Aryl Hydrocarbon Receptor Involvement in the Sodium-Dependent Glutamate/Aspartate Transporter Regulation in Cerebellar Bergmann Glia Cells.

Glutamate, the major excitatory neurotransmitter in the vertebrate brain, exerts its functions through the activation of specific plasma membrane receptors and transporters. Overstimulation of glutamate receptors results in neuronal cell death through a process known as excitotoxicity. A family of sodium-dependent glutamate plasma membrane transporters is responsible for the removal of glutamate from the synaptic cleft, preventing an excitotoxic insult. Glial glutamate transporters carry out more than 90% of the brain glutamate uptake activity and are responsible for glutamate recycling through the GABA/Glutamate/Glutamine shuttle. The aryl hydrocarbon receptor is a ligand-dependent transcription factor that integrates environmental clues through its ability to heterodimerize with different transcription factors. Taking into consideration the fundamental role of glial glutamate transporters in glutamatergic synapses and that these transporters are regulated at the transcriptional, translational, and localization levels in an activity-dependent fashion, in this contribution, we explored the involvement of the aryl hydrocarbon receptor, as a model of environmental integrator, in the regulation of the glial sodium-dependent glutamate/aspartate transporter. Using the model of chick cerebellar Bergmann glia cells, we report herein that the aryl hydrocarbon receptors exert a time-dependent decrease in the transporter mRNA levels and a diminution of its uptake activity. The nuclear factor kappa light chain enhancer of the activated B cell signaling pathway is involved in this regulation. Our results favor the notion of an environmentally dependent regulation of glutamate removal in glial cells and therefore strengthen the notion of the involvement of glial cells in xenobiotic neurotoxic effects.

Kynurenine attenuates mitochondrial depolarization and neuronal cell death induced by rotenone exposure independently of AhR-mediated parkin induction in SH-SY5Y differentiated cells.

Rotenone is a pesticide commonly used in agriculture that is associated with the risk of developing Parkinson's disease (PD) by inducing mitochondrial damage. As a protective cell response to different challenges, they activate mitophagy, which involves parkin activity. Parkin is an E3 ubiquitin ligase necessary in the initial steps of mitophagy, and its overexpression protects against parkinsonian effects in different models. Recent studies have reported that the aryl hydrocarbon receptor (AHR), a ligand-dependent transcription factor, induces parkin expression. Kynurenine, an endogenous AHR ligand, promotes neuroprotection in chronic neurodegenerative disorders, such as PD, although its neuroprotective mechanism needs to be fully understood. Therefore, we evaluated whether the overexpression of parkin by AHR activation with kynurenine promotes autophagy and reduces the neurotoxicity induced by rotenone in SH-SY5Y cells differentiated to dopaminergic neurons. SH-SY5Y neurons were treated with rotenone or pretreated with kynurenine or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and parkin levels, apoptosis, mitochondrial potential membrane, and autophagy were determined. The results showed that kynurenine and TCDD treatments induced parkin expression in an AHR-dependent manner. Kynurenine pretreatment inhibited rotenone-induced neuronal apoptosis in 17%, and the loss of mitochondrial membrane potential in 30% when compare to rotenone alone, together with a decrease in autophagy. By contrast, although TCDD treatment increased parkin levels, non-neuroprotective effects were observed. The kynurenine protective activity was AHR independent, suggesting that parkin induction might not be related to this effect. On the other hand, kynurenine treatment inhibited alpha amine-3-hydroxy-5-methyl-4-isoxazol propionic acid and N-methyl-D-aspartate receptors, which are well-known excitotoxicity mediators activated by rotenone exposure.

Synergistic effect of antagonists to KRas4B/PDE6 molecular complex in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) has the worst prognosis among all human cancers as it is highly resistant to chemotherapy. K-Ras mutations usually trigger the development and progression of PDAC. We hypothesized that compounds stabilizing the KRas4B/PDE6δ complex could serve as PDAC treatments. Using in silico approaches, we identified the small molecules C14 and P8 that reduced K-Ras activation in primary PDAC cells. Importantly, C14 and P8 significantly prevented tumor growth in patient-derived xenotransplants. Combined treatment with C14 and P8 strongly increased cytotoxicity in PDAC cell lines and primary cultures and showed strong synergistic antineoplastic effects in preclinical murine PDAC models that were superior to conventional therapeutics without causing side effects. Mechanistically, C14 and P8 reduced tumor growth by inhibiting AKT and ERK signaling downstream of K-RAS leading to apoptosis, specifically in PDAC cells. Thus, combined treatment with C14 and P8 may be a superior pharmaceutical strategy to improve the outcome of PDAC.

Methylated and ethylated dialkylphosphate metabolites of organophosphate pesticides: DNA damage in bone marrow cells of Balb/c mice.

Dialkylphosphates (DAPs), metabolites of organophosphate (OP) pesticides, are widely distributed in the environment and are often used as biomarkers of OP exposure. Recent reports indicate that DAPs may be genotoxic, both in vitro and in vivo. We have examined the genotoxicity of the methylated DAPs dimethyldithiophosphate (DMDTP) and dimethylphosphate (DMTP) and the ethylated DAPs diethyldithiophosphate (DEDTP) and diethylphosphate (DETP), in comparison with their parental compounds, malathion and terbufos, respectively, in bone marrow polychromatic erythrocytes (PCE) of male and female Balb/c mice. We also compared DNA damage (comet assay) induced by DMDTP and dimethyl phosphate (DMP) in human cell lines. Both DMDTP and DMP caused DNA damage in peripheral blood mononuclear cells, HeLa cells, and the hepatic cell lines HepG2 and WRL-68. In the in vivo micronucleus assay, methylated and ethylated DAPs increased micronucleated PCE cells in both male and female mice. Female mice were more susceptible to DNA damage. In comparison to their parental compounds, methylated DAPs, particularly DMTP, were more genotoxic than malathion; DEDTP, DETP, and terbufos were similar in potency. These results suggest that DAPs may contribute to DNA damage associated with OP pesticide exposure.

Methylated dialkylphosphate metabolites of the organophosphate pesticide malathion modify actin cytoskeleton arrangement and cell migration via activation of Rho GTPases Rac1 and Cdc42.

The non-cholinergic molecular targets of organophosphate (OP) compounds have recently been investigated to explain their role in the generation of non-neurological diseases, such as immunotoxicity and cancer. Here, we evaluated the effects of malathion and its dialkylphosphate (DAP) metabolites on the cytoskeleton components and organization of RAW264.7 murine macrophages as non-cholinergic targets of OP and DAPs toxicity. All OP compounds affected actin and tubulin polymerization. Malathion, dimethyldithiophosphate (DMDTP) dimethylthiophosphate (DMTP), and dimethylphosphate (DMP) induced elongated morphologies and the formation of pseudopods rich in microtubule structures, and increased filopodia formation and general actin disorganization in RAW264.7 cells and slightly reduced stress fibers in the human fibroblasts GM03440, without significantly disrupting the tubulin or vimentin cytoskeleton. Exposure to DMTP and DMP increased cell migration in the wound healing assay but did not affect phagocytosis, indicating a very specific modification in the organization of the cytoskeleton. The induction of actin cytoskeleton rearrangement and cell migration suggested the activation of cytoskeletal regulators such as small GTPases. We found that DMP slightly reduced Ras homolog family member A activity but increased the activities of Ras-related C3 botulinum toxin substrate 1 (Rac1) and cell division control protein 42 (Cdc42) from 5 min to 2 h of exposure. Chemical inhibition of Rac1 with NSC23766 reduced cell polarization and treatment with DMP enhanced cell migration, but Cdc42 inhibition by ML-141 completely inhibited the effects of DMP. These results suggest that methylated OP compounds, especially DMP, can modify macrophage cytoskeleton function and configuration via activation of Cdc42, which may represent a potential non-cholinergic molecular target for OP compounds.

Inhibition of acetylation, is it enough to fight cancer?

Acetylation is a reversible post-translational modification (PTM) that regulates important cellular processes such as proliferation, DNA damage repair and cell cycle progress. When the balance is broken, these processes are affected and lead to carcinogenesis. Therefore, the study of acetylation has led to its proposal as a target pathway for anticancer therapies. Here, we discuss how acetylation regulates the cell cycle process, how it is modified in cancer cells and which are the key proteins in the regulation of apoptosis induction in cancer cells that can become targets to fight cancer. The inhibition of acetylation has been proposed as an emergent therapy against cancer, compounds such as 6-Penthadecyl salicylic acid, Curcumin, Garcinol and C646, among others, are currently studied because they show antitumor activity related to the inhibition of acetylation. Recently, the use of the acetylomics research tool has improved the study of acetylation as a target against tumor cells, but still the thresholds between promoting DNA instability and regulating gene expression by acetylation are not clear in many cell types.

Characterisation of Macrophage Polarisation in Mice Infected with Ninoa Strain of Trypanosoma cruzi.

Macrophages (MΦ) play a key role in the development of the protective immune response against Trypanosoma cruzi infection. To determine the role of MΦ subtypes M1 and M2 in the development of immunity against the Mexican strain of T. cruzi (Ninoa strain), we have analysed in a time course the infection and characterised the M1 and M2 subtypes in two mouse models, BALB/c and C57BL/6. After infection, BALB/c mice developed an increased blood parasite load and the parasites were cleared from the blood one week later than in C57BL/6 mice. However, similar cellular infiltrate and cardiac alterations were observed between BALB/c and C57BL/6 mice. At 36 days, the T. cruzi infection differentially modulated the expression of immune cells, and both the BALB/c and C57BL/6 mice significantly reduced TCD4+ cells. However, BALB/c mice produced significantly more TCD8+ than C57BL/6 mice in the spleen and lymph nodes. Furthermore, BALB/c mice produce significantly more MΦ in the spleen, while C57BL/6 produce similar levels to uninfected mice. The M1 MΦ ratio increased significantly at 3-5 days post-infection (dpi), but then decreased slightly. On the contrary, the M2 MΦ were low at the beginning of the infection, but the proportion of M1 and M2 MΦ at 36 dpi was similar. Importantly, the MΦ subtypes M2c and M2d significantly increased the induction of tissue repair by the end of the acute phase of the infection. These results indicate that the Ninoa strain has developed strategies to modulate the immune response, with fine differences depending on the genetic background of the host.

The cytoskeleton as a non-cholinergic target of organophosphate compounds.

Current organophosphate (OP) toxicity research now considers potential non-cholinergic mechanisms for these compounds, since the inhibition of acetylcholinesterase (AChE) cannot completely explain all the adverse biological effects of OP. Thanks to the development of new strategies for OP detection, some potential molecular targets have been identified. Among these molecules are several cytoskeletal proteins, including actin, tubulin, intermediate filament proteins, and associated proteins, such as motor proteins, microtubule-associated proteins (MAPs), and cofilin. in vitro, ex vivo, and some in vivo reports have identified alterations in the cytoskeleton following OP exposure, including cell morphology defects, cells detachments, intracellular transport disruption, aberrant mitotic spindle formation, modification of cell motility, and reduced phagocytic capability, which implicate the cytoskeleton in OP toxicity. Here, we reviewed the evidence indicating the cytoskeletal targets of OP compounds, including their strategies, the potential effects of their alterations, and their possible participation in neurotoxicity, embryonic development, cell division, and immunotoxicity related to OP compounds exposure.

Anacardic Acids from Amphipterygium adstringens Confer Cytoprotection against 5-Fluorouracil and Carboplatin Induced Blood Cell Toxicity While Increasing Antitumoral Activity and Survival in an Animal Model of Breast Cancer.

Amphipterygium adstringens (cuachalalate) contains anacardic acids (AAs) such as 6-pentadecyl salicylic acid (6SA) that show immunomodulatory and antitumor activity with minimal or no secondary adverse effects. By contrast, most chemotherapeutic agents, such as 5-fluorouracil (5-FU) and carboplatin (CbPt), induce myelosuppression and leukopenia. Here, we investigated the myeloprotective and antineoplastic potential of an AA extract or the 6SA as monotherapy or in combination with commonly used chemotherapeutic agents (5-FU and CbPt) to determine the cytoprotective action of 6SA on immune cells. Treatment of Balb/c breast tumor-bearing female mice with an AA mixture or 6SA did not induce the myelosuppression or leukopenia observed with 5-FU and CbPt. The co-administration of AA mixture or isolated 6SA with 5-FU or CbPt reduced the apoptosis of circulating blood cells and bone marrow cells. Treatment of 4T1 breast tumor-bearing mice with the AA mixture or 6SA reduced tumor growth and lung metastasis and increased the survival rate compared with monotherapies. An increased effect was observed in tumor reduction with the combination of 6SA and CbPt. In conclusion, AAs have important myeloprotective and antineoplastic effects, and they can improve the efficiency of chemotherapeutics, thereby protecting the organism against the toxic effects of drugs such as 5-FU and CbPt.

Regulation of Parkin expression as the key balance between neural survival and cancer cell death.

Parkin is a cytosolic E3 ubiquitin ligase that plays an important role in neuroprotection by targeting several proteins to be degraded by the 26S proteasome. Its dysfunction has been associated not only with Parkinson's disease (PD) but also with other neurodegenerative pathologies, such as Alzheimer's disease and Huntington's disease. More recently, Parkin has been identified as a tumor suppressor gene implicated in cancer development. Due to the important roles that this E3 ubiquitin ligase plays in cellular homeostasis, its expression, activity, and turnover are tightly regulated. Several reviews have addressed Parkin regulation; however, genetic and epigenetic regulation have been excluded. In addition to posttranslational modifications (PTMs), this review examines the regulatory mechanisms that control Parkin function through gene expression, epigenetic regulation, and degradation. Furthermore, the consequences of disrupting these regulatory processes on human health are discussed.

Association Between Socio-Affective Symptoms and Glutathione and CD4 and CD8 Lymphocytes in College Students.

Background: The prevalence of anxiety and depression in young students is associated with biosocial factors and scholastic stress. However, few studies have evaluated emotional-affective symptoms that are related to the immune system and antioxidant parameters in young individuals without diagnoses of affective disorders. Aim: This study aims to assess the relationship between emotional-affective symptoms and glutathione concentrations and CD4 and CD8 lymphocyte counts in college students. Methods: College students (n = 177) completed standardized psychometric instruments, including the Perceived Stress Scale, Hamilton Anxiety Scale, Beck Depression Inventory, Familiar Social and Friends Support Scale, and Rosenberg Scale. Blood samples were biochemically analyzed. Analyses of variance were conducted between four groups according to symptom severity. Results: A considerable prevalence of stress, anxiety, and depression symptoms was observed and negatively correlated with self-esteem and socio-familiar support. Perceived stress was sexually dimorphic. Although biochemical parameters were within reference ranges, glutathione, CD4, and CD8 tended to be lower in participants with anxiety and depression symptoms, which may be of predictive value. Conclusion: The relationship between antioxidant/immune parameters and socio-affective scores is latent in undiagnosed college students who might develop affective disorders. The findings suggest that during the initial development of affective disorders, stress management strategies should be implemented to help college students cope with the academic load and monitor negative changes in their physiological state.

Anacardic 6-pentadecyl salicylic acid induces apoptosis in breast cancer tumor cells, immunostimulation in the host and decreases blood toxic effects of taxol in an animal model.

Many antineoplastic agents induce myelosuppression and leukopenia as secondary effects in patients. The development of anticancer agents that simultaneously provoke antitumor immune response represents an important therapeutic advance. The administration of 6-pentadecyl salicylic acid (6SA) contributes to the antitumor immunity using 4T1 breast cancer cells in Balb/c female mice, with Taxol as a positive control and in cotreatment with 6SA (6SA + Taxol; CoT). Our results show that 6SA reduces tumor volume and size by inducing caspase-8-mediated apoptosis without reducing tumor infiltrated lymphocytes. Also, 6SA reduced lung metastasis and increased the proportion of immune cells in blood, lymph nodes and bone marrow; more evidently, in the proportion of tumor-infiltrated natural killer (NK) cells and cytotoxic T lymphocytes. Taxol reduces helper and cytotoxic lymphocytes causing systemic immunosuppression and myelosuppression in bone marrow, whereas 6SA does not decrease any immune cell subpopulations in circulating blood and lymph nodes. More importantly, the CoT decreased the Taxol-induced cytotoxicity in circulating T cells and bone marrow. Treatment with 6SA increases the secretion of IL-2, IL-12, GM-CSF, TNF-α and IFN-γ and significantly reduces IL-10 and IL-17 secretion, suggesting that the reduction of regulatory T cells and tumor-associated macrophages contribute to the host control of tumor development. Finally, 6SA has an effective antineoplastic activity against breast cancer cells in an immunocompetent animal, reduces the myelosuppression and leukopenia that Taxol produces, improves the antitumoral immunological microenvironment and increases the overall survival of the animals improving the quality of life of patients with cancer.

Genotoxicity of the organophosphate pesticide malathion and its metabolite dimethylthiophosphate in human cells in vitro.

Organophosphate (OP) pesticides are biotransformed into metabolites such as dialkylphosphates (DAPs). We have evaluated the genotoxicity of malathion and its metabolite dimethylthiophosphate (DMTP) in the human hepatic cell lines HepG2 and WRL-68 and in peripheral blood mononuclear cells (PBMC). In the Cytokinesis-Block Micronucleus assay (CBMN), malathion and DMTP increased the frequencies of micronuclei (MN) and nucleoplasmic bridges (NPB). Malathion was primarily clastogenic whereas DMTP was aneuploidogenic. When HepG2 or WRL-68 cells were treated with DMTP in the presence of sulconazole, a non-specific cytochrome P450 inhibitor, MN frequency was reduced, indicating that DMTP genotoxicity requires P450-cataliyzed metabolism.

Increased heart fibrosis and acute infection in a murine Chagas disease model associated with organophosphorus pesticide metabolite exposure.

Some reports suggest that exposure to organophosphorus (OP) pesticides increases the incidence of infections. Ethylated dialkylphosphates (EtDAPs) are metabolites of OP pesticides widely distributed with immunomodulatory potential. Chagas disease is produced by Trypanosoma cruzi parasites, and resolution of this infection requires the activation of inflammatory macrophages (MΦ), which results in cardiac fibrosis. Some reports indicate that EtDAPs increase the amount of the anti-inflammatory alternatively activated MΦ (M2; CD206+F4/80+). Therefore, we analyzed the course of T. cruzi infection, MΦ profiles from peritoneal exudate cells (PECs), inflammatory cell infiltration and fibrosis in the heart of BALB/c mice exposed to diethyldithiophosphate (DEDTP), diethylthiophosphate (DETP) or diethylphosphate (DEP, 0.01 g/kg), common DAPs produced by OP pesticides, 24 h before infection with T. cruzi. We found that DEDTP increased the parasite burden in blood by 99% at the peak of the infection and enhanced the myocardial damage due to an increase in infiltrated inflammatory cells (induced by DEDTP or DETP) and fibrosis (induced by EtDAPs). In the PECs, exposure to EtDAPs increased the proportion of the MΦ subpopulations of M2a, M2b and M2d, which are associated with tissue repair. These results indicate that exposure to EtDAPs can exacerbate the acute phase of a parasitic infection and increase the long-term damage to the heart.

Parkin is transcriptionally regulated by the aryl hydrocarbon receptor: Impact on α-synuclein protein levels.

Parkin (PRKN) is a ubiquitin E3 ligase that catalyzes the ubiquitination of several proteins. Mutations in the human Parkin gene, PRKN, leads to degeneration of dopaminergic (DA) neurons, resulting in autosomal recessive early-onset parkinsonism and the loss of PRKN function is linked to sporadic Parkinson's disease (PD). Additionally, several in vitro studies have shown that overexpression of exogenous PRKN protects against the neurotoxic effects induced by a wide range of cellular stressors, emphasizing the need to study the mechanism(s) governing PRKN expression and induction. Here, Prkn was identified as a novel target gene of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor and member of the bHLH/PAS (basic helix-loop-helix/Per-Arnt-Sim) superfamily. AhR binds and transactivates the Prkn gene promoter. We also demonstrated that AhR is expressed in DA neurons and that its activation upregulates Prkn mRNA and protein levels in the mouse ventral midbrain. Additionally, the AhR-dependent increase in PRKN levels is associated with a decrease in the protein levels of its target substrate, α-synuclein, in an AhR-dependent manner, because this effect is not observed in Ahr-null mice. These results suggest that treatments designed to induce PRKN expression through the use of nontoxic AhR agonist ligands may be novel strategies to prevent and delay PD.

The antineoplastic agent anacardic 6-pentadecyl salicylic acid produces immunomodulation in vivo via the activation of MAPKs.

Anacardic 6-pentadecyl salicylic acid (6SA) is the active component of Amphipterygium adstringens, a plant used in traditional medicine for the treatment of malaria and vascular diseases and as an anti-bacterial and immune-modulatory agent. However, the effect of 6SA on the immune system remains unclear. In this study, we examined the immune-stimulatory activity of 6SA in 6-8-week-old female BALB/c mice. We found that treatment with 2 mg/kg of 6SA increased the proportions of macrophages after 7 and 14 days of treatment and of natural killer (NK) cells after 14 days of treatment in circulating blood. In lymph nodes, treatment with 6SA for 14 days increased the number of macrophages. In addition, 6SA increases in the systemic levels of pro-inflammatory cytokines such as tumour necrosis factor (TNF)-α, interleukin (IL)-2, IL-12, IL-6 and IL-1ß and of nitric oxide (NO). We observed an increase in the secretion of Granulocyte/Macrophage Colony Stimulation Factor (GM-CSF) that could explain the increase in the proportion of macrophages. Moreover, 6SA induced the classical activation of macrophages by increasing their expression of MHC-II and their production of TNF-α. These M1-polarised macrophages presented enhanced phagocytosis and NO secretion. This activation was due to induction of the phosphorylation of MAPKs such as ERK, JNK and p38 because specific inhibitors of the phosphorylation of these MAPKs reduced the 6SA-induced phagocytosis and NO and particularly, the secretion of GM-CSF in macrophages by inhibition of ERK. Despite these effects on macrophages, 6SA does not have any direct effect on the proportion of lymphocytes.

The neurotoxin diethyl dithiophosphate impairs glutamate transport in cultured Bergmann glia cells.

Glutamate, the main excitatory neurotransmitter in the vertebrate Central Nervous System, is involved in almost every aspect of brain physiology, and its signaling properties are severely affected in most neurodegenerative diseases. This neurotransmitter has to be efficiently removed from the synaptic cleft in order to prevent an over-stimulation of glutamate receptors that leads to neuronal death. Specific sodium-dependent membrane transporters, highly enriched in glial cells, elicit the clearance of glutamate. Once internalized, it is metabolized to glutamine by the glia-enriched enzyme Glutamine synthetase. Accumulated glutamine is released into the extracellular space for its uptake into pre-synaptic neurons and its conversion to glutamate that is packed into synaptic vesicles completing the glutamate/glutamine cycle. Diverse chemical compounds, like organophosphates, directly affect brain chemistry by altering levels of neurotransmitters in the synaptic cleft. Organophosphate compounds are widely used as pesticides, and all living organisms are continuously exposed to these substances, either in a direct or indirect manner. Its metabolites, like the diethyl dithiophosphate, are capable of causing brain damage through diverse mechanisms including perturbation of neuronal-glial cell interactions and have been associated with attention-deficit disorders and other mental illness. In order to characterize the neurotoxic mechanisms of diethyl dithiophosphate, we took advantage of the well characterized model of chick cerebellar Bergmann glia cultures. A significant impairment of [3H] d-Aspartate transport was found upon exposure to the metabolite. These results indicate that glia cells are targets of neurotoxic substances such as pesticides and that these cells might be critically involved in the associated neuronal death.

Exploring an animal model of amodiaquine-induced liver injury in rats and mice.

Amodiaquine (AQ) is associated with a relatively high incidence of idiosyncratic drug-induced liver injury (IDILI) and agranulocytosis. A previous study reported that a combination of high dose AQ and glutathione (GSH) depletion led to liver injury. However, the characteristics of this toxicity were very different from AQ-induced liver injury in humans. We developed a model of AQ-induced liver injury with characteristics similar to the injury in humans by treating mice with lower doses of AQ for several weeks. In this study we found that not only did GSH depletion not increase AQ covalent binding to hepatic proteins at this lower dose, but also it paradoxically prevented the liver injury. We extended the model to rats and found AQ treatment led to a mild delayed onset liver injury that resolved despite continued treatment with AQ. Immunohistochemistry indicated the presence of Kupffer cell activation, apoptosis and hepatocyte proliferation in the liver. There was also an increase in serum IL-2, IL-5, IL-9, IL-12, MCP-1 and TGFß, but a decrease in leptin. Coincident with the elevated serum ALT, the number of liver CD4(+) T-cells, IL-17 secreting cells and TH17/Treg cells increased at Week 3 and decreased during continued treatment. Increases in NK1.1+ cells and activated M2 macrophages were also observed during liver injury. These results suggest that the outcome of the liver injury was determined by the balance between effector and regulatory cells. Co-treatment with cyclosporin prevented AQ-induced liver injury, which supports an immune mechanism. Retinoic acid (RA), which has been reported to enhance natural killer (NK) cell activity, exacerbated AQ-induced liver injury. These results suggest that AQ-induced IDILI is immune mediated and the subsequent adaptation appears to represent immune tolerance.

Aryl hydrocarbon receptor influences nitric oxide and arginine production and alters M1/M2 macrophage polarization.

AIMS: The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates the toxicity of environmental pollutants. It is also implicated in the regulation of the immune system. Ahr-null macrophages overproduce several proinflammatory cytokines following LPS-mediated stimulation, suggesting that AHR affects the balance between the inflammatory M1 and anti-inflammatory M2 phenotypes. Therefore, the present study aimed to examine whether the loss of AHR modifies macrophage polarization. MATERIALS AND METHODS: Peritoneal macrophages from wild-type and Ahr-null mice were differentiated into M1 or M2 phenotype by treatment with LPS/IFNγ or IL-4, and several M1 and M2 markers were determined by qPCR and ELISA assays. Macrophage phagocytic capacity was determined through phagocytosis of yeast and Leishmania major infection assays. Nitric oxide (NO) and urea production, and arginase activity were also determined. KEY FINDINGS: When macrophages were polarized to the M1 phenotype, Ahr-null cells presented a mixed response; higher levels of IL-1ß, IL-6, IL-12, and TNFα were observed after IFNγ- and LPS-mediated activation. However, Ahr-null cells also exhibited decreased NO production and phagocytic capacity. When macrophage was polarized to the M2 phenotype, Ahr-null cells exhibited lower levels of Fizz1, Ym1, and IL-10. In contrast, arginase activity was increased when compared to wild-type macrophages. In addition, macrophages from Ahr-null mice were more susceptible to L. major infection. SIGNIFICANCE: Disruption of the Ahr gene alters macrophage polarization when compared to WT macrophage. These changes may affect the development and resolution of several diseases such as bacterial or parasitic infections.

The anacardic 6-pentadecyl salicylic acid induces macrophage activation via the phosphorylation of ERK1/2, JNK, P38 kinases and NF-κB.

Amphipterygium adstringens is a plant traditionally used to treat gingivitis, gastric ulcer and even gastric cancer but the mechanism involved in the regulation of the immune response is not elucidated yet. The 6-pentadecylsalicylic acid (6SA) is the main anacardic acid found in A. adstringens. In order to evaluate the immune-modulatory abilities of 6SA, we used mouse splenocytes and determined the phosphorylation of the transcription factor NF-κB and MAP kinases ERK1/2, JNK and p38 in helper and cytotoxic T cells, natural killer (NK) cells and F4/80(+) macrophages. Treatment with 6SA was not cytotoxic as measured by both trypan blue exclusion and tetrazolium salts (MTT) tests. Additionally, 6SA did not alter the proportion of helper and cytotoxic T lymphocytes, NK cells or macrophages. Moreover, 6SA treatment significantly increased the phosphorylation of ERK1/2, JNK, P38 and NF-κB mainly in macrophages. In this cells (peritoneal macrophages), treatment with 6SA increased the secretion of nitric oxide (NO), interleukin (IL)-6 and tumour necrosis factor (TNF)-α and decreased the secretion of IL-4 and IL-10 depending on MAPK and NF-κB phosphorylation. In addition, 6SA increased the migration and phagocytic activity of macrophages also depending on the phosphorylation of different kinases. These data suggest that 6SA induces the classical activation pathway in macrophages via the phosphorylation of MAP kinases and NF-κB thus activating the adaptive immune system.