OBJECTIVES: Sepsis-associated acute lung injury (S-ALI) is one of the major causes of death in intensive care unit (ICU) patients, yet its mechanisms remain incompletely understood and effective therapies are lacking. Lytic cell death of macrophages is a key driver of the inflammatory cascade in S-ALI. PANoptosis, a newly recognized form of lytic cell death characterized by PANoptosome assembly and activation, involves plasma membrane rupture (PMR) mediated by ninjurin-1 (NINJ1), a recently identified pore-forming protein. Itaconic acid is known for its anti-inflammatory effects, but its role in macrophage PANoptosis during S-ALI is unclear. This study aims to investigate the protective effect of itaconic acid on macrophage PANoptosis in S-ALI to provide new therapeutic insights. METHODS: Male specific-pathogen-free C57BL/6J mice (6-8 weeks, 18-20 g) received intraperitoneal lipopolysaccharide (LPS) to establish a classical S-ALI model. Western blotting was used to assess PANoptosome-related proteins and enzymes involved in the itaconic acid metabolic pathway, while real-time reverse transcription polymerase chain reaction and metabolomics quantified itaconic acid levels. Primary peritoneal macrophages (PMs) were pretreated with the itaconate derivative 4-octyl itaconate (4-OI) and then exposed to tumor necrosis factor alpha (TNF-α) plus interferon gamma (IFN-γ) to induce PANoptosis. Cell viability was evaluated by cell counting kit-8 (CCK-8) assay. Western blotting was employed to quantify enzymes of the itaconate-metabolic pathway in PANoptotic macrophages, to evaluate the impact of 4-OI on PANoptosome-associated proteins, and to determine NINJ1 abundance in lung tissues from S-ALI mice and in PANoptotic macrophages. Fluorescent dye FM_4-64 was used to visualize 4-OI-mediated changes in PMR, whereas immunofluorescence staining mapped the effect of 4-OI on both the expression level and membrane localization of NINJ1 in PANoptotic macrophages. The effect of 4-OI on lactate dehydrogenase (LDH) release in culture supernatants and peripheal blood serum was assessed using a LDH assay kit, and non-denataring polyacylamide gel electrophoresis was used to assess the expression of NINJ1 in S-ALI mouse lung tissues and the impact of 4-OI on the expression of PANoptosis-associated NINJ1 multimeric reflected protein in macropahges. RESULTS: In S-ALI mouse lungs, PANoptosome components [NOD-like receptor thermal protein domain associated protein 3 (NLRP3), Gasdermin D (GSDMD), Caspase-1, Z-DNA binding protein (ZBP1), and Caspase-3] and phosphorylated mixed lineage kinase domain-like protein (MLKL) S345 were significantly upregulated (all P<0.05), while metabolomics showed compensatory increases in itaconic acid and its key enzymes [aconitate decarboxylase 1 (ACOD1)/immunoresponsive gene 1 (IRG1)]. In macrophages, 4-OI obviously suppressed PANoptosome protein expression, reduced LDH release, restored plasma membrane integrity, and inhibited NINJ1 expression and oligomerization at the membrane (P<0.05). CONCLUSIONS Itaconic acid may alleviate macrophage PANoptosis in S-ALI by inhibiting NINJ1-mediated plasma membrane rupture. Targeting NINJ1 or enhancing itaconate pathways may offer a novel therapeutic strategy for S-ALI.
Animals ; Acute Lung Injury/pathology* ; Succinates/pharmacology* ; Sepsis/complications* ; Mice, Inbred C57BL ; Male ; Mice ; Macrophages/pathology* ; Cell Membrane/metabolism* ; Lipopolysaccharides ; Hydro-Lyases

Objective To observe the clinical effect of Huangqi Yixin Decoction combined with sacubitril/valsartan in treatment of patients with heart failure with preserved ejection fraction(HFpEF)of Qi deficiency and blood stasis type.Methods A total of 86 patients with HFpEF of Qi deficiency and blood stasis type were randomly divided into western medicine group and combination group using the random number table method,with 43 cases in each group.The western medicine group was trea-ted with sacubitril/valsartan sodium tablets on the basis of conventional anti-heart failure therapy,while the combination group was additionally treated with Huangqi Yixin Decoction on the basis of the western medicine group's treatment.The therapeutic efficacy,traditional Chinese medicine(TCM)syndrome scores,cardiac function indicators[Tei index,N-terminal pro-brain natriuretic peptide(NT-proBNP)],exercise tolerance[6-minute walk distance(6MWD),maximum exercise heart rate,metabolic equivalent(MET)],and the occurrence of adverse reactions were compared between the two groups.Results The overall clinical effective rate in the combination group was 93.02％,which was higher than that in the western medicine group(76.74％),and the difference was statistically significant(P＜0.05).After treatment,the TCM syndrome scores in both groups were lower than those before treatment,and the scores in the combination group were lower than those in the western medicine group,with a statistically significant difference(P＜0.05).After treatment,the levels of Tei index and NT-proBNP in both groups were lower than those before treatment,and the levels in the combination group were lower than those in the western medicine group,with a statistically sig-nificant difference(P＜0.05).After treatment,the 6MWD,maximum exercise heart rate,and MET in both groups were higher than those before treatment,and the values in the combination group were higher than those in the western medicine group,with a statistically significant difference(P＜0.05).The incidence of adverse reactions in the western medicine group was 18.60％,which was higher than that in the combination group(4.65％),and the difference was statistically signifi-cant(P＜0.05).Conclusion Huangqi Yixin Decoction combined with sacubitril/valsartan has a good therapeutic effect on HFpEF of Qi deficiency and blood stasis type.It can reduce the TCM syn-drome scores of patients,improve exercise tolerance,and has good safety.

Objective:To explore the association between cognitive function and the level of gamma-amino-butyric acid(GABA)in anterior cingulate cortex(ACC)before and after treatment in patients with major depres-sion disorder.Methods:Totally 31 medication-naive patients with major depression disorder meeting the criteria of the Diagnostic and Statistical Manual of Mental Disorders,Fifth Edition(DSM-5)and 33 normal controls were col-lected.Each eligible patient received treatment with selective serotonin reuptake inhibitor agents for 8 weeks.The MATRICS Consensus Cognitive Battery(MCCB)was used to evaluate the cognitive function.By means of 1H magnetic resonance spectroscopy,anterior cingulate cortex GABA concentrations were measured.Results:At base-line,the concentration of ACC GABA relative to water(GABA+/W)was lower in the patient group than in the control group(P＜0.05)and increased after treatment(P＜0.05).ACC GABA+/W was negatively associated with verbal learning and visual memory score in patient group at baseline(correlation coefficient and P value were r=-0.40,P＜0.05;r=-0.42,P＜0.05,respectively).The ACC GABA+/W difference resulted of treatment in patient group was positively associated with the difference of working memory score and the difference of reasoning and problem-solving score(correlation coefficient and P value were r=0.58,P＜0.05;r=0.66,P＜0.05,respec-tively).Conclusion:The cognitive dysfunction of patients with major depression disorder may not be related to the degree of depression and anxiety.And improvement of cognitive function may be associated with increase of ACC GABA concentrations.

Objective:To assess if metagenomic next-generation sequencing (mNGS) of periprosthetic joint tissue can provide an alternative rapid and sensitive tool for the diagnosis of prosthetic joint infection (PJI), especially compared to microbiological culture.Methods:A total of 33 eligible patients who underwent revision arthroplasty from June 2019 to June 2020 in orthopedic surgery department of the first affiliated hospital of Zhengzhou University were retrospectively analyzed. Twenty-one patients were included in PJI group according to the American Academy of Musculoskeletal Infection diagnostic criteria, with 17 cases of knee and 4 cases of hip, including 9 cases of male and 12 cases of female, with an average age of 59.14±14.55 years old (range from 28 to 84), and an average BMI of 23.7±2.8 kg/m 2 (range from 17.7 to 29.4 kg/m 2). Twelve patients were included in aseptic loosening group (control group), with 4 cases of knee and 8 cases of hip, including 4 cases of male and 8 cases of female, with an average age of 53.08±10.05 years old (range from 39 to 70), and an average BMI of 25.2±2.9 kg/m 2 (range from 18.3 to 31.2 kg/m 2 ). Microbiological culture results of synovial fluid and periprosthetic joint tissue and mNGS results of periprosthetic joint tissue were collected. The sensitivity and specificity of mNGS and microbiological culture were calculated and compared. The species of pathogenic microorganismsdetected by the two techniques were summarized. In addition, the impact of antibiotic use on the efficacy of both techniques were compared. Results:mNGS detected 13 positive cases and microbiological culture detected 6 positive cases in the PJI group. In the aseptic loosening group, 1 case was determined positive by mNGS, and all the microbiological culture results were negative. In the diagnosis of PJI, mNGS showed significantly higher sensitivity than that of culture (61.9% vs 28.6%, χ2=4.71, P=0.03), while no statistical difference was observed in terms of specificity (91.7% vs 100%, χ2=1.04, P=0.31). In the PJI cases with prior exposure to antibiotics within two weeks, the sensitivity of mNGS was significantly higher than that of culture (53.8% vs 15.4%, χ2=4.25, P=0.04). However, there was no significant difference in the sensitivity between mNGS and culture in patients without antibioticsexposure (66.7% vs 44.4%, χ2=0.90, P=0.34). In the detection of pathogenic microorganism, mNGS detected 9 kinds of bacteria (Staphylococcus aureus, Staphylococcus family, Moraxella Oslo, Propionibacterium acnes, Streptococcus acnes, Staphylococcus epidermidis, Mycobacterium tuberculosis, Staphylococcus Lyons, Bacteroides fragilis) and 2 kinds of fungi (Aspergillus fumigatus, Candida parapsilosis), while microbiological culture detected 3 kinds of bacteria (Staphylococcus aureus, Moraxella catarrhalis, Mycobacterium tuberculosis) and one kind of fungi (Candida parapsilosis). mNGS and microbiological culture were both positive in 5 cases, among which 3 cases had completely matched results (Staphylococcus aureus, Mycobacterium tuberculosis, Candida parapsilosis), one case had partly matched results (mNGS detected more bacteria than culture) and one case had totally mismatched results. Additionally, in the diagnosis of the 3 included tuberculous PJI, mNGS showed 100% specificity and sensitivity. Conclusion:mNGS of periprosthetic joint tissue is a more powerful tool for diagnosis and pathogen detection of PJI compared to microbiological culture, especially in the diagnosis of tuberculosis PJI. Besides, mNGS is more resistant to antibiotic exposure than culture.

Objective To study the effects of human insulin-like growth factor 1(hIGF-1) gene transfection on the proliferation of NIH3T3 fibroblasts.Methods The plasmid of pcDNA3.1-hIGF-1 was transfected into NIH3T3 fibroblasts by using Lipofectin method.The positive cell clones were selected with G418 and cultured for 4 weeks.The stable expression of hIGF-1 in the positive cells was determined by in situ hybridization and immunocytochemical analysis.MTT assay and flow cytometer analysis were used to observe the proliferation of NIH3T3 fibroblasts.Results hIGF-1 mRNA and protein expressed in NIH3T3 fibroblasts transfected with pcDNA3.1-hIGF-1 by in situ hybridization and immunocytochemical analysis.MTT assay showed the A value of transfected NIH3T3 fibroblasts rose,compared with untransfected NIH3T3 fibroblasts group,the difference was significant(P