BackgroundRegulators of G-protein signaling (RGS) are major physiological modulators of G-protein-coupled receptors (GPCR) signaling. Several GPCRs expressed in both neurons and astrocytes participate in the central control of pain processing, and the reduced efficacy of analgesics in neuropathic pain conditions may rely on alterations in RGS function. The expression and the regulation of RGS in astrocytes is poorly documented, and we herein hypothesized that neuroinflammation which is commonly observed in neuropathic pain could influence RGS expression in astrocytes.MethodsIn a validated model of neuropathic pain, the spared nerve injury (SNI), the regulation of RGS2, RGS3, RGS4, and RGS7 messenger RNA (mRNA) was examined up to 3 weeks after the lesion. Changes in the expression of the same RGS were also studied in cultured astrocytes exposed to defined activation protocols or to inflammatory cytokines.ResultsWe evidenced a differential regulation of these RGS in the lumbar spinal cord of animals undergoing SNI. In particular, RGS3 appeared upregulated at early stages after the lesion whereas expression of RGS2 and RGS4 was decreased at later stages. Decrease in RGS7 expression was already observed after 3 days and outlasted until 21 days after the lesion. In cultured astrocytes, we observed that changes in the culture conditions distinctly influenced the constitutive expression of these RGS. Also, brief exposures (4 to 8 h) to either interleukin-1β, interleukin-6, or tumor necrosis factor α caused rapid changes in the mRNA levels of the RGS, which however did not strictly recapitulate the regulations observed in the spinal cord of lesioned animals. Longer exposure (48 h) to inflammatory cytokines barely influenced RGS expression, confirming the rapid but transient regulation of these cell signaling modulators.ConclusionChanges in the environment of astrocytes mimicking the inflammation observed in the model of neuropathic pain can affect RGS expression. Considering the role of astrocytes in the onset and progression of neuropathic pain, we propose that the inflammation-mediated modulation of RGS in astrocytes constitutes an adaptive mechanism in a context of neuroinflammation and may participate in the regulation of nociception.

IL-1α is an intracellular danger signal (DAMP) released by macrophages contributing to the development of silica-induced lung inflammation. The exact molecular mechanism orchestrating IL-1α extracellular release from particle-exposed macrophages is still unclear. To delineate this process, murine J774 and bone-marrow derived macrophages were exposed to increasing concentrations (1–40 cm2/ml) of a set of amorphous and crystalline silica particles with different surface chemical features. In particular, these characteristics include the content of nearly free silanols (NFS), a silanol population responsible for silica cytotoxicity recently identified. We first observed de novo stocks of IL-1α in macrophages after silica internalization regardless of particle physico-chemical characteristics and cell stress. IL-1α intracellular production and accumulation were observed by exposing macrophages to biologically-inert or cytotoxic crystalline and amorphous silicas. In contrast, only NFS-rich reactive silica particles triggered IL-1α release into the extracellular milieu from necrotic macrophages. We demonstrate that IL-1α is actively secreted through the formation of gasdermin D (GSDMD) pores in the plasma membrane and not passively released after macrophage plasma membrane lysis. Our findings indicate that the GSDMD pore-dependent secretion of IL-1α stock from macrophages solely depends on cytotoxicity induced by NFS-rich silica. This new regulated process represents a key first event in the mechanism of silica toxicity, suitable to refine the existing adverse outcome pathway (AOP) for predicting the inflammatory activity of silicas.

Although epidemiological studies have suggested an association between asbestos exposure and systemic autoimmunity, the underlying mechanisms are poorly understood. Short asbestos fibers are often considered less harmful than long fibers but such a statement is still a matter of debates. This study aimed to compare the effects of short (SFA) and long (LFA) amosite fiber exposure on lung damage, autoimmunity and macrophage phenotype. Four months after lung exposure to 0.1 mg of fibers, BAL levels of lactate dehydrogenase, free DNA, CCL2, TIMP-1 and immunoglobulins A of LFA-exposed C57Bl/6 mice were increased when compared to fluids from control- and SFA-exposed mice. Effects in LFA-exposed mice were associated with lung fibrosis and autoimmunity including anti-double-strand DNA antibody production. Human monocyte-derived macrophages (MDMs) exposed to SFA or LFA at 20 µg/cm2 have a pro-inflammatory phenotype characterized by a significant increase of TNFα and IL-6 secretion. A decrease of efferocytosis capacities was also noted after SFA and LFA, whereas macrophage abilities to phagocyte fluorescent beads were unchanged when compared to control MDMs. SFA exposure induced IL-6 secretion and reduced the percentage of MDMs expressing MHCII and CD86 markers involved in antigen and T-lymphocyte stimulation. By contrast, NLRP3 inflammasome activation, evaluated through quantification of caspase-1 activity and IL-1β secretion is rather associated to LFA than SFA exposure. Our results demonstrated that only long-term exposure to LFA has induced significant lung damages and autoimmune effects supporting a worsened health effects of LFA in comparison to SFA.

<p>Exposure to asbestos is known for inducing inflammation, pulmonary fibrosis, lung cancer and malignant mesothelioma. Although several novel biomarkers and treatments are being tested, these chronic disorders are currently incurable, and the prognosis is particularly poor. The development of asbestos-induced cancer results from mutations, cell transformation and proliferation caused by reactive oxygen species and elevated levels of pro-inflammatory cytokines. Fiber-induced chronic inflammation also explains asbestosis, an interstitial lung disease characterized by uncontrolled matrix protein deposition leading to detrimental lung fibrosis. The investigation of the in-vivo molecular mechanisms driving asbestos pathogenicity is still a matter of debate. In this context, it is accepted that the physico-chemical properties of the fibers play a crucial role in causing adverse effects, and long fibers are still considered more toxic than short ones. Indeed, when fibers reach the alveolar space and migrate to the pleural/peritoneal cavity, long and thin fibers shows stronger inflammogenic, fibrogenic and tumorigenic effects than short fibers, in the long term. It is largely hold that short asbestos fibers are more easily cleared from the lungs and elicit a lower reactional and/or inflammatory effect. To further investigate this paradigm of toxicity, we compared the pro-inflammatory and pro-fibrogenic potential of short and long amosite fibers in <em>in-vitro</em> (J774 murine macrophages) and <em>in-vivo</em> (C57BL/6 mice) models. Surprisingly, our results demonstrated that short fibers were more prone to induce in-vitro cytotoxicity, accompanied by the release of pro-inflammatory biomarkers, in comparison to long fibers. On the contrary, the long fibers were significantly more inflammogenic and fibrogenic in the lungs of treated mice, while the short fibers were almost inert and did not induce acute and chronic inflammation and fibrosis. Furthermore, we observed that, while the long fibers were still present in the lungs of the animals 4 months after the exposure, the short amosite was substantially absent. These findings imply that the pulmonary deposition and a defect of clearance of the fibers play a crucial role in the development of in-vivo detrimental effects associated to long asbestos. This effect overcomes the mere in-vitro cytotoxic and inflammogenic potential of short fibers. Present results provide new insights into the mechanisms that drive asbestos toxicity, opening new perspectives for the development of reliable in vitro tests that fully predict health adverse effects associated to inorganic mineral fibers.</p>