interleukin-8 and nickel-sulfate

In vitro safety tests may be used as replacements for animal tests owing to their accuracy and high-throughput performance. However, several in vitro skin sensitization tests produce false-negative results such as acid anhydride. Here, we investigated the relationship between false-negative results of acid anhydride and its hydrolysis by aqueous vehicle. Differences in the pattern of hydrolysis for phthalic anhydride (PAH) due to addition of 1 drop of stock solution of PAH in liquid paraffin (LP) dispersion medium and PAH in DMSO were analyzed in a cell-free system. The results showed that use of LP dispersion medium stabilized the concentration of PAH in water over 5min by sustained-release, although almost all PAH converted to phthalic acid in water within 5min using DMSO. Additionally, treatment of THP-1 cells with PAH and phthalic acid using LP dispersion medium for 5min resulted in a 32-fold increase in IL-8 release for PAH as compared with that in the vehicle control. In contrast, for PAH using aqueous vehicle and phthalic acid using LP dispersion medium, there were no significant increases in IL-8 release. Similarly, using LP dispersion medium, trimellitic anhydride significantly increased IL-8 release was observed.

Topics: Allergens; Anhydrides; Cell Line, Tumor; Cell Survival; Humans; Hydrolysis; Interleukin-8; Mineral Oil; Nickel; Phthalic Acids; Skin Tests; Sodium Dodecyl Sulfate

Look for an in vitro test method to evaluate sensitization using THP-1 cells by the changes of the expression of cytokines to provide more reliable markers of the identification of sensitization.. The monocyte-like THP-1 cells were induced and differentiated into THP-1-macrophages with PMA (0.1 microg/ml). The changes of expression of cytokines at different time points after the cells being treated with five known allergens, 2,4-dinitrochlorobenzene (DNCB), nickel sulfate (NiSO4), phenylene diamine (PPDA) potassium dichromate (K2Cr2O7) and toluene diisocyanate (TDI) and two non-allergens sodium dodecyl sulfate (SDS) and isopropanol (IPA) at various concentrations were evaluated. The IL-6 and TNF-alpha production was measured by ELISA. The secretion of IL-1beta and IL-8 was analyzed by Cytometric Bead Array (CBA).. The section of the IL-6, TNF-alpha, IL-1beta and IL-8 were the highest when THP-1 cells were exposed to NiSO4, DNCB and K2Cr2O7 for 6h, PPDA and TDI for 12h. The production of IL-6 were approximately 40, 25, 20, 50 and 50 times for five kinds chemical allergens NiSO4, DNCB, K2Cr2O7, PPDA and TDI respectively at the optimum time points and the optimal concentration compared to the control group. The expression of TNF-alpha were 20, 12, 20, 8 and 5 times more than the control group respectively. IL-1beta secretion were 30, 60, 25, 30 and 45 times respectively compared to the control group. The production of IL-8 were approximately 15, 12, 15, 12 and 7 times respectively compared to the control group. Both non-allergens SDS and IPA significantly induced IL-6 secretion in a dose-dependent manner however SDS cause a higher production levels, approximately 20 times of the control. Therefore IL-6 may not be a reliable marker for identification of allergens. TNF-alpha, IL-1beta and IL-8 expressions did not change significantly after exposed to the two non-allergens.. The test method using THP-1 cells by detecting the productions of cytokines (TNF-alpha, IL-1beta and IL-8) can effectively distinguish chemical allergens and non-allergens. The three cytokines may be reliable markers for the identification of potential sensitizing chemicals.

Topics: Allergens; Cytokines; Dinitrochlorobenzene; Humans; Interleukin-1beta; Interleukin-6; Interleukin-8; Monocytes; Nickel; Sodium Dodecyl Sulfate; Tumor Necrosis Factor-alpha

Numerous epidemiological studies have linked exposure to particulate matter (PM) air pollution with acute respiratory infection and chronic respiratory and cardiovascular diseases. We have previously shown that soluble nickel (Ni), a common component of PM, alters the release of CXC chemokines from cultured human lung fibroblasts (HLF) in response to microbial stimuli via a pathway dependent on disrupted prostaglandin (PG)E2 signaling. The current study sought to identify the molecular events underlying Ni-induced alterations in PGE2 signaling and its effects on IL-8 production. PGE2 synergistically enhances Ni-induced IL-8 release from HLF in a concentration-dependent manner. The effects of PGE2 were mimicked by butaprost and PGE1-alcohol and inhibited with antagonists AH6809 and L-161,982, indicating PGE2 signals via PGE2 receptors 2 and 4. PGE2 and forskolin stimulated cAMP, but it was only in the presence of Ni-induced hypoxia-inducible factor 1, α subunit (HIF1A) that these agents stimulated IL-8 release. The Ni-induced HIF1A DNA binding was enhanced by PGE2 and mediated, in part, by activation of p38 MAPK. Negation of cAMP-response element binding protein 1 or HIF1A using short interfering RNA blocked the synergistic interactions between Ni and PGE2. The results of the current study provide novel information on the ability of atmospheric hypoxia-mimetic metals to disrupt the release of immune-modulating chemokines by HLF in response to PGE2. Moreover, in the presence of HIF1A, cAMP-mediated signaling pathways may be altered to exacerbate inflammatory-like processes in lung tissue, imparting a susceptibility of PM-exposed populations to adverse respiratory health effects.

Topics: Alprostadil; Biomimetics; Cells, Cultured; Cyclic AMP; Cyclic AMP Response Element-Binding Protein; Dinoprostone; Drug Synergism; Fibroblasts; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Inflammation; Interleukin-8; Lung; Nickel; p38 Mitogen-Activated Protein Kinases; Receptors, Prostaglandin E; Receptors, Prostaglandin E, EP2 Subtype; Receptors, Prostaglandin E, EP4 Subtype; Signal Transduction; Xanthones

Contact allergens induce in vitro and in vivo the activation of dendritic cells (DC) and Langerhans cells (LC), which includes the up-regulation of surface marker expression (e.g. CD86, CD54) and cytokine production (e.g. TNF-alpha, IL-1beta, IL-8). The mitogen-activated protein kinase (MAPK) pathway also has a crucial role in this activation. However, the extent of MAPK involvement in the IL-8 production during DC/LC activation is not well understood. Earlier, we reported that contact allergens activated THP-1 cells, human monocytic cell line, like LC/DC in vitro. In this study, we further characterize the mechanism of IL-8 production using THP-1 cells as surrogate DCs. First, we evaluated the potential of 23 chemicals with different skin sensitization potencies to predominantly induce IL-8 production in vitro. Next we investigated the role of MAPK signaling and TNF-alpha, which is known to have autocrine effects on DC activation (e.g., IL-8 production). Inhibition of extracellular signal-regulated kinase (ERK), one of the MAPK pathways, suppressed the IL-8 production induced by both 2,4-dinitrochlorobenzene (DNCB) and nickel sulfate (NiSO(4)), and inhibition of p38 MAPK, a second MAPK pathway, significantly suppressed IL-8 production induced by only DNCB. Additionally, neutralization of TNF-alpha activity suppressed IL-8 production in THP-1 cells exposed to DNCB and NiSO(4). In conclusion, IL-8 production was predominantly induced in THP-1 cells following allergen stimulation, and MAPK pathways and TNF-alpha were involved in the IL-8 production induced by DNCB and NiSO(4). A better understanding of the mechanism of DC activation in vitro might lead to the clarification of the in vivo skin sensitization mechanism.

Topics: Allergens; Cell Line; Dinitrochlorobenzene; Humans; Interleukin-8; Mitogen-Activated Protein Kinases; Nickel; Tumor Necrosis Factor-alpha

Particulate matter air pollution (PM) has been linked with chronic respiratory diseases. Real-life exposures are likely to involve a mixture of chemical and microbial stimuli, yet little attention has been paid to the potential interactions between PM components (e.g., Ni) and microbial agents on the development of inflammatory-like conditions in the lung. Using the Toll-like receptor (TLR)-2 agonist MALP-2 as a lipopeptide relevant to microbial colonization, we hypothesized that nickel sensitizes human lung fibroblasts (HLF) for microbial-driven chemokine release through modulation of TLR signaling pathways. NiSO(4) (200 muM) synergistically enhanced CXCL8, yet antagonized CXCL10 mRNA expression and protein release from HLF in response to MALP-2. RT(2)-PCR pathway-focused array results indicated that NiSO(4) exposure did not alter the expression of TLRs or their downstream signaling mediators, yet significantly increased the expression of cyclooxygenase 2 (COX-2). Moreover, when NiSO(4) was given in combination with MALP-2, there was an amplified induction of COX-2 mRNA and protein along with its metabolic product, PGE2, in HLF. The COX-2 inhibitor, NS-398, attenuated NiSO(4) and MALP-2-induced PGE2 and CXCL8 release and partially reversed the NiSO(4)-dependent inhibition of MALP-2-induced CXCL10 release from HLF. These data indicate that NiSO(4) alters the pattern of TLR-2-dependent chemokine release from HLF via a COX-2-mediated pathway. The quantitative and qualitative effects of NiSO(4) on microbial-driven chemokine release from HLF shed new light on how PM-derived metals can exacerbate respiratory diseases.

Topics: Cells, Cultured; Chemokine CXCL10; Chemokines; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Fibroblasts; Gene Expression Profiling; Humans; Interleukin-8; Lipopeptides; Lung; Nickel; Oligopeptides; Toll-Like Receptor 2

The detection of the sensitizing potential of chemicals is of great importance to industry. A promising in vitro alternative to the currently applied animal assays for sensitization testing makes use of dendritic cells (DCs) that have the capability to process and present antigens to naive T cells and induce their proliferation. Here, we studied changes in gene expression profiles after exposing DCs to the contact allergen nickel sulfate. CD34+-progenitor-derived DCs, initiated from 3 different donors, were exposed to 60 microM nickel sulfate, during 0.5, 1, 3, 6, 12 and 24 h. cDNA microarrays were used to assess the transcriptional activity of about 11,000 genes. Significant changes in the expression of 283 genes were observed; 178 genes were up-regulated and 93 down-regulated. These genes were involved in metabolism, cell structure, immune response, transcription, signal transduction, transport, and apoptosis. No functional information was found for 74 genes. Real-time RT-PCR was used to confirm the microarray results of 12 genes. In addition, 3 DC maturation markers not present on the microarrays (DEC205, DC LAMP and CCR7) were analyzed using real-time RT-PCR and found to be up-regulated at several time points. Our data indicate that a broad range of biological processes is influenced by nickel. Some processes are clearly linked to the immune response and DC maturation, others may indicate a toxic effect of nickel.

Topics: Allergens; Antigens, CD34; Apoptosis Regulatory Proteins; Dendritic Cells; Dose-Response Relationship, Drug; Down-Regulation; Female; Gene Expression Profiling; Gene Expression Regulation; Humans; Infant, Newborn; Interleukin-1beta; Interleukin-6; Interleukin-8; Nickel; Oligonucleotide Array Sequence Analysis; Pregnancy; Receptors, Interleukin-1 Type II; Reproducibility of Results; Reverse Transcriptase Polymerase Chain Reaction; Stem Cells; Time Factors; Up-Regulation

The immune system is called into action by alarm signals generated from injured tissues. We examined the nature of these alarm signals after exposure of skin residential cells to contact allergens (nickel sulfate and potassium dichromate) and a contact irritant [sodium dodecyl sulfate (SDS)]. Nickel sulfate, potassium dichromate, and SDS were applied topically to the stratum corneum of human skin equivalents. A similar concentration-dependent increase in chemokine (CCL20, CCL27, and CXCL8) secretion was observed for all three chemicals. Exposure to nickel sulfate and SDS was investigated in more detail: similar to chemokine secretion, no difference was observed in the time- and concentration-dependent increase in pro-inflammatory cytokine [interleukin-1alpha (IL-1alpha) and tumor necrosis factor-alpha (TNF-alpha)] secretion. Maximal increase in IL-1alpha secretion occurred within 2 h after exposure to both nickel sulfate and SDS and prior to increased chemokine secretion. TNF-alpha secretion was detectable 8 h after chemical exposure. After allergen or irritant exposure, increased CCL20 and CXCL8, but not CCL27, secretion was inhibited by neutralizing human antibodies to either IL-1alpha or TNF-alpha. Our data show that alarm signals consist of primary and secondary signals. IL-1alpha and TNF-alpha are released as primary alarm signals, which trigger the release of secondary chemokine (CCL20 and CXCL8) alarm signals. However, some chemokines, for example, CCL27 can be secreted in an IL-1alpha and TNF-alpha independent manner. Our data suggest that skin residential cells respond to both allergen and irritant exposure by releasing mediators that initiate infiltration of immune responsive cells into the skin.

Topics: Allergens; Caustics; Cells, Cultured; Chemokine CCL20; Chemokine CCL27; Chemokines, CC; Cytokines; Enzyme-Linked Immunosorbent Assay; Fibroblasts; Humans; Interleukin-1; Interleukin-8; Irritants; Keratinocytes; Macrophage Inflammatory Proteins; Nickel; Potassium Dichromate; Recombinant Proteins; Skin; Sodium Dodecyl Sulfate; Surface-Active Agents; Time Factors; Tumor Necrosis Factor-alpha

The identification of potential damage due to chemical exposure in the workplace is a major health and regulatory concern. Traditional tests that measure both sensitization and elicitation responses require the use of animals. An alternative to this widespread use of experimental animals could have a crucial impact on risk assessment, especially for the preliminary screening of new molecules. We developed an in vitro model for the screening of potential toxic compounds. Human keratinocytes (HaCat) were used as target cells while matrix metalloproteinases (MMP) were selected as responders because they are key enzymes involved in extracellular matrix (ECM) degradation in physiological and pathological conditions. Chemical exposure was performed using nickel sulphate as a positive tester. Nickel contact induced upregulation of MMP-2 and IL-8 mRNA production. Molecular activation occurred even at very low nickel concentrations even though no phenotypic changes were observed. MMP-9 accumulation was found in the medium of treated cells with respect to controls. These observations led to the hypothesis that even minimal exposure can accumulate transcriptional activity resulting in long-term clinical signs after contact. Our simple in vitro model can be applied as a useful preliminary complement to the animal studies to screen the effects of new potential toxic compounds.

Topics: Animal Testing Alternatives; Biomarkers; Cell Line; Cell Survival; Dermatitis, Contact; Dermatitis, Occupational; Enzyme-Linked Immunosorbent Assay; Humans; Interleukin-8; Keratinocytes; Matrix Metalloproteinases; Models, Biological; Nickel; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Toxicity Tests; Up-Regulation

Exposure to ambient air particulate matter (PM) is associated with increased mortality and morbidity in susceptible populations. The epidemiological data also suggest a relationship between PM air pollution and impairment of cardiopulmonary function. The mechanisms that may be responsible for these effects are not fully understood and are likely related to perturbations of cellular and molecular functions. One type of PM, residual oil fly ash (ROFA), is of particular interest. ROFA does not contain much organic material, but does contain relatively high quantities of transition metals, predominantly nickel, vanadium, and iron, as well as black carbon and sulfates. In this study, we investigated the effect of two metals (iron and nickel) on the induction of "hypoxia-like" stress and the production of interleukins (ILs) in minimally transformed human airway epithelial cells (1HAEo(-)). We found that exposure to soluble nickel sulfate results in the induction of hypoxia-inducible genes and IL-8 production by the 1HAEo(-) cells. The simultaneous addition of iron in either ferric or ferrous form and nickel completely inhibited IL-8 production and had no effect on "hypoxia-like" stress caused by nickel, suggesting the existence of two different pathways for the induction "hypoxia-like" stress and IL-8 production. The effect of nickel was not related to the blocking of iron entry into cells since the level of intracellular iron was not affected by co-exposure with nickel. The obtained data indicate that nickel can induce different signaling pathways with or without interference with iron metabolism. Our observations suggest that in some cases the excess of iron in PM can cancel the effects of nickel.

Topics: Air Pollutants; Blotting, Western; Carbon; Cell Cycle Proteins; Chlorides; Coal Ash; Deferoxamine; Epithelial Cells; Ferric Compounds; Humans; Hypoxia; Interleukin-8; Intracellular Signaling Peptides and Proteins; Iron Chelating Agents; Lung; Nickel; Particulate Matter

The potential of organic dust to induce inflammation in vitro can be viewed as a crude measure of the total biologically active compounds in a dust sample. The purpose of this study was to further develop an in vitro screening method for evaluation of potential hazard related to low doses of dust exposure using two monocytic cell lines (U937 and THP-1). Dust was obtained from schools in Copenhagen. U937 and THP-1 cells were stimulated with dust for 24 h and interleukin-8 secretion was measured. The initial slopes of the dose-response curves were used to calculate the inflammatory potential, or potency factor (PF), of the samples. In characterization of the method, lipopolysaccharide from Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Salmonella enteritidis were tested together with three glucans, nickel sulfate (NiSO(4)), methyl methacrylate (MMA), formaldehyde, and four surfactants. The PF values of LPSs in both monocytic assays ranked as follows: S. enteritidis> E. coli>K. pneumoniae/P. aeruginosa. The PF values of NiSO(4), MMA, formaldehyde, and the surfactants were zero or below. Using the THP-1 cell line, the PF values of dust samples were 30 times higher than when using the U937 cell line, and 7 times higher than when using the lung epithelial cell line (A549). The high sensitivity of the THP-1 bioassay makes it potentially useful as a screening tool for hazard evaluation of dust from, e.g., the indoor environment.

Topics: Biological Assay; Cell Line; Dose-Response Relationship, Drug; Dust; Formaldehyde; Glucans; Humans; Inflammation; Interleukin-8; Lipopolysaccharides; Methylmethacrylate; Monocytes; Nickel; Reproducibility of Results; Respiratory Hypersensitivity; Sensitivity and Specificity; Stimulation, Chemical; Surface-Active Agents; U937 Cells