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Alveolar epithelial cell (AEC) necroptosis is critical to disrupt the alveolar barrier and provoke acute lung injury (ALI). Here, we define calcitonin gene-related peptide (CGRP), the most abundant endogenous neuropeptide in the lung, as a novel modulator of AEC necroptosis in lipopolysaccharide (LPS)-induced ALI. Upon LPS-induced ALI, overexpression of Cgrp significantly mitigates the inflammatory response, alleviates lung tissue damage, and decreases AEC necroptosis. Similarly, CGRP alleviated AEC necroptosis under the LPS challenge in vitro. Previously, we identified that long optic atrophy 1 (L-OPA1) deficiency mediates mitochondrial fragmentation, leading to AEC necroptosis. In this study, we discovered that CGRP positively regulated mitochondrial fusion through stabilizing L-OPA1. Mechanistically, we elucidate that CGRP activates AMP-activated protein kinase (AMPK). Furthermore, the blockade of AMPK compromised the protective effect of CGRP against AEC necroptosis following the LPS challenge. Our study suggests that CRGP-mediated activation of the AMPK/L-OPA1 axis may have potent therapeutic benefits for patients with ALI or other diseases with necroptosis.
Assuntos
Lesão Pulmonar Aguda , Animais , Masculino , Camundongos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/tratamento farmacológico , Células Epiteliais Alveolares/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Proteínas Quinases Ativadas por AMP/metabolismo , Peptídeo Relacionado com Gene de Calcitonina/genética , Peptídeo Relacionado com Gene de Calcitonina/farmacologia , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Linhagem Celular , GTP Fosfo-Hidrolases/metabolismo , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/metabolismo , Pulmão/metabolismo , Camundongos Endogâmicos C57BL , Necroptose , Transdução de SinaisRESUMO
Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity, mortality, and health care use worldwide with heterogeneous pathogenesis. Mitochondria, the powerhouses of cells responsible for oxidative phosphorylation and energy production, play essential roles in intracellular material metabolism, natural immunity, and cell death regulation. Therefore, it is crucial to address the urgent need for fine-tuning the regulation of mitochondrial quality to combat COPD effectively. Mitochondrial quality control (MQC) mainly refers to the selective removal of damaged or aging mitochondria and the generation of new mitochondria, which involves mitochondrial biogenesis, mitochondrial dynamics, mitophagy, etc. Mounting evidence suggests that mitochondrial dysfunction is a crucial contributor to the development and progression of COPD. This article mainly reviews the effects of MQC on COPD as well as their specific regulatory mechanisms. Finally, the therapeutic approaches of COPD via MQC are also illustrated.
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Mitocôndrias , Doença Pulmonar Obstrutiva Crônica , Humanos , Mitocôndrias/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Envelhecimento , MitofagiaRESUMO
BACKGROUND: Necroptosis of macrophages is a necessary element in reinforcing intrapulmonary inflammation during acute lung injury (ALI). However, the molecular mechanism that sparks macrophage necroptosis is still unclear. Triggering receptor expressed on myeloid cells-1 (TREM-1) is a pattern recognition receptor expressed broadly on monocytes/macrophages. The influence of TREM-1 on the destiny of macrophages in ALI requires further investigation. METHODS: TREM-1 decoy receptor LR12 was used to evaluate whether the TREM-1 activation induced necroptosis of macrophages in lipopolysaccharide (LPS)-induced ALI in mice. Then we used an agonist anti-TREM-1 Ab (Mab1187) to activate TREM-1 in vitro. Macrophages were treated with GSK872 (a RIPK3 inhibitor), Mdivi-1 (a DRP1 inhibitor), or Rapamycin (an mTOR inhibitor) to investigate whether TREM-1 could induce necroptosis in macrophages, and the mechanism of this process. RESULTS: We first observed that the blockade of TREM-1 attenuated alveolar macrophage (AlvMs) necroptosis in mice with LPS-induced ALI. In vitro, TREM-1 activation induced necroptosis of macrophages. mTOR has been previously linked to macrophage polarization and migration. We discovered that mTOR had a previously unrecognized function in modulating TREM-1-mediated mitochondrial fission, mitophagy, and necroptosis. Moreover, TREM-1 activation promoted DRP1Ser616 phosphorylation through mTOR signaling, which in turn caused surplus mitochondrial fission-mediated necroptosis of macrophages, consequently exacerbating ALI. CONCLUSION: In this study, we reported that TREM-1 acted as a necroptotic stimulus of AlvMs, fueling inflammation and aggravating ALI. We also provided compelling evidence suggesting that mTOR-dependent mitochondrial fission is the underpinning of TREM-1-triggered necroptosis and inflammation. Therefore, regulation of necroptosis by targeting TREM-1 may provide a new therapeutic target for ALI in the future.
Assuntos
Lesão Pulmonar Aguda , Lipopolissacarídeos , Animais , Camundongos , Receptor Gatilho 1 Expresso em Células Mieloides , Lipopolissacarídeos/farmacologia , Dinâmica Mitocondrial , Necroptose , Serina-Treonina Quinases TOR , Macrófagos , InflamaçãoRESUMO
Necroptosis, a recently described form of programmed cell death, is the main way of alveolar epithelial cells (AECs) death in acute lung injury (ALI). While the mechanism of how to trigger necroptosis in AECs during ALI has been rarely evaluated. Long optic atrophy protein 1 (L-OPA1) is a crucial mitochondrial inner membrane fusion protein, and its deficiency impairs mitochondrial function. This study aimed to investigate the role of L-OPA1 deficiency-mediated mitochondrial dysfunction in AECs necroptosis. We comprehensively investigated the detailed contribution and molecular mechanism of L-OPA1 deficiency in AECs necroptosis by inhibiting or activating L-OPA1. First, our data showed that L-OPA1 expression was downregulated in the lungs and AECs under the lipopolysaccharide (LPS) challenge. Furthermore, inhibition of L-OPA1 aggravated the pathological injury, inflammatory response, and necroptosis in the lungs of LPS-induced ALI mice. In vitro, inhibition of L-OPA1 induced necroptosis of AECs, while activation of L-OPA1 alleviated necroptosis of AECs under the LPS challenge. Mechanistically, inhibition of L-OPA1 aggravated necroptosis of AECs by inducing mitochondrial fragmentation and reducing mitochondrial membrane potential. While activation of L-OPA1 had the opposite effects. In summary, these findings indicate for the first time that L-OPA1 deficiency mediates mitochondrial fragmentation, induces necroptosis of AECs, and exacerbates ALI in mice.
Assuntos
Lesão Pulmonar Aguda , Células Epiteliais Alveolares , GTP Fosfo-Hidrolases/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/metabolismo , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Animais , GTP Fosfo-Hidrolases/genética , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Mitocôndrias/metabolismo , NecroptoseRESUMO
BACKGROUND: Uncontrolled inflammation is an important factor in the occurrence and development of acute lung injury (ALI). Fibroblast growth factor-inducible 14 (Fn14), a plasma membrane-anchored receptor, takes part in the pathological process of a variety of acute and chronic inflammatory diseases. However, the role of Fn14 in ALI has not yet been elucidated. This study aimed to investigate whether the activation of Fn14 exacerbated lipopolysaccharide (LPS)-induced ALI in mice. METHODS: In vivo, ALI was induced by intratracheal LPS-challenge combined with/without Fn14 receptor blocker aurintricarboxylic acid (ATA) treatment in C57BL/6J mice. Following LPS administration, the survival rate, lung tissue injury, inflammatory cell infiltration, inflammatory factor secretion, oxidative stress, and NLRP3 inflammasome activation were assessed. In vitro, primary murine macrophages were used to evaluate the underlying mechanism by which Fn14 activated the NLRP3 inflammasome. Lentivirus was used to silence Fn14 to observe its effect on the activation of NLRP3 inflammasome in macrophages. RESULTS: In this study, we found that Fn14 expression was significantly increased in the lungs of LPS-induced ALI mice. The inhibition of Fn14 with ATA downregulated the protein expression of Fn14 in the lungs and improved the survival rate of mice receiving a lethal dose of LPS. ATA also attenuated lung tissue damage by decreasing the infiltration of macrophages and neutrophils, reducing inflammation, and suppressing oxidative stress. Importantly, we found that ATA strongly inhibited the activation of NLRP3 inflammasome in the lungs of ALI mice. Furthermore, in vitro, TWEAK, a natural ligand of Fn14, amplified the activation of NLRP3 inflammasome in the primary murine macrophage. By contrast, inhibition of Fn14 with shRNA decreased the expression of Fn14, NLRP3, Caspase-1 p10, and Caspase-1 p20, and the production of IL-1ß and IL-18. Furthermore, the activation of Fn14 promoted the production of reactive oxygen species and inhibited the activation of Nrf2-HO-1 in activated macrophages. CONCLUSIONS: Our study first reports that the activation of Fn14 aggravates ALI by amplifying the activation of NLRP3 inflammasome. Therefore, blocking Fn14 may be a potential way to treat ALI.
Assuntos
Lesão Pulmonar Aguda , Inflamassomos , Receptor de TWEAK/metabolismo , Lesão Pulmonar Aguda/patologia , Animais , Caspase 1/metabolismo , Inflamassomos/metabolismo , Inflamação/metabolismo , Lipopolissacarídeos/farmacologia , Pulmão , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismoRESUMO
Idiopathic pulmonary fibrosis (IPF) is a progressive disease characterized by excessive deposition of extracellular matrix (ECM) and chronic inflammation with limited therapeutic options. Psoralen, a major active component extracted from Psoralea corylifolia L. seed, has several biological effects. However, the role of psoralen in IPF is still unclear. Here, we hypothesized that psoralen played an essential role in IPF in the inhibition of fibroblast proliferation and inflammatory response. A murine model of IPF was established by injecting bleomycin (BLM) intratracheally, and psoralen was administered for 14 days from the 7th to 21st day after BLM injection. Our results demonstrated that psoralen treatment reduced body weight loss and improved the survival rate of mice with IPF. Histological and immunofluorescent examination showed that psoralen alleviated BLM-induced lung parenchymal inflammatory and fibrotic alteration. Furthermore, psoralen inhibited proliferation and collagen synthesis of mouse fibroblasts and partially reversed BLM-induced expression of α-smooth muscle actin at both the tissue and cell level. Moreover, psoralen decreased the expression of transforming growth factor-ß1, interleukin-1ß, and tumor necrosis factor-α in the lungs of BLM-stimulated mice. Our results reveale for the first time that psoralen exerts therapeutic effects against IPF in a BLM-induced murine model.
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Gluconic metabolic reprogramming, immune response, and inflammation are intimately linked. Glycolysis involves in the pathologic progress in acute and chronic inflammatory diseases. However, the involvement of glycolysis in the acute lung injury (ALI) is still unclear. This study investigated the role of glycolysis in an animal model of ALI. First, we found that lactate content in serum was remarkably increased in ALI patients and a murine model induced by intratracheal administration of lipopolysaccharide (LPS). The key proteins involving in glycolysis were robustly elevated, including HK2, PKM2, and HIF-1α. Intriguingly, inhibition of glycolysis by 2-deoxyglucose (2-DG) pronouncedly attenuated the lung tissue pathological injury, accumulation of neutrophil, oxidative stress, expression of proinflammatory factors in the lung of ALI mice induced by LPS. The 2-DG treatment also strongly suppressed the activation of the NOD-like receptor (NLR) family and pyrin domain-containing protein 3 (NLRP3) inflammasome. Furthermore, we investigated the role of glycolysis in the inflammatory response of primary murine macrophages activated by LPS in vitro. We found that the 2-DG treatment remarkably reduced the expression of proinflammatory factors induced by LPS, including tumor necrosis factor-α messenger RNA (mRNA), pro-interleukin (IL)-1ß mRNA, pro-IL-18 mRNA, NLRP3 mRNA, caspase-1 mRNA, and IL-1ß protein. Altogether, these data provide a novel link between gluconic metabolism reprogramming and uncontrolled inflammatory response in ALI. This study suggests glycolytic inhibition as an effective anti-inflammatory strategy in treating ALI.
Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Anti-Inflamatórios/farmacologia , Desoxiglucose/farmacologia , Glicólise/efeitos dos fármacos , Lipopolissacarídeos , Pulmão/efeitos dos fármacos , Macrófagos Peritoneais/efeitos dos fármacos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/patologia , Animais , Estudos de Casos e Controles , Células Cultivadas , Modelos Animais de Doenças , Humanos , Mediadores da Inflamação/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Macrófagos Peritoneais/metabolismo , Macrófagos Peritoneais/patologia , Masculino , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Infiltração de Neutrófilos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Fatores de TempoRESUMO
BACKGROUND: Breast lesions classified as BI-RADS-US 3 are probably benign and observation was recommended, while a considerable number of BI-RADS-US 4 lesions were benign, resulting in excessive biopsies. We focus exclusively on BI-RADS-US 3 and 4 lesions and hypothesize that improved diagnostic performance can be achieved by integrating real-time elastography (strain ratio) into the BI-RADS-US classification system. METHOD: From April 2010 to September 2015, 1071 lesions were included in the final analysis. After the conventional ultrasound examination, the BI-RADS-US (2013) classification was used to evaluate the lesions. Then the strain ratios were calculated, and the final diagnosis was made on the basis of histological results. The sensitivity, specificity, accuracy, PPV and NPV were calculated and the AUCs were compared. Additionally, an analysis of the diagnostic performance expressed by the pretest and posttest probability of disease (POD) was performed in BI-RADS-US 3 and 4A lesions. RESULTS: With the cutoff point of 2.98, the sensitivity, specificity and accuracy of the strain ratio method were 86.9â%, 86.6â% and 82.6â%, respectively. In BI-RADS-US 3 lesions, a suspicious strain ratio significantly modified the POD from 1.3â% to a posttest POD of 29.8â%. In BI-RADS-US 4A lesions, a suspicious strain ratio significantly modified the POD from 8.5â% to a posttest POD of 48.7â%. CONCLUSION: Ultrasonographic elastography (strain ratio) yields additional diagnostic information in the evaluation of BI-RADS-US 3 and 4 breast lesions. The strain ratios should be integrated into the BI-RADS-US classification system and into daily practice.
Assuntos
Neoplasias da Mama , Técnicas de Imagem por Elasticidade , Neoplasias da Mama/diagnóstico por imagem , Diagnóstico Diferencial , Feminino , Humanos , Sensibilidade e Especificidade , Ultrassonografia MamáriaRESUMO
The activation of NLRP3 inflammasome in microglia is critical for neuroinflammation during postoperative cognitive dysfunction (POCD) induced by sevoflurane. However, the molecular mechanism by which sevoflurane activates the NLRP3 inflammasome in microglia remains unclear. The cGAS-STING pathway is an evolutionarily conserved inflammatory defense mechanism. The role of the cGAS-STING pathway in sevoflurane-induced NLRP3 inflammasome-dependent neuroinflammation and the underlying mechanisms require further investigation. We found that prolonged anesthesia with sevoflurane induced cognitive dysfunction and triggered the neuroinflammation characterized by the activation of NLRP3 inflammasome in vivo. Interestingly, the cGAS-STING pathway was activated in the hippocampus of mice receiving sevoflurane. While the blockade of cGAS with RU.521 attenuated cognitive dysfunction and NLRP3 inflammasome activation in mice. In vitro, we found that sevoflurane treatment significantly activated the cGAS-STING pathway in microglia, while RU.521 pre-treatment robustly inhibited sevoflurane-induced NLRP3 inflammasome activation. Mechanistically, sevoflurane-induced mitochondrial fission in microglia and released mitochondrial DNA (mtDNA) into the cytoplasm, which could be abolished with Mdivi-1. Blocking the mtDNA release via the mPTP-VDAC channel inhibitor attenuated sevoflurane-induced mtDNA cytosolic escape and reduced cGAS-STING pathway activation in microglia, finally inhibiting the NLRP3 inflammasome activation. Therefore, regulating neuroinflammation by targeting the cGAS-STING pathway may provide a novel therapeutic target for POCD.
Assuntos
Inflamassomos , Complicações Cognitivas Pós-Operatórias , Camundongos , Animais , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , DNA Mitocondrial/metabolismo , Sevoflurano , Doenças Neuroinflamatórias , Nucleotidiltransferases/metabolismoRESUMO
The triggering receptor expressed on myeloid cells-1 (TREM-1) is a pro-inflammatory immune receptor potentiating acute lung injury (ALI). However, the mechanism of TREM-1-triggered inflammation response remains poorly understood. Here, we showed that TREM-1 blocking attenuated NOD-, LRR- and pyrin domain-containing 3 (NLRP3) inflammasome activation and glycolysis in LPS-induced ALI mice. Then, we observed that TREM-1 activation enhanced glucose consumption, induced glycolysis, and inhibited oxidative phosphorylation in macrophages. Specifically, inhibition of glycolysis with 2-deoxyglucose diminished NLRP3 inflammasome activation of macrophages triggered by TREM-1. Hypoxia-inducible factor-1α (HIF-1α) is a critical transcriptional regulator of glycolysis. We further found that TREM-1 activation facilitated HIF-1α accumulation and translocation to the nucleus via the phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway. Inhibiting mTOR or HIF-1α also suppressed TREM-1-induced metabolic reprogramming and NLRP3/caspase-1 activation. Overall, the mTOR/HIF-1α/glycolysis pathway is a novel mechanism underlying TREM-1-governed NLRP3 inflammasome activation. Therapeutic targeting of the mTOR/HIF-1α/glycolysis pathway in TREM-1-activated macrophages could be beneficial for treating or preventing inflammatory diseases, such as ALI.
Assuntos
Lesão Pulmonar Aguda , Inflamassomos , Animais , Camundongos , Receptor Gatilho 1 Expresso em Células Mieloides/metabolismo , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Fosfatidilinositol 3-Quinases/metabolismo , Camundongos Endogâmicos NOD , Macrófagos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/metabolismo , Glicólise , Lipopolissacarídeos , Camundongos Endogâmicos C57BL , Mamíferos/metabolismoRESUMO
Alveolar epithelial cell (AEC) senescence is implicated in the pathogenesis of pulmonary fibrosis (PF). However, the exact mechanism underlying AEC senescence during PF remains poorly understood. Here, we reported an unrecognized mechanism for AEC senescence during PF. We found that, in bleomycin (BLM)-induced PF mice, the expressions of isocitrate dehydrogenase 3α (Idh3α) and citrate carrier (CIC) were significantly down-regulated in the lungs, which could result in mitochondria citrate (citratemt) accumulation in our previous study. Notably, the down-regulation of Idh3α and CIC was related to senescence. The mice with AECs-specific Idh3α and CIC deficiency by adenoviral vector exhibited spontaneous PF and senescence in the lungs. In vitro, co-inhibition of Idh3α and CIC with shRNA or inhibitors triggered the senescence of AECs, indicating that accumulated citratemt triggers AEC senescence. Mechanistically, citratemt accumulation impaired the mitochondrial biogenesis of AECs. In addition, the senescence-associated secretory phenotype from senescent AECs induced by citratemt accumulation activated the proliferation and transdifferentiation of NIH3T3 fibroblasts into myofibroblasts. In conclusion, we show that citratemt accumulation would be a novel target for protection against PF that involves senescence.
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Alveolar epithelial cell (AEC) senescence is a key driver of a variety of chronic lung diseases. It remains a challenge how to alleviate AEC senescence and mitigate disease progression. Our study identified a critical role of epoxyeicosatrienoic acids (EETs), downstream metabolites of arachidonic acid (ARA) by cytochrome p450 (CYP), in alleviating AEC senescence. In vitro, we found that 14,15-EET content was significantly decreased in senescent AECs. Exogenous EETs supplementation, overexpression of CYP2J2, or inhibition of EETs degrading enzyme soluble epoxide hydrolase (sEH) to increase EETs alleviated AECs' senescence. Mechanistically, 14,15-EET promoted the expression of Trim25 to ubiquitinate and degrade Keap1 and promoted Nrf2 to enter the nucleus to exert an anti-oxidant effect, thereby inhibiting endoplasmic reticulum stress (ERS) and alleviating AEC senescence. Furthermore, in D-galactose (D-gal)-induced premature aging mouse model, inhibiting the degradation of EETs by Trifluoromethoxyphenyl propionylpiperidin urea (TPPU, an inhibitor of sEH) significantly inhibited the protein expression of p16, p21, and γH2AX. Meanwhile, TPPU reduced the degree of age-related pulmonary fibrosis in mice. Our study has confirmed that EETs are novel anti-senescence substances for AECs, providing new targets for the treatment of chronic lung diseases.
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Células Epiteliais Alveolares , Senescência Celular , Eicosanoides , Estresse do Retículo Endoplasmático , Fator 2 Relacionado a NF-E2 , Animais , Camundongos , Células Epiteliais Alveolares/efeitos dos fármacos , Células Epiteliais Alveolares/fisiologia , Eicosanoides/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Proteína 1 Associada a ECH Semelhante a Kelch , Fator 2 Relacionado a NF-E2/genética , Fibrose Pulmonar , Senescência Celular/efeitos dos fármacosRESUMO
Alveolar epithelial cell (AEC) senescence is considered to be a universal pathological feature of many chronic pulmonary diseases. Our previous study found that epoxyeicosatrienoic acids (EETs), produced from arachidonic acid (ARA) through the cytochrome P450 cyclooxygenase (CYP) pathway, have significant negative regulatory effects on cellular senescence in AECs. However, the exact mechanisms by which EETs alleviate the senescence of AECs still need to be further explored. In the present study, we observed that bleomycin (BLM) induced enhanced mitophagy accompanied by increased mitochondrial ROS (mito-ROS) content in the murine alveolar epithelial cell line MLE12. While EETs reduced BLM-induced mitophagy and mito-ROS content in MLE12 cells, and the mechanism was related to the regulation of NOX4/Nrf2-mediated redox imbalance. Furthermore, we found that inhibition of EETs degradation could significantly inhibit mitophagy and regulate NOX4/Nrf2 balance to exert anti-oxidant effects in D-galactose-induced premature aging mice. Collectively, these findings may provide new ideas for treating age-related pulmonary diseases by targeting EETs to improve mitochondrial dysfunction and reduce oxidative stress.
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Células Epiteliais Alveolares , Pneumopatias , Camundongos , Animais , Células Epiteliais Alveolares/metabolismo , Mitofagia , Fator 2 Relacionado a NF-E2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Senescência CelularRESUMO
Our previous study showed that triggering receptors expressed on myeloid cell-1 (TREM-1) was upregulated in bleomycin (BLM)-induced pulmonary fibrosis (PF) mouse model. However, the role of TREM-1 in the development of PF and its underlying mechanism remain unclear. Herein, we report that the prophylactical blockade of TREM-1 using a decoy peptide dodecapeptide (LR12) exerted protective effects against BLM-induced PF in mice, with a higher survival rate, attenuated tissue injury, and less extracellular matrix deposition. Interestingly, therapeutic blockade of TREM-1 at the early stage of fibrosis also attenuated BLM-induced PF, suggesting a non-inflammatory effect. More importantly, we observed that TREM-1 blockade with LR12 significantly reduced the expression of the senescence-relative protein, including p16, p21, p53, and γ-H2AX in the lungs of PF mice. Notably, TREM-1 was upregulated in alveolar epithelial cells (AECs) and correlated with the levels of senescence markers in BLM-treated mice. In vitro, activating TREM-1 with an agonistic antibody exacerbated BLM-induced senescence in MLE12 cells, a murine AEC cell line. Furthermore, prophylactic or therapeutic blockade of TREM-1 protected MLE12 cells from senescence induced by BLM or H2O2. In conclusion, our findings elucidate a pro-fibrotic effect of TREM-1 by inducing AECs senescence in PF, providing a potential strategy for fibrotic disease treatment.
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Células Epiteliais Alveolares , Fibrose Pulmonar , Receptor Gatilho 1 Expresso em Células Mieloides , Animais , Camundongos , Células Epiteliais Alveolares/patologia , Bleomicina/toxicidade , Peróxido de Hidrogênio/metabolismo , Células Mieloides , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/fisiopatologia , Receptor Gatilho 1 Expresso em Células Mieloides/metabolismoRESUMO
RATIONALE: Alveolar epithelial cell death, inflammation, and oxidative stress are typical features of acute lung injury (ALI). Aloperine (Alo), an alkaloid isolated from Sophora alopecuroides, has been reported to display various biological effects, such as anti-inflammatory, immunoregulatory, and anti-oxidant properties. In this study, we investigated the effects and mechanisms of Alo in treating a lipopolysaccharide (LPS)-induced ALI in a murine model. METHODS: The effects of Alo in LPS-induced ALI were investigated in C57BL/6 mice. The RIPK1 inhibitor (Nec-1) and the RIPK3 inhibitor (GSK'872) were used to evaluate the relationship of necroptosis, NF-κB activation, and PDC subunits in LPS-treated mouse alveolar epithelial cells (MLE-12). Then the effects of Alo on necroptosis, inflammation, and oxidative stress of LPS-stimulated MLE-12 cells were evaluated. RESULTS: Alo significantly attenuated histopathological lung injuries and reduced lung wet/dry ratio in LPS-induced ALI mice. Alo also remarkedly reduced total protein and neutrophils recruitment in bronchoalveolar lavage fluid of ALI mice. Meanwhile, Alo ameliorated the LPS-induced necroptosis in the lungs of ALI mice. The RIPK3 inhibitor GSK'872, but not the RIPK1 inhibitor Nec-1, reversed LPS-induced p65 phosphorylation and translocation to the nucleus in MLE-12 cells. GSK'872 also reversed the LPS-induced increase in ROS and binding of RIPK3 and PDC subunits in MLE-12 cells. Moreover, Alo down-regulated the levels of p-RIPK1, p-RIPK3, p-MLKL, p-p65, the translocation of p65 to the nucleus, and reduced the expression of IL-6 and IL-8 in LPS-stimulated MLE-12 cells. Alo also inhibited the binding of RIPK3 and PDC-E1α, PDC-E1ß, PDC-E2, and PDC-E3 and the ROS production in LPS-treated MLE-12 cells. CONCLUSION: The present study validated the beneficial effects of Alo on LPS-induced ALI , suggesting Alo may be a new drug candidate against ALI.
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Lesão Pulmonar Aguda , Lipopolissacarídeos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Animais , Modelos Animais de Doenças , Inflamação/tratamento farmacológico , Inflamação/patologia , Lipopolissacarídeos/farmacologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Necroptose , Estresse Oxidativo , Piperidinas/farmacologia , Quinolizidinas , Espécies Reativas de OxigênioRESUMO
Necroptosis is the major cause of death in alveolar epithelial cells (AECs) during acute lung injury (ALI). Here, we report a previously unrecognized mechanism for necroptosis. We found an accumulation of mitochondrial citrate (citratemt) in lipopolysaccharide (LPS)-treated AECs because of the downregulation of Idh3α and citrate carrier (CIC, also known as Slc25a1). shRNA- or inhibitor-mediated inhibition of Idh3α and Slc25a1 induced citratemt accumulation and necroptosis in vitro. Mice with AEC-specific Idh3α and Slc25a1 deficiency exhibited exacerbated lung injury and AEC necroptosis. Interestingly, the overexpression of Idh3α and Slc25a1 decreased citratemt levels and rescued AECs from necroptosis. Mechanistically, citratemt accumulation induced mitochondrial fission and excessive mitophagy in AECs. Furthermore, citratemt directly interacted with FUN14 domain-containing protein 1 (FUNDC1) and promoted the interaction of FUNDC1 with dynamin-related protein 1 (DRP1), leading to excessive mitophagy-mediated necroptosis and thereby initiating and promoting ALI. Importantly, necroptosis induced by citratemt accumulation was inhibited in FUNDC1-knockout AECs. We show that citratemt accumulation is a novel target for protection against ALI involving necroptosis.
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Lesão Pulmonar Aguda , Células Epiteliais Alveolares , Camundongos , Animais , Células Epiteliais Alveolares/metabolismo , Lipopolissacarídeos/efeitos adversos , Necroptose , Ácido Cítrico/efeitos adversos , Ácido Cítrico/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/genética , Proteínas Mitocondriais/metabolismo , Proteínas de Membrana/metabolismoRESUMO
Citrate has a prominent role as a substrate in cellular energy metabolism. Recently, citrate has been shown to drive inflammation. However, the role of citrate in lipopolysaccharide (LPS)-induced acute lung injury (ALI) remains unclear. Here, we aimed to clarify whether extracellular citrate aggravated the LPS-induced ALI and the potential mechanism. Our findings demonstrated that extracellular citrate aggravated the pathological lung injury induced by LPS in mice, characterized by up-regulation of pro-inflammatory factors and over-activation of NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome in the lungs. In vitro, we found that citrate treatment significantly augmented the expression of NLRP3 and pro-IL-1ß and enhanced the translocation of NF-κB/p65 into the nucleus. Furthermore, extracellular citrate plus adenosine-triphosphate (ATP) significantly increased the production of reactive oxygen species (ROS) in primary murine macrophages. Inhibiting the production of ROS with a ROS scavenger N-acetyl-L-cysteine (NAC) attenuated the activation of NLRP3 inflammasome. Altogether, we conclude that extracellular citrate may serve as a damage-associated molecular pattern (DAMP) and aggravates LPS-induced ALI by activating the NLRP3 inflammasome.
Assuntos
Alarminas/metabolismo , Ácido Cítrico/metabolismo , Lipopolissacarídeos/toxicidade , Lesão Pulmonar/induzido quimicamente , Ativação de Macrófagos/fisiologia , Macrófagos/efeitos dos fármacos , Trifosfato de Adenosina , Animais , Citocinas/genética , Citocinas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Lesão Pulmonar/metabolismo , Lesão Pulmonar/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Distribuição AleatóriaRESUMO
NOD-, LRR- and pyrin domain-containing 3 (NLRP3) inflammasome and triggering receptor expressed on myeloid cells-1 (TREM-1) are considered critical orchestrators of the inflammatory response in acute lung injury (ALI). However, few assumptions are based on the relationship between them. Here, we investigated the effect of NLRP3 inflammasome activation on the TREM-1 expression in lipopolysaccharide (LPS)-induced ALI and macrophages. We found that inhibition of the NLRP3 inflammasome reduced the TREM-1 expression and pathological lung injury in mice with ALI. Then, primary murine macrophages were used to dissect the underlying mechanistic events of the activation NLRP3 inflammasome involved in the TREM-1 expression. Our results demonstrated that the conditioned medium (CM) from NLRP3 inflammasome-activated-macrophages up-regulated the TREM-1 expression in macrophages, while this effect was reversed by an NLRP3 inflammasome inhibitor MCC950. Furthermore, neutralizing antibodies anti-IL-18 and anti-HMGB1 reduced the TREM-1 expression induced by NLRP3 inflammasome activation. Mechanistically, we found that CM from NLRP3 inflammasome-activated-macrophages increased the level of inhibitor κB kinase protein phosphorylation (p-IκBα) and reactive oxygen species (ROS) content, and promoted IκBα protein degradation in macrophages. While the inhibition of nuclear factor kappa-B (NF-κB) and scavenging ROS eliminated the up-regulation of TREM-1 induced by the NLRP3 inflammasome activation in macrophages. In summary, our study confers NLRP3 inflammasome as a new trigger of TREM-1 signing, which allows additional insight into the pathological of the inflammatory response in ALI.
Assuntos
Proteína HMGB1/metabolismo , Inflamassomos/metabolismo , Interleucina-18/metabolismo , Macrófagos Peritoneais/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Receptor Gatilho 1 Expresso em Células Mieloides/metabolismo , Animais , Células Cultivadas , Furanos , Regulação da Expressão Gênica/efeitos dos fármacos , Proteína HMGB1/genética , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Indenos , Lipopolissacarídeos/toxicidade , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pneumopatias/induzido quimicamente , Pneumopatias/metabolismo , Pneumopatias/patologia , Macrófagos Peritoneais/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Sulfonamidas , Sulfonas/farmacologia , Receptor Gatilho 1 Expresso em Células Mieloides/genética , Regulação para Cima/efeitos dos fármacosRESUMO
Pulmonary fibrosis (PF) is a senescence-associated disease with poor prognosis. Currently, there is no effective therapeutic strategy for preventing and treating the disease process. Mounting evidence suggests that arachidonic acid (ARA) metabolites are involved in the pathogenesis of various fibrosis. However, the relationship between the metabolism of ARA and PF is still elusive. In this study, we observed a disorder in the cyclooxygenase-2/cytochrome P450 (COX-2/CYP) metabolism of ARA in the lungs of PF mice induced by bleomycin (BLM). Therefore, we aimed to explore the role of COX-2/CYP-derived ARA metabolic disorders in PF. PTUPB, a dual COX-2 and soluble epoxide hydrolase (sEH) inhibitor, was used to restore the balance of COX-2/CYP metabolism. sEH is an enzyme hydrolyzing epoxyeicosatrienoic acids derived from ARA by CYP. We found that PTUPB alleviated the pathological changes in lung tissue and collagen deposition, as well as reduced senescence marker molecules (p16Ink4a and p53-p21Waf1/Cip1 ) in the lungs of mice treated by BLM. In vitro, we found that PTUPB pretreatment remarkably reduced the expression of senescence-related molecules in the alveolar epithelial cells (AECs) induced by BLM. In conclusion, our study supports the notion that the COX-2/CYP-derived ARA metabolic disorders may be a potential therapeutic target for PF via inhibiting the cellular senescence in AECs.
Assuntos
Envelhecimento/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Fibrose Pulmonar/tratamento farmacológico , Células A549 , Animais , Ácido Araquidônico/metabolismo , Bleomicina , Senescência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/administração & dosagem , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Epóxido Hidrolases/metabolismo , Humanos , Injeções Subcutâneas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Arachidonic acid can be metabolized to prostaglandins and epoxyeicosatrienoic acids (EETs) by cyclooxygenase-2 (COX-2) and cytochrome P450 (CYP), respectively. While protective EETs are degraded by soluble epoxide hydrolase (sEH) very fast. We have reported that dual inhibition of COX-2 and sEH with specific inhibitor PTUPB shows anti-pulmonary fibrosis and renal protection. However, the effect of PTUPB on cecal ligation and puncture (CLP)-induced sepsis remains unclear. The current study aimed to investigate the protective effects of PTUPB against CLP-induced sepsis in mice and the underlying mechanisms. We found that COX-2 expressions were increased, while CYPs expressions were decreased in the liver, lung, and kidney of mice undergone CLP. PTUPB treatment significantly improved the survival rate, reduced the clinical scores and systemic inflammatory response, alleviated liver and kidney dysfunction, and ameliorated the multiple-organ injury of the mice with sepsis. Besides, PTUPB treatment reduced the expression of hypoxia-inducible factor-1α in the liver, lung, and kidney of septic mice. Importantly, we found that PTUPB treatment suppressed the activation of NLRP3 inflammasome in the liver and lung of septic mice. Meanwhile, we found that PTUPB attenuated the oxidative stress, which contributed to the activation of NLRP3 inflammasome. Altogether, our data, for the first time, demonstrate that dual inhibition of COX-2 and sEH with PTUPB ameliorates the multiple organ dysfunction in septic mice.