Critical role of mitochondrial oxidative stress in acid aspiration induced ALI in mice.
线粒体氧化应激在酸吸入诱导的小鼠 ALI 中的关键作用。
- 作者列表："Puri G","Naura AS
:Acute lung injury (ALI) is a pulmonary inflammatory disorder which causes significant mortality in critically ill patients. Intracellular oxidative stress has been considered to be the major component in the pathogenesis of ALI but exact source of intracellular ROS is not clearly known. The present study has been designed to elucidate the role of NADPH oxidase system and/or mitochondrial oxidative stress and its downstream pathway NLRP3 inflammasomes in mouse model of acid aspiration mediated ALI. Our data showed that acid aspiration induced lung inflammation was associated with enhanced oxidative stress as evident by data on MDA levels, nitrite levels and redox imbalance. Further acid aspiration resulted in elevation of expression of NADPH oxidase subunits (gp91 phox/p22 phox/p67 phox) as well as mitochondrial oxidative stress as reflected by aconitase activity, mitochondrial ROS levels. Interestingly, NADPH oxidase inhibitor, apocynin did not alter lung inflammation upon HCl instillation. Conversely, mitochondrial antioxidant mito-tempo resulted in significant amelioration of lung inflammation as indicated by suppression of pulmonary neutrophils and inflammatory cytokines namely IL-1β, TNF-α, IL-6 in BALF. Analysis of mitochondrial enzymes aconitase/mitochondrial ROS/Mn-SOD confirmed that reduction in lung inflammation by mito-tempo was associated with normalization of oxidative stress in mitochondria. Further, mito-tempo administration blunted phosphorylation of p65- NF-κB at Ser 536. Finally, mito-tempo downregulated HCl-induced NF-κB-dependent pro-inflammatory cytokines (IL-1β, TNF-α, IL-6) drastically at mRNA levels. Overall, our data support that mitochondrial oxidative stress is crucial in modulating the HCl induced lung inflammation and identifies mitochondrial-targeted antioxidant as a potential therapeutic agent.
急性肺损伤 (ALI) 是一种肺部炎症性疾病，可导致危重患者的严重死亡。细胞内氧化应激已被认为是 ALI 发病机制中的主要成分，但细胞内 ROS 的确切来源尚不清楚。本研究旨在阐明 NADPH 氧化酶系统和/或线粒体氧化应激及其下游通路 NLRP3 炎症小体在酸吸入介导的 ALI 小鼠模型中的作用。我们的数据表明，酸吸入诱导的肺部炎症与氧化应激增强有关，从 MDA 水平、亚硝酸盐水平和氧化还原失衡的数据可以明显看出。进一步的酸抽吸导致 NADPH 氧化酶亚单位 (gp91 phox/p22 phox/p67 phox) 的表达升高，以及由乌头酸酶活性、线粒体 ROS 水平反映的线粒体氧化应激。有趣的是，NADPH 氧化酶抑制剂 apocynin 在 HCl 滴注后没有改变肺部炎症。相反，线粒体抗氧化剂 mito-tempo 可显著改善肺部炎症，表现为抑制肺中性粒细胞和 BALF 中的炎性细胞因子，即 il-1 β 、 TNF-α 、 IL-6。线粒体酶乌头酸酶/线粒体 ROS/Mn-SOD 的分析证实，mito-tempo 减少肺部炎症与线粒体氧化应激正常化相关。此外，mito-tempo 给药在 Ser 536 时钝化了 p65-NF-κ b 的磷酸化。最后，mito-tempo 在 mRNA 水平显著下调 HCl 诱导的 NF-κ b 依赖性促炎细胞因子 (il-1 β 、 TNF-α 、 IL-6)。总的来说，我们的数据支持线粒体氧化应激在调节 HCl 诱导的肺部炎症中至关重要，并确定线粒体靶向抗氧化剂是一种潜在的治疗药物。
METHODS:BACKGROUND AND PURPOSE:A critical role for sphingosine kinase/sphingosine-1-phosphate (S1P) pathway in the control of airway function has been demonstrated in respiratory diseases. Here, we address S1P contribution in a mouse model of mild chronic obstructive pulmonary disease (COPD). EXPERIMENTAL APPROACH:C57BL/6J mice have been exposed to room air or cigarette smoke up to 11 months and killed at different time points. Functional and molecular studies have been performed. KEY RESULTS:Cigarette smoke caused emphysematous changes throughout the lung parenchyma coupled to a progressive collagen deposition in both peribronchiolar and peribronchial areas. The high and low airways showed an increased reactivity to cholinergic stimulation and α-smooth muscle actin overexpression. Similarly, an increase in airway reactivity and lung resistances following S1P challenge occurred in smoking mice. A high expression of S1P, Sph-K2 , and S1P receptors (S1P2 and S1P3 ) has been detected in the lung of smoking mice. Sphingosine kinases inhibition reversed the increased cholinergic response in airways of smoking mice. CONCLUSIONS AND IMPLICATIONS:S1P signalling up-regulation follows the disease progression in smoking mice and is involved in the development of airway hyperresponsiveness. Our study defines a therapeutic potential for S1P inhibitors in management of airways hyperresponsiveness associated to emphysema in smokers with both asthma and COPD.
METHODS::The interim results from this 90-day multi-dose, inhalation toxicology study with life-time post-exposure observation has shown an important fundamental difference in persistence and pathological response in the lung between brake dust derived from brake-pads manufactured with chrysotile, TiO2 or chrysotile alone in comparison to the amphiboles, crocidolite and amosite asbestos. In the brake dust exposure groups no significant pathological response was observed at any time. Slight macrophage accumulation of particles was noted. Wagner-scores, were from 1 to 2 (1 = air-control group) and were similar to the TiO2 group. Chrysotile being biodegradable, shows a weakening of its matrix and breaking into short fibers & particles that can be cleared by alveolar macrophages and continued dissolution. In the chrysotile exposure groups, particle laden macrophage accumulation was noted leading to a slight interstitial inflammatory response (Wagner-score 1-3). There was no peribronchiolar inflammation and occasional very slight interstitial fibrosis. The histopathology and the confocal analyses clearly differentiate the pathological response from amphibole asbestos, crocidolite and amosite, compared to that from the brake dust and chrysotile. Both crocidolite and amosite induced persistent inflammation, microgranulomas, and fibrosis (Wagner-scores 4), which persisted through the post exposure period. The confocal microscopy of the lung and snap-frozen chestwalls quantified the extensive inflammatory response and collagen development in the lung and on the visceral and parietal surfaces. The interim results reported here, provide a clear basis for differentiating the effects from brake dust exposure from those following amphibole asbestos exposure. The subsequent results through life-time post-exposure will follow.
METHODS::The respiratory tract is lined by a pseudo-stratified epithelium from the nose to terminal bronchioles. This first line of defense of the lung against external stress includes five main cell types: basal, suprabasal, club, goblet and multiciliated cells, as well as rare cells such as ionocytes, neuroendocrine and tuft/brush cells. At homeostasis, this epithelium self-renews at low rate but is able of fast regeneration upon damage. Airway epithelial cell lineages during regeneration have been investigated in the mouse by genetic labeling, mainly after injuring the epithelium with noxious agents. From these approaches, basal cells have been identified as progenitors of club, goblet and multiciliated cells, but also of ionocytes and neuroendocrine cells. Single-cell RNA sequencing, coupled to lineage inference algorithms, has independently allowed the establishment of comprehensive pictures of cell lineage relationships in both mouse and human. In line with genetic tracing experiments in mouse trachea, studies using single-cell RNA sequencing (RNAseq) have shown that basal cells first differentiate into club cells, which in turn mature into goblet cells or differentiate into multiciliated cells. In the human airway epithelium, single-cell RNAseq has identified novel intermediate populations such as deuterosomal cells, 'hybrid' mucous-multiciliated cells and progenitors of rare cells. Novel differentiation dynamics, such as a transition from goblet to multiciliated cells have also been discovered. The future of cell lineage relationships in the respiratory tract now resides in the combination of genetic labeling approaches with single-cell RNAseq to establish, in a definitive manner, the hallmarks of cellular lineages in normal and pathological situations.