The transient receptor potential (TRP) vanilloid subtype 4 (V4) is a nonselective cation channel that exhibits polymodal activation and is expressed in the endothelium, where it contributes to intracellular Ca 2ϩ homeostasis and regulation of cell volume. The purpose of the present study was to evaluate the systemic cardiovascular effects of GSK1016790A, a novel TRPV4 activator, and to examine its mechanism of action. In three species (mouse, rat, and dog), the i.v. administration of GSK1016790A induced a dose-dependent reduction in blood pressure, followed by profound circulatory collapse. In contrast, GSK1016790A had no acute cardiovascular effects in the TRPV4 Ϫ/Ϫ null mouse. Hemodynamic analyses in the dog and rat demonstrate a profound reduction in cardiac output. However, GSK1016790A had no effect on rate or contractility in the isolated, buffer-perfused rat heart, and it produced potent endothelial-dependent relaxation of rodent-isolated vascular ring segments that were abolished by nitric-oxide synthase (NOS) inhibition (N-nitro-L-arginine methyl ester; L-NAME), ruthenium red, and endothelial NOS (eNOS) gene deletion. However, the in vivo circulatory collapse was not altered by NOS inhibition (L-NAME) or eNOS gene deletion but was associated with (concentration and time appropriate) profound vascular leakage and tissue hemorrhage in the lung, intestine, and kidney. TRPV4 immunoreactivity was localized in the endothelium and epithelium in the affected organs. GSK1016790A potently induced rapid electrophysiological and morphological changes (retraction/condensation) in cultured endothelial cells. In summary, inappropriate activation of TRPV4 produces acute circulatory collapse associated with endothelial activation/injury and failure of the pulmonary microvascular permeability barrier. It will be important to determine the role of TRPV4 in disorders associated with edema and microvascular congestion.Evidence suggests that the transient receptor potential (TRP) vanilloid subtype 4 (V4), a member of the TRP family, is a thermo/osmo/mechanosensitive cationic channel that regulates intracellular Ca 2ϩ -homeostasis and cell volume (for review, see Plant and Strotmann, 2007). The TRPV4 message is expressed in cardiovascular tissues (heart and blood vessels), and evidence of functional expression has been demonstrated in vascular smooth muscle and endothelial cells (Earley, 2006;Inoue et al., 2006;Yang et al., 2006). In the endothelium, activation of TRPV4 by ligands or shearstress triggers nitric oxide (NO)-dependent vasorelaxation (Kohler et al., 2006). These studies suggest that TRPV4 activation is linked mechanistically to NO generation during the process of endothelial mechanotransduction.TRPV4 also seems to play a role in fluid distribution and integrity of endothelial/epithelial barriers. It is important to note that TRPV4 activation in the lung microvasculature Article, publication date, and citation information can be found at

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Englerin A is a structurally unique natural product reported to selectively inhibit growth of renal cell carcinoma cell lines. A large scale phenotypic cell profiling experiment (CLiP) of englerin A on ¬over 500 well characterized cancer cell lines showed that englerin A inhibits growth of a subset of tumor cell lines from many lineages, not just renal cell carcinomas. Expression of the TRPC4 cation channel was the cell line feature that best correlated with sensitivity to englerin A, suggesting the hypothesis that TRPC4 is the efficacy target for englerin A. Genetic experiments demonstrate that TRPC4 expression is both necessary and sufficient for englerin A induced growth inhibition. Englerin A induces calcium influx and membrane depolarization in cells expressing high levels of TRPC4 or its close ortholog TRPC5. Electrophysiology experiments confirmed that englerin A is a TRPC4 agonist. Both the englerin A induced current and the englerin A induced growth inhibition can be blocked by the TRPC4/C5 inhibitor ML204. These experiments confirm that activation of TRPC4/C5 channels inhibits tumor cell line proliferation and confirms the TRPC4 target hypothesis generated by the cell line profiling. In selectivity assays englerin A weakly inhibits TRPA1, TRPV3/V4, and TRPM8 which suggests that englerin A may bind a common feature of TRP ion channels. In vivo experiments show that englerin A is lethal in rodents near doses needed to activate the TRPC4 channel. This toxicity suggests that englerin A itself is probably unsuitable for further drug development. However, since englerin A can be synthesized in the laboratory, it may be a useful chemical starting point to identify novel modulators of other TRP family channels.

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We cloned a rat vascular chymase (RVCH) from smooth muscle cells (SMCs) that converts angiotensin I to II and is up-regulated in SMC from spontaneously hypertensive vs. normotensive rats. To determine whether increased activity of RVCH is sufficient to cause hypertension, transgenic mice were generated with targeted conditional expression of RVCH to SMC, with the use of the tetracyclinecontrolled transactivator (tTA). We confirmed conditional expression of RVCH by mRNA, protein, and chymase activity in the absence, but not in the presence, of dietary doxycycline. The systolic blood pressure (mmHg), measured by carotid artery cannulation at 10 -12 weeks of age, was higher in tTA؉͞RVCH؉ mice than in nonbinary transgenic littermates (136 ؎ 4 vs. 109 ؎ 3) (P < 0.05), as were the diastolic and mean pressures. Hypertension was completely reversed by doxycycline, suggesting a causal link with chymase expression. Medial thickening of mesenteric arteries from tTA؉͞RVCH؉ mice vs. littermates (0.82 ؎ 0.1 vs. 0.42 ؎ 0.02) (P < 0.05) was associated with increased SMC proliferation, as judged by positive immunoreactivity, with the use of an antibody to the proliferating cell nuclear antigen. These structural changes were prevented by doxycycline. Perfusion myography of mesenteric arteries from tTA؉͞RVCH؉ mice also revealed increased vasoconstriction in response to phenylephrine and impaired metacholine-induced vasodilatation when compared with littermate controls or with the doxycyline-treated group. Our studies suggest that up-regulation of this vascular chymase is sufficient to cause a hypertensive arteriopathy, and that RVCH may be a candidate gene and a therapeutic target in patients with high blood pressure.smooth muscle cells C hymases are serine proteinases that were thought to be produced exclusively by mast cells in blood vessels and the myocardium. These enzymes generate angiotensin (Ang) II in large and resistance vessels (1-3) and in the heart (4) and also can convert big-endothelin-1 to endothelin-1 in vitro (5) and inactivate the vasodilator bradykinin (6). That chymase-dependent Ang II formation can occur in the intact animal is evident from studies in the baboon, where it was shown that Ang-converting enzyme inhibitors could not prevent the hemodynamic response to [Pro11, Da1a12]Ang I, which produces Ang II upon incubation with chymase (7). In addition to blood pressure regulation, both Ang II and endothelin-1 are involved in the stimulation of vascular smooth muscle cell (SMC) growth and proliferation, vascular remodeling, and cardiac hypertrophy (8, 9). Chymases also can influence structural remodeling through their ability to degrade extracellular matrix proteins, including fibronectin and collagen type IV, possibly through activation of matrix metalloproteinases (MMPs) (MMP-1, MMP-3, and MMP-9) (10-12). Chymases also stimulate collagen fibirillogenesis by cleavage of type I procollagen (13).Several studies have provided compelling data to suggest that chymases could play a central role in cardiovascular di...