OBJECTIVE To explore the molecular mechanism of Xijiao Dihuang Decoction(XJDHT)in inhibiting inflammatory response in sepsis based on network pharmacology,molecular docking and in vitro and in vivo experiments.METHODS Active com-ponents of XJDHT were screened using the TCMSP and HERB databases.Sepsis-related targets and histone H3K36 trimethylation(H3K36me3)-associated targets were retrieved from GeneCard,OMIM,and DisGeNet databases.A protein-protein interaction(PPI)network was constructed using the STRING database,and core targets were identified via Cytoscape 3.9.1.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses were performed to predict potential mechanisms.Mo-lecular docking validated ligand-receptor binding affinity.A cecal ligation and puncture(CLP)model was established in mice to evalu-ate 24-hour sepsis scores(MSS)and survival rates.Blood routine parameters were analyzed using an automated hematology analyzer.Serum IL-1β and TNF-α levels were measured by ELISA.Liver histopathology was assessed via HE staining,and H3K36me3 expression in Kupffer cells was detected by immunofluorescence.In vitro,LPS-induced THP-1 cells were used as an in-flammatory model.ChIP-qPCR evaluated H3K36me3 enrichment at IL-1β and TNF-α gene loci.Western blot analyzed HIF-1α,EGFR,and AKT1 pathway proteins.RESULTS A total of 28 active components of XJDHT were identified,corresponding to 987 gene targets,with 189 overlapping sepsis-related targets.Core targets included TNF,IL1B,and GAPDH.Enriched pathways includ-ed EGFR tyrosine kinase inhibitor resistance.Molecular docking confirmed strong binding between core components and key targets.In vivo,compared to the sham group,the CLP group exhibited significantly reduced 24-hour survival(P<0.01),elevated MSS(P<0.01),immune imbalance,and increased serum IL-1β and TNF-α levels(P<0.01).High-and low-dose XJDHT intervention im-proved survival(P<0.01),reduced MSS(P<0.01),restored immune homeostasis,and dose-dependently suppressed IL-1β and TNF-α(P<0.01).CLP mice showed elevated H3K36me3 in Kupffer cells and severe hepatic inflammation,while XJDHT dose-de-pendently reduced H3K36me3(P<0.05)and attenuated liver injury.In peritoneal macrophages,CLP upregulated H3K36me3,IL-1β,TNF-α,HIF-1α,p-AKT1/AKT1,and EGFR(P<0.01),which were reversed by XJDHT(P<0.05,P<0.01).In vitro,LPS increased H3K36me3 and IL-1β and TNF-α expression(P<0.01),with ChIP-qPCR confirming H3K36me3 enrichment at IL-1β lo-ci(P<0.01).Treatment with 15%XJDHT-containing serum for 24 h reduced H3K36me3(P<0.01),diminished its recruitment to IL-1β loci(P<0.01),and inhibited LPS-induced activation of EGFR,HIF-1α,and p-AKT1/AKT1(P<0.05,P<0.01).HIF-1α agonist Dimethyloxallyl Glycine(DMOG)further validated that XJDHT suppressed H3K36me3-mediated epigenetic remodeling via HIF-1α-related pathways.CONCLUSION XJDHT inhibits inflammatory responses,regulates immune homeostasis,and improves sepsis prognosis,potentially by modulating H3K36me3 epigenetic modifications at IL-1β loci.

Objective To analyze clinical effect of digital template combined with limited incision reduction and fixation in treatment of ankle fractures with degree Ⅳ supination and external rotation.Methods A total of 73 patients with posterior ankle rotation fractures admitted to Jinhua Hospital of Traditional Chinese Medicine,Zhejiang Chinese Medical University from January 2020 to June 2023 were selected as study subjects.According to the surgical method,32 patients treated with conventional open reduction and internal fixation were selected as control group,and 41 patients treated with digital template combined with limited open reduction and fixation technique were selected as observation group.Perioperative related indexes,ankle function and complications were compared between two groups.Results Time of operation and length of incision in observation group were shorter than those in control group,time of getting out of bed,time of hospitalization in observation group were shorter than those in control group(P＜0.01).The Kofoed scores at 1 and 6 months after operation in observation group were higher than those in control group(P＜0.05).The angles of ankle dorsiflexion and plantar flexion at 1 and 6 months after operation in observation group were higher than those in control group(P＜0.05).The score of arthritis quality of life measurement scale 2-short vol in observation group was higher than that in control group at 12 months(P＜0.05).Conclusion The digital template combined with limited open reduction and fixation in the treatment of ankle fracture of degree Ⅳ supination and external rotation can promote the recovery of patient's ankle function in a short time and improve the quality of life.

ObjectiveTo investigate the effects of Xijiao Dihuangtang （XJDHT） on mice with sepsis and cellular models of sepsis and explore its molecular mechanism in alleviating sepsis-induced inflammatory responses via regulating pyruvate kinase M2 （PKM2）-mediated one-carbon metabolism pathway. MethodsForty C57BL/6N mice were randomly divided into four groups： normal group， model group， low-dose XJDHT group （7.7 g·kg^-1）， and high-dose XJDHT group （15.4 g·kg^-1）. After one week of continuous gavage， sepsis was induced using cecal ligation and puncture （CLP） in groups except the normal group. 24 h after the surgery， mortality rates in all groups were recorded， and serum cytokines were measured by enzyme linked immunosorbent assay （ELISA）. Lung histopathology was examined by hematoxylin-eosin （HE） staining. During the in vitro experiment， the human monocytic leukemia cell line （THP-1） was exposed to various concentrations of XJDHT and treated with lipopolysaccharide （LPS） at a final concentration of 2 mg·L^-1 for 24 h. Cell apoptosis was detected using terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） assay. Protein levels of tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， B-cell lymphoma 2 （Bcl-2）， and Bcl-2-associated X protein （Bax） were measured by Western blot. Transcriptome sequencing was performed to analyze differentially expressed genes in all groups and conduct gene ontology （GO） enrichment. Key genes in the one-carbon metabolism pathway， including pyruvate kinase M2 （PKM2）， 5-methyltetrahydrofolate-homocysteine methyltransferase （MTR）， and phosphoglycerate dehydrogenase （PHGDH）， were verified by Western blot. A PKM2 inhibition model was established using shikonin for further protein expression analysis. ResultsAnimal experiments showed that compared with the normal group， the model group exhibited significantly elevated body temperature and lung pathology （P<0.01） and increased serum TNF-α and IL-1β levels （P<0.01）. High-dose XJDHT reduced body temperature and lung tissue damage （P<0.01） and significantly decreased serum TNF-α and IL-1β levels （P<0.01）. Low-dose XJDHT treatment showed no significant temperature change （P<0.01） but reduced serum TNF-α and IL-1β levels （P<0.01）. Transcriptome sequencing and Western blot revealed significant differences in the expression of TNF-α， IL-1β， and one-carbon metabolism genes （PKM2， MTR， and PHGDH） （P<0.01）. Cell experiments demonstrated that compared to the normal group， the model group showed elevated protein expressions of TNF-α and IL-1β in THP-1 cells （P<0.01）， decreased Bcl-2/Bax ratio， and increased apoptosis （P<0.01）. Transcriptome sequencing and Western blot revealed significant differences in the expression of TNF-α， IL-1β， and one-carbon metabolism genes （PKM2， MTR， and PHGDH） （P<0.01）. Compared to the model group， high-dose XJDHT significantly increased Bax/Bcl-2 ratio and PHGDH protein expression （P<0.01） and effectively reduced cell apoptosis （P<0.01） while down-regulating protein expressions of TNF-α， IL-1β， PKM2， and MTR （P<0.01）. Low-dose XJDHT moderately increased Bax/Bcl-2 ratio and PHGDH protein expression （P<0.05）， reduced apoptosis （P<0.05）， and decreased IL-1β and MTR protein levels （P<0.05， P<0.01）， but there were no significant changes in TNF-α and PKM2 expression. After PKM2 inhibition by shikonin in THP-1 cells， the expression of protein related to one-carbon metabolism was detected. Compared with the blank group， the LPS-induced model group showed significantly upregulated PKM2 and MTR protein expression （P<0.01） and downregulated PHGDH expression （P<0.01）. Compared with the model group， shikonin treatment significantly reduced PKM2 expression （P<0.05）， increased PHGDH expression （P<0.01）， and decreased MTR expression （P<0.05）. ConclusionXJDHT can inhibit the release of inflammatory factors in sepsis， and its mechanism is related to the intervention of the PKM2-regulated one-carbon metabolism pathway in macrophages.

OBJECTIVE To explore the molecular mechanism of Xijiao Dihuang Decoction(XJDHT)in inhibiting inflammatory response in sepsis based on network pharmacology,molecular docking and in vitro and in vivo experiments.METHODS Active com-ponents of XJDHT were screened using the TCMSP and HERB databases.Sepsis-related targets and histone H3K36 trimethylation(H3K36me3)-associated targets were retrieved from GeneCard,OMIM,and DisGeNet databases.A protein-protein interaction(PPI)network was constructed using the STRING database,and core targets were identified via Cytoscape 3.9.1.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses were performed to predict potential mechanisms.Mo-lecular docking validated ligand-receptor binding affinity.A cecal ligation and puncture(CLP)model was established in mice to evalu-ate 24-hour sepsis scores(MSS)and survival rates.Blood routine parameters were analyzed using an automated hematology analyzer.Serum IL-1β and TNF-α levels were measured by ELISA.Liver histopathology was assessed via HE staining,and H3K36me3 expression in Kupffer cells was detected by immunofluorescence.In vitro,LPS-induced THP-1 cells were used as an in-flammatory model.ChIP-qPCR evaluated H3K36me3 enrichment at IL-1β and TNF-α gene loci.Western blot analyzed HIF-1α,EGFR,and AKT1 pathway proteins.RESULTS A total of 28 active components of XJDHT were identified,corresponding to 987 gene targets,with 189 overlapping sepsis-related targets.Core targets included TNF,IL1B,and GAPDH.Enriched pathways includ-ed EGFR tyrosine kinase inhibitor resistance.Molecular docking confirmed strong binding between core components and key targets.In vivo,compared to the sham group,the CLP group exhibited significantly reduced 24-hour survival(P<0.01),elevated MSS(P<0.01),immune imbalance,and increased serum IL-1β and TNF-α levels(P<0.01).High-and low-dose XJDHT intervention im-proved survival(P<0.01),reduced MSS(P<0.01),restored immune homeostasis,and dose-dependently suppressed IL-1β and TNF-α(P<0.01).CLP mice showed elevated H3K36me3 in Kupffer cells and severe hepatic inflammation,while XJDHT dose-de-pendently reduced H3K36me3(P<0.05)and attenuated liver injury.In peritoneal macrophages,CLP upregulated H3K36me3,IL-1β,TNF-α,HIF-1α,p-AKT1/AKT1,and EGFR(P<0.01),which were reversed by XJDHT(P<0.05,P<0.01).In vitro,LPS increased H3K36me3 and IL-1β and TNF-α expression(P<0.01),with ChIP-qPCR confirming H3K36me3 enrichment at IL-1β lo-ci(P<0.01).Treatment with 15%XJDHT-containing serum for 24 h reduced H3K36me3(P<0.01),diminished its recruitment to IL-1β loci(P<0.01),and inhibited LPS-induced activation of EGFR,HIF-1α,and p-AKT1/AKT1(P<0.05,P<0.01).HIF-1α agonist Dimethyloxallyl Glycine(DMOG)further validated that XJDHT suppressed H3K36me3-mediated epigenetic remodeling via HIF-1α-related pathways.CONCLUSION XJDHT inhibits inflammatory responses,regulates immune homeostasis,and improves sepsis prognosis,potentially by modulating H3K36me3 epigenetic modifications at IL-1β loci.

Objective To analyze clinical effect of digital template combined with limited incision reduction and fixation in treatment of ankle fractures with degree Ⅳ supination and external rotation.Methods A total of 73 patients with posterior ankle rotation fractures admitted to Jinhua Hospital of Traditional Chinese Medicine,Zhejiang Chinese Medical University from January 2020 to June 2023 were selected as study subjects.According to the surgical method,32 patients treated with conventional open reduction and internal fixation were selected as control group,and 41 patients treated with digital template combined with limited open reduction and fixation technique were selected as observation group.Perioperative related indexes,ankle function and complications were compared between two groups.Results Time of operation and length of incision in observation group were shorter than those in control group,time of getting out of bed,time of hospitalization in observation group were shorter than those in control group(P＜0.01).The Kofoed scores at 1 and 6 months after operation in observation group were higher than those in control group(P＜0.05).The angles of ankle dorsiflexion and plantar flexion at 1 and 6 months after operation in observation group were higher than those in control group(P＜0.05).The score of arthritis quality of life measurement scale 2-short vol in observation group was higher than that in control group at 12 months(P＜0.05).Conclusion The digital template combined with limited open reduction and fixation in the treatment of ankle fracture of degree Ⅳ supination and external rotation can promote the recovery of patient's ankle function in a short time and improve the quality of life.

Antineoplastic Agents, Immunological ; immunology ; pharmacology ; Cell Line, Tumor ; GPI-Linked Proteins ; antagonists & inhibitors ; immunology ; Humans ; Neoplasm Proteins ; analysis ; immunology ; Pancreatic Neoplasms ; drug therapy ; immunology ; pathology

Objective To investigate the activation of the autophagy and the signal transduction pathway in the scrapie 139A infected mice brain tissues.Methods The Western Blot was used to evaluate the expression level of AMPK,ULK1,p-AMPK,P-ULKI and LC3 Ⅱ in the normal and scrapie 139A infected mice brain tissues.Results The expression levels of AMPK,ULK1 and their phosphorylated form p-AMPK (Thr172),p-ULK1 (Ser555) are markedly up-regulated in the brains of the mice infected with scrapie agent 139A.Furthermore,the expression of LC3 Ⅱ was also found increased in the brains of 139A infected mice,but not in normal mice.Conclusion AMPK-ULK1 pathway activated and contributed to autophagy in 139A infected mice brain.

Objective To assess the influence between managements in emergency room(ER) andoutcome of severe traumatic brain injury (TBI),in order to provide inference for treatment.Methods A retrospective analysis was performed in severe TBI patients and recorded next indexes.(1) The managements in ER,including endotracheal intubation and oxygenation,fluid resuscitation,and mannitol intake.(2) The vital signs arriving at ICU,including systolic pressure and blood oxygen saturation.(3) Prognostic indicators including inhospital mortality and days during ICU,the scores of Glasgow outcome scale (GOS) at discharge and 6 months after injury.Results In 140 severe TBI patients,65 patients (46.4％) died during ICU.The mortality of patients with endotracheal intubation [65.0％ (39/60)] was significantly higher than that without endotracheal intubation [32.5％(26/80)](P＜ 0.01).The mortality in whether fluid resuscitation and using mannitol had no significant difference [44.7％ (46/103) vs.51.4％ (19/37),49.2％ (31/63) vs.44.2％ (34/77)] (P ＞0.05).In days during ICU,there was no significant difference among the three treatment measures (P＞ 0.05).In GOS grade at discharge and 6 months after injury,the proportion of 4 and 5 grade were 8.3％ (5/60) and 25.0％ (15/60) in patients with endotracheal intubation,while 27.5％ (22/80) and 52.5％ (42/80) in patients without endotraeheal intubation (P ＜ 0.01).In fluid resuscitation and using mannitol patients,there were no significant difference(P ＞ 0.05).Conclusion Treating severe TBI patients in ER,endotracheal intubation should be carefully chosen,fluid resuscitation and mannitol may not be given.