Silica-induced Inflammation Research Articles

Glycolytic reprogramming governs crystalline silica-induced pyroptosis and inflammation through promoting lactylation modification

Prolonged inhalation of environmental crystalline silica (CS) can cause silicosis, characterized by persistent pulmonary inflammation and irreversible fibrosis, but the mechanism has not been elucidated. To uncover the role and underlying mechanism of glycolytic reprogramming in CS-induced pulmonary inflammation, the mouse silicosis models and glycolysis inhibition models were established in vivo. And the CS-induced macrophage activation models were utilized to further explore the underlying mechanism in vitro. The results showed that CS induced lung inflammation accompanied by glycolytic reprogramming and pyroptosis. The application of glycolysis inhibitor (2-DG) suppressed CS-induced pyroptosis and alleviated lung inflammation. In vitro, 2-DG effectively impeded CS-induced macrophage pyroptosis and inflammatory response. Mechanistically, 2-DG suppressed pyroptosis by inhibiting NLRP3 inflammasome activation both in vivo and in vitro. Furtherly, metabolite lactate facilitated NLRP3-dependent pyroptosis synergistically with CS particles, while blocking the source of lactate largely alleviated NLRP3 inflammasome activation and subsequent pyroptosis triggered by CS. More profoundly, the increment of lactate induced by CS might drive NLRP3-dependent pyroptosis by increasing histone lactylation levels. In conclusion, our findings demonstrated inhibiting glycolytic reprogramming could alleviate CS-induced inflammatory response through suppressing NLRP3 -dependent pyroptosis. Increased glycolytic metabolite lactate and protein lactylation modifications might represent significant mechanisms during CS-induced NLRP3 activation and macrophage pyroptosis.

Differential regulation of lung homeostasis and silicosis by the TAM receptors MerTk and Axl.

TAM receptor-mediated efferocytosis plays an important function in immune regulation and may contribute to antigen tolerance in the lungs, a site with continuous cellular turnover and generation of apoptotic cells. Some studies have identified failures in efferocytosis as a common driver of inflammation and tissue destruction in lung diseases. Our study is the first to characterize the in vivo function of the TAM receptors, Axl and MerTk, in the innate immune cell compartment, cytokine and chemokine production, as well as the alveolar macrophage (AM) phenotype in different settings in the airways and lung parenchyma. We employed MerTk and Axl defective mice to induce acute silicosis by a single exposure to crystalline silica particles (20 mg/50 μL). Although both mRNA levels of Axl and MerTk receptors were constitutively expressed by lung cells and isolated AMs, we found that MerTk was critical for maintaining lung homeostasis, whereas Axl played a role in the regulation of silica-induced inflammation. Our findings imply that MerTk and Axl differently modulated inflammatory tone via AM and neutrophil recruitment, phenotype and function by flow cytometry, and TGF-β and CXCL1 protein levels, respectively. Finally, Axl expression was upregulated in both MerTk-/- and WT AMs, confirming its importance during inflammation. This study provides strong evidence that MerTk and Axl are specialized to orchestrate apoptotic cell clearance across different circumstances and may have important implications for the understanding of pulmonary inflammatory disorders as well as for the development of new approaches to therapy.

Macrophage-derived exosomal HMGB3 regulates silica-induced pulmonary inflammation by promoting M1 macrophage polarization and recruitment

BackgroundChronic inflammation and fibrosis are characteristics of silicosis, and the inflammatory mediators involved in silicosis have not been fully elucidated. Recently, macrophage-derived exosomes have been reported to be inflammatory modulators, but their role in silicosis has not been explored. The purpose of the present study was to investigate the role of macrophage-derived exosomal high mobility group box 3 (HMGB3) in silica-induced pulmonary inflammation.MethodsThe induction of the inflammatory response and the recruitment of monocytes/macrophages were evaluated by immunofluorescence, flow cytometry and transwell assays. The expression of inflammatory cytokines was examined by RT–PCR and ELISA, and the signalling pathways involved were examined by western blot analysis.ResultsHMGB3 expression was increased in exosomes derived from silica-exposed macrophages. Exosomal HMGB3 significantly upregulated the expression of inflammatory cytokines, activated the STAT3/MAPK (ERK1/2 and p38)/NF-κB pathways in monocytes/macrophages, and promoted the migration of these cells by CCR2.ConclusionsExosomal HMGB3 is a proinflammatory modulator of silica-induced inflammation that promotes the inflammatory response and recruitment of monocytes/macrophages by regulating the activation of the STAT3/MAPK/NF-κB/CCR2 pathways.

Lysosomal BK channels facilitate silica-induced inflammation in macrophages

Background Lysosomal ion channels are proposed therapeutic targets for a number of diseases, including those driven by NLRP3 inflammasome-mediated inflammation. Here, the specific role of the lysosomal big conductance Ca2+-activated K+ (BK) channel was evaluated in a silica model of inflammation in murine macrophages. A specific-inhibitor of BK channel function, paxilline (PAX), and activators NS11021 and NS1619 were utilized to evaluate the role of lysosomal BK channel activity in silica-induced lysosomal membrane permeabilization (LMP) and NLRP3 inflammasome activation resulting in IL-1β release. Methods Murine macrophages were exposed in vitro to crystalline silica following pretreatment with BK channel inhibitors or activators and LMP, cell death, and IL-1β release were assessed. In addition, the effect of PAX treatment on silica-induced cytosolic K+ decrease was measured. Finally, the effects of BK channel modifiers on lysosomal pH, proteolytic activity, and cholesterol transport were also evaluated. Results PAX pretreatment significantly attenuated silica-induced cell death and IL-1β release. PAX caused an increase in lysosomal pH and decrease in lysosomal proteolytic activity. PAX also caused a significant accumulation of lysosomal cholesterol. BK channel activators NS11021 and NS1619 increased silica-induced cell death and IL-1β release. BK channel activation also caused a decrease in lysosomal pH and increase in lysosomal proteolytic function as well as a decrease in cholesterol accumulation. Conclusion Taken together, these results demonstrate that inhibiting lysosomal BK channel activity with PAX effectively reduced silica-induced cell death and IL-1β release. Blocking cytosolic K+ entry into the lysosome prevented LMP through the decrease of lysosomal acidification and proteolytic function and increase in lysosomal cholesterol.

Anti-inflammatory protein TSG-6 secreted by BMSCs attenuates silica-induced acute pulmonary inflammation by inhibiting NLRP3 inflammasome signaling in macrophages

Silicosis is a severe occupational lung disease caused by inhalation of silica particles. Unfortunately, there are currently limited treatment options available for silicosis. Recent advances have indicated that bone marrow mesenchymal stem cells (BMSCs) have a therapeutic effect on silicosis, but their efficacy and underlying mechanisms remain largely unknown. In this study, we focused on the early phase of silica-induced lung injury to investigate the therapeutic effect of BMSCs. Our findings demonstrated that BMSCs attenuated silica-induced acute pulmonary inflammation by inhibiting NLRP3 inflammasome pathways in lung macrophages. To further understand the mechanisms involved, we utilized RNA sequencing to analyze the transcriptomes of BMSCs co-cultured with silica-stimulated bone marrow-derived macrophages (BMDMs). The results clued tumor necrosis factor-stimulated gene 6 (TSG-6) might be a potentially key paracrine secretion factor released from BMSCs, which exerts a protective effect. Furthermore, the anti-inflammatory and inflammasome pathway inhibition effects of BMSCs were attenuated when TSG-6 expression was silenced, both in vivo and in vitro. Additionally, treatment with exogenous recombinant mouse TSG-6 (rmTSG-6) demonstrated similar effects to BMSCs in attenuating silica-induced inflammation. Overall, our findings suggested that BMSCs can regulate the activation of inflammasome in macrophages by secreting TSG-6, thereby protecting against silica-induced acute pulmonary inflammation both in vivo and in vitro.

Crystalline silica-exposed human lung epithelial cells presented enhanced anchorage-independent growth with upregulated expression of BRD4 and EZH2 in autocrine and paracrine manners.

Crystalline silica-induced inflammation possibly facilitates carcinogenesis. Here, we investigated its effect on lung epithelium damage. We prepared conditioned media of immortalized human bronchial epithelial cell lines (hereinafter bronchial cell lines) NL20, BEAS-2B, and 16HBE14o- pre-exposed to crystalline silica (autocrine crystalline silica conditioned medium), a phorbol myristate acetate-differentiated THP-1 macrophage line, and VA13 fibroblast line pre-exposed to crystalline silica (paracrine crystalline silica conditioned medium). As cigarette smoking imposes a combined effect on crystalline silica-induced carcinogenesis, a conditioned medium was also prepared using the tobacco carcinogen benzo[a]pyrene diol epoxide. Crystalline silica-exposed and growth-suppressed bronchial cell lines exhibited enhanced anchorage-independent growth in autocrine crystalline silica and benzo[a]pyrene diol epoxide conditioned medium compared with that in unexposed control conditioned medium. Crystalline silica-exposed nonadherent bronchial cell lines in autocrine crystalline silica and benzo[a]pyrene diol epoxide conditioned medium showed increased expression of cyclin A2, cdc2, and c-Myc, and of epigenetic regulators and enhancers, BRD4 and EZH2. Paracrine crystalline silica and benzo[a]pyrene diol epoxide conditioned medium also accelerated the growth of crystalline silica-exposed nonadherent bronchial cell lines. Culture supernatants of nonadherent NL20 and BEAS-2B in crystalline silica and benzo[a]pyrene diol epoxide conditioned medium had higher EGF concentrations, whereas those of nonadherent 16HBE14o- had higher TNF-α levels. Recombinant human EGF and TNF-α promoted anchorage-independent growth in all lines. Treatment with EGF and TNF-α neutralizing antibodies inhibited cell growth in crystalline silica conditioned medium. Recombinant human TNF-α induced BRD4 and EZH2 expression in nonadherent 16HBE14o-. The expression of γH2AX occasionally increased despite PARP1 upregulation in crystalline silica-exposed nonadherent lines with crystalline silica and benzo[a]pyrene diol epoxide conditioned medium. Collectively, crystalline silica- and benzo[a]pyrene diol epoxide-induced inflammatory microenvironments comprising upregulated EGF or TNF-α expression may promote crystalline silica-damaged nonadherent bronchial cell proliferation and oncogenic protein expression despite occasional γH2AX upregulation. Thus, carcinogenesis may be cooperatively aggravated by crystalline silica-induced inflammation and genotoxicity.

Cholesterol-dependent molecular mechanisms contribute to cationic amphiphilic drugs’ prevention of silica-induced inflammation

Silicosis is considered an irreversible chronic inflammatory disease caused by the inhalation of crystalline silica (cSiO2). The cycle of inflammation that drives silicosis and other particle-caused respiratory diseases is mediated by NLRP3 inflammasome activity in macrophages resulting in the release of IL-1β. Lysosomal membrane permeability (LMP) initiated by inhaled particles is the key regulatory step in leading to NLRP3 activity. In addition to its role in LMP, the lysosome is crucial to cellular cholesterol trafficking. Lysosomal cholesterol has been demonstrated to regulate LMP while cationic amphiphilic drugs (CADs) reduce cholesterol trafficking from lysosomes and promote endolysosomal cholesterol accumulation as seen in Niemann Pick disease. Using a bone marrow derived macrophage (BMdM) model, four CADs were examined for their potential to reduce cSiO2-induced inflammation. Here we found that FDA-approved CAD drugs imipramine, hydroxychloroquine, fluvoxamine, and fluoxetine contributed to reduced LMP and IL-1β release in cSiO2 treated BMdM. These drugs inhibited lysosomal enzymatic activity of acid sphingomyelinase, decreased lysosomal proteolytic function, and increased lysosomal pH. CADs also demonstrated a significant increase in lysosomal-associated free cholesterol. Increased lysosomal cholesterol was associated with a significant reduction in cSiO2 induced LMP and IL-1β release. In contrast, reduced lysosomal cholesterol significantly exacerbated cSiO2-induced IL-1β release and reduced the protective effect of CADs on IL-1β release following cSiO2 exposure. Taken together, these results suggest that CAD modification of lysosomal cholesterol may be used to reduce LMP and cSiO2-induced inflammation and could prove an effective therapeutic for silicosis and other particle-caused respiratory diseases.

Human umbilical cord mesenchymal stem cell-derived extracellular vesicles alleviated silica induced lung inflammation and fibrosis in mice via circPWWP2A/miR-223–3p/NLRP3 axis

Silicosis is a progressive inflammatory disease with poorly defined mechanisms and limited therapeutic options. Recent studies found that microRNAs (miRNAs) and circular RNAs (circRNAs) were involved in the development of respiratory diseases; however, the function of non-coding RNAs in silicosis was still needed to be further explored. We found that miR-223–3p was significantly decreased in macrophages and lung tissues of mice after silica treatment, which were consistent with the results of GEO database microarray analysis. Notably, NLRP3 is a target gene downstream of miR-223–3p. And circular RNA PWWP2A (circPWWP2A) was significantly elevated after silica stimulation. To elucidate the role of these RNAs in silica-induced inflammation in macrophages and lung tissues, we investigated the upstream molecular mechanisms of circPWWP2A on the inflammatory response. The inhibitory effect of miR-223–3p on its target NLRP3 was suppressed by circPWWP2A, which led to lung fibrosis. Our study found that circPWWP2A could adsorb miR-223–3p to regulate NLRP3 after silica stimulation in pulmonary fibrosis. And our results revealed that the circPWWP2A–miR-223–3p–NLRP3 axis was potentially instrumental in managing silica-induced inflammation and fibrosis. Previous studies have demonstrated that human umbilical cord mesenchymal stem cell-derived extracellular vesicles (hucMSC-EVs) exhibit anti-inflammatory and anti-fibrotic effects in multiple organs. However, the potential effectiveness of hucMSC-EVs against silicosis or the underlying mechanisms of their biological outcomes remains unclear. Therefore, we used 3D culture technology to extract hucMSC-EVs and observed their effects in macrophages and lung tissues, respectively. According to the EVmiRNA database, miR-223–3p was abundant in MSC-EVs. In addition, hucMSC-EVs may modulate lung function, reduce the secretion of inflammatory factors (NLRP3, IL-1β, IL-18 and cleaved Caspase-1) and attenuate the deposition of fibrosis-related factors (Collagen Ⅰ, Collagen Ⅲ, fibronectin and α-SMA). In vitro results evinced that hucMSC-EVs reduced the inflammatory response of macrophages and restricted the activation and proliferation of fibroblasts. Moreover, our study showed that hucMSCs-EVs acted as a mediator to transfer miR-223–3p to suppress circPWWP2A, thereby alleviating pulmonary fibrosis through the NLRP3 signaling pathway. These data may provide potentially novel strategies for investigating the pathogenesis of silicosis and developing novel treatments for this disease.

Effects of early dexamethasone treatment on several markers of inflammation and fibrosis in an animal model of lung silicosis in rats – A pilot study

Abstract Lung silicosis is primarily caused by inhalation of particles of silicon oxide (silica). Despite a huge progress in understanding the interactions among the pathomechanisms of lung silicosis in the last years, there is a lack of effective therapy. With respect to a wide therapeutic action of corticosteroids, the purpose of this pilot study was to evaluate early effects of dexamethasone on several markers of inflammation and lung fibrosis in a rat model of silicosis. The silicosis model was induced by a single transoral intratracheal instillation of silica (50 mg/ml/animal), while the controls received an equivalent volume of sterile saline. The treatment with intraperitoneal dexamethasone initiated the next day after the silica instillation and was given 2-times a week at a dose of 1 mg/kg, while the controls received an equivalent volume of saline. The animals were euthanized 14 or 28 days after the treatment onset. Total and differential counts of leukocytes in the blood and bronchoalveolar lavage (BAL) fluid were determined. The presence of collagen in the bronchioles and lung vessels was detected by Sirius red staining and a smooth muscle mass was detected by smooth muscle actin. In comparison to saline, the instillation of silica increased the total count of circulating leukocytes after 14 and 28 days of the experiment (both p<0.05), which was associated with higher counts of lymphocytes (p<0.05 after 14 days, p>0.05 after 28 days) and slight but non-significant increases in neutrophils and eosinophils (both p>0.05). Although the total cell count in the BAL fluid did not change significantly, the percentages and absolute counts of neutrophils, eosinophils, and lymphocytes (p<0.05, p<0.01 or p<0.001) elevated after 14 and 28 days of the experiment. Silica induced an accumulation of collagen in the bronchioles (p<0.001 after both 14 and 28 days) and pulmonary vessels (p<0.01 after both 14 and 28 days) and elevated a formation of smooth muscle mass (p<0.05 after 14 days, p<0.01 or p<0.001 after 28 days). Treatment with dexamethasone decreased circulating leukocytes (p<0.01) and lymphocytes (p<0.001) and increased neutrophils (p<0.05), which was associated with a slightly decreased total cell count in BAL (p>0.05), decline in lymphocytes (p<0.01), and slight decreases in neutrophils and eosinophils after 28 days of the treatment. Moreover, dexamethasone reduced the accumulation of collagen (p<0.01 after 14 days and p<0.001 after 28 days) and the formation of smooth muscle mass (p<0.01 for bronchioles and p>0.05 for vessels after 24 days, p<0.001 for both bronchioles and vessels after 28 days). In conclusion, early dexamethasone treatment mitigated silica-induced granulocytic-lymphocytic inflammation and decreased a formation of collagen and smooth muscle mass in the bronchiolar and vascular walls, demonstrating a therapeutic potential of dexamethasone in the lung silicosis.

Silica exposure activates non-canonical inflammasome complex in intratracheal instilled rat model.

Inhalation of silica crystals in occupational settings is a main cause of silicosis, a chronic irreversible pulmonary disorder. Our prior studies demonstrated the activation of inflammasome sensors AIM2 and NLRP3, effector protein caspase-1, and significant increase in IL-1β in silica exposed rats, suggesting that the canonical inflammasome activation may be associated with silica-induced tissue damage and inflammation. In our current study using the same animal model system, we further evaluated the components of non-canonical inflammasome, including NEK7, caspase-11, and GSDMD following silica exposure. We demonstrated sustained NEK7 elevation in the rat lung epithelial cells and macrophages following 1- and 3-day exposure. Enhanced NEK7 expression was also detected in lung homogenate by western blot. Similarly, caspase-11 expression was induced by silica exposure in lung sections and homogenate. Elevated GSDMD was observed both in lung sections by immunohistochemical staining and in lung tissue homogenate by western blot. In summary, our current study demonstrated increase in NEK7, caspase-11, and GSDMD in silica exposed rats, indicating activation of non-canonical inflammasome complex, thereby providing a broad inflammasome activation pathway caused by silica exposure.

Protective effects of intranasal curcumin on silica-induced lung damage

ObjectiveBeing anti-inflammatory and an antioxidant in nature, curcumin has been studied for its anti-asthmatic effects, but its impact on silicosis has not been investigated before. It is a form of occupational lung illness caused by inhaling crystalline silica. It is particularly common among those who work in construction-related sectors. Therefore, present study has been undertaken to investigate impact of intranasal curcumin on silica induced lung damage in mice model of silicosis. Materials and methodsMice model of silicosis was developed by intranasal silica instillation (2.5 mg/mice) for different durations mainly 7, 14 and 21 days, where the longest duration of silica exposure (21 days) mimics chronic occupational exposure of silica dust leading to silicosis. Curcumin (5 mg/kg,i.n) and /or dexamethasone, a known corticosteroid (10 mg/kg,i.p) was administered an hour prior to silica administration. ResultsPresent study revealed silica induced lung damage in the mice model of silicosis characterized by airway inflammation, collagen deposition and enhanced expression of fibrosis markers (MMP-9, α-SMA, Hydroxyproline), which were significantly reduced in curcumin treatment groups. Inhibitory effects of curcumin were compared with standard drug, dexamethasone, a corticosteroid and was found better in protecting structural alterations in the lung.Damaged and abnormal mitochondria (enlarged and irregular shapes) were observed in silicosis group which were reduced in curcumin and dexamethasone treatment groups as revealed in transmission electron microscopic studies. ConclusionsPresent study shows protective effects of intranasal curcumin on silica-induced airway inflammation and structural changes thereby lung damage. Hence, it can be considered as an alternative and complementary medication for silicosis.

Alveolar macrophage-derived progranulin mediated pro-inflammatory Il-6 expression via regulating Creb1 in silicosis model

Progranulin (PGRN) is a secreted factor involved in inflammatory diseases. However, the function of PGRN in silica-induced lung inflammation has not been elucidated. In this study, we demonstrated that PGRN in serum and lung tissues was markedly increased in silicosis mouse model. And immunohistochemistry results showed that PGRN was mainly expressed in alveolar macrophages, which was further confirmed in silica-treated alvelar macrophages cell line (MH-S) in vitro. PGRN promoted pro-inflammatory cytokines transcription such as interleukin (Il)-6, tumor necrosis factor-α (Tnf-α) and Il-1β in MH-S cells, and the increasing of Il-6 was most obvious. Knockdown of PGRN blocked the silica-induced elevation of intracellular Il-6 in MH-S cells. Furthermore, we also found that PGRN could increase the phosphorylation of Cyclic AMP-responsive element-binding protein 1 (Creb1), a transcriptional regulator of Il-6. Inhibition of p-Creb1 by the phosphorylation inhibitor of Creb1 (666–15) decreased PGRN-induced intracellular Il-6 production in MH-S cells. In conclusion, PGRN was highly increased in silicosis mouse model and upregulated inflammatory cytokines expression. These findings suggested that PGRN might be a key mediator in silica-induced inflammation and provided a new clue for the diagnosis and drug therapy of silicosis.

Tetracycline ameliorates silica-induced pulmonary inflammation and fibrosis via inhibition of caspase-1

BackgroundInhalation of dust containing silica particles is associated with severe pulmonary inflammation and lung injury leading to chronic silicosis including fibrotic remodeling of the lung. Silicosis represents a major global health problem causing more than 45.000 deaths per year. The inflammasome-caspase-1 pathway contributes to the development of silica-induced inflammation and fibrosis via IL-1β and IL-18 production. Recent studies indicate that tetracycline can be used to treat inflammatory diseases mediated by IL-1β and IL-18. Therefore, we hypothesized that tetracycline reduces silica-induced lung injury and lung fibrosis resulting from chronic silicosis via limiting IL-1β and IL-18 driven inflammation.MethodsTo investigate whether tetracycline is a therapeutic option to block inflammasome-caspase-1 driven inflammation in silicosis, we incubated macrophages with silica alone or combined with tetracycline. The in vivo effect of tetracycline was determined after intratracheal administration of silica into the mouse lung.ResultsTetracycline selectively blocks IL-1β production and pyroptotic cell death via inhibition of caspase-1 in macrophages exposed to silica particles. Consistent, treatment of silica-instilled mice with tetracycline significantly reduced pulmonary caspase-1 activation as well as IL-1β and IL-18 production, thereby ameliorating pulmonary inflammation and lung injury. Furthermore, prolonged tetracycline administration in a model of chronic silicosis reduced lung damage and fibrotic remodeling.ConclusionsThese findings suggest that tetracycline inhibits caspase-1-dependent production of IL-1β in response to silica in vitro and in vivo. The results were consistent with tetracycline reducing silica-induced pulmonary inflammation and chronic silicosis in terms of lung injury and fibrosis. Thus, tetracycline could be effective in the treatment of patients with silicosis as well as other diseases involving silicotic inflammation.

Herbal Compounds in the Treatment of Pulmonary Silicosis

Herbal compounds including those already well-established in traditional Chinese medicine have been increasingly tested in the treatment of various diseases. Recent studies have shown that herbal compounds can be of benefit also for pulmonary silicosis as they can diminish changes associated with silica-induced inflammation, fibrosis, and oxidative stress. Due to a lack of effective therapeutic strategies, development of novel approaches which may be introduced particularly in the early stage of the disease, is urgently needed. This review summarizes positive effects of several alternative plant-based drugs in the models of experimental silicosis with a potential for subsequent clinical investigation and use in future.

Herbal compounds in the treatment of pulmonary silicosis.

Herbal compounds including those already well-established in traditional Chinese medicine have been increasingly tested in the treatment of various diseases. Recent studies have shown that herbal compounds can be of benefit also for pulmonary silicosis as they can diminish changes associated with silica-induced inflammation, fibrosis, and oxidative stress. Due to a lack of effective therapeutic strategies, development of novel approaches which may be introduced particularly in the early stage of the disease, is urgently needed. This review summarizes positive effects of several alternative plant-based drugs in the models of experimental silicosis with a potential for subsequent clinical investigation and use in future.

Caveolin-1 negatively regulates inflammation and fibrosis in silicosis.

Inhalation of crystalline silica causes silicosis, the most common and serious occupational disease, which is characterized by progressive lung inflammation and fibrosis. Recent studies revealed the anti‐inflammatory and anti‐fibrosis role of Caveolin‐1 (Cav‐1) in lung, but this role in silicosis has not been investigated. Thus, this study evaluated Cav‐1 regulatory effects in silicosis. It was found that Cav‐1 levels were significantly reduced in the lung from silicosis patients and silicotic mice. The silicosis models were established in C57BL/6 (wild‐type) and Cav‐1 deficiency (Cav‐1 −/−) mice, and Cav‐1 −/− mice displayed wider alveolar septa, increased collagen deposition and more silicotic nodules. The mice peritoneal‐derived macrophages were used to explore the role of Cav‐1 in silica‐induced inflammation, which plays a central role in mechanism of silicosis. Cav‐1 inhibited silica‐induced infiltration of inflammatory cells and secretion of inflammatory factors in vitro and in vivo, partly by downregulating NF‐κB pathway. Additionally, silica uptake and expression of 4‐hydroxynonenal in silicotic mice were observed, and it was found that Cav‐1 absence triggered excessive silica deposition, causing a stronger oxidative stress response. These findings demonstrate the protective effects of Cav‐1 in silica‐induced lung injury, suggesting its potential therapeutic value in silicosis.

Necroptosis in pulmonary macrophages promotes silica-induced inflammation and interstitial fibrosis in mice

Silicosis is a disease characterized by extensive lung nodules and fibrosis caused by the prolonged inhalation of silica in occupational settings. However, the molecular mechanism of silicosis development is complex and not fully understood. Furthermore, the role of necroptosis, a death receptor-mediated and caspase-independent mode of inflammatory cell death, is not well understood in silicosis. Here, we demonstrate that the necroptotic signaling pathway of macrophages is significantly activated in the lungs of silicosis mouse models. Meanwhile, increased M1 macrophage infiltration and up-regulation of pro-inflammatory cytokines (TNF-α, IL-6) were observed in our silicosis model. Notably, the expression of the pro-fibrotic factor, TGF-β1, and fibrosis biomarkers α-SMA and collagen I were also unregulated; however, these phenomena were recovered by Nec-1, an inhibitor specific for RIP1 kinase-dependent necroptosis. We conclude that macrophage-mediated necroptosis promotes the progression of silicosis by enhancing lung inflammatory responses and fibrogenesis in a mouse model of silicosis. These findings provide new insights for drug discovery and clinical treatment of silicosis.

New Insights into Pathomechanisms and Treatment Possibilities for Lung Silicosis

Inhalation of silica particles is an environmental and occupational cause of silicosis, a type of pneumoconiosis. Development of the lung silicosis is a unique process in which the vicious cycle of ingestion of inhaled silica particles by alveolar macrophages and their release triggers inflammation, generation of nodular lesions, and irreversible fibrosis. The pathophysiology of silicosis is complex, and interactions between the pathomechanisms have not been completely understood. However, elucidation of silica-induced inflammation cascades and inflammation-fibrosis relations has uncovered several novel possibilities of therapeutic targeting. This article reviews new information on the pathophysiology of silicosis and points out several promising treatment approaches targeting silicosis-related pathways.

Triiodothyronine Attenuates Silica-Induced Oxidative Stress, Inflammation, and Apoptosis via Thyroid Hormone Receptor α in Differentiated THP-1 Macrophages.

Alveolar macrophage (AM) injury and inflammatory response are key processes in pathological damage caused by silica. However, the role of triiodothyronine (T3) in silica-induced AM oxidative stress, inflammation, and mitochondrial apoptosis remained unknown. To investigate the possible effects and underlying mechanism of T3 in silica-induced macrophage damage, differentiated human acute monocytic leukemia cells (THP-1) were exposed to different silica concentrations (0, 50, 100, 200, and 400 μg/mL) for 24 h. Additionally, silica-activated THP-1 macrophages were treated with gradient-dose T3 (0, 5, 10, 20, and 40 nM) for 24 h. To illuminate the potential mechanism, we used short hairpin RNA to knock down the thyroid hormone receptor α (TRα) in the differentiated THP-1 macrophages. The results showed that T3 decreased lactate dehydrogenase and reactive oxygen species levels, while increasing cell viability and superoxide dismutase in silica-induced THP-1 macrophages. In addition, silica increased the expression of interleukin 1 beta (IL-1β), interleukin 6 (IL-6), and tumor necrosis factor-α (TNF-α), and T3 treatment reduced those pro-inflammatory cytokines secretion. Compared with silica-alone treated groups, cells treated with silica and T3 restored the mitochondrial membrane potential loss and had reduced levels of cytochrome c and cleaved caspase-3 expressions. Lastly, we observed that TRα-knockdown inhibited the protective effects of T3 silica-induced THP-1 macrophages. Together, these findings revealed that T3 could serve as a potential therapeutic target for protection against silica-induced oxidative stress, inflammatory response, and mitochondrial apoptosis, which are mediated by the activation of the T3/TRα signal pathway.

The Role of CTGF in Inflammatory Responses Induced by Silica Particles in Human Bronchial Epithelial Cells.

Prolonged exposure to crystalline silica leads to persistent pulmonary inflammation and progressive fibrosis. Connective tissue growth factor (CTGF) has emerged as a potent proinflammatory and profibrotic regulator to participate in a variety of chronic inflammatory diseases. However, the role of CTGF in silica-induced pulmonary inflammation remains poorly understood. To explore the effect of CTGF on inflammatory responses caused by silica particles, human bronchial epithelial cells (16HBE) were transfected with CTGF siRNA and exposed to silica particles at concentrations of 0, 12.5, 25, 50, 100μg/ml for 48h. Intracellular CTGF mRNA and protein expressions were determined by RT-PCR and Western blotting, respectively. The levels of inflammatory cytokines including IL-8, TNF-α, IL-6, IL-1β, IL-17A and TGF-β1 were measured by ELISA kits. Silica particles induce significantly elevated intracellular CTGF mRNA expression in 16HBE cells in a dose-dependent manner when compared with blank control group (P < 0.05). The secretions of IL-8, TNF-α, IL-6 and IL-17A were also significantly increased by silica particles (P < 0.05). After exposure to 25 or 50μg/ml silica particles, the expression of intracellular CTGF mRNA was significantly inhibited in 16HBE cells when transfected with CTGF siRNA (P < 0.05). The secreted levels of IL-8, TNF-α, IL-6 and IL-17A induced by silica particles were also significantly lower from CTGF siRNA-transfected cells than that from normal 16HBE cells (P < 0.05). Inhibition of CTGF gene attenuates silica-induced inflammatory responses in bronchial epithelial cells, suggesting that CTGF could be a pivotal regulator in the development of silica-induced inflammation.