing pressure was diminished by ML-7; however, the phasic contraction amplitude was not significantly altered by ML-7 either in the absence or presence of SP. These data provide the first evidence that tonic contraction strength and phasic contraction amplitude of the lymphatics can be differentially regulated, whereby the increase in MLC(20) phosphorylation produces an activation in the tonic contraction without significant changes in the phasic contraction amplitude. Thus, tonic contraction of rat mesenteric lymphatics appears to be MLC kinase dependent.
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