Effects of Endocrine Disruptor Compounds, Alone or in Combination, on Human Macrophage-Like THP-1 Cell Response.

The aim of the present study was to evaluate the immunological effects on human macrophages of four endocrine disruptor compounds (EDCs) using the differentiated human THP-1 cell line as a model. We studied first the effects of these EDCs, including Bisphenol A (BPA), di-ethylhexyl-phthalate (DEHP), dibutyl phthalate (DBP) and 4-tert-octylphenol (4-OP), either alone or in combination, on cytokine secretion, and phagocytosis. We then determined whether or not these effects were mediated by estrogen receptors via MAPK pathways. It was found that all four EDCs studied reduced strongly the phagocytosis of the differentiated THP-1 cells and that several of these EDCs disturbed also TNF-α, IL-1 ß and IL-8 cytokine secretions. Furthermore, relative to control treatment, decreased ERK 1/2 phosphorylation was always associated with EDCs treatments-either alone or in certain combinations (at 0.1 µM for each condition). Lastly, as treatments by an estrogen receptor antagonist suppressed the negative effects on ERK 1/2 phosphorylation observed in cells treated either alone with BPA, DEHP, 4-OP or with the combined treatment of BPA and DEHP, we suggested that estrogen receptor-dependent pathway is involved in mediating the effects of EDCs on human immune system. Altogether, these results advocate that EDCs can disturb human immune response at very low concentrations.

Regulation of LRP-1 expression: make the point.

The low-density lipoprotein receptor-related protein-1 (LRP-1) is a membrane receptor displaying both scavenging and signaling functions. The wide variety of extracellular ligands and of cytoplasmic scaffolding and signaling proteins interacting with LRP-1 gives it a major role not only in physiological processes, such as embryogenesis and development, but also in critical pathological situations, including cancer and neurological disorders. In this review, we describe the molecular mechanisms involved at distinct levels in the regulation of LRP-1, from its expression to the proper location and stability at the cell surface.

Effects of 4-nonylphenol and/or diisononylphthalate on THP-1 cells: impact of endocrine disruptors on human immune system parameters.

The aim of the present work is to investigate the link between two endocrine disruptor compounds (EDCs), which are chemicals that interfere with the hormone system in human and wildlife, and the human immune response through a study of their effects on the THP-1 human cell line which was used as a model for macrophages. We used two EDCs, diisononylphthalate (DiP) and 4-n-nonylphenol (NP) alone or in combination in order to evaluate the effects of these compounds on several parameters of the immune response - cytokine secretion, phagocytosis and the putative implication of the estrogen receptors - by studying the level of MAPK activation. NP and DiP strongly reduced phagocytosis and modify cytokine secretions. Indeed, differentiated THP-1 cell exposures (i) to 5 and 10 microM of combination of NP and DiP induced an IL-8 level in the medium respectively of 28.9 and 45 percent higher than the level obtained for the control (untreated cells), (ii) to combination of NP and DiP at 10 microM induced an increase of IL-1ß level in comparison to the control level, (iii) to combination of NP and DiP induced an increase of TNF-α level whatever the concentration of EDCs tested (between 0 and 10 microM). Lastly, differentiated THP-1 cell exposure to NP, DiP alone or in combination (2 microM for each condition) induced a decrease of ERK1/2 phosphorylation in comparison to ERK1/2 phosphorylation level of the control. Moreover, differentiated THP-1 cell treatments by ICI-182780 (an estrogen receptor antagonist) supressed the EDC effects on ERK1/2 phosphorylation level which indicates an estrogen receptor-dependent pathway. These results suggest that EDCs have the ability to alter the human immune function, maybe by interfering with endocrine balance.

Oxidative stress induction by nanoparticles in THP-1 cells with 4-HNE production: stress biomarker or oxidative stress signalling molecule?

The aim of this study was to investigate whether carbon black (CB) nanoparticles might induce toxicity to monocytic cells in vitro via an oxidative stress mechanism involving formation of the lipid peroxidation product 4-hydroxynonenal (4-HNE) and the subsequent role of 4-HNE in inducing further cytotoxic effects. ROS production in cells by CB nanoparticles was shown by the oxidation of DCFH after a short time exposure. These particles induced the formation of 4-HNE-protein adducts and significant modification of glutathione content corresponding to an increase of oxidized glutathione form (GSSG) and a decrease of total glutathione (GSX) content. These results attest to an oxidative stress induced by the carbon black nanoparticles, although no induction of HO-1 protein expression was detected. Concerning the effects of a direct exposure to 4-HNE, our results showed that 4-HNE is not cytotoxic for concentrations lower than 12.5 microM. By contrast, it provokes a very high cytotoxicity for concentrations above 25 microM. An induction of HO-1 expression was observed from concentrations above 5 microM of 4-HNE. Finally, glutathione content decreased significantly from 5 microM of 4-HNE but no modification was observed under this concentration. The discrepancy between effects of carbon black nanoparticles and 4-HNE on the intracellular markers of oxidative stress suggests that 4-HNE is not directly implied in the signalling of oxidative toxicity of nanoparticles but is an effective biomarker of oxidative effects of nanoparticles.

Effect of polycyclic aromatic hydrocarbons and carbon black particles on pro-inflammatory cytokine secretion: impact of PAH coating onto particles.

It has been suggested that the organic fraction of particulate matter in air pollution has a major role in the toxicity of this pollutant, notably via its effects on inflammation. The major organic compounds adsorbed onto these particles are polycyclic aromatic hydrocarbons (PAH), among which benzo[a]pyrene (B[a]P), benzo[b]fluoranthene (B[b]F), and pyrene (Pyr) are quantitatively the most important. Generally, cells or organisms are exposed to organic extracts of the particles rather than the native particles in order to study the effects of these PAH. In this study, B[a]P, B[b]F, and Pyr were tested alone and/or adsorbed onto carbon black (CB) particles differing in size in order to evaluate their impact on cytokine production (with or without LPS stimulation) by THP-1 macrophage-like cells. PAH induced significant secretion of IL-1beta, IL-8, and IL-12 after 24 or 48 hr of treatment, an effect reinforced by LPS stimulation; no effect on IL-10 secretion was noted. Fine CB particles (260 nm diameter) induced secretion of each cytokine. In general, coating the CB with PAH did not modify the effect of the CB alone; the exception was that LPS-induced IL-1beta secretion was reduced. In contrast, ultrafine CB (14 nm diameter: ufCB) caused a decrease in cytokine secretion; this effect was modified by PAH coating. For example, PAH coating on ufCB amplified the inhibitory effect of ufCB against IL-1beta secretion but did not modify IL-8 formation. Moreover, PAH coating on ufCB tended to minimize the effect of LPS stimulation; this included (i) inhibition of the decrease in IL-12 secretion induced by uncoated ufCB and (ii) stimulation of IL-10 production. It was concluded that adsorption of PAH onto these particles could decrease their bioavailability and so their abilities to affect cell cytokine production. The results also showed that when PAH were adsorbed onto the fine particles, any observed increases in cytokine secretion consistently appeared to be due to the particles themselves. In contrast, while ufCB alone almost uniformly led to decreases in cytokine formation by the cells, the added presence of the test PAHs led to variable effects - depending on whether stimulation with LPS took place or not. Thus, while some PAHs likely to be associated with PM are clearly immunomodulants, their ultimate effects in situ will likely depend on the properties of the particles themselves, in particular, their size.

Oxidative stress induction by short time exposure to ozone on THP-1 cells.

Ozone is a major component of air pollution mainly formed by photochemical reactions of nitrogen oxides with volatile organic compounds and/or carbon monoxide. Numerous studies have shown the association between ozone exposure with pulmonary injuries. This pollutant is a strong oxidant exerting its biological action either by direct reaction with target molecules or by generating reactive oxygen species which result in its biological effects and its toxicity. In order to study the effects of an induced oxidative stress by ozone on THP-1 cell, a human macrophage-like cell line, we used an in vitro system which has been previously used to study the rapid responses to ozone exposure. Using this system, THP-1 cells were subjected to short time exposure (30 min) followed by different incubation times ranging from 4 to 24 h. Our results show that ozone exposure provokes an alteration of the cell membrane translating an induction of lipid peroxidation resulting in a 3.2-fold increase of thiobarbituric reactive substances (TBARS), an increase by 35% of heme oxygenase-1 (HO-1) expression, and significant modifications of the redox status evaluated by glutathione measurement and of antioxidant enzyme activities in THP-1 cells. Our in vitro model constitutes a very interesting tool for the measurement of ozone effect on rapid modifications induced by this pollutant as well as intracellular modifications due to an oxidative stress.

Inhibition by transmembrane peptides of chimeric insulin receptors.

Receptor tyrosine kinases play essential roles in cell proliferation and differentiation. We have recently shown that peptides corresponding to the transmembrane domains of the epidermal growth factor (EGF) and ErbB2 receptors inhibit their corresponding receptor activation in cancer cell lines. We extend this observation to cells transfected with chimeric insulin receptors where the transmembrane domain has been replaced by that of the EGF receptor or a mutated Erb2 domain. Peptides corresponding to the transmembrane domains of the EGF receptor and ErbB2 are able to inhibit specifically the autophosphorylation of insulin receptors with the corresponding domain. This inhibitory effect is correlated with the propensity of the different transmembrane domains to self-associate in a genetic reporter assay. Thus, our data strengthen the notion that transmembrane domains are involved in erbB receptor activation, and that these receptors can be modulated by inhibiting protein-protein interactions within the membrane.

Xaliproden (SR57746A) induces 5-HT1A receptor-mediated MAP kinase activation in PC12 cells.

Neurotrophic growth factors are involved in cell survival. However, natural growth factors have a very limited therapeutic use because of their short half-life. In the present study, we investigated the mechanism of action of a non-peptidic neurotrophic drug, Xaliproden, a potential molecule for the treatment of motoneuron diseases, since the transduction pathways of this synthetic 5-HT1A agonist are very poorly understood. Xaliproden does not activate the Trk receptor but causes a rapid increase in the activities of the ERK1 and ERK2 isoforms of MAP kinase, which then rapidly decrease to the basal level. We demonstrate that isoforms of the SHC adapter protein are phosphorylated independently of each other and are probably not the source of the Xaliproden-induced MAP kinases activation. The inhibitor of Ras farnesylation, FPT-1, and the protein kinase C inhibitors, GF 109203X and chelerythrine, inhibited the Xaliproden-induced MAP kinase activation, suggesting p21Ras and PKC involvement. Moreover, the observations that the 5-HT1A antagonist, pindobind, and pertussis toxin abolished the Xaliproden-induced ERK stimulation suggested that Xaliproden activates the MAP kinase pathways by stimulating the G protein-coupled receptor, 5-HT1A. These results demonstrate clearly that the non-peptidic compound, Xaliproden, exerts its neurotrophic effects through a mechanism of action differing from that of neurotrophins. These findings suggest that this compound does not involve MAPK activation by TrkA receptor stimulation but acts by MAP kinase pathway by a pertussis toxin-sensitive mechanism involving 5-HT1A receptors, p21 Ras and MEK-1 and by PKC and Akt pathways.