NLRP3 inflammasome activation determines the fibrogenic potential of PM(2.5) air pollution particles in the lung.

Airborne fine particulate matter (PM2.5) is known to cause respiratory inflammation such as chronic obstructive pulmonary disease and lung fibrosis. NLRP3 inflammasome activation has been implicated in these diseases; however, due to the complexity in PM2.5 compositions, it is difficult to differentiate the roles of the components in triggering this pathway. We collected eight real-life PM2.5 samples for a comparative analysis of their effects on NLRP3 inflammasome activation and lung fibrosis. In vitro assays showed that although the PM2.5 particles did not induce significant cytotoxicity at the dose range of 12.5 to 100 µg/mL, they induced potent TNF-α and IL-1β production in PMA differentiated THP-1 human macrophages and TGF-β1 production in BEAS-2B human bronchial epithelial cells. At the dose of 100 µg/mL, PM2.5 induced NLRP3 inflammasome activation by inducing lysosomal damage and cathepsin B release, leading to IL-1β production. This was confirmed by using NLRP3- and ASC-deficient cells as well as a cathepsin B inhibitor, ca-074 ME. Administration of PM2.5 via oropharyngeal aspiration at 2 mg/kg induced significant TGF-β1 production in the bronchoalveolar lavage fluid and collagen deposition in the lung at 21 days post-exposure, suggesting PM2.5 has the potential to induce pulmonary fibrosis. The ranking of in vitro IL-1β production correlates well with the in vivo total cell count, TGF-β1 production, and collagen deposition. In summary, we demonstrate that the PM2.5 is capable of inducing NLRP3 inflammasome activation, which triggers a series of cellular responses in the lung to induce fibrosis.

Cao W ，Wang X ，Li J ，Yan M ，Chang CH ，Kim J ，Jiang J ，Liao YP ，Tseng S ，Kusumoputro S ，Lau C ，Huang M ，Han P ，Lu P ，Xia T ... -  《Journal of Environmental Sciences》

NLRP3 inflammasome activation and lung fibrosis caused by airborne fine particulate matter.

Airborne fine particulate matter (PM2.5) has been known capable of causing lung inflammation and fibrosis, as a result of a series of chronic respiration diseases. Although NLRP3 inflammasome activation is essential for development of many chronic diseases, the relationship between PM2.5-induced toxicological effect and NLRP3 inflammasome activation is rarely investigated. Since PM2.5 contains a large population of nanosized materials and many types of nanomaterials can activate NLRP3 inflammasome, the NLRP3 inflammasome activation and lung fibrosis induced by PM2.5 were investigated in the present study. PM2.5 was found capable of causing weak cell death but potent IL-1β secretion in THP-1 cells, which was involved in NLRP3 inflammasome activation as evidenced by Z-YVAD-FMK inhibited IL-1β secretion and overexpressed ASC and NLRP3 protein in PM2.5 treated cells. PM2.5 could be internalized into cells through multiple endocytosis processes, such as phagocytosis and pinocytosis (macropinocytosis, clathrin- and caveolin-mediated endocytosis), and activate NLRP3 inflammasome through cathepsin B release, ROS production, and potassium efflux. After 21 days of exposure to PM2.5 through oropharyngeal aspiration, Balb/c mice showed increased IL-1β and TGF-β1 levels in the bronchoalveolar lavage fluid (BALF) of lung and significant collagen deposition around small airways of mice, suggesting potential lung inflammation and pulmonary fibrosis.

Zheng R ，Tao L ，Jian H ，Chang Y ，Cheng Y ，Feng Y ，Zhang H ... -  《-》

Molecular mechanisms underlying NLRP3 inflammasome activation and IL-1β production in air pollution fine particulate matter (PM(2.5))-primed macrophages.

Exposure to air pollution fine particulate matter (PM2.5) aggravates respiratory and cardiovascular diseases. It has been proposed that PM2.5 uptake by alveolar macrophages promotes local inflammation that ignites a systemic response, but precise underlying mechanisms remain unclear. Here, we demonstrate that PM2.5 phagocytosis leads to NLRP3 inflammasome activation and subsequent release of the pro-inflammatory master cytokine IL-1β. Inflammasome priming and assembly was time- and dose-dependent in inflammasome-reporter THP-1-ASC-GFP cells, and consistent across PM2.5 samples of variable chemical composition. While inflammasome activation was promoted by different PM2.5 surrogates, significant IL-1β release could only be observed after stimulation with transition-metal rich Residual Oil Fly Ash (ROFA) particles. This effect was confirmed in primary human monocyte-derived macrophages and murine bone marrow-derived macrophages (BMDMs), and by confocal imaging of inflammasome-reporter ASC-Citrine BMDMs. IL-1β release by ROFA was dependent on the NLRP3 inflammasome, as indicated by lack of IL-1β production in ROFA-exposed NLRP3-deficient (Nlrp3-/-) BMDMs, and by specific NLRP3 inhibition with the pharmacological compound MCC950. In addition, while ROFA promoted the upregulation of pro-inflammatory gene expression and cytokines release, MCC950 reduced TNF-α, IL-6, and CCL2 production. Furthermore, inhibition of TNF-α with a neutralizing antibody decreased IL-1β release in ROFA-exposed BMDMs. Using electron tomography, ROFA particles were observed inside intracellular vesicles and mitochondria, which showed signs of ultrastructural damage. Mechanistically, we identified lysosomal rupture, K+ efflux, and impaired mitochondrial function as important prerequisites for ROFA-mediated IL-1β release. Interestingly, specific inhibition of superoxide anion production (O2•-) from mitochondrial respiratory Complex I, but not III, blunted IL-1β release in ROFA-exposed BMDMs. Our findings unravel the mechanism by which PM2.5 promotes IL-1β release in macrophages and provide a novel link between innate immune response and exposure to air pollution PM2.5.

Caceres L ，Abogunloko T ，Malchow S ，Ehret F ，Merz J ，Li X ，Sol Mitre L ，Magnani N ，Tasat D ，Mwinyella T ，Spiga L ，Suchanek D ，Fischer L ，Gorka O ，Colin Gissler M ，Hilgendorf I ，Stachon P ，Rog-Zielinska E ，Groß O ，Westermann D ，Evelson P ，Wolf D ，Marchini T ... -  《-》

Resveratrol alleviates chronic "real-world" ambient particulate matter-induced lung inflammation and fibrosis by inhibiting NLRP3 inflammasome activation in mice.

Inhalation of fine particulate matter (PM2.5) induces the occurrence of lung inflammation and fibrosis, but its molecular mechanism remains unclear. Resveratrol (RES) is known to have anti-inflammatory properties in many pulmonary diseases. Here, we aimed to investigate the effect of long-term "real-world" ambient PM exposure on lung inflammation and fibrosis and further explore the protective effect and mechanism of RES. RES (50 and 100 mg/kg.bw) was administered to C57BL/6J mice that were exposed to ambient PM for 5 months. The control group breathed filtered air without RES, and the PM group was exposed to PM without RES. The inflammatory cytokine levels in bronchoalveolar lavage fluid (BALF) and lung fibrosis were evaluated by enzyme-linked immune sorbent assay (ELISA) kits and Masson's trichrome staining. The real-time PCR and Western blot analysis were used to determine the signal pathway. In vivo, PM exposure markedly elevated the levels of inflammatory cytokines and TGF-β1 in BALF, induced lung fibrosis. Meanwhile, PM exposure triggered autophagy process and activated the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome in lung. Also, RES treatment abolished PM-induced lung inflammation and fibrosis, and inhibited autophagic process and NLRP3 inflammasome activation. In vitro, PM2.5-induced cytotoxicity in BEAS-2B cells dose-dependently. Besides, RES alleviated PM2.5-induced cytotoxicity, inhibited autophagic process and NLRP3 inflammasome activity and decreased IL-1β production in BEAS-2B cells. Long-term PM exposure induced lung inflammation and fibrosis, and RES intervention alleviated these adverse effects via inhibiting autophagy-related NLRP3 inflammasome activation.

Ding S ，Wang H ，Wang M ，Bai L ，Yu P ，Wu W ... -  《-》

High glucose enhances the activation of NLRP3 inflammasome by ambient fine particulate matter in alveolar macrophages.

Epidemiological studies have demonstrated that individuals with preexisting conditions, including diabetes mellitus (DM), are more susceptible to air pollution. However, the underlying mechanisms remain unclear. In this study, we proposed that a high glucose setting enhances ambient fine particulate matter (PM2.5)-induced macrophage activation and secretion of the proinflammatory cytokine, IL-1β, through activation of the NLRP3 inflammasome, altering the balance between matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs). Exposure of mouse alveolar macrophages to non-cytotoxic doses of PM2.5 led to upregulation of IL-1β, activation of the NLRP3 inflammasome, increased nuclear translocation of the transcription factor NF-κB, increased generation of reactive oxygen species (ROS), and increased expression and enzymatic activity of MMP-9; these effects were enhanced when cells were pretreated with high glucose. However, pretreatment in a high glucose setting alone did not induce significant changes. ROS generation following PM2.5 exposure was abolished when cells were pretreated with ROS scavengers such as Trolox and superoxide dismutase (SOD), or with an NADPH oxidase inhibitor, DPI. Pretreatment of cells with DPI attenuated the effects of a high glucose setting on PM2.5-induced upregulation of IL-1β, activation of the NLRP3 inflammasome, and nuclear translocation of NF-κB. In addition, enhancement of PM2.5-induced expression and enzymatic activity of MMP-9 following high glucose pretreatment was not observed in primary alveolar macrophages obtained from NLRP3 or IL-1R1 knockout (KO) mice, where pro-IL-1β cannot be cleaved to IL-1β or cells are insensitive to IL-1β, respectively. This study demonstrated that exposure of mouse alveolar macrophages to PM2.5 in a high glucose setting enhanced PM2.5-induced production of IL-1β through activation of the NLRP3 inflammasome and nuclear translocation of NF-κB due to PM2.5-induced oxidative stress, leading to MMP-9 upregulation. The key role of NADPH oxidase in PM2.5-induced ROS generation and activation of the IL-1β secretion pathway and the importance of IL-1β secretion and signaling in PM2.5-induced increases in MMP-9 enzymatic activity were also demonstrated. This study provides a further understanding of the potential mechanisms underlying the susceptibility of individuals with DM to air pollution and suggests potential therapeutic targets.

Mo Y ，Mo L ，Zhang Y ，Zhang Y ，Yuan J ，Zhang Q ... -  《-》