Sepsis-induced acute lung injury (ALI) is a complex and life-threatening condition characterized by excessive inflammatory responses, ferroptosis, and oxidative stress. A comprehensive investigation and effective therapeutic strategies are crucial for managing this condition. In this study, we established in vivo sepsis models using lipopolysaccharide (LPS) in wild-type (WT) mice and triggering receptor expressed on myeloid cells 2 (TREM2) knockout (TREM2-KO) mice to assess lung morphology, oxidative stress, and ferroptosis. In vitro, RAW264.7 cells with TREM2 overexpression (TREM2-OE) or knockdown (TREM2-SiRNA) were utilized to assess oxidative stress and ferroptosis. RNA sequencing of LPS-stimulated cells transfected with either vector or TREM2-OE revealed significant differences in inflammation- and ferroptosis-related pathways. LPS-induced lung injury and ferroptosis were exacerbated in TREM2-KO mice and TREM2-SiRNA cells but alleviated by the ferroptosis inhibitor ferrostatin-1 (Fer-1). Mechanistically, TREM2-KO led to SHP1 downregulation and STAT3-P upregulation, which were reversed by the SHP1 agonist SC-43. These findings highlight the role of TREM2 in the SHP1/STAT3 signaling pathway and its regulatory effects on ferroptosis. Our study demonstrates that TREM2, via the SHP1/STAT3 pathway, suppresses oxidative stress and ferroptosis, thereby significantly mitigating sepsis-induced ALI. These results underscore the pivotal role of TREM2 in modulating inflammatory responses and immunity, providing a theoretical foundation for developing therapeutic strategies.

Lung ischemia/reperfusion injury (LIRI) is a significant complication following lung transplantation driven by neutrophil extracellular traps (NETs) associated with mitochondrial oxidative stress. However, the intercellular signaling mechanisms mediating oxidative stress remain unresolved. Here, we elucidated a mitochondrial reactive oxygen species (mtROS) amplification mechanism driven by extracellular vesicles (EVs). In this mechanism, EVs derived from oxygen-glucose deprivation/reperfusion (OGD/R)-activated macrophages transferred endothelial monocyte-activating polypeptide-II (EMAP-II) to neutrophils, suppressed PI3K/AKT signaling, and thereby induced mitochondrial oxidative stress that drove pathological NETs formation. Proteomic profiling identified EMAP-II as a key signaling molecule enriched in EVs secreted by OGD/R-activated macrophages. Pharmacological inhibition of mtROS or AKT activation abolished NETs formation, confirming the PI3K/AKT/mtROS as the central redox-sensitive pathway. Crucially, shRNA-mediated EMAP-II knockdown in macrophages abolished the ability of OGD/R-EVs to induce mtROS and NETs formation, mitigating pulmonary inflammation and tissue injury in mice. This study establishes EMAP-II from macrophage-derived EVs as transcellular drivers of neutrophil mitochondrial oxidative stress. We propose EMAP-II blockade as a therapeutic strategy to disrupt the pathogenic cascade in LIRI, wherein macrophage-derived EVs trigger NETs formation through PI3K/AKT/mtROS.

Introduction: Dexamethasone (DEX), as an important enduring-effect glucocorticoid (GC), holds great promise in the field of lung ischemia-reperfusion injury (LIRI) comprehensive therapy owing to its immunomodulatory properties, such as inducing apoptosis and cell cycle distribution. However, its potent anti-inflammatory application is still restricted because of multiple internal physiologic barriers. Methods: Herein, we developed upconversion nanoparticles (UCNPs) coated with photosensitizer/capping agent/fluorescent probe-modified mesoporous silica (UCNPs@mSiO2[DEX]-Py/β-CD/FITC, USDPFs) for precise DEX release synergistic LIRI comprehensive therapy. The UCNPs were designed by covering an inert YOF:Yb shell on the YOF:Yb, Tm core to achieve high-intensity blue and red upconversion emission upon Near-Infrared (NIR) laser irradiation. Results: Under suitable compatibility conditions, the molecular structure of photosensitizer can be damaged along with capping agent shedding, which endowed USDPFs with an outstanding capability to carry out DEX release controlling and fluorescent indicator targeting. Furthermore, the hybrid encapsulating of DEX significantly increased utilization of nano-drugs, improving the water solubility and bioavailability, which was conducive to developing the anti-inflammatory performance of USDPFs in the complex clinical environment. Discussion: The response-controlled release of DEX in the intrapulmonary microenvironment can reduce normal cell damage, which can effectively avoid the side effects of nano-drugs in anti-inflammatory application. Meanwhile, the multi-wavelength of UCNPs endowed nano-drugs with the fluorescence emission imaging capacity in an intrapulmonary microenvironment, providing precise guidance for LIRI.

Lung ischemia-reperfusion injury (LIRI) is a complex "aseptic" inflammatory response, macrophage play a pivotal role in the pathogenesis of LIRI. Galectin-3 (Gal3), a lectin implicated inflammation, has received limited attention in LIRI. Studies have reported Gal3 as a ligand for triggering receptor expressed on myeloid cell 2 (TREM2) in macrophages in Alzheimer's disease. Hence, we established LIRI C57BL/6 mice model and hypoxia/glucose deprivation and reoxygenation (OGD/R) model to investigate the relationship among Gal3, TREM2, and macrophage polarization. Our result demonstrated inhibition of Gal3 significantly reduced M1-type macrophage polarization while markedly increased M2-type in LIRI. In addition, we observed colocalization of Gal3 and TREM2 in macrophages, inhibition of Gal3 could recover the downregulation of TREM2 induced by LIRI while promoting TREM2 expression could attenuate lung injury in LIRI. In summary, our findings suggest Gal3 as an upstream factor of TREM2, play a crucial role in LIRI by regulating macrophage polarization.

Polarization of alveolar macrophages (AMs) into the M1 phenotype contributes to inflammatory responses and tissue damage that occur during lung ischemia-reperfusion injury (LIRI). Programmed cell death factor-1 (PD-1) regulates polarization of macrophages, but its role in LIRI is unknown. We examined the role of PD-1 in AM polarization in models of LIRI in vivo and in vitro. Adult Sprague-Dawley rats were subjected to ischemia-reperfusion with or without pretreatment with a PD-1 inhibitor, SHP1/2 inhibitor, or Akt activator. Lung tissue damage and infiltration by M1-type AMs were assessed. As an in vitro complement to the animal studies, rat alveolar macrophages in culture were subjected to oxygen/glucose deprivation and reoxygenation. Levels of SHP1/2 and Akt proteins were evaluated using Western blots, while levels of pro-inflammatory cytokines were measured using enzyme-linked immunosorbent assays. Injury upregulated PD-1 both in vivo and in vitro. Inhibiting PD-1 reduced the number of M1-type AMs, expression of SHP1 and SHP2, and levels of inflammatory cytokines. At the same time, it partially restored Akt activation. Similar results were observed after inhibition of SHP1/2 or activation of the PI3K/Akt pathway. PD-1 promotes polarization of AMs to the M1 phenotype and inflammatory responses through the SHP1/2-PI3K/Akt axis. Inhibiting PD-1 may be an effective therapeutic strategy to limit LIRI.

为提升稀土基氟氧化物的荧光发射能力及细胞吞噬效果,采用双惰性层包覆工艺,同时利用功能性亲水聚合物进行表面修饰,制备获得YOF:Yb3+,Tm3+@YOF:Yb3+@mSiO2-Py-FITC-β-CD纳米颗粒.使用透射电镜(TEM,含能谱分析)、紫外(UV-Vis)及荧光(PL)分光光度计、傅里叶变换红外光谱仪(FT-IR)及共聚焦激光扫描显微镜(CLSM)分别对试样的成分、形貌、荧光及细胞吞噬能力等进行表征.结果表明:双惰性层包覆的纳米颗粒尺寸范围30～35 nm,且具有较强的上转换荧光锐峰发射,在475 nm处(蓝色)荧光寿命可达58.696μs.样品在浓度较低时(50μg/mL)易被RAW264.7细胞大量吞噬.

Abstract Background: Lung ischemia-reperfusion injury (LIRI) is a common and complex pathophysiological process. Activation of alveolar macrophages (AMs) is involved in the inflammatory response and tissue damage during LIRI. Programmed cell death factor-1 (PD-1) plays an important role in immunomodulatory participants in diseases progression by regulating apoptosis and polarization of macrophages. In this study, we investigated the function of PD-1 on macrophage polarization during pulmonary ischemia-reperfusion.Methods: We conducted experiments both in a rat model of lung ischemia/reperfusion injury and in NR8383 cells (a rat alveolar macrophage line) model of oxygen-glucose deprivation-reoxygenation (OGD/R). The extent of lung injury was measured by pathohistological lung tissue damage, wet/dry ratios of pulmonary tissue, levels of inflammatory factors. Western blot was used to detect the relative expression of PD-1, SHP1, SHP2, p-Akt, and Akt. Expression of surface markers on macrophages was measured by immunohistochemistry, flow cytometry, and immunofluorescence staining.Results: The results showed that the expression of PD-1 increased in both in vivo and in vitro models, which promoted the polarization of M1 macrophages, resulting in the aggravation of the pulmonary ischemia-reperfusion injury. These effects were at least partially attributable to PD-1 inhibiting the PI3K/Akt pathway by regulating SHP1/2. Conclusion: PD-1 may exacerbate LIRI by promoting macrophage polarization, and inhibition of PD-1 may be an important role and therapeutic target of acute lung injury.

目的:探讨半乳凝集素3(Ga13)在小鼠肺缺血再灌注损伤(LIRI)模型中的表达情况.方法:将20只C57BL/6J雄性小鼠随机分为假手术组和4个实验组(即LIRI后2 h组、LIRI后6 h组、LIRI后12 h组、LIRI后24 h组),每组4只.假手术组只开胸不夹闭肺门,实验组夹闭肺门60min并分别于松开血管夹后2h、6h、12h、24h收集左肺标本.采用苏木精—伊红(HE)染色法和肺组织湿干重比(W/D)观察肺组织病理学变化,酶联免疫吸附法(ELISA)检测支气管肺泡灌洗液(BALF)炎性因子白细胞介素-1β(IL-1β)和肿瘤坏死因子α(TNF-a)的表达,分别采用实时荧光定量PCR(RT-qPCR)、western blotting、免疫荧光染色法检测Gal3 mRNA与Ga13蛋白的表达水平.结果:与假手术组相比,LIRI后各实验组出现肺组织损伤,组织结构破坏明显,肺损伤评分和W/D均升高,炎症因子IL-1β和TNF-a水平升高(均P＜0.05),在LIRI后6h肺组织病理学损伤程度和炎性因子水平达到峰值(P＜0.05),随后逐渐降低.与假手术组相比,LIRI后各实验组Ga13 mRNA及其蛋白表达水平升高,Ga13免疫荧光染色表达水平升高(P＜0.05),均于LIRI后6 h达到高峰(P＜0.05),然后逐渐下降.结论:小鼠LIRI后Ga13表达升高,高水平的Ga13可能与LIRI后的肺组织损伤和炎症爆发有关.

Fei Lin (  linfei@gxmu.edu.cn ) Guangxi Medical University Cancer Hospital Xiaojing He Guangxi Medical University Cancer Hospital Jingyuan Xiao Guangxi Medical University Cancer Hospital Jinyuan Lin Guangxi Medical University Cancer Hospital Zhen Liu Guangxi Medical University Cancer Hospital Yubing Liang Guangxi Medical University Cancer Hospital Huijun Dai Guangxi Medical University Cancer Hospital Ren Jing Guangxi Medical University Cancer Hospital Zhao Li Guangxi Medical University Cancer Hospital

Lung ischemia-reperfusion injury (LIRI) is a severe multifaceted pathological condition that can lead to poor patient outcome where oxidative stress and the resulting inflammatory response can trigger and exacerbate tissue damage in LIRI patients. Sirtuin3 (SIRT3), a member of the sirtuin family, protects against oxidative stress-related diseases. However, it remains unclear if and how SIRT3 alleviates lung injury induced by ischemia/reperfusion (I/R). Our previous study showed that lung tissue structures were severely damaged at 6 h after lung I/R in mice, however, repair of the injured lung tissue was significant at 24 h. In this study, we found that both SIRT3 mRNA and protein levels were markedly increased at 24 h after lung I/R in vivo. Meanwhile, inhibition of SIRT3 aggravated lung injury and inflammation, augmented mitochondrial fission and oxidative stress and increased Hypoxia-inducible factor-1α (HIF-1α) expression in vivo. The results suggest that SIRT3 may be an upstream regulator of HIF-1α expression. Knockdown of SIRT3 resulted in excessive mitochondrial fission and increased oxidative stress in vitro, and we found that knocking down the expression of HIF-1α alleviated these changes. This suggests that the SIRT3-HIF-1α signaling pathway is involved in regulating mitochondrial function and oxidative stress. Furthermore, inhibition of dynamin-related protein 1 (Drp-1) by the inhibitor of mitophagy, Mdivi-1, blocked mitochondrial fission and alleviated oxidative stress in vitro. Taken together, our results demonstrated that downregulation of SIRT3 aggravates LIRI by increasing mitochondrial fission and oxidative stress. Activation of SIRT3 inhibits mitochondrial fission and this mechanism may serve as a new therapeutic strategy to treat LIRI.

Lung ischemia reperfusion injury (LIRI) is a complex pathophysiological process with high morbidity and mortality. An important pathophysiological characteristic of LIRI is endothelial barrier dysfunction, although the mechanism involved in this process remains unclear. VX765, a specific caspase-1 inhibitor, has been shown to have a protective effect against several diseases including sepsis, atherosclerosis, and glial inflammatory disease. The objective of this study was to determine whether VX765 had a protective effect in LIRI. The results showed that lung ischemia/reperfusion (I/R) and oxygen/glucose deprivation and reoxygenation (OGD/R) induced endothelial pyroptosis and barrier dysfunction characterized by an inflammatory response. Treatment with VX765 successfully alleviated I/R- and OGD/R-induced endothelial pyroptosis and barrier dysfunction by inhibiting caspase-1 in vivo and in vitro. In conclusion, these findings showed that VX765 provided effective protection against lung I/R-induced endothelial pyroptosis and barrier dysfunction.

目的:探讨热休克蛋白60(HSP60)调控2型髓系细胞触发受体(TREM2)对肺缺血再灌注损伤(LIRI)小鼠的保护作用.方法:将24只C57BL/6小鼠随机分成对照组、HSP60组、LIRI组、LIRI+HSP60组,每组6只.LIRI+HSP60组于术前1 h腹腔注射HSP60(5 μg/g).HSP60组腹腔注射等剂量的HSP60;对照组不做任何处理.LIRI组和LIRI+HSP60组小鼠建立LIRI模型,术后6 h收集左肺组织标本,苏木精—伊红(HE)染色法观察肺组织病理学变化,透视电镜观察肺组织超微结构改变,酶联免疫吸附法(ELISA)检测肺组织中炎性因子白细胞介素-1β(IL-1β)、白细胞介素-18(IL-18)的表达,实时荧光定量PCR(qPCR)法与Western blotting检测TREM2 mRNA及TREM2蛋白的相对表达水平.结果:与对照组和HSP60组比较,LIRI组肺组织损伤严重,肺组织结构改变明显,肺损伤评分明显增加(P＜0.05),炎症因子IL-1β和IL-18水平均明显升高(均P＜0.05),TREM2 mRNA及其蛋白相对表达水平均明显降低(均P＜0.05).与LIRI组比较,LIRI+HSP60组肺组织病理学损伤明显改善,肺损伤评分明显降低(P＜0.05),炎症因子IL-1β和IL-18表达明显减少(均PP＜0.05),TREMM2 mRNA及其蛋白的相对表达水平均明显升高(均P＜0.05).与对照组相比较,HSP60组TREM2mRNA及蛋白的相对表达水平均明显升高(P＜0.05).结论:HSP60可以减轻LIRI,其机制可能与上调TREM2的表达有关.

Objective:To investigate the role and regulatory mechanism of triggering receptor expressed on myeloid cell 2 (TREM2) in mice lung ischemia/reperfusion injury (LIRI).Methods:Thirty-six healthy male C57BL/6 mice were divided into six groups according to the random number method ( n = 6): normal control group, and LIRI 2, 6, 12, 24, 48 hours group. Mice LIRI models were established by clamping the left hilum. The wet/dry weight ratio (W/D) of left lung tissue was measured. Lung injury was observed and evaluated by hematoxylin-eosin (HE) staining and electron microscopy. The levels of interleukins (IL-1β, IL-18) in lung tissue were detected by enzyme linked immunosorbent assay (ELISA). The mRNA expressions of TREM2 and caspase-1 were determined by polymerase chain reaction (PCR). The protein expressions of TREM2, caspase-1, Gasdermin-D (GSDMD) were determined by Western blotting. Results:At 2 hours after LIRI, lung injury began to appear, the lung ultrastructure changed, and the lung injury score increased; at 6 hours, the degree of lung injury was the most serious; after 12 hours, the lung injury gradually reduced and the lung injury score gradually decreased. Compared with the normal control group, lung W/D ratio and lung injury score of LIRI 2, 6, 12, 24, 48 hours groups were significantly higher, the differences were statistically significant (lung W/D ratio: 7.06±0.52, 8.34±0.17, 6.42±0.35, 5.34±0.25, 5.59±0.45 vs. 4.69±0.23; lung injury score: 5.50±0.54, 9.75±0.89, 5.88±0.84, 3.63±0.74, 4.13±0.64 vs. 1.13±0.35, all P < 0.05). Compared with the normal control group, the levels of IL-1β and IL-18 in lung tissue were significantly increased at 2 hours after LIRI, reached a peak at 6 hours [IL-1β (ng/L): 502.76±12.25 vs. 56.50±8.07, IL-18 (ng/L): 414.02±10.75 vs. 81.63±5.29, both P < 0.05], then decreased gradually, and were still significantly higher than the normal control group at 48 hours. The PCR and Western blotting showed that the expression of TREM2 was significantly lower than that in the normal control group at 2 hours after LIRI, and reached a valley at 6 hours [TREM2 mRNA (2 -ΔΔCt): 0.47±0.05 vs. 1.02±0.05, TREM2/GAPDH: 0.23±0.13 vs. 0.48±0.17, both P < 0.05], then gradually increased, and reached the peak at 24 hours [TREM2 mRNA (2 -ΔΔCt): 3.98±0.15 vs. 1.02±0.05, TREM2/GAPDH: 0.71±0.17 vs. 0.48±0.17, both P < 0.05]. The trend of expression of caspase-1 and GSDMD were opposite to that of TREM2, which increased at first and then decreased, and reached a peak at 6 hours after reperfusion [caspase-1 mRNA (2 -ΔΔCt): 2.20±0.13 vs. 1.01±0.02, caspase-1/GAPDH: 0.64±0.02 vs. 0.20±0.06, GSDMD/GAPDH: 1.23±0.01 vs. 0.87±0.02, all P < 0.05]. Conclusions:TREM2 might be involved in LIRI in mice. The mechanism may be related to the effect of TREM2 on caspase-1-mediated pyroptosis.