Cyclooxygenase-2-linked attenuation of hypoxia-induced pulmonary hypertension and intravascular thrombosis

Exogenous prostacyclin is effective in reducing pulmonary vascular resistance in some forms of human pulmonary hypertension (PH). To explore whether endogenous prostaglandins played a similar role in pulmonary hypertension, we examined the effect of deleting cyclooxygenase (COX)-gene isoforms in a chronic hypoxia model of PH. Pulmonary hypertension, examined by direct measurement of right ventricular end systolic pressure (RVESP), right ventricular hypertrophy (n = 8), and hematocrit (n = 3), was induced by 3 weeks of hypobarichypoxia in wild-type and COX-knockout (KO) mice. RVESP was increased in wild-type hypoxic mice compared with normoxic controls (24.4 ± 1.4 versus 13.8 ± 1.9 mm Hg; n = 8; p < 0.05). COX-2 KO mice showed a greater increase in RVESP following hypoxia (36.8 ± 2.7 mm Hg; p < 0.05). Urinary thromboxane (TX)B2 excretion increased following hypoxia (44.6 ± 11.1 versus 14.7 ± 1.8 ng/ml; n = 6; p < 0.05), an effect that was exacerbated by COX-2 gene disruption (54.5 ± 10.8 ng/ml; n = 6). In contrast, the increase in 6-keto-prostacyclin1α excretion following hypoxia was reduced by COX-2 gene disruption (29 ± 3 versus 52 ± 4.6 ng/ml; p < 0.01). Tail cut bleed times were lower following hypoxia, and there was evidence of intravascular thrombosis in lung vessels that was exacerbated by disruption of COX-2 and reduced by deletion of COX-1. The TXA2/endoperoxide receptor antagonist ifetroban (50 mg/kg/day) offset the effect of deleting the COX-2 gene, attenuating the hypoxia-induced rise in RVESP and intravascular thrombosis. COX-2 gene deletion exacerbates pulmonary hypertension, enhances sensitivity to TXA2, and induces intravascular thrombosis in response to hypoxia. The data provide evidence that endogenous prostaglandins modulate the pulmonary response to hypoxia. Copyright © 2008 by The American Society for Pharmacology and Experimental Therapeutics.

Angiogenesis and non-small cell lung cancer

Lung cancer is the commonest cause of cancer death in the western world. Recent evidence suggests that angiogenesis is related to poor prognosis in many solid tumours including non-small cell lung cancer. Angiogenesis is controlled by a complex interaction between growth and apoptotic factors, proteases and adhesion molecules. The angiogenic process may prove a target for novel therapies such as matrix metalloproteinase inhibitors, growth factor antisense RNA, growth factor receptor antagonists and naturally occurring antiangiogenic peptides. These agents may be used alone or in combination with traditional chemotherapy, radiotherapy and surgery. (C) 2000 Elsevier Science Ireland Ltd.

The effects and interactions of GABAergic and dopaminergic agents in the prevention of form deprivation myopia by brief periods of normal vision

Intravitreal injections of GABA antagonists, dopamine agonists and brief periods of normal vision have been shown separately to inhibit form-deprivation myopia (FDM). Our study had three aims: (i) establish whether GABAergic agents modify the myopia protective effect of normal vision, (ii) investigate the receptor sub-type specificity of any observed effect, and (iii) consider an interaction with the dopamine (DA) system. Prior to the period of normal vision GABAergic agents were applied either (i) individually, (ii) in combination with other GABAergic agents (an agonist with an antagonist), or (iii) in combination with DA agonists and antagonists. Water injections were given to groups not receiving drug treatments so that all experimental eyes received intravitreal injections. As shown previously, constant form-deprivation resulted in high myopia and when diffusers were removed for 2 h per day the period of normal vision greatly reduced the FDM that developed. GABA agonists inhibited the protective effect of normal vision whereas antagonists had the opposite effect. GABAA/C agonists and D2 DA antagonists when used in combination were additive in suppressing the protective effect of normal vision. A D2 DA agonist restored some of the protective effect of normal vision that was inhibited by a GABA agonist (muscimol). The protective effect of normal vision against form-deprivation is modifiable by both the GABAergic and DAergic pathways.

Advances in the systemic therapy of malignant pleural mesothelioma

Malignant pleural mesothelioma is an aggressive thoracic malignancy associated with exposure to asbestos, and its incidence is anticipated to increase during the first half of this century. Chemotherapy is the mainstay of treatment, yet sufficiently robust evidence to substantiate the current standard of care has emerged only in the past 5 years. This Review summarizes the evidence supporting the clinical activity of chemotherapy, discusses the use of end points for its assessment and examines the influence of clinical and biochemical prognostic factors on the natural history of malignant pleural mesothelioma. Early-phase clinical trials of second-line and novel agents are emerging from an increased understanding of mesothelioma cell biology. Coupled with high-quality translational research, such developments have real potential to improve the outlook of patients at a time of increasing incidence.

Cardiac safety concerns for ondansetron, an antiemetic commonly used for nausea linked to cancer treatment and following anaesthesia

Introduction: Ondansetron is a 5-HT3 receptor antagonist commonly used as an anti-emetic to prevent the nausea and vomiting associated with anti-cancer drugs, cancer radiotherapy, or postoperatively. Recently, the US Food and Drug Administration (FDA) issued a warning for ondansetron due to a potential for prolongation of the QT interval of the electrocardiogram (ECG), a phenomenon that is associated with an increased risk of the potentially fatal arrhythmia torsade de pointes. Areas covered: We undertook a review of the cardiac safety of ondansetron. Our primary sources of information were PubMed (with downloading of full articles), and the internet. Expert opinion: The dose of ondansetron that the FDA has concerns about is 32 mg iv (or several doses that are equivalent to this), which is only used in preventing nausea and vomiting associated with cancer chemotherapy. This suggests that ondansetron may be safe in the lower doses used to prevent the nausea and vomiting in radiation treatment or postoperatively. However, as there is a report that a lower dose of ondansetron prolonged the QT interval in healthy volunteers, this needs to be clarified by the FDA. More research needs to be undertaken of the relationship between QT prolongation and torsades in order that the FDA can produce clear-cut evidence of pro-arrhythmic risk when introducing warnings for this.

Cardiac safety concerns for domperidone, an antiemetic and prokinetic, and galactogogue medicine

Introduction: Domperidone is a dopamine D2-receptor antagonist developed as an antiemetic and prokinetic agents. Oral domperidone is not approved in the US, but is used in many countries to treat nausea and vomiting, gastroparesis, and as a galactogogue (to promote lactation). The US Food and Drug Administration (FDA) have issued a warning about the cardiac safety of domperidone. Areas covered: The authors undertook a review of the cardiac safety of oral domperidone. Expert opinion: The data from preclinical studies are unambiguous in identifying domperidone as able to produce marked hERG channel inhibition and action potential prolongation at clinically relevant concentrations. The compound’s propensity to augment instability of action potential duration and action potential triangulation are also indicative of proarrhythmic potential. Domperidone should not be administered to subjects with pre-existing QT prolongation/LQTS, subjects receiving drugs that inhibit CYP3A4, subjects with electrolyte abnormalities or with other risk factors for QT-prolongation. With these provisos, it is possible that domperidone may be used as a galactogogue without direct risk to healthy breast feeding women but more safety information should be sought in this situation. Also, more safety information is required regarding risk to breast feeding infants or before domperidone is routinely used in gastroparesis or gastroesphageal reflux in children.

Acquired differential regulation of caspase-8 in cisplatin-resistant non-small-cell lung cancer

Failure to efficiently induce apoptosis contributes to cisplatin resistance in non-small-cell lung cancer (NSCLC). Although BCL-2-associated X protein (BAX) and BCL-2 antagonist killer (BAK) are critical regulators of the mitochondrial apoptosis pathway, their requirement has not been robustly established in relation to cisplatin. Here, we show that cisplatin can efficiently bypass mitochondrial apoptosis block caused by loss of BAX and BAK, via activation of the extrinsic death receptor pathway in some model cell lines. Apoptosis resistance following cisplatin can only be observed when both extrinsic and intrinsic pathways are blocked, consistent with redundancy between mitochondrial and death receptor pathways in cisplatin-induced apoptosis. In H460 NSCLC cells, caspase-8 cleavage was shown to be induced by cisplatin and is dependent on death receptor 4, death receptor 5, Fas-associated protein with death domain, acid sphingomyelinase and ceramide synthesis. In contrast, cisplatin-resistant cells fail to activate caspase-8 via this pathway despite conserving sensitivity to death ligand-driven activation. Accordingly, caspase-8 activation block acquired during cisplatin resistance, can be bypassed by death receptor agonism.

Gelatine methacrylamide-based hydrogels : an alternative three-dimensional cancer cell culture system

Modern cancer research requires physiological, three-dimensional (3-D) cell culture platforms, wherein the physical and chemical characteristics of the extracellular matrix (ECM) can be modified. In this study, gelatine methacrylamide (GelMA)-based hydrogels were characterized and established as in vitro and in vivo spheroid-based models for ovarian cancer, reflecting the advanced disease stage of patients, with accumulation of multicellular spheroids in the tumour fluid (ascites). Polymer concentration (2.5-7% w/v) strongly influenced hydrogel stiffness (0.5±0.2kPa to 9.0±1.8kPa) but had little effect on solute diffusion. The diffusion coefficient of 70kDa fluorescein isothiocyanate (FITC)-labelled dextran in 7% GelMA-based hydrogels was only 2.3 times slower compared to water. Hydrogels of medium concentration (5% w/v GelMA) and stiffness (3.4kPa) allowed spheroid formation and high proliferation and metabolic rates. The inhibition of matrix metalloproteinases and consequently ECM degradability reduced spheroid formation and proliferation rates. The incorporation of the ECM components laminin-411 and hyaluronic acid further stimulated spheroid growth within GelMA-based hydrogels. The feasibility of pre-cultured GelMA-based hydrogels as spheroid carriers within an ovarian cancer animal model was proven and led to tumour development and metastasis. These tumours were sensitive to treatment with the anti-cancer drug paclitaxel, but not the integrin antagonist ATN-161. While paclitaxel and its combination with ATN-161 resulted in a treatment response of 33-37.8%, ATN-161 alone had no effect on tumour growth and peritoneal spread. The semi-synthetic biomaterial GelMA combines relevant natural cues with tunable properties, providing an alternative, bioengineered 3-D cancer cell culture in in vitro and in vivo model systems.

Ghrelin receptor (GHS-R1A) antagonism suppresses both operant alcohol self-administration and high alcohol consumption in rats

The mechanisms involved in alcohol use disorders are complex. It has been shown that ghrelin is an important signal for the control of body weight homeostasis, preferably by interacting with hypothalamic circuits, as well as for drug reward by activating the mesolimbic dopamine system. The ghrelin receptor (GHS-R1A) has been shown to be required for alcohol-induced reward. Additionally, ghrelin increases and GHR-R1A antagonists reduce moderate alcohol consumption in mice, and a single nucleotide polymorphism in the GHS-R1A gene has been associated with high alcohol consumption in humans. However, the role of central ghrelin signaling in high alcohol consumption is not known. Therefore, the role of GHS-R1A in operant self-administration of alcohol in rats as well as for high alcohol consumption in Long-Evans rats and in alcohol preferring [Alko alcohol (AA)] rats was studied here. In the present study, the GHS-R1A antagonist, JMV2959, was found to reduce the operant self-administration of alcohol in rats and to decrease high alcohol intake in Long-Evans rats as well as in AA rats. These results suggest that the ghrelin receptor signaling system, specifically GHS-R1A, is required for operant self-administration of alcohol and for high alcohol intake in rats. Therefore, the GHS-R1A may be a therapeutic target for treatment of addictive behaviors, such as alcohol dependence.

Mifepristone in the central nucleus of the amygdala reduces yohimbine stress-induced reinstatement of ethanol-seeking

Chronic ethanol exposure leads to dysregulation of the hypothalamic-pituitary-adrenal axis, leading to changes in glucocorticoid release and function that have been proposed to maintain pathological alcohol consumption and increase vulnerability to relapse during abstinence. The objective of this study was to determine whether mifepristone, a glucocorticoid receptor antagonist, plays a role in ethanol self-administration and reinstatement. Male, Long-Evans rats were trained to self-administer either ethanol or sucrose in daily 30 min operant self-administration sessions using a fixed ratio 3 schedule of reinforcement. Following establishment of stable baseline responding, we examined the effects of mifepristone on maintained responding and yohimbine-induced increases in responding for ethanol and sucrose. Lever responding was extinguished in separate groups of rats and animals were tested for yohimbine-induced reinstatement and corticosterone release. We also investigated the effects of local mifepristone infusions into the central amygdala (CeA) on yohimbine-induced reinstatement of ethanol- and sucrose-seeking. In addition, we infused mifepristone into the basolateral amygdala (BLA) in ethanol-seeking animals as an anatomical control. We show that both systemic and intra-CeA (but not BLA) mifepristone administration suppressed yohimbine-induced reinstatement of ethanol-seeking, while only systemic injections attenuated sucrose-seeking. In contrast, baseline consumption, yohimbine-induced increases in responding, and circulating CORT levels were unaffected. The data indicate that the CeA plays an important role in the effects of mifepristone on yohimbine-induced reinstatement of ethanol-seeking. Mifepristone may be a valuable pharmacotherapeutic strategy for preventing relapse to alcohol use disorders and, as it is FDA approved, may be a candidate for clinical trials in the near future.

PPARγ-independent induction of growth arrest and apoptosis in prostate and bladder carcinoma

Background Although PPARγ antagonists have shown considerable pre-clinical efficacy, recent studies suggest PPARγ ligands induce PPARγ-independent effects. There is a need to better define such effects to permit rational utilization of these agents. Methods We have studied the effects of a range of endogenous and synthetic PPARγ ligands on proliferation, growth arrest (FACS analysis) and apoptosis (caspase-3/7 activation and DNA fragmentation) in multiple prostate carcinoma cell lines (DU145, PC-3 and LNCaP) and in a series of cell lines modelling metastatic transitional cell carcinoma of the bladder (TSU-Pr1, TSU-Pr1-B1 and TSU-Pr1-B2). Results 15-deoxy-prostaglandin J2 (15dPGJ2), troglitazone (TGZ) and to a lesser extent ciglitazone exhibited inhibitory effects on cell number; the selective PPARγ antagonist GW9662 did not reverse these effects. Rosiglitazone and pioglitazone had no effect on proliferation. In addition, TGZ induced G0/G1 growth arrest whilst 15dPGJ2 induced apoptosis. Conclusion Troglitazone and 15dPGJ2 inhibit growth of prostate and bladder carcinoma cell lines through different mechanisms and the effects of both agents are PPARγ-independent.

Frizzled-7 receptor ectodomain expression in a colon cancer cell line induces morphological change and attenuates tumor growth

Frizzled (FZD) receptors have a conserved N-terminal extracellular cysteine-rich domain that interacts with Wnts and co-expression of the receptor ectodomain can antagonize FZD-mediated signalling. Using the ectodomain as an antagonist we have modulated endogenous FZD7 signalling in the moderately differentiated colon adenocarcinoma cell line, SK-CO-1. Unlike the parental cell line, which grows as tightly associated adherent cell clusters, the FZD7 ectodomain expressing cells display a spread out morphology and grow as a monolayer in tissue culture. This transition in morphology was associated with decreased levels of plasma membrane-associated E-cadherin and β-catenin, localized increased levels of vimentin and redistribution of α6 integrin to cellular processes in the FZD7 ectodomain expressing cells. The morphological and phenotype changes induced by FZD7 ectodomain expression in SK-CO-1 cells is thus consistent with the cells undergoing an epithelial-to-mesenchymal-like transition. Furthermore, initiation of tumor formation in a xenograft tumor growth assay was attenuated in the FZD7 ectodomain expressing cells. Our results indicate a pivotal role for endogenous FZD7 in morphology transitions that are associated with colon tumor initiation and progression.

Epidermal growth factor-induced epithelio-mesenchymal transition in human breast carcinoma cells

PMC42-LA cells display an epithelial phenotype: the cells congregate into pavement epithelial sheets in which E-cadherin and β-catenin are localized at cell-cell borders. They abundantly express cytokeratins, although 5% to 10% of the cells also express the mesenchymal marker vimentin. Stimulation of PMC42-LA cells with epidermal growth factor (EGF) leads to epithelio-mesenchymal transition-like changes including up-regulation of vimentin and down-regulation of E-cadherin. Vimentin expression is seen in virtually all cells, and this increase is abrogated by treatment of cells with an EGF receptor antagonist. The expression of the mesenchyme-associated extracellular matrix molecules fibronectin and chondroitin sulfate proteoglycan also increase in the presence of EGF. PMC42-LA cells adhere rapidly to collagen I, collagen IV, and laminin-1 substrates and markedly more slowly to fibronectin and vitronectin. EGF increases the speed of cell adhesion to most of these extracellular matrix molecules without altering the order of adhesive preference. EGF also caused a time-dependent increase in the motility of PMC42-LA cells, commensurate with the degree of vimentin staining. The increase in motility was at least partly chemokinetic, because it was evident both with and without chemoattractive stimuli. Although E-cadherin staining at cell-cell junctions disappeared in response to EGF, β-catenin persisted at the cell periphery. Further analysis revealed that N-cadherin was present at the cell-cell junctions of untreated cells and that expression was increased after EGF treatment. N- and E-cadherin are not usually coexpressed in human carcinoma cell lines but can be coexpressed in embryonic tissues, and this may signify an epithelial cell population prone to epithelio-mesenchymal-like responses.

Role of serotonin via 5-HT2B receptors in the reinforcing effects of MDMA in mice

The amphetamine derivative 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) reverses dopamine and serotonin transporters to produce efflux of dopamine and serotonin, respectively, in regions of the brain that have been implicated in reward. However, the role of serotonin/dopamine interactions in the behavioral effects of MDMA remains unclear. We previously showed that MDMA-induced locomotion, serotonin and dopamine release are 5-HT(2B) receptor-dependent. The aim of the present study was to determine the contribution of serotonin and 5-HT(2B) receptors to the reinforcing properties of MDMA. We show here that 5-HT(2B) (-/-) mice do not exhibit behavioral sensitization or conditioned place preference following MDMA (10 mg/kg) injections. In addition, MDMA-induced reinstatement of conditioned place preference after extinction and locomotor sensitization development are each abolished by a 5-HT(2B) receptor antagonist (RS127445) in wild type mice. Accordingly, MDMA-induced dopamine D1 receptor-dependent phosphorylation of extracellular regulated kinase in nucleus accumbens is abolished in mice lacking functional 5-HT(2B) receptors. Nevertheless, high doses (30 mg/kg) of MDMA induce dopamine-dependent but serotonin and 5-HT(2B) receptor-independent behavioral effects. These results underpin the importance of 5-HT(2B) receptors in the reinforcing properties of MDMA and illustrate the importance of dose-dependent effects of MDMA on serotonin/dopamine interactions.

Atmospheric gas plasma–induced ROS production activates TNF-ASK1 pathway for the induction of melanoma cancer cell apoptosis

Atmospheric gas plasmas (AGPs) are able to selectively induce apoptosis in cancer cells, offering a promising alternative to conventional therapies that have unwanted side effects such as drug resistance and toxicity. However, the mechanism of AGP-induced cancer cell death is unknown. In this study, AGP is shown to up-regulate intracellular reactive oxygen species (ROS) levels and induce apoptosis in melanoma but not normal melanocyte cells. By screening genes involved in apoptosis, we identify tumor necrosis factor (TNF)-family members as the most differentially expressed cellular genes upon AGP treatment of melanoma cells. TNF receptor 1 (TNFR1) antagonist-neutralizing antibody specifically inhibits AGP-induced apoptosis signal, regulating apoptosis signal-regulating kinase 1 (ASK1) activity and subsequent ASK1-dependent apoptosis. Treatment of cells with intracellular ROS scavenger N-acetyl-l-cysteine also inhibits AGP-induced activation of ASK1, as well as apoptosis. Moreover, depletion of intracellular ASK1 reduces the level of AGP-induced oxidative stress and apoptosis. The evidence for TNF-signaling dependence of ASK1-mediated apoptosis suggests possible mechanisms for AGP activation and regulation of apoptosis-signaling pathways in tumor cells.