No abstract

Plant volatile organic compounds, which are generated in a tissue-specific manner, play important ecological roles in the interactions between plants and their environments, including the well-known functions of attracting pollinators and protecting plants from herbivores/fungi attacks. However, to date, there have not been reports of holistic volatile profiling of the various tissues of a single plant species, even for the model plant species. In this study, we qualitatively and quantitatively analyzed 85 volatile chemicals, including 36 volatile terpenes, in 23 different tissues of cucumber (Cucumis sativus) plants using solid-phase microextraction combined with gas chromatography-mass spectrometry. Most volatile chemicals were found to occur in a highly tissue-specific manner. The consensus transcriptomes for each of the 23 cucumber tissues were generated with RNA sequencing data and used in volatile organic compound-gene correlation analysis to screen for candidate genes likely to be involved in cucumber volatile biosynthetic pathways. In vitro biochemical characterization of the candidate enzymes demonstrated that TERPENE SYNTHASE11 (TPS11)/TPS14, TPS01, and TPS15 were responsible for volatile terpenoid production in the roots, flowers, and fruit tissues of cucumber plants, respectively. A functional heteromeric geranyl(geranyl) pyrophosphate synthase, composed of an inactive small subunit (type I) and an active large subunit, was demonstrated to play a key role in monoterpene production in cucumber. In addition to establishing a standard workflow for the elucidation of plant volatile biosynthetic pathways, the knowledge generated from this study lays a solid foundation for future investigations of both the physiological functions of cucumber volatiles and aspects of cucumber flavor improvement.

Common chromosomal fragile sites FRA3B and FRA16D are frequent sites of DNA instability in cancer, but their contribution to cancer cell biology is not yet understood. Genes that span these sites (FHIT and WWOX, respectively) are often perturbed (either increased or decreased) in cancer cells and both are able to suppress tumour growth. While WWOX has some tumour suppressor characteristics, its normal role and functional contribution to cancer has not been fully determined. We find that a significant proportion of Drosophila Wwox interactors identified by proteomics and microarray analyses have roles in aerobic metabolism. Functional relationships between Wwox and either CG6439/isocitrate dehydrogenase (Idh) or Cu–Zn superoxide dismutase (Sod) were confirmed by genetic interactions. In addition, altered levels of Wwox resulted in altered levels of endogenous reactive oxygen species. Wwox (like FHIT) contributes to pathways involving aerobic metabolism and oxidative stress, providing an explanation for the ‘non-classical tumour suppressor’ behaviour of WWOX. Fragile sites, and the genes that span them, are therefore part of a protective response mechanism to oxidative stress and likely contributors to the differences seen in aerobic glycolysis (Warburg effect) in cancer cells.

Cerebral ischemia that develops after subarachnoid hemorrhage (SAH) carries high morbidity and mortality. Inflammatory mediators are involved in the development of cerebral ischemia through activation of the mitogen-activated protein kinase pathway. We hypothesized that blockade of the MAPkinase/ERK (MEK)/extracellular signal-regulated kinase (ERK) pathway upstream with a specific raf inhibitor would prevent SAH-induced activation of the cerebrovascular inflammatory response. The raf inhibitor SB-386023-b was injected intracisternally in our rat model at 0, 6, or 12 hours after the SAH. After 48 hours, cerebral arteries were harvested, and iNOS, interleukin (IL)-6, IL-1b, matrix metalloproteinase (MMP)-9, tissue inhibitors of metalloproteinase (TIMP)-1, and phosphorylated ERK1/2 were investigated by immunofluorescence, real-time polymerase chain reaction (PCR), and Western blot analysis. Cerebral blood flow (CBF) was measured using autoradiography. Protein levels of MMP-9, TIMP-1, iNOS, IL-6, and IL-1b were increased after SAH, as were mRNA levels of IL-6, MMP-9, and TIMP-1. After SAH, pERK1/2 was increased, but CBF was reduced. Treatment with SB-386023-b at 0 or 6 hours after SAH normalized CBF and prevented SAH-induced upregulation of MMPs, pro-inflammatory cytokines, and pERK1/2 proteins. These results suggested that inhibition of MEK/ERK signal transduction by a specific raf inhibitor administered up to 6 hours after SAH normalized the expression of pro-inflammatory mediators and extracellular matrix-related genes.

BackgroundTumour necrosis factor-α (TNF-α) is a pleiotropic pro-inflammatory cytokine, which is rapidly upregulated in the brain after injury. TNF-α acts by binding to its receptors, TNF-R1 (p55) and TNF-R2 (p75), on the cell surface. The aim of this study was first to investigate if there is altered expression of TNF-α and TNF-α receptors in cerebral artery walls following global or focal ischemia, and after organ culture. Secondly, we asked if the expression was regulated via activation of the MEK-ERK1/2 pathway.MethodsThe hypothesis was tested in vivo after subarachnoid hemorrhage (SAH) and middle cerebral artery occlusion (MCAO), and in vitro by organ culture of isolated cerebral arteries. The localization and amount of TNF-α, TNF-α receptor 1 and 2 proteins were analysed by immunohistochemistry and western blot after 24 and 48 h of organ culture and at 48 h following SAH or MCAO. In addition, cerebral arteries were incubated for 24 or 48 h in the absence or presence of a B-Raf inhibitor (SB386023-b), a MEK- inhibitor (U0126) or an NF-κB inhibitor (IMD-0354), and protein expression evaluated.ResultsImmunohistochemistry revealed enhanced expression of TNF-α, TNF-R1 and TNF-R2 in the walls of cerebral arteries at 48 h after MCAO and SAH compared with control. Co-localization studies showed that TNF-α, TNF-R1 and TNF-R2 were primarily localized to the cell membrane and the cytoplasm of the smooth muscle cells (SMC). There was, in addition, some expression of TNF-R2 in the endothelial cells. Immunohistochemistry and western blot analysis showed that these proteins were upregulated after 24 and 48 h in culture, and this upregulation reached an apparent maximum at 48 h of organ culture. Treatment with U0126 significantly reduced the enhanced SMC expression of TNF-α, TNF-R1 and TNF-R2 immunoreactivities after 24 and 48 h of organ culture. The Raf and NF-κB inhibitors significantly reduced organ culture induced TNF-α expression while they had minor effects on the TNF-α receptors.ConclusionThe present study shows that cerebral ischemia and organ culture induce expression of TNF-α and its receptors in the walls of cerebral arteries and that upregulation is transcriptionally regulated via the MEK/ERK pathway.

Two GFPPS linked (+)-thalianatriene (1) and (-)-retigeranin B (2) sesterterpene synthase genes were identified from the genome of Arabidopsis thaliana. 1 possesses an unprecedented 11-6-5 tricyclic ring system, while 2 contains a characteristic 5-5-5-6-5 pentacyclic ring system. Their structures were determined by extensive NMR spectroscopy, chemical derivatization, and X-ray crystallography. The variable-temp NMR measurement of 3, a diepoxy-bearing derivative of 1, enables us to completely assign the NMR signals of the two conformers as 3a (67%, UUU) and 3b (33%, UUD). A plausible biosynthesis mechanism of 1 was proposed.

Background: Cerebral ischemia is usually characterized by a reduction in local blood flow and metabolism and by disruption of the blood-brain barrier in the infarct region. The formation of oedema and opening of the blood-brain barrier in stroke is associated with enhanced expression of metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1).

Cigarette smoke exposure is a strong risk factor for cardiovascular and respiratory diseases. However, the knowledge about how cigarette smoke induces damage to vasculature and airway is limited. The present study was designed to examine the effects of cigarette smoke particles extracted by heptane (heptane-soluble smoke particles, HSP), by water (watersoluble smoke particles, WSP) and by DMSO (DMSO-soluble smoke particles, DSP), which represent lipophilic, hydrophilic and ambiphoteric constituents from the cigarette smoke, respectively. Human aortic smooth muscle cell (HASMC) proliferation was assessed in cell culture. Rat resistance artery and airway contractile responses to serotonin, U46619, phenylephrine, noradrenaline, acetylcholine, des-Arg 9 -bradykinin, bradykinin, sarafotoxin 6c and endothelin-1 were monitored by a sensitive myograph system. Immunocytochemistry and cell-based phosphoELISA assay were used to demonstrate activation of extracellular signal-regulated kinases 1 ⁄ 2 (ERK1 ⁄ 2). For the first time, our results demonstrate that although all the three extracts promote HASMC proliferation, the HSP and DSP effects occur earlier. HSP and DSP, but not WSP, increase the contractile responses to sarafotoxin 6c, U46619 or bradykinin in rat mesenteric artery and ⁄ or in bronchi. ERK1 ⁄ 2 is activated by HSP and DSP in HASMCs and inhibition of ERK1 ⁄ 2 abrogated the smoke extracts-induced HASMC proliferation, while blockage of nicotinic receptors had no effects, suggesting that the toxic effects of the smoke extracts occur via activation of intracellular ERK1 ⁄ 2 signalling, but not nicotinic receptors.Exposure to cigarette smoke is strongly associated with cardiovascular diseases like atherosclerosis, coronary heart disease, stroke, myocardial infarction, aortic aneurysm and peripheral vascular disease [1,2]. In airways, cigarette smoke, via direct smoking or second-hand inhalation, exacerbates chronic obstructive pulmonary disease (COPD), chronic bronchitis and asthma and also causes lung cancer. An estimated 1.6 million cardiovascular deaths worldwide are attributable to smoking [3] and about 73% of the COPD mortality is related to cigarette smoke exposure [4]. However, our knowledge about how cigarette smoke induces cardiovascular and airway diseases is very limited.Cigarette smoke contains over 4000 different compounds. Among them, 158 chemical constituents of smoke are identified to be toxic hazards. Of the 17 non-cancer compounds, based on a non-cancer risk index, 15 individual chemical constituents contribute to the respiratory irritation and the cardiovascular dysfunction [5]. Previously, we have revealed that DMSO-soluble smoke particles (DSP), but not nicotine or water-soluble cigarette smoke particles (WSP), induce dysfunctions of vascular smooth muscle cells [6] suggesting that it is the lipid-soluble particles from cigarette smoke that are responsible for the toxic effects. Furthermore, DSP exerts a direct toxic effect on vascular endothelial cells and reduces endothelium-depende...