Human fibroblasts release reactive oxygen species in response to interleukin-1 or tumour necrosis factor-alpha.
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ABSTRACT:
SUBMITTER: Meier B
PROVIDER: S-EPMC1133461 | biostudies-other | 1989 Oct
REPOSITORIES: biostudies-other
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Project description:In this study, we show that reactive oxygen species production induced by tumour necrosis factor alpha (TNF-alpha) in L929 cells was associated with a decrease in the steady-state mRNA levels of the mitochondrial transcript ATPase 6-8. Simultaneously, the transcript levels of two nuclear-encoded glycolytic enzymes, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphofructokinase, were increased. These changes were associated with decreased protein levels of the ATPase subunit a (encoded by the mitochondrial ATPase 6 gene) and cytochrome c oxidase subunit II, and increased protein levels of phosphofructokinase. Since TNF-alpha had no effect on the amount of mitochondrial DNA, the results suggested that TNF-alpha acted at the transcriptional and/or post-transcriptional level. Reactive oxygen species scavengers, such as butylated hydroxianisole and butylated hydroxytoluene, blocked the production of free radicals, prevented the down-regulation of ATPase 6-8 transcripts, preserved the protein levels of ATPase subunit a and cytochrome c oxidase subunit II, and attenuated the cytotoxic response to TNF-alpha, indicating a direct link between these two phenomena.
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Project description:Childhood trauma confers higher risk of adulthood physical and mental illness; however, the biological mechanism mediating this association remains largely unknown. Recent research has suggested dysregulation of the immune system as a possible biological mediator. The present paper conducted a meta-analysis to establish whether early-life adversity contributes to potentially pathogenic pro-inflammatory phenotypes in adult individuals. A systematic search of Pubmed, PsycINFO, EMBASE, Scopus and Medline identified 25 articles for the meta-analysis, including 18 studies encompassing a sample of 16 870 individuals for C-reactive protein (CRP), 15 studies including 3751 individuals for interleukin-6 (IL-6) and 10 studies including 881 individuals for tumour necrosis factor-α (TNF-α). Random-effects meta-analysis showed that individuals exposed to childhood trauma had significantly elevated baseline peripheral levels of CRP (Fisher's z=0.10, 95% confidence interval (CI)=0.05-0.14), IL-6 (z=0.08, 95% CI=0.03-0.14) and TNF-α (z=0.23, 95% CI=0.14-0.32). Subgroup analyses for specific types of trauma (sexual, physical or emotional abuse) revealed that these impact differentially the single inflammatory markers. Moreover, meta-regression revealed greater effect sizes in clinical samples for the association between childhood trauma and CRP but not for IL-6 or TNF-α. Age, body mass index (BMI) and gender had no moderating effects. The analysis demonstrates that childhood trauma contributes to a pro-inflammatory state in adulthood, with specific inflammatory profiles depending on the specific type of trauma.
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Project description:In a variety of non-phagocytic cell types, there is a marked increase in intracellular levels of reactive oxygen species (ROS), including superoxide and H2O2, after ligand stimulation. We demonstrate that in NIH 3T3 cells transient expression of constitutively activated forms of the small GTP-binding proteins Ras or Rac1 leads to a significant increase in intracellular ROS. An increase in intracellular ROS is also demonstrated after growth factor [platelet-derived growth factor (PDGF) or epidermal growth factor (EGF)] or cytokine [tumour necrosis factor-alpha (TNF-alpha) or interleukin (IL)-1 beta] stimulation of NIH 3T3 cells. Expression of a dominant negative allele of Rac1 inhibits the rise in ROS seen after Ras expression or after stimulation by either growth factors or cytokines. These results provide the first demonstration of the pathway by which ligand stimulation of ROS occurs in non-phagocytic cells and suggest that the family of Ras-related small GTP-binding proteins may function as regulators of the intracellular redox state.
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Project description:There is abundant evidence that H2O2 can act as an endothelium-derived hyperpolarizing factor in the resistance vasculature. However, whilst scavenging H2O2 can abolish endothelial dependent hyperpolarization (EDH) and the associated vascular relaxation in some arteries, EDH-dependent vasorelaxation can often be mimicked only by using relatively high concentrations of H2O2. We have examined the role of H2O2 in EDH-dependent vasodilatation by simultaneously measuring vascular diameter and changes in endothelial cell (EC) [Ca(2+)]i during the application of H2O2 or carbachol, which triggers EDH. Carbachol (10µM) induced dilatation of phenylephrine-preconstricted rat cremaster arterioles was largely (73%) preserved in the presence of indomethacin (3µM) and l-NAME (300µM). This residual NO- and prostacyclin-independent dilatation was reduced by 89% upon addition of apamin (0.5µM) and TRAM-34 (10µM), and by 74% when an extracellular ROS scavenging mixture of SOD and catalase (S&C; 100Uml(-1) each) was present. S&C also reduced the carbachol-induced EC [Ca(2+)]i increase by 74%. When applied in Ca(2+)-free external medium, carbachol caused a transient increase in EC [Ca(2+)]i. This was reduced by catalase, and was enhanced when 1µM H2O2 was present in the bath. H2O2 -induced dilatation, which occurred only at concentrations ?100µM, was reduced by a blocking antibody to TRPM2, which had no effect on carbachol-induced responses. Similarly, iberotoxin and Rp-8bromo cGMP reduced the vasodilatation induced by H2O2, but not by carbachol. Inhibiting PLC, PLA2 or CYP450 2C9 each greatly reduced the carbachol-induced increase in EC [Ca(2+)]i and vasodilatation, but adding 10µM H2O2 during PLA2 or CYP450 2C9 inhibition completely restored both responses. The nature of the effective ROS species was investigated by using Fe(2+) chelators to block the formation of ?OH. A cell permeant chelator was able to inhibit EC Ca(2+) store release, but cell impermeant chelators reduced both the vasodilatation and EC Ca(2+) influx, implying that ?OH is required for these responses. The results indicate that rather than mediating EDH by acting directly on smooth muscle, H2O2 promotes EDH by acting within EC to enhance Ca(2+) release.
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Project description:Neutrophils (PMNs) are a vital part of host defense and are the principal leukocyte in innate immunity. Interleukin (IL)-18 is a proinflammatory cytokine with roles in both innate and adaptive immunity. We hypothesize that PMNs contain preformed IL-18, which is released in response to specific inflammatory stimuli. Isolated PMNs were stimulated with a battery of chemoattractants (5 min to 24 h), and IL-18 release was measured. PMNs were also separated into subcellular fractions and immunoblotted with antibodies against IL-18 or were fixed and probed with antibodies to IL-18 as well as to the contents of granules, intracellular organelles, and filamentous actin (F-actin), incubated with fluorescent secondary antibodies, and examined by digital microscopy. Quiescent PMNs contained IL-18 in the cytoplasm, associated with F-actin, as determined by positive fluorescence resonance energy transfer (FRET+). In turn, TNF-alpha stimulation disrupted the association of IL-18 with F-actin, induced a FRET+ interaction of IL-18 with lipid rafts, and elicited IL-18 release. Manipulation of F-actin status confirmed the relationship between IL-18 and F-actin in resting PMNs. Consequently, incubation with monomeric IL-18 binding protein inhibited TNF-alpha-mediated priming of the PMN oxidase. We conclude that human PMNs contain IL-18 associated with F-actin in the cytoplasm and TNF-alpha stimulation causes dissociation of IL-18 from F-actin, association with lipid rafts, and extracellular release. Extracellular IL-18 participates in TNF-alpha priming of the PMN oxidase as demonstrated by inhibition with the IL-18 binding protein.
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Project description:A new micelle drug carrier that consists of a diblock polymer of propylene sulfide (PS) and N,N-dimethylacrylamide (poly(PS??-b-DMA???)) has been synthesized and characterized for site-specific release of hydrophobic drugs to sites of inflammation. Propylene sulfide was first polymerized using a thioacyl group transfer (TAGT) method with the RAFT chain transfer agent (CTA) 4-cyano-4-(ethylsulfanylthiocarbonylsulfanyl) pentanoic acid (CEP), and the resultant poly(PS??-CEP) macro-CTA was used to polymerize a second polymer block of DMA using reversible addition-fragmentation chain transfer (RAFT). The formation of the poly(PS??-b-DMA???) diblock polymer was confirmed by ¹H NMR spectra and gel permeation chromatography (GPC). Poly(PS??-b-DMA???) formed 100 nm micelles in aqueous media as confirmed by dynamic light scattering (DLS) and transmission electron microscopy (TEM). Micelles loaded with the model drugs Nile red and DiO were used to demonstrate the ROS-dependent drug release mechanism of these micelles following treatment with hydrogen peroxide (H?O?), 3-morpholinosydnonimine (SIN-1), and peroxynitrite. These oxidants were found to oxidize the micelle PPS core, making it more hydrophilic and triggering micelle disassembly and cargo release. Delivery of poly(PS??-b-DMA???) micelles dual-loaded with the Förster Resonance Energy Transfer (FRET) fluorophore pair DiI and DiO was used to prove that endogenous oxidants generated by lipopolysaccharide (LPS)-treated RAW 264.7 macrophages significantly increased release of nanocarrier contents relative to macrophages that were not activated. In vitro studies also demonstrated that the poly(PS??-b-DMA???) micelles were cytocompatible across a broad range of concentrations. These combined data suggest that the poly(PS??-b-DMA???) micelles synthesized using a combination of TAGT and RAFT have significant potential for site-specific drug delivery to tissues with high levels of oxidative stress.
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Project description:AimTo determine if a regimen with prandial + basal insulin compared with basal insulin attenuates post-meal inflammatory and glycative biomarkers in patients with Type 2 diabetes.MethodsThis test-meal sub-study in the USA is from a previously reported clinical trial comparing the effect on glycaemic control of 24 weeks of thrice-daily pre-meal insulin lispro mix 50 (50% insulin lispro, 50% insulin lispro protamine suspension) or bedtime insulin glargine, both plus metformin. In the sub-study, glucose, insulin, triglycerides, high-sensitivity C-reactive protein, tumour necrosis factor α, interleukin-6, methylglyoxal and 3-deoxyglucosone were measured during the post-meal period of a mixed-meal breakfast at the final visit. Prandial + basal (n = 25) and basal (n = 21) insulin were administered at the same times as during the previous 24 weeks.ResultsPost-meal, the prandial + basal insulin group had significantly higher insulin, lower glucose and triglycerides, as well as lower high-sensitivity C-reactive protein, tumour necrosis factor α and interleukin-6, than the basal insulin group. Glucose incremental area under the concentration curve significantly correlated with high-sensitivity C-reactive protein, tumour necrosis factor α, interleukin-6, methylglyoxal and 3-deoxyglucosone incremental area under the concentration curve. Insulin incremental area under the concentration curve correlated inversely with high-sensitivity C-reactive protein and tumour necrosis factor α incremental area under the concentration curve. However, after adjusting for glucose incremental area under the concentration curve, these inverse correlations were no longer significant. Triglyceride incremental area under the concentration curve was not correlated with any biomarker incremental area under the concentration curve.ConclusionsControlling post-meal hyperglycaemia with prandial + basal insulin in patients with Type 2 diabetes attenuates meal-induced increases in high-sensitivity C-reactive protein, interleukin-6 and tumour necrosis factor α compared with basal insulin. The rise in post-meal glucose, but not triglycerides, significantly correlated with the rise in post-meal inflammatory and glycative biomarkers.
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2010-05-18 | E-GEOD-16146 | biostudies-arrayexpress
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