Dataset Information

5,14-HEDGE, a 20-HETE mimetic, reverses hypotension and improves survival in a rodent model of septic shock: contribution of soluble epoxide hydrolase, CYP2C23, MEK1/ERK1/2/IKK?/I?B-?/NF-?B pathway, and proinflammatory cytokine formation.

ABSTRACT: We have previously demonstrated that a stable synthetic analog of 20-HETE, N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-HEDGE), restores vascular reactivity, blood pressure, and heart rate in endotoxemic rats. The aim of this study was to determine whether decreased renal expression and activity of soluble epoxide hydrolase (sEH), MEK1, ERK1/2, IKK?, I?B-?, and NF-?B as well as systemic and renal proinflammatory cytokine production associated with increased expression and activity of CYP2C23 contributes to the effect of 5,14-HEDGE to prevent hypotension, tachycardia, inflammation, and mortality in response to systemic administration of lipopolysaccharide (LPS). Blood pressure fell by 33 mmHg and heart rate rose by 57 beats/min in LPS (10 mg/kg, i.p.)-treated rats. Administration of LPS also increased mRNA and protein expression of sEH associated with a decrease in CYP2C23 mRNA and protein expression. Increased activity of sEH and p-MEK1, p-ERK1/2, p-I?B-?, NF-?B, and p-NF-?B protein levels as well as TNF-? and IL-8 production by LPS were also associated with a decreased activity of AA epoxygenases. These effects of LPS were prevented by 5,14-HEDGE (30 mg/kg, s.c.; 1 h after LPS). Treatment of endotoxemic mice with 5,14-HEDGE also raised the survival rate of animals from 84% to 98%. A competitive antagonist of vasoconstrictor effects of 20-HETE, 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid, 20-HEDE (30 mg/kg, s.c.; 1 h after LPS) prevented the effects of 5,14-HEDGE on blood pressure, heart rate, expression and/or activity of sEH, CYP2C23, and ERK1/2 as well as TNF-? and IL-8 levels in rats treated with LPS. These results suggest that decreased expression and/or activity of sEH and MEK1/ERK1/2/IKK?/I?B-?/NF-?B pathway as well as proinflammatory cytokine production associated with increased CYP2C23 expression and antiinflammatory mediator formation participate in the protective effect of 5,14-HEDGE against hypotension, tachycardia, inflammation, and mortality in the rodent model of septic shock.

SUBMITTER: Tunctan B

PROVIDER: S-EPMC3739859 | biostudies-literature | 2013 Apr-May

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:We have previously demonstrated that a stable synthetic analog of 20-hydroxyeicosatetraenoic acid (20-HETE), N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-HEDGE), prevents vascular hyporeactivity, hypotension, tachycardia, and inflammation in rats treated with lipopolysaccharide (LPS) and mortality in endotoxemic mice. These changes were attributed to decreased production of inducible nitric oxide (NO) synthase (iNOS)-derived NO, cyclooxygenase (COX)-2-derived vasodilator prostanoids, and proinflammatory mediators associated with increased cyctochrome P450 (CYP) 4A1-derived 20-HETE and CYP2C23-dependent antiinflammatory mediator formation. The aim of this study was to determine whether decreased expression and activity of iNOS, soluble guanylyl cyclase (sGC), protein kinase G (PKG), COX-2, gp91(phox) (NOX2; a superoxide generating NOX enzyme), and peroxynitrite production associated with increased expression of COX-1 and CYP4A1 and 20-HETE formation in renal and cardiovascular tissues of rats contributes to the effect of 5,14-HEDGE to prevent vasodilation, hypotension, tachycardia, and inflammation in response to systemic administration of LPS. Mean arterial pressure fell by 28mmHg and heart rate rose by 47beats/min in LPS (10mg/kg, i.p.)-treated rats. Administration of LPS also increased mRNA and protein expression of iNOS and COX-2 associated with a decrease in COX-1 and CYP4A1 mRNA and protein expression. Increased NOS activity, iNOS-heat shock protein 90 complex formation (an index for iNOS activity), protein expression of phosphorylated vasodilator stimulated phosphoprotein (an index for PKG activity), gp91(phox), p47(phox) (NOXO2; organizer subunit of gp91(phox)), and nitrotyrosine (an index for peroxynitrite production) as well as cGMP (an index for sGC activity), 6-keto-PGF1α (a stable metabolite PGI2) and PGE2 levels (indexes for COX activity), and nitrotyrosine levels by LPS were also associated with decreased CYP hydroxylase activity as measured by 20-HETE formation from arachidonic acid in renal microsomes of LPS-treated rats. These effects of LPS, except iNOS mRNA and COX-1 protein expression, were prevented by 5,14-HEDGE (30mg/kg, s.c.; 1h after LPS). A competitive antagonist of vasoconstrictor effects of 20-HETE, 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (30mg/kg, s.c.; 1h after LPS) reversed the effects of 5,14-HEDGE, except iNOS and COX-1 mRNA and protein expression as well as expression of CYP4A1 mRNA. These results suggest that increased CYP4A1 expression and 20-HETE formation associated with suppression of iNOS/sGC/PKG pathway, COX-2, and gp91(phox) participate in the protective effect of 5,14-HEDGE against vasodilation, hypotension, tachycardia, and inflammation in the rat model of septic shock.

Project description:Mechanical compression is a common abnormality of brain tumors that has been shown to be responsible for the severe neurological defects of brain cancer patients representing a negative prognostic factor. Indeed, it is of note that patients that undergo resection exhibited higher survival rates than those subjected to biopsy only, suggesting that compressive forces generated during brain tumor growth play a key role in tumor progression. Despite the importance of mechanical compression in brain tumors, there is a lack of studies examining its direct effects on brain cancer cells and the mechanisms involved. In the present study, we used two brain cancer cell lines with distinct metastatic potential, the less aggressive H4 and the highly aggressive A172 cell lines, in order to study the effect of compression on their proliferative and migratory ability. Specifically, we used multicellular tumor spheroids (MCS) embedded in agarose matrix to show that compression strongly impaired their growth. Using mathematical modeling, we estimated the levels of compressive stress generated during the growth of brain MCS and then we applied the respective stress levels on brain cancer cell monolayers using our previously established transmembrane pressure device. By performing a scratch assay, we found that compression strongly induced the migration of the less aggressive H4 cells, while a less pronounced effect was observed for A172 cells. Analysis of the gene expression profile of both cell lines revealed that GDF15 and small GTPases are strongly regulated by mechanical compression, while GDF15 was further shown to be necessary for cells to migrate under compression. Through a phospho-proteomic screening, we further found that compressive stimulus is transmitted through the MEK1/Erk1 signaling pathway, which is also necessary for the migration of brain cancer cells. Finally, our results gave the first indication that GDF15 could regulate and being regulated by MEK1/Erk1 signaling pathway in order to facilitate the compression-induced brain cancer cell migration, rendering them along with small GTPases as potential targets for future anti-metastatic therapeutic innovations to treat brain tumors.

Project description:Primary effusion lymphoma (PEL) is a lymphogenic disorder associated with Kaposi's sarcoma-associated herpesvirus (KSHV) infection. Key to the survival and proliferation of PEL is the canonical NF-?B pathway, which becomes constitutively activated following overexpression of the viral oncoprotein KSHV vFLIP (ks-vFLIP). This arises from its capacity to form a complex with the modulatory subunit of the I?B kinase (IKK) kinase, IKK? (or NEMO), resulting in the overproduction of proteins that promote cellular survival and prevent apoptosis, both of which are important drivers of tumorigenesis. Using a combination of cell-based and biophysical assays together with structural techniques, we showed that the observed resistance to cell death is largely independent of autophagy or major death receptor signaling pathways and demonstrated that direct targeting of the ks-vFLIP-IKK? interaction both in cells and in vitro can be achieved using IKK?-mimetic peptides. Our results further reveal that these peptides not only induce cell killing but also potently sensitize PEL to the proapoptotic agents tumor necrosis factor alpha and etoposide and are the first to confirm ks-vFLIP as a tractable target for the treatment of PEL and related disorders.IMPORTANCE KSHV vFLIP (ks-vFLIP) has been shown to have a crucial role in cellular transformation, in which it is vital for the survival and proliferation of primary effusion lymphoma (PEL), an aggressive malignancy associated with infection that is resistant to the majority of chemotherapeutic drugs. It operates via subversion of the canonical NF-?B pathway, which requires a physical interaction between ks-vFLIP and the IKK kinase modulatory subunit IKK?. While this interaction has been directly linked to protection against apoptosis, it is unclear whether the suppression of other cell death pathways implicated in ks-vFLIP pathogenesis is an additional contributor. We demonstrate that the interaction between ks-vFLIP and IKK? is pivotal in conferring resistance to apoptosis. Additionally, we show that the ks-vFLIP-IKK? complex can be disrupted using peptides leading to direct killing and the sensitization of PEL cells to proapoptotic agents. Our studies thus provide a framework for future therapeutic interventions.

Project description:Purposeinhibitors of apoptosis proteins (IAPs) have been shown to contribute to resistance of neoplastic cells to chemotherapy and to biologic antineoplastic agents. Consequently, new agents are being developed targeting this family of proteins. In a panel of bladder cancer cell lines, we evaluated a Smac mimetic that antagonizes several IAPs for its suitability for bladder cancer therapy. Experimental design: A panel of seven bladder cancer cell lines were evaluated for sensitivity to the Smac mimetic compound-A alone, TRAIL alone, chemotherapy alone, compound-A plus TRAIL, and compound-A plus chemotherapy by DNA fragmentation analysis. IAP levels and caspase activation were examined by western blotting. Release of caspase-3 from X-linked inhibitor of apoptosis protein (XIAP), the most effective IAP, was assessed by immunoprecipitation and western blotting. Finally, siRNA knockdown of XIAP was correlated with the sensitivity of cells to apoptosis induced by compound-A plus TRAIL by DNA fragmentation and western blotting.Resultssingle-agent compound-A had little effect, but compound-A augmented TRAIL- and chemotherapy-induced apoptosis. Immunoblotting showed that combination treatment with compound-A and TRAIL resulted in cleavage of procaspase-3 and procaspase-7, activation of which irreversibly commits cells to apoptosis. Immunoprecipitation of XIAP showed displacement of active caspase-3 fragments from XIAP, supporting the proposed mechanism of action. Furthermore, siRNA-mediated silencing of XIAP similarly sensitized these cells to apoptosis.Experimental designa panel of seven bladder cancer cell lines were evaluated for sensitivity to the Smac mimetic compound-Alone, TRAIL alone, Chemotherapy alone, compound-A plus TRAIL and compound-A plus chemotherapy by DNA fragmentation analysis. IAP levels and caspase activation were examined by western blotting. Release of caspase-3 from X-linked inhibitor of apoptosis protein (XIAP), the most effective IAP, was assessed by immunoprecipitation and western blotting. Finally siRNA knockdown of XIAP was correlated with the sensitivity of cells to apoptosis induced by compound-A plus TRAIL by DNA fragmentation and western blotting.Conclusionour results suggest that targeting of XIAP with the Smac mimetic compound-A has the potential to augment the effects of a variety of chemotherapeutic and biologic therapies in bladder cancer.

Dataset Information

5,14-HEDGE, a 20-HETE mimetic, reverses hypotension and improves survival in a rodent model of septic shock: contribution of soluble epoxide hydrolase, CYP2C23, MEK1/ERK1/2/IKK?/I?B-?/NF-?B pathway, and proinflammatory cytokine formation.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets