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BACKGROUND: Pediatric sepsis is a common and complex syndrome characterized by a dysregulated immune response to infection. Aberrations in the renin-angiotensin system (RAS) are factors in several infections of adults. However, the precise impact of RAS dysregulation in pediatric sepsis remains unclear. METHODS: Serum samples were collected from a derivation cohort (58 patients with sepsis, 14 critically ill control subjects, and 37 healthy controls) and validation cohort (50 patients with sepsis, 37 critically ill control subjects, and 46 healthy controls). Serum RAS levels on day of PICU admission were determined and compared with survival status and organ dysfunction. RESULTS: In the derivation cohort, the serum renin concentration was significantly higher in patients with sepsis (3678 ± 4746) than that in healthy controls(635.6 ± 199.8) (p < 0.0001). Meanwhile, the serum angiotensin (1-7) was significantly lower in patients with sepsis (89.7 ± 59.7) compared to that in healthy controls(131.4 ± 66.4) (p < 0.01). These trends were confirmed in a validation cohort. Non-survivors had higher levels of renin (8207 ± 7903) compared to survivors (2433 ± 3193) (p = 0.0001) and lower levels of angiotensin (1-7) (60.9 ± 51.1) compared to survivors (104.0 ± 85.1) (p < 0.05). A combination of renin, angiotensin (1-7) and procalcitonin achieved a model for diagnosis with an area under the receiver operating curve (AUROC) of 0.87 (95% CI: 0.81-0.92). CONCLUSION: Circulating renin and angiotensin (1-7) have predictive value in pediatric sepsis.
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The role of inflammatory cytokines in children with moderate to severe TBI (m-sTBI) is still incompletely understood. We aimed to investigate the associations between early plasma expression profiles of inflammatory cytokines and clinical outcomes in children with m-sTBI. We prospectively recruited children admitted to the intensive care unit (ICU) of a tertiary pediatric hospital due to m-sTBI from November 2022 to May 2023. Plasma interleukin (IL)-1ß, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, interferon (IFN)-α, IFN-γ and tumor necrosis factor (TNF)-α concentrations were detected by flow cytometry on admission and on days 5 to 7. The primary outcome was in-hospital mortality. The secondary outcome was the 6-month functional outcome assessed by the Glasgow Outcome Scale Extended-Pediatrics (GOS-E Peds) score, dichotomized as favorable (1-4) or unfavorable (5-8). Fifty patients and 20 healthy controls were enrolled. Baseline IL-6, IL-8 and IL-10 levels were significantly higher in TBI patients than in healthy controls. Twelve patients died in the hospital. Compared with survivors, nonsurvivors had significantly increased baseline IL-6 and IL-8 levels. Baseline IL-5, IL-6 and IL-8 levels were also significantly greater in children with unfavorable versus favorable outcomes. The area under the receiver operating characteristic curve (AUC) of the IL-6 and IL-8 levels and motor Glasgow Coma Scale (GCS) score for predicting in-hospital mortality was 0.706, 0.754, and 0.776, respectively. Baseline IL-1ß, IL-2, IL-4, IL-10, IL-12p70, IL-17A, IFN-γ, IFN-α and TNF-α levels were not associated with in-hospital mortality or an unfavorable 6-month outcome. On days 5 to 7, the IL-6 and IL-8 levels were significantly decreased in survivors but increased in nonsurvivors compared to their respective baselines. CONCLUSION: After m-sTBI, the plasma profiles of inflammatory cytokines are markedly altered in children. The trends of IL-6 and IL-8 expression vary among m-sTBI children with different outcomes. Elevated plasma IL-6 and IL-8 levels are related to in-hospital mortality and unfavorable 6-month outcomes. TRIAL REGISTRATION: This trial was registered in the Chinese Clinical Trial Registry (Registration number: ChiCTR2200065505). Registered November 7, 2022. WHAT IS KNOWN: ⢠Inflammation is an important secondary physiological response to TBI. WHAT IS NEW: ⢠The plasma profiles of inflammatory cytokines are markedly altered in children with m-sTBI. Elevated IL-6 and IL-8 levels are related to mortality and unfavorable outcomes.
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Lesões Encefálicas Traumáticas , Citocinas , Humanos , Masculino , Feminino , Estudos Prospectivos , Criança , Citocinas/sangue , Lesões Encefálicas Traumáticas/sangue , Lesões Encefálicas Traumáticas/mortalidade , Pré-Escolar , Biomarcadores/sangue , Mortalidade Hospitalar , Estudos de Casos e Controles , Adolescente , Prognóstico , Lactente , Escala de Resultado de GlasgowRESUMO
Adipose-derived stromal cells (ADSCs) are promising stem cell sources for tissue engineering and cell-based therapy. However, long-term in vitro expansion of ADSCs impedes stemness maintenance, which is partly attributed to deprivation of their original microenvironment. Incompetent cells limit the therapeutic effects of ADSC-based clinical strategies. Therefore, reconstructing a more physiologically and physically relevant niche is an ideal strategy to address this issue and therefore facilitates the extensive application of ADSCs. Here, we transplanted separated ADSCs into local subcutaneous adipose tissues of nude mice as an in vivo cell culture model. We found that transplanted ADSCs maintained their primitive morphology and showed improved proliferation and delayed senescence compared to those of cells cultured in an incubator. Significantly increased expression of stemness-related markers and multilineage differentiation abilities were further observed in in vivo cultured ADSCs. Finally, sequencing revealed that genes whose expression differed between ADSCs obtained under in vivo and in vitro conditions were mainly located in the extracellular matrix and extracellular space and that these genes participate in regulating transcription and protein synthesis. Moreover, we found that an Egr1 signaling pathway might exert a crucial impact on controlling stemness properties. Our findings might collectively pave the way for ADSC-based applications.
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Diferenciação Celular , Proliferação de Células , Animais , Ratos , Diferenciação Celular/fisiologia , Camundongos , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Camundongos Nus , Células Estromais/metabolismo , Células Estromais/citologia , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/citologia , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Masculino , Células CultivadasRESUMO
OBJECTIVES: To describe the epidemiological characteristics of pediatric sepsis in Southwest China PICUs. DESIGN: A prospective, multicenter, and observational study. SETTING: Twelve PICUs in Southwest China. PATIENTS: The patients admitted to the PICU from April 1, 2022, to March 31, 2023. The age ranged from 28 days to 18 years. All patients met the criteria of severe sepsis or septic shock. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Of the 31 PICUs invited to participate, 12 PICUs (capacity of 292 beds) enrolled patients in the study. During the study period, 11,238 children were admitted to the participating PICUs, 367 (3.3%) of whom met the diagnosis of severe sepsis or septic shock. The most prevalent sites of infection were the respiratory system (55%) and the digestive system (15%). The primary treatments administered to these patients included antibiotics (100%), albumin (61.3%), invasive mechanical ventilation (58.7%), glucocorticoids (55.6%), blood products (51%), gammaglobulin (51%), and vasoactive medications (46.6%). Sepsis-related mortality in the PICU was 11.2% (41/367). Nearly half of the sepsis deaths occurred within the first 3 days of PICU admission (22/41, 53.7%). The mortality rate of septic shock (32/167, 19.2%) was significantly higher than that of severe sepsis (9/200, 4.5%; p < 0.001). The outcomes of a multivariate logistic regression analysis suggested that a higher pediatric Sequential Organ Failure Assessment score, and the use of invasive mechanical ventilation and vasoactive medications were independently associated with PICU mortality in children with sepsis. CONCLUSIONS: This report updates the epidemiological data of pediatric sepsis in PICUs in Southwest China. Sepsis is still a life-threatening disease in children.
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Unidades de Terapia Intensiva Pediátrica , Sepse , Humanos , Estudos Prospectivos , Pré-Escolar , China/epidemiologia , Criança , Lactente , Masculino , Feminino , Adolescente , Unidades de Terapia Intensiva Pediátrica/estatística & dados numéricos , Sepse/epidemiologia , Recém-Nascido , Mortalidade Hospitalar , Choque Séptico/epidemiologiaRESUMO
BACKGROUND: Sepsis is a severe condition characterized by acute organ dysfunction resulting from an imbalanced host immune response to infections. Apolipoprotein H (APOH) is a critical plasma protein that plays a crucial role in regulating various biological processes. However, the precise role of APOH in the immunopathology of paediatric sepsis remains unclear. METHODS: In this study, we evaluated the concentration of APOH in paediatric patients with sepsis and healthy individuals. In an experimental sepsis model of caecal ligation and puncture (CLP), the impact of APOH on survival, organ injury, and inflammation was measured. Furthermore, the anti-inflammatory effects of APOH were investigated across diverse immune cell types, encompassing peripheral blood mononuclear cells (PBMCs), peritoneal macrophages (PMs), bone marrow-derived macrophages (BMDMs), and RAW 264.7 macrophages. RESULTS: In the pilot cohort, the relative abundance of APOH was found to be decreased in patients with sepsis (2.94 ± 0.61) compared to healthy controls (1.13 ± 0.84) (p < 0.001), non-survivors had lower levels of APOH (0.50 ± 0.37) compared to survivors (1.45 ± 0.83) (p < 0.05). In the validation cohort, the serum concentration of APOH was significantly decreased in patients with sepsis (202.0 ± 22.5 ng/ml) compared to healthy controls (409.5 ± 182.9 ng/ml) (p < 0.0001). The application of recombinant APOH protein as a therapeutic intervention significantly lowered the mortality rate, mitigated organ injury, and suppressed inflammation in mice with severe sepsis. In contrast, neutralizing APOH with an anti-APOH monoclonal antibody increased the mortality rate, exacerbated organ injury, and intensified inflammation in mice with non-severe sepsis. Intriguingly, APOH exhibited minimal effects on the bacterial burden, neutrophil, and macrophage counts in the sepsis mouse model, along with negligible effects on bacterial phagocytosis and killing during Pseudomonas aeruginosa infection in PMs, RAW 264.7 cells, and PBMCs. Mechanistic investigations in PMs and RAW 264.7 cells revealed that APOH inhibited M1 polarization in macrophages by suppressing toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signalling pathway. CONCLUSION: This proof-of-concept study demonstrated that APOH has a protective role in the host defense response to sepsis, highlighting the potential therapeutic value of APOH in sepsis treatment.
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Leucócitos Mononucleares , Sepse , Animais , Criança , Humanos , Camundongos , beta 2-Glicoproteína I , Inflamação , Leucócitos Mononucleares/metabolismo , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , NF-kappa B/farmacologia , NF-kappa B/uso terapêutico , Fagocitose , Apolipoproteínas/metabolismoRESUMO
OBJECTIVE: To investigate lncRNAs and their roles in regulating the pulmonary inflammatory response under dexamethasone (Dex) treatment. METHODS: IL-1ß (10 ng/mL) and LPS (1 µg/mL) was used to construct inflammatory cell models with A549 cells; IL-1ß performed better against LPS. Different concentrations of Dex were used to attenuate the inflammation induced by IL-1ß, and its effect was assessed via RT-PCR to detect inflammatory cytokine-related mRNA levels, including those of IKß-α, IKKß, IL-6, IL-8, and TNF-α. Furthermore, ELISA was used to detect the levels of the inflammatory cytokines TNF-α, IL-6, and IL-8. RT-PCR was used to quantify the levels of lncRNAs, including lncMALAT1, lncHotair, lncH19, and lncNeat1. LncH19 was most closely associated with the inflammatory response, which was induced by IL-1ß and attenuated by Dex. Among the lncRNAs, the level of lncH19 showed the highest increase following treatment with 1 and 10 µM Dex. Therefore, lncH19 was selected for further functional studies. LncH19 expression was inhibited by shRNA transduced with lentivirus. Cell assays for cell proliferation and apoptosis as well as RT-PCR, western blot, and ELISA for inflammatory genes were conducted to confirm the functions of lncH19. The predicted target miRNAs of lncH19 were hsa-miR-346, hsa-miR-324-3p, hsa-miR-18a-3p, hsa-miR-18b-5p, hsa-miR-146b-3p, hsa-miR-19b-3p, and hsa-miR-19a-3p. Following estimation via RT-PCR, hsa-miR-346, hsa-miR-18a-3p, and hsa-miR-324-3p showed consistent patterns in A549 NC and A549 shlncH19. An miRNA inhibitor was transfected into A549 NC and A549 shlncH19 cells, and the expression levels were determined via RT-PCR. hsa-miR-324-3p was inhibited the most compared with hsa-miR-346 and hsa-miR-18a-3p and was subjected to further functional studies. RT-PCR, ELISA, and western blotting for inflammatory gene detection were conducted to validate the functions of the target hsa-miR-324-3p. RESULTS: Treatment with 1 and 10 µM Dex could effectively attenuate the inflammatory response. During this process, lncH19 expression significantly increased (P < 0.05). Therefore, treatment with 1 µM Dex was used for further study. Under IL-1ß treatment with or without Dex, lncH19 inhibition led to an increase in cell proliferation; a decrease in cell apoptosis; an increase in the protein levels of inflammatory genes; phosphorylation of P65, ICAM-1, and VCAM-1; and increase inflammatory cytokines. Prediction of the targets of lncH19 and validation via RT-PCR revealed that miR-346, miR-18a-3p, and miR-324-3p negatively correlate with lncH19. Additionally, Dex increased the lncH19 expression but reduced that of the miRNAs. Among the miRNAs, miR-324-3p was the most markedly downregulated miRNA following treatment of miRNA inhibitors. The MTS assay and cell apoptosis assay showed that the miR-324-3p inhibitor inhibited cell proliferation and induced cell apoptosis, thereby significantly attenuating the inflammatory response, which reversed the effect of lncH19 in regulating cell proliferation and the secretion of inflammatory cytokines (P < 0.05). Therefore, lncH19 might regulate miR-324-3p in pulmonary inflammatory response under Dex treatment. CONCLUSION: Dex can attenuate the pulmonary inflammatory response by regulating the lncH19/miR-324-3p cascade.
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Sepsis or septic shock is often accompanied by organ dysfunction, among which acute kidney injury (AKI) is the most frequent event that appears early during sepsis. To harness urinary metabolic profiling to discover potential biomarkers of septic acute kidney injury in pediatric patients at intensive care units, we collected urine samples from 27 septic children with AKI and 30 septic children without AKI. We used ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS) for profiling and multiple regression analysis to explore the potential biomarkers of sepsis with AKI. We identified a clear distinction in the UPLC-QTOF/MS results for septic children with and without AKI after the development of sepsis, specifically 18 and 17 metabolites with different levels at 12 and 24 h, respectively. Metabolic pathways associated with septic AKI included lipid metabolism, particularly processes involving glycerophospholipid metabolism. L-Histidine, DL-indole-3-lactic acid, trimethylamine N-oxide, and caprylic acid were uncovered as potential biomarkers of septic AKI at 12 h, while gentisaldehyde, 3-ureidopropionate, N4-acetylcytidine, and 3-methoxy-4-hydroxyphenylglycol sulfate were identified as potential candidates at 24 h. We further found that combinations of metabolites were more effective diagnostic marker compared with individual metabolites, with an area under the receiver operating characteristics curve of 0.905 and 0.97 at 12 and 24 h, respectively. Our results indicated that metabolomic analysis could be a promising approach for identifying diagnostic biomarkers of pediatric septic AKI and helped elucidate the pathological mechanisms involved.
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Injúria Renal Aguda/diagnóstico , Injúria Renal Aguda/metabolismo , Unidades de Terapia Intensiva , Sepse/diagnóstico , Sepse/metabolismo , Espectrometria de Massas em Tandem/métodos , Biomarcadores/metabolismo , Biomarcadores/urina , Criança , Pré-Escolar , Cromatografia Líquida de Alta Pressão/métodos , Feminino , Humanos , Masculino , Metabolômica/métodos , Projetos Piloto , Estudos ProspectivosRESUMO
OBJECTIVES: To estimate the prevalence, management, and outcomes of pediatric severe sepsis in the main PICUs in Southwest China. DESIGN: A prospective, observational, and multicenter study. SETTING: Eight PICUs in Southwest China with 19 (13-24) beds and 1,322 (1,066-1,452) annual admissions each. PATIENTS: A total of 10,598 patients (29 d to 18 yr old) were consecutively admitted between September 1, 2016, and August 31, 2017. All patients were screened and evaluated for severe sepsis or septic shock. Of them, 10,353 patients were excluded due to incomplete data or not meeting the consensus criteria for severe sepsis or septic shock; 245 patients were included with complete data. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Finally, 245 patients who were diagnosed with severe sepsis or septic shock were included in the study, with an incidence rate of 2.3%. Of them, 64.0% of the enrolled patients were male with 80.8% being less than 5 years old and 60.8% being from rural areas. The respiratory system was the most common organ system in which dysfunction was observed (76.7%) as well as the most frequently infected site (37.6%). The primary therapies were antibiotics (99.0%), immunoglobulin (88.3%), mechanical ventilation (78.4%), vasoactive infusions (59.6%), and corticosteroids (46.1%). Among the 188 patients who had respiratory dysfunction, 173(92%) required mechanical ventilation and 39 (20.7%) met the criteria for pediatric acute respiratory distress syndrome. Seven of the patients with pediatric acute respiratory distress syndrome died (7/39, 17.9%). The median durations for mechanical ventilation and vasoactive medications were 123.5 hours (35.25-226.00 hr) and 2 days (1-5 d), respectively. Eighty-six percent of patients had multiple organ dysfunction syndrome at the point at which severe sepsis was recognized, and 31% had underlying conditions. The hospital mortality rate was 18.8%. CONCLUSIONS: This report is the first to present the prevalence, treatment, and outcomes of pediatric severe sepsis in the main PICU centers in Southwest China. The mortality rate remains high; therefore, improved clinical management and implementation of large-scale clinical trials are necessary to improve early diagnoses and treatment.
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Unidades de Terapia Intensiva Pediátrica/estatística & dados numéricos , Sepse/epidemiologia , Sepse/fisiopatologia , Adolescente , Corticosteroides/uso terapêutico , Antibacterianos/uso terapêutico , Criança , Pré-Escolar , China/epidemiologia , Comorbidade , Feminino , Humanos , Imunoglobulinas/uso terapêutico , Lactente , Masculino , Escores de Disfunção Orgânica , Respiração Artificial , Sepse/mortalidade , Sepse/terapia , Choque Séptico/epidemiologia , Choque Séptico/fisiopatologiaRESUMO
OBJECTIVE: To investigate the damage mechanism of type II alveolar epithelial cells (AEC II) after hyperoxia exposure by proteomics. METHODS: The primary AEC II of preterm Sprague-Dawley (SD) rats were divided into normoxia and hyperoxia groups, and cultured in room air (21% O2) or hyperoxia (95% O2) condition, respectively. The cell morphology change was observed under an inverted contrast microscope; the protein expressions of Bcl-2 and caspase-3 were detected by Western Blot to ensure a successful model. Total protein in AEC II was collected, and mass spectrometry-based tandem mass tag (TMT)-labeled quantitative proteomics were used to detect the change of protein profile. Proteins with changes greater than 1.5-fold and P < 0.05 were considered differentially expressed, and bioinformatics analysis was performed. According to the proteomic results, AEC II were divided into three groups: normoxia group, hyperoxia group and hyperoxia+MW167 group (γ-secretase inhibitor MW167 was added to culture medium 30 minutes before they were placed into the chamber). The cell viability was detected by the cell proliferation and toxicity kit (CCK-8), and the expressions of Hes1, Bax mRNA were detected by real-time fluorescence quantitative reverse transcription-polymerase chain reaction (qRT-PCR). RESULTS: (1) The cells in the normoxia group proliferated and prolonged significantly, and the cytoplasmic particulate matter was abundant. In the hyperoxia group, nucleus pyknosis and cytoplasmic particulate matter decreased significantly. Compared with the normoxia group, the expression of caspase-3 in the hyperoxia group was significantly increased, and the expression of Bcl-2 was significantly decreased (caspase-3/GAPDH: 1.352±0.086 vs. 0.769±0.080, Bcl-2/GAPDH: 0.614±0.060 vs. 1.361±0.078, both P < 0.01). (2) A total of 162 differentially expressed proteins were identified between normoxia and hyperoxia groups, the proteins up-regulated by hyperoxia were commonly associated with response processes to various stimuli, and located in the extracellular region; the proteins down-regulated by hyperoxia were commonly associated with synthesis of substances, and located in the cellular matrix. KEGG Pathway analyses suggested that metabolism by cytochrome P450, oxidative phosphorylation, and Notch signaling pathway were associated with the mechanism of hyperoxia injury on AEC II. (3) Compared with the normoxia group, the viability of cells in the hyperoxia group was significantly decreased, and the expressions of Hes1 and Bax mRNA were significantly increased [cell viability (A value): 0.060±0.003 vs. 1.058± 0.017, Hes1 mRNA (2-ΔΔCt): 2.235±0.606 vs. 1.144±0.107, Bax mRNA (2-ΔΔCt): 2.210±0.240 vs. 1.084±0.096, all P < 0.05]. Compared with the hyperoxia group, the viability of cells in the hyperoxia+MW167 group was significantly increased, and the expressions of Hes1 and Bax mRNA were significantly decreased [cell viability (A value): 0.271±0.025 vs. 0.060±0.003, Hes1 mRNA (2-ΔΔCt): 0.489±0.046 vs. 2.235±0.606, Bax mRNA (2-ΔΔCt): 1.289±0.041 vs. 2.210±0.240, all P < 0.05]. CONCLUSIONS: The mechanism of hyperoxia injury on AECII may be related to the metabolism by cytochrome P450, oxidative phosphorylation and activation of Notch signaling pathway.
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Células Epiteliais Alveolares/metabolismo , Hiperóxia/complicações , Lesão Pulmonar/etiologia , Proteoma/metabolismo , Animais , Proteômica , Ratos , Ratos Sprague-DawleyRESUMO
Purpose/Aim: Exposure to hyperoxia leads to lung injury both in vivo and in vitro, molecular hydrogen has been reported to protect against hyperoxia-induced lung injury; however, the underlying molecular mechanisms remain largely unknown. The objective of this study was to characterize differentially regulated proteins and biological processes in hydrogen-treated hyperoxic primary type II alveolar epithelial cells (AECIIs) to elucidate the protective mechanism of hydrogen using quantitative proteomics. Materials and Methods: AECIIs were divided into three groups that were cultured for 24 h in three different conditions: control (21% oxygen), hyperoxia (95% oxygen), and hyperoxia + hydrogen. Morphologic examination, flow cytometric analysis, cell viability assessment and analysis of the expression of apoptosis-associated proteins Bax and Bcl-2 as well as AECI markers (AQP5, T1α) and an AECII marker (SP-C) were performed for each group. The TMT labeling quantitative proteome technique was used to detect changes in the protein expression profile, and bioinformatics analysis was performed. Results: Hydrogen plays a protective role in hyperoxia-induced damage in AECIIs, as evidenced by reduced apoptosis, increased viability and survival, improved morphology, and enhanced transdifferentiation of AECIIs into AECIs. A total of 5782 proteins were identified in our study, of which 162 were significantly altered in abundance after hyperoxia exposure, and 97 were significantly altered in abundance in response to hydrogen treatment. The Gene Ontology and KEGG enrichment analyses identified a large number of proteins and biological processes that may responsible for the protective effect of hydrogen, including VEGFA, PDGFB, IGFBP3, EDN1, NADPH oxidase, the coagulation cascade, etc. Conclusions: Molecular hydrogen protects AECIIs from hyperoxic injury by complex mechanisms involving a variety of proteins and biological processes, such as VEGFA, PDGFB, IGFBP3, EDN1, NADPH oxidase and the coagulation cascade. These findings suggest novel pathways that need to be investigated as possible therapeutic targets for hyperoxia-induced lung injury.