P75NTR+CD64+ neutrophils promote sepsis-induced acute lung injury.

Patients suffering from sepsis-induced acute lung injury (ALI) exhibit a high mortality rate, and their prognosis is closely associated with infiltration of neutrophils into the lungs. In this study, we found a significant elevation of CD64+ neutrophils, which highly expressed p75 neurotrophin receptor (p75NTR) in peripheral blood of mice and patients with sepsis-induced ALI. p75NTR+CD64+ neutrophils were also abundantly expressed in the lung of ALI mice induced by lipopolysaccharide. Conditional knock-out of the myeloid lineage's p75NTR gene improved the survival rates, attenuated lung tissue inflammation, reduced neutrophil infiltration and enhanced the phagocytic functions of CD64+ neutrophils. In vitro, p75NTR+CD64+ neutrophils exhibited an upregulation and compromised phagocytic activity in blood samples of ALI patients. Blocking p75NTR activity by soluble p75NTR extracellular domain peptide (p75ECD-Fc) boosted CD64+ neutrophils phagocytic activity and reduced inflammatory cytokine production via regulation of the NF-κB activity. The findings strongly indicate that p75NTR+CD64+ neutrophils are a novel pathogenic neutrophil subpopulation promoting sepsis-induced ALI.

Circular RNA circVAPA modulates macrophage pyroptosis in sepsis-induced acute lung injury through targeting miR-212-3p/Sirt1/Nrf2/NLRP3 axis.

Sepsis-induced acute lung injury (ALI) is an inflammatory condition involving the pyroptosis of macrophages. This study investigated the role of circular RNA hsa_circ_0006990 (circVAPA) in regulating macrophage pyroptosis in ALI and the underlying mechanisms. The expression pattern of circVAPA was examined in the mouse model of ALI and in the LPS-treated RAW264.7 macrophage cell line. Lung tissue damage was evaluated by haematoxylin and eosin staining, immunohistochemistry and a myeloperoxidase activity assay. The molecular mechanisms were investigated by luciferase reporter assay, western blot, RT-qPCR and ELISA. circVAPA was down-regulated in the lung tissues of ALI mice and LPS-induced RAW264.7 cells. circVAPA over-expression alleviated lung tissue injury and dampened LPS-induced pyroptosis and Th17-associated inflammatory responses. miR-212-3p was identified as a target of circVAPA, and miR-212-3p negatively regulated the expression of Sirt1. Sirt1 knockdown largely abolished the effect of circVAPA over-expression on pyroptosis. CircVAPA/miR-212-3p/Sirt1 axis also regulates Nrf2 and NLRP3 expression upon LPS challenge. By targeting miR-212-3p, circVAPA over-expression negatively regulates the expression of Sirt1 and pyroptosis-related factors (Nrf2 and NLRP3), which alleviates the inflammatory damages in sepsis-induced ALI.

Elucidating the role of Rhodiola rosea L. in sepsis-induced acute lung injury via network pharmacology: emphasis on inflammatory response, oxidative stress, and the PI3K-AKT pathway.

CONTEXT: Sepsis-induced acute lung injury (ALI) is associated with high morbidity and mortality. Rhodiola rosea L. (Crassulaceae) (RR) and its extracts have shown anti-inflammatory, antioxidant, immunomodulatory, and lung-protective effects. OBJECTIVE: This study elucidates the molecular mechanisms of RR against sepsis-induced ALI. MATERIALS AND METHODS: The pivotal targets of RR against sepsis-induced ALI and underlying mechanisms were revealed by network pharmacology and molecular docking. Human umbilical vein endothelial cells (HUVECs) were stimulated by 1 µg/mL lipopolysaccharide for 0.5 h and treated with 6.3, 12.5, 25, 50, 100, and 200 µg/mL RR for 24 h. Then, the lipopolysaccharide-stimulated HUVECs were subjected to cell counting kit-8 (CCK-8), enzyme-linked immunosorbent, apoptosis, and Western blot analyses. C57BL/6 mice were divided into sham, model, low-dose (40 mg/kg), mid-dose (80 mg/kg), and high-dose (160 mg/kg) RR groups. The mouse model was constructed through caecal ligation and puncture, and histological, apoptosis, and Western blot analyses were performed for further validation. RESULTS: We identified six hub targets (MPO, HRAS, PPARG, FGF2, JUN, and IL6), and the PI3K-AKT pathway was the core pathway. CCK-8 assays showed that RR promoted the viability of the lipopolysaccharide-stimulated HUVECs [median effective dose (ED50) = 18.98 µg/mL]. Furthermore, RR inhibited inflammation, oxidative stress, cell apoptosis, and PI3K-AKT activation in lipopolysaccharide-stimulated HUVECs and ALI mice, which was consistent with the network pharmacology results. DISCUSSION AND CONCLUSION: This study provides foundational knowledge of the effective components, potential targets, and molecular mechanisms of RR against ALI, which could be critical for developing targeted therapeutic strategies for sepsis-induced ALI.

Irf7 regulates the expression of Srg3 and ferroptosis axis aggravated sepsis-induced acute lung injury.

OBJECTIVE: To investigate the mechanism of action of Srg3 in acute lung injury caused by sepsis. METHODS: First, a sepsis-induced acute lung injury rat model was established using cecal ligation and puncture (CLP). RNA sequencing (RNA-seq) was used to screen for highly expressed genes in sepsis-induced acute lung injury (ALI), and the results showed that Srg3 was significantly upregulated. Then, SWI3-related gene 3 (Srg3) was knocked down using AAV9 vector in vivo, and changes in ALI symptoms in rats were analyzed. In vitro experiments were conducted by establishing a cell model using lipopolysaccharide (LPS)-induced BEAS-2B cells and coculturing them with phorbol 12-myristate 13-acetate (PMA)-treated THP-1 cells to analyze macrophage polarization. Next, downstream signaling pathways regulated by Srg3 and transcription factors involved in regulating Srg3 expression were analyzed using the KEGG database. Finally, gain-of-loss functional validation experiments were performed to analyze the role of downstream signaling pathways regulated by Srg3 and transcription factors involved in regulating Srg3 expression in sepsis-induced acute lung injury. RESULTS: Srg3 was significantly upregulated in sepsis-induced acute lung injury, and knocking down Srg3 significantly improved the symptoms of ALI in rats. Furthermore, in vitro experiments showed that knocking down Srg3 significantly weakened the inhibitory effect of LPS on the viability of BEAS-2B cells and promoted alternative activation phenotype (M2) macrophage polarization. Subsequent experiments showed that Srg3 can regulate the activation of the NF-κB signaling pathway and promote ferroptosis. Specific activation of the NF-κB signaling pathway or ferroptosis significantly weakened the effect of Srg3 knockdown. It was then found that Srg3 can be transcriptionally activated by interferon regulatory factor 7 (Irf7), and specific inhibition of Irf7 significantly improved the symptoms of ALI. CONCLUSIONS: Irf7 transcriptionally activates the expression of Srg3, which can promote ferroptosis and activate classical activation phenotype (M1) macrophage polarization by regulating the NF-κB signaling pathway, thereby exacerbating the symptoms of septic lung injury.

SRPK1 promotes sepsis-induced acute lung injury via regulating PI3K/AKT/FOXO3 signaling.

OBJECTIVE: Sepsis is the most common cause of death in the intensive care unit. Moreover, sepsis is the leading cause of acute lung injury (ALI). Serine-arginine protein kinase 1 (SRPK1) was demonstrated to promote the development of ALI. However, the potentials of SRPK1 in sepsis-induced ALI are still unknown. This study aimed to investigate the potentials of SRPK1 in sepsis-induced ALI and the underlying mechanisms. METHODS: Cecal ligation and puncture (CLP) was performed to establish a sepsis-induced ALI model in vivo. Primary human pulmonary microvascular endothelial cells (HPMECs) were exposed to lipopolysaccharide (LPS) to construct a sepsis-induced ALI model in vitro. Gene expression was detected using western blot and qRT-PCR. The interaction between forkhead box O3 (FOXO3) and NOD-like receptor thermal protein domain associated protein 3 (NLRP3) was detected using luciferase and Chromatin immunoprecipitation (ChIP) assay. Cellular functions were CCK-8, colony formation, PI staining, and flow cytometry assay. RESULTS: SRPK1 was downregulated in patients with sepsis-induced ALI. Overexpression of SRPK1 suppressed the pyroptosis of HPMECs as well as promoted cell proliferation. Additionally, SRPK1 overexpression alleviated sepsis-induced ALI in vivo. SRPK1 activated phosphatidylinositol3-kinase (PI3K) signaling pathways. Blocking the activation of PI3K degraded the cellular functions of HPMECs. Moreover, FOXO3 transcriptionally inactivated NLRP3 and suppressed its mRNA and protein expression. CONCLUSION: Taken together, SRPK1 suppressed sepsis-induced ALI via regulating PI3K/AKT/FOXO3/NLRP3 signaling. SRPK1 may be the potential biomarker for sepsis-induced ALI.

Inhibition of STAT3 phosphorylation by colchicine regulates NLRP3 activation to alleviate sepsis-induced acute lung injury.

The pharmacotherapeutic mechanism of colchicine, a tricyclic, lipid-soluble alkaloid extracted from the plant of the Lily family Colchicum autumnale, has not been fully understood in diverse disorders, including sepsis-induced acute lung injury (ALI). The study aimed at exploring the impact of colchicine on sepsis-induced ALI and the relevant mechanisms. Colchicine significantly attenuated ALI in mice caused by sepsis by alleviating respiratory dysfunction and pulmonary edema in mice, inhibiting NLRP3 inflammasome formation, and reducing oxidative stress, pyroptosis, and apoptosis of murine alveolar macrophage (J774A.1) cells. The targets of colchicine were predicted in the superPRED database and intersected with the differentially expressed genes in the GSE5883 and GSE129775 datasets. The major targets were subjected to protein-protein interaction network generation and Kyoto Encyclopedia of Genes and Genomes enrichment analysis. It was thus found that colchicine inhibited STAT3 phosphorylation but did not alter STAT3 total protein expression. Phosphorylated STAT3 recruited EP300 to form a complex to promote histone H3 acetylation and histone H4 acetylation of NLRP3 promoter, leading to pyroptosis of J774A.1 cells. In conclusion, inhibition of STAT3 phosphorylation by colchicine represses NLRP3 promoter acetylation via the STAT3/EP300 complex, thereby alleviating ALI caused by sepsis.

Upregulation of endothelial cell-derived exosomal microRNA-125b-5p protects from sepsis-induced acute lung injury by inhibiting topoisomerase II alpha.

OBJECTIVE: Emerging evidence has revealed that exosomal microRNAs (miRNAs) are implicated in human diseases. However, role of exosomal miR-125b-5p in sepsis-induced acute lung injury (ALI) remains further explored. We focused on the effect of exosomal miR-125b-5p on ALI progression via targeting topoisomerase II alpha (TOP2A). METHODS: The ALI mouse models were established by cecal ligation and perforation, which were then treated with miR-125b-5p agomir or overexpressed TOP2A. Next, the pathological structure of ALI mouse lung tissues were observed, miR-125b-5p, TOP2A and vascular endothelial growth factor (VEGF) expression was determined, and the lung water content, inflammatory response, protein content in bronchoalveolar lavage fluid (BALF) and cell apoptosis in ALI mouse lung tissues were assessed. Exosomes were extracted from endothelial cells (ECs) and identified, which were then injected into the modeled mice to observe their roles in ALI. The targeting relationship between miR-125b-5p and TOP2A was confirmed. RESULTS: MiR-125b-5p was downregulated while TOP2A was upregulated in ALI mice. MiR-125b-5p elevation or ECs-derived exosomes promoted VEGF expression, improved pathological changes and restrained lung water content, inflammatory response, protein content in BALF and cell apoptosis in lung tissues ALI mice. TOP2A overexpression reversed the repressive role of miR-125b-5p upregulation in ALI, while downregulated miR-125b-5p abrogated the effect of ECs-derived exosomes on ALI. TOP2A was confirmed as a direct target gene of miR-125b-5p. CONCLUSION: Our study indicates that ECs-derived exosomes overexpressed miR-125b-5p to protect from sepsis-induced ALI by inhibiting TOP2A, which may contribute to ALI therapeutic strategies.

Norisoboldine Attenuates Sepsis-Induced Acute Lung Injury by Modulating Macrophage Polarization via PKM2/HIF-1α/PGC-1α Pathway.

This study aimed to investigate the effect of norisopoldine (NOR) on acute lung injury in septic mice. Lipopolysaccharide (LPS) was used to establish sepsis induced acute lung injury (ALI) in mice. The dry and wet weight of mice lung was detected, and the pathological changes of lung were observed by hematoxylin and eosin (H&E) staining. Bronchoalveolar lavage fluid (BALF) was detected. Inflammatory factors in BALF were detected by enzyme-linked immunosorbent assay (ELISA). The polarization of macrophages in lung tissue was detected by flow cytometry. The markers of M1 and M2 macrophages were detected by RT-PCR. LPS induced RAW264.7 cells were treated with NOR. Inflammatory response, macrophage polarization, glycolysis, and M2 pyruvate kinase (PKM2)/hypoxia inducible factor-1α (HIF-1α)/peroxisome proliferator activated receptor-γ co-activator 1-α (PGC-1α) signaling pathway were detected. NOR could effectively alleviate sepsis induced ALI, and reduce the number of total cells, total protein concentration, neutrophils, macrophages in BALF. NOR decreased the level of inflammatory factors and promoted macrophages from M1 to M2 type in vivo and vitro. Moreover, NOR could activated PKM2, and inhibited PKM2 from cytoplasm to nuclear, attenuated HIF-1α expression, and increased PGC-1α and peroxisome proliferator-activated receptor (PPAR)-γ expression. In addition, NOR inhibited glycolysis and promoted oxidative phosphorylation in RAW264.7 cells. Furthermore, PKM2 inhibitors could reverse the effect of NOR on PKM2/HIF-1α/PGC-1α signaling pathway in RAW264.7 cells. NOR alleviated sepsis induced AIL in mice, inhibited the inflammatory response, promote M2 polarization of macrophages through regulating PKM2/HIF-1α/PGC-1α signaling pathway.

The Mechanisms Involved in Mesenchymal Stem Cell Alleviation of Sepsis-Induced Acute Lung Injury in Mice: A Pilot Study.

BACKGROUND: Acute lung injury is a common complication of sepsis in intensive care unit patients. Inflammation is among the main mechanisms of sepsis. Therefore, suppression of inflammation is an important mechanism for sepsis treatment. Mesenchymal stem cells (MSCs) have been reported to exhibit antimicrobial properties. OBJECTIVE: The present study investigated the effects of MSCs on sepsis-induced acute lung injury. METHODS: Male C57BL/6 mice underwent a cecal ligation and puncture (CLP) operation to induce sepsis and then received either normal saline or MSCs (1 × 106 cells intravenously) at 3 hours after surgery. Survival after surgery was assessed. Lung injury was assessed by histology score, the presence of lung edema, vascular permeability, inflammatory cell infiltration, and cytokine levels in bronchoalveolar lavage fluid. Finally, we tested nuclear factor kappa-light-chain-enhancer of activated B cells activation in lung tissue. RESULTS: As expected, CLP caused lung injury as indicated by significant increases in the histopathology score, lung wet to dry weight ratio, and total protein concentration. However, mice treated with MSCs had amelioration of the lung histopathologic changes, lung wet to dry weight ratio, and total protein concentration. The levels of cytokines tumor necrosis factor alpha, interleukin 6, interleukin 1ß, and interleukin 17 in bronchoalveolar lavage fluid were dramatically decreased after MSCs treatment. In contrast, expression of interleukin 10 was increased after MSCs treatment. Moreover, mice treated with MSCs had a higher survival rate than the CLP group. Neutrophil infiltration into bronchoalveolar lavage fluid was attenuated after MSCs injection, but the amounts of macrophages observed in the MSC group showed no significant differences compared with the CLP group. In addition, MSCs treatment significantly reduced nuclear factor kappa-light-chain-enhancer of activated B cells activation in lung tissue. CONCLUSIONS: Based on the above findings, treatment with MSCs dampened the inflammatory response and inhibited nuclear factor kappa-light-chain-enhancer of activated B cells activation in the mouse CLP model. Thus, MSCs may be a potential new agent for the treatment of sepsis-induced acute lung injury. (Curr Ther Res Clin Exp. 2020; 81:XXX-XXX).

BMAL1 plays a crucial role in immune homeostasis during sepsis-induced acute lung injury.

Sepsis is a widespread and life-threatening disease characterised by infection-triggered immune hyperactivation and cytokine storms, culminating in tissue damage and multiple organ dysfunction syndrome. BMAL1 is a pivotal transcription factor in the circadian clock that plays a crucial role in maintaining immune homeostasis. BMAL1 dysregulation has been implicated in inflammatory diseases and immunodeficiency. However, the mechanisms underlying BMAL1 disruption in sepsis-induced acute lung injury (ALI) remain poorly understood. In vitro, we used THP1 and mouse peritoneal macrophages to elucidate the potential mechanism of BMAL1 function in sepsis. In vivo, an endotoxemia model was used to investigate the effect of BMAL1 on sepsis and the therapeutic role of targeting CXCR2. We showed that BMAL1 significantly affected the regulation of innate immunity in sepsis-induced ALI. BMAL1 deficiency in the macrophages exacerbated systemic inflammation and sepsis-induced ALI. Mechanistically, BMAL1 acted as a transcriptional suppressor and regulated the expression of CXCL2. BMAL1 deficiency in macrophages upregulated CXCL2 expression, increasing the recruitment of polymorphonuclear neutrophils and the formation of neutrophil extracellular traps (NETs) by binding to the chemokine receptor CXCR2, thereby intensifying lung injury in a sepsis model. Furthermore, a selective inhibitor of CXCR2, SB225002, exerted promising therapeutic effects by markedly reducing neutrophil infiltration and NETs formation and alleviating lung injury. Importantly, CXCR2 blockade mitigated multiple organ dysfunction. Collectively, these findings suggest that BMAL1 controls the CXCL2/CXCR2 pathway, and the therapeutic efficacy of targeting CXCR2 in sepsis has been validated, presenting BMAL1 as a potential therapeutic target for lethal infections.

Exosomal Tenascin-C primes macrophage pyroptosis amplifying aberrant inflammation during sepsis-induced acute lung injury.

Sepsis-induced acute lung injury (ALI) is a serious complication of sepsis and the predominant cause of death. Exosomes released by lung tissue cells critically influence the progression of ALI during sepsis by modulating the inflammatory microenvironment. However, the molecular mechanisms by which exosome-mediated intercellular signaling exacerbates ALI in septic infection remain undefined. Our study found increased levels of exosomal Tenascin-C (TNC) in the plasma of both patients and mice with ALI, showing a strong association with disease progression. By integrating exosomal proteomics with transcriptome sequencing and experimental validation, we elucidated that LPS induce unresolved endoplasmic reticulum stress (ERs) in alveolar epithelial cells (AECs), ultimately leading to the release of exosomal TNC through the activation of PERK-eIF2α and the transcription factor CHOP. In the sepsis mouse model with TNC knockout, we noted a marked reduction in macrophage pyroptosis. Our detailed investigations found that exosomal TNC binds to TLR4 on macrophages, resulting in an augmented production of ROS, subsequent mitochondrial damage, activation of the NF-κB signaling pathway, and induction of DNA damage response. These interconnected events culminate in macrophage pyroptosis, thereby amplifying the release of inflammatory cytokines. Our findings demonstrate that exosomal Tenascin-C, released from AECs under unresolved ER stress, exacerbates acute lung injury by intensifying sepsis-associated inflammatory responses. This research provides new insights into the complex cellular interactions underlying sepsis-induced ALI.

LL37-mtDNA regulates viability, apoptosis, inflammation, and autophagy in lipopolysaccharide-treated RLE-6TN cells by targeting Hsp90aa1.

Sepsis-induced acute lung injury is associated with lung epithelial cell injury. This study analyzed the role of the antimicrobial peptide LL37 with mitochondrial DNA (LL37-mtDNA) and its potential mechanism of action in lipopolysaccharide (LPS)-treated rat type II alveolar epithelial cells (RLE-6TN cells). RLE-6TN cells were treated with LPS alone or with LL37-mtDNA, followed by transcriptome sequencing. Differentially expressed and pivotal genes were screened using bioinformatics tools. The effects of LL37-mtDNA on cell viability, inflammation, apoptosis, reactive oxygen species (ROS) production, and autophagy-related hallmark expression were evaluated in LPS-treated RLE-6TN cells. Additionally, the effects of Hsp90aa1 silencing following LL37-mtDNA treatment were investigated in vitro. LL37-mtDNA further suppressed cell viability, augmented apoptosis, promoted the release of inflammatory cytokines, increased ROS production, and elevated LC3B expression in LPS-treated RLE-6TN cells. Using transcriptome sequencing and bioinformatics, ten candidate genes were identified, of which three core genes were verified to be upregulated in the LPS + LL37-mtDNA group. Additionally, Hsp90aa1 downregulation attenuated the effects of LL37-mtDNA on LPS-treated RLE-6TN cells. Hsp90aa1 silencing possibly acted as a crucial target to counteract the effects of LL37-mtDNA on viability, apoptosis, inflammation, and autophagy activation in LPS-treated RLE-6TN cells.

ANXA3 interference inactivates ERK/ELK1 pathway to mitigate inflammation and apoptosis in sepsis-associated acute lung injury.

Acute lung injury (ALI) is a prevailing and deadly complication of sepsis coupled with increasing incidence and fatality rate. Annexin A3 (ANXA3) has been unraveled to be upregulated during sepsis. This study purposed to assess the role and the mechanism of ANXA3 in sepsis-induced ALI. After the construction of mouse model of sepsis, the pathological changes of mice lung tissues were estimated by H&E staining. ANXA3 expression in mice lung tissues and serum was examined. The degree of pulmonary edema and the levels of inflammatory factors in bronchoalveolar lavage fluid (BALF) were analyzed. In lipopolysaccharide (LPS)-induced mouse ALI model in vitro, CCK-8 assay measured cell viability and flow cytometry analysis detected cell apoptosis. Besides, ELISA assay detected the release of inflammatory cytokines. Western blot analyzed the expression of proteins associated with inflammation, apoptosis and extracellular-signal-regulated kinase (ERK)/ETS-like gene 1 (ELK1) signaling. Results revealed that ANXA3 was overexpressed in the lung tissues and serum of septic mice. Following the knockdown of ANXA3, sepsis-induced lung injury was alleviated, manifested as reduced lung edema, decreased inflammatory cell infiltration and inhibited cell apoptosis. Additionally, ANXA3 silence blocked ERK/ELK1 signaling both in sepsis mouse models and in vitro model of ALI induced by lipopolysaccharide (LPS). Moreover, the inhibitory effects of ANXA3 silencing on ERK/ELK1 signaling activation, the viability damage, inflammation and apoptosis in LPS-induced mouse ALI model in vitro were partially reversed by ERK activator. Collectively, depletion of ANXA3 exerted suppressive effects on the inflammation and apoptosis in sepsis-induced ALI through blocking ERK/ELK1 signaling.

Ginsenoside Rg1 Alleviates Sepsis-Induced Acute Lung Injury by Reducing FBXO3 Stability in an m6A-Dependent Manner to Activate PGC-1α/Nrf2 Signaling Pathway.

BACKGROUND: Sepsis-induced acute lung injury (ALI) is one of the serious life-threatening complications of sepsis and is pathologically associated with mitochondrial dysfunction. Ginsenoside Rg1 has good therapeutic effects on ALI. Herein, the pharmacological effects of Rg1 in sepsis-induced ALI were investigated. METHODS: Sepsis-induced ALI models were established by CLP operation and LPS treatment. HE staining was adopted to analyze lung pathological changes. The expression and secretion of cytokines were measured by RT-qPCR and ELISA. Cell viability and apoptosis were assessed by MTT assay, flow cytometry and TUNEL staining. ROS level and mitochondrial membrane potential (MMP) were analyzed using DHE probe and JC-1 staining, respectively. FBXO3 m6A level was assessed using MeRIP assay. The interactions between FBXO3, YTHDF1, and PGC-1α were analyzed by Co-IP or RIP. RESULTS: Rg1 administration ameliorated LPS-induced epithelial cell inflammation, apoptosis, and mitochondrial dysfunction in a dose-dependent manner. Mechanically, Rg1 reduced PGC-1α ubiquitination modification level by inhibiting FBXO3 expression m6A-YTHDF1 dependently. As expected, Rg1's mitigative effect on LPS-induced inflammation, apoptosis and mitochondrial dysfunction in lung epithelial cells was abolished by FBXO3 overexpression. Moreover, FBXO3 upregulation eliminated the restoring effect of Rg1 on CLP-induced lung injury in rats. CONCLUSION: Rg1 activated PGC-1α/Nrf2 signaling pathway by reducing FBXO3 stability in an m6A-YTHDF1-dependent manner to improve mitochondrial function in lung epithelial cells during sepsis-induced ALI progression.

Andrographolide ameliorates sepsis-induced acute lung injury by promoting autophagy in alveolar macrophages via the RAGE/PI3K/AKT/mTOR pathway.

Autophagy in alveolar macrophages (AMs) is an important mechanism for maintaining immune homeostasis and normal lung tissue function, and insufficient autophagy in AMs may mediate the development of sepsis-induced acute lung injury (SALI). Insufficient autophagy in AMs and the activation of the NLRP3 inflammasome were observed in a mouse model with SALI induced by cecal ligation and puncture (CLP), resulting in the release of a substantial quantity of proinflammatory factors and the formation of SALI. However, after andrographolide (AG) intervention, autophagy in AMs was significantly promoted, the activation of the NLRP3 inflammasome was inhibited, the release of proinflammatory factors and pyroptosis were suppressed, and SALI was then ameliorated. In the MH-S cell model stimulated with LPS, insufficient autophagy was discovered to promote the overactivation of the NLRP3 inflammasome. AG was found to significantly promote autophagy, inhibit the activation of the NLRP3 inflammasome, and attenuate the release of proinflammatory factors. The primary mechanism of AG promoting autophagy was to inhibit the activation of the PI3K/AKT/mTOR pathway by binding RAGE to the membrane. In addition, it inhibited the activation of the NLRP3 inflammasome to ameliorate SALI. Our findings suggest that AG promotes autophagy in AMs through the RAGE/PI3K/AKT/mTOR pathway to inhibit the activation of the NLRP3 inflammasome, remodel the functional homeostasis of AMs in SALI, and exert anti-inflammatory and lung-protective effects. It has also been the first to suggest that RAGE is likely a direct target through which AG regulates autophagy, providing theoretical support for a novel therapeutic strategy in sepsis.

Extracellular Vesicles Derived from Human Adipose-Derived Mesenchymal Stem Cells Alleviate Sepsis-Induced Acute Lung Injury through a MicroRNA-150-5p-Dependent Mechanism.

Extracellular vesicles (EVs) derived from human adipose mesenchymal stem cells (hADSCs) may exert a therapeutic benefit in alleviating sepsis-induced organ dysfunction by delivering cargos that include RNAs and proteins to target cells. The current study aims to explore the protective effect of miR-150-5p delivered by hADSC-EVs on sepsis-induced acute lung injury (ALI). We noted low expression of miR-150-5p in plasma and bronchoalveolar lavage fluid samples from patients with sepsis-induced ALI. The hADSC-EVs were isolated and subsequently cocultured with macrophages. It was established that hADSC-EVs transferred miR-150-5p to macrophages, where miR-150-5p targeted HMGA2 to inhibit its expression and, consequently, inactivated the MAPK pathway. This effect contributed to the promotion of M2 polarization of macrophages and the inhibition of proinflammatory cytokines. Further, mice were made septic by cecal ligation and puncture in vivo and treated with hADSC-EVs to elucidate the effect of hADSC-EVs on sepsis-induced ALI. The in vivo experimental results confirmed a suppressive role of hADSC-EVs in sepsis-induced ALI. Our findings suggest that hADSC-EV-mediated transfer of miR-150-5p may be a novel mechanism underlying the paracrine effects of hADSC-EVs on the M2 polarization of macrophages in sepsis-induced ALI.

The Scutellaria-Coptis herb couple and its active small-molecule ingredient wogonoside alleviate cytokine storm by regulating the CD39/NLRP3/GSDMD signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: A drug pair is a fundamental aspect of traditional Chinese medicine prescriptions. Scutellaria baicalensis Georgi and Coptis chinensis Franch, commonly used as an herb couple (SBCC), are representative heat-clearing and dampness-drying drugs. They possess functions such as clearing heat, drying dampness, purging fire, and detoxifying. These herbs are used in both traditional and modern medicine for treating inflammation. AIM OF THE STUDY: This study investigated the effects of SBCC on cytokine storm syndrome (CSS) and explored its potential regulatory mechanism. MATERIALS AND METHODS: We assessed the impact of SBCC in a sepsis-induced acute lung injury mouse model by administering an intraperitoneal injection of LPS (15 mg/kg). The cytokine levels in the serum and lungs, the wet-to-dry ratio of the lungs, and lung histopathological changes were evaluated. The macrophages in the lung tissue were examined through transmission electron microscopy. Western blot was used to measure the levels of the CD39/NLRP3/GSDMD pathway-related proteins. Immunofluorescence imaging was used to assess the activation of pro-caspase-1 and ASC and their interaction. AMP-Glo™ assay was used to screen for active ingredients in SBCC targeting CD39. One of the ingredients was selected, and its effect on cell viability was assessed. We induced inflammation in macrophages using LPS + ATP and detected the levels of proinflammatory factors. The images of cell membrane large pores were captured using scanning electron microscopy, the interaction between NLRP3 and ASC was detected using immunofluorescence imaging, and the levels of CD39/NLRP3/GSDMD pathway-related proteins were assessed using Western blot. RESULTS: SBCC administration effectively mitigated LPS-induced cytokine storm, pulmonary edema and lung injury. Furthermore, it repressed the programmed death of lung tissue macrophages by inhibiting the NLRP3/GSDMD pyroptosis pathway and regulating the CD39 purinergic pathway. Based on the results of the AMP-Glo™ assay, we selected wogonoside for further valuation. Wogonoside alleviated LPS + ATP-induced inflammatory damage by regulating the inhibiting the NLRP3/GSDMD pyroptosis pathway and regulating the CD39 purinergic pathway. However, its effect on NLRP3 is not mediated though CD39. CONCLUSION: SBCC and its active small-molecule ingredient, wogonoside, improved CSS by regulating the NLRP3/GSDMD pyroptosis pathway and its upstream CD39 purinergic pathway. It is essential to note that the regulatory effect of wogonoside on NLRP3 is not mediated by CD39.

Monotropein alleviates sepsis-elicited acute lung injury via the NF-κB pathway.

OBJECTIVES: To address the effect and mechanism of Monotropein (Mon) on sepsis-induced acute lung injury (ALI). METHODS: ALI model was established by lipopolysaccharide (LPS)-stimulated mouse lung epithelial cell lines (MLE-12) and cecal ligation and puncture (CLP)-treated mice, respectively. The function of Mon was examined by cell counting kit-8 (CCK-8), pathological staining, the pulmonary function examination, flow cytometry, enzyme-linked immunosorbent assay, terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labellingand western blot. RESULTS: Mon increased the LPS-reduced viability but decreased the LPS-evoked apoptosis rate in MLE-12 cells. Mon suppressed the concentrations and protein expressions of proinflammatory factors, and the expressions of fibrosis-related proteins in LPS-challenged MLE-12 cells compared with LPS treatment alone. Mechanically, Mon downregulated the levels of NF-κB pathway, which was confirmed with the application of the receptor activator of nuclear factor-κB ligand (RANKL). Correspondingly, RANKL reversed the ameliorative effect of Mon on the proliferation, apoptosis, inflammation and fibrosis. Moreover, Mon improved the pathological manifestations, apoptosis, the W/D ratio and pulmonary function indicators in CLP-treated mice. Consistently, Mon attenuated inflammation, fibrosis and NF-κB pathway in CLP-treated mice. CONCLUSION: Mon inhibited apoptosis, inflammation and fibrosis to alleviate sepsis-evoked ALI via the NF-κB pathway.

Chromofungin, a chromogranin A-derived peptide, protects against sepsis-induced acute lung injury by inhibiting LBP/TLR4-dependent inflammatory signaling.

Chromofungin (CHR) is a biologically active peptide derived from chromogranin A that exhibits anti-inflammatory effects. However, it remains unclear whether and how CHR protects against sepsis-induced acute lung injury (ALI). A murine model of sepsis-induced ALI was established through cecal ligation and puncture, with intraperitoneal injection of CHR. Lung inflammation and macrophage polarization were examined by measuring the levels of cytokines and markers of M1 (CD86, inducible nitric oxide synthase [iNOS]) or M2 macrophages (arginase-1 [Arg1], resistin-like molecule α1 [Fizz1] and CD206). In vitro, mouse MH-S cells pretreated with CHR was employed to explore the interplay between the lipopolysaccharide-binding protein (LBP)/toll-like receptor 4 (TLR4) signaling pathway and M1/M2 polarity. The results revealed CHR's ability to enhance the 7-day survival rate and protect lung pathological injury in sepsis-induced ALI. CHR increased the expression of interleukin-4 and interleukin-10 but decreased the expression of tumour necrosis factor-α and interleukin-1ß. In addition, CHR notably facilitated M2 macrophage polarization, while significantly suppressingM1 polarization of alveolar macrophages. Mechanistic investigations delineated CHR's role in macrophage polarization by downregulating nuclear factor-κB expression through modulation of the LBP/TLR4 signaling pathway. Therefore, CHR may represent a novel strategy for the prevention of sepsis-induced ALI.

METTL3-mediated N6-methyladenosine exacerbates ferroptosis via m6A-IGF2BP2-dependent mitochondrial metabolic reprogramming in sepsis-induced acute lung injury.

Neutrophil extracellular traps (NETs), released by polymorphonuclear neutrophils (PMNs), exert a robust antimicrobial function in infectious diseases such as sepsis. NETs also contribute to the pathogenesis and exacerbation of sepsis. Although the lung is highly vulnerable to infections, few studies have explored the role of NETs in sepsis-induced acute lung injury (SI-ALI). We demonstrate that NETs induce SI-ALI via enhanced ferroptosis in alveolar epithelial cells. Our findings reveal that the excessive release of NETs in patients and mice with SI-ALI is accompanied by upregulation of ferroptosis depending on METTL3-induced m6A modification of hypoxia-inducible factor-1α (HIF-1α) and subsequent mitochondrial metabolic reprogramming. In addition to conducting METTL3 overexpression and knockdown experiments in vitro, we also investigated the impact of ferroptosis on SI-ALI caused by NETs in a caecum ligation and puncture (CLP)-induced SI-ALI model using METTL3 condition knockout (CKO) mice and wild-type mice. Our results indicate the crucial role of NETs in the progression of SI-ALI via NET-activated METTL3 m6A-IGF2BP2-dependent m6A modification of HIF-1α, which further contributes to metabolic reprogramming and ferroptosis in alveolar epithelial cells.