OBJECTIVETo investigate the dual role of daphnetin in suppressing high mobility group box-1 protein (HMGB1) release and blocking HMGB1-induced inflammatory response.

METHODSMurine macrophage RAW264.7 cells were cultured in the presence of daphnetin, lipopolysaccharide (LPS), or both. HMGB1 release from the cells was determined using ELISA, and phosphorylations of JAK1/2 and of STAT1 were detected by Western blotting. Human monocytic THP-1 cells exposed to daphnetin, rhHMGB1, or both were examined for NO production using a NO detection kit, for the release of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and prostaglandin E2 (PGE2) using ELISA, and for expressions of iNOS, COX-2 and phosphorylated p38, ERK, and JNK with Western blotting.

RESULTSDaphnetin dose-dependently reduced the release of HMGB1 in RAW264.7 cells and suppressed rhHMGB1-induced iNOS and COX-2 expressions and release of TNF-α, IL-6, PGE2, and NO in THP-1 cells. Western blotting revealed that daphnetin significantly down-regulated the phosphorylations of JAK-STAT1 pathway in LPS-stimulated RAW264.7 cells but did not suppress the phosphorylations of MAPKs signaling pathway induced by rhHMGB1 in THP-1 cells.

CONCLUSIONDaphnetin can reduce the release of HMGB1 and suppress HMGB1-induced inflammatory response. In RAW264.7 cells, daphnetin inhibited LPS induced HMGB1 release is at least partly mediated by suppressing JAK-STAT1 signaling pathway activation.

Animals ; Cell Line ; Cyclooxygenase 2 ; metabolism ; Dinoprostone ; metabolism ; HMGB1 Protein ; metabolism ; Humans ; Inflammation ; metabolism ; Interleukin-6 ; metabolism ; Janus Kinase 1 ; metabolism ; Lipopolysaccharides ; Macrophages ; drug effects ; Mice ; Monocytes ; drug effects ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase Type II ; metabolism ; RAW 264.7 Cells ; STAT1 Transcription Factor ; metabolism ; Signal Transduction ; drug effects ; Tumor Necrosis Factor-alpha ; metabolism ; Umbelliferones ; pharmacology

Objective To investigate the molecular mechanism by which salidroside protects PC12 cells from H2O2-induced apoptosis.Methods PC12 cells cultured in DMEM supplemented with 10％ horse serum and 5％ fetal bovine serum were pretreated with different doses of salidroside for 2 h and then stimulated with H2O2 for different lengths of time.The expression levels of PARP and caspase 3 and the phosphorylation of p38,ERK and JNK were determined with Western blotting.The cell nuclear morphology was observed after DAPI staining.The production of ROS was detected using a ROS detection kit,and the levels of gp91phox and p47phox in the membrane and cytoplasm were detected by membrane-cytoplasm separation experiment;the binding between gp91phox and p47phox was assayed by coimmunoprecipitation experiment.Results Salidroside dose-dependently suppressed cell apoptosis,lowered phosphorylation levels of p38,ERK and JNK,inhibited the production of ROS,reduced the binding between gp91phox and p47phox,and inhibited the activity of NOX2 in PC12 cells exposed to H2O2.Conclusion Salidroside protects PC12 cells from H2O2-induced apoptosis at least partly by suppressing NOX2-ROS-MAPKs signaling pathway.

Objective To investigate the molecular mechanism by which salidroside protects PC12 cells from H2O2-induced apoptosis.Methods PC12 cells cultured in DMEM supplemented with 10％ horse serum and 5％ fetal bovine serum were pretreated with different doses of salidroside for 2 h and then stimulated with H2O2 for different lengths of time.The expression levels of PARP and caspase 3 and the phosphorylation of p38,ERK and JNK were determined with Western blotting.The cell nuclear morphology was observed after DAPI staining.The production of ROS was detected using a ROS detection kit,and the levels of gp91phox and p47phox in the membrane and cytoplasm were detected by membrane-cytoplasm separation experiment;the binding between gp91phox and p47phox was assayed by coimmunoprecipitation experiment.Results Salidroside dose-dependently suppressed cell apoptosis,lowered phosphorylation levels of p38,ERK and JNK,inhibited the production of ROS,reduced the binding between gp91phox and p47phox,and inhibited the activity of NOX2 in PC12 cells exposed to H2O2.Conclusion Salidroside protects PC12 cells from H2O2-induced apoptosis at least partly by suppressing NOX2-ROS-MAPKs signaling pathway.

OBJECTIVETo evaluate the therapeutic effect of MHSP65-TCL on melanoma and its effect on the activity of the immunocytes.

METHODSMHSP65-TCL was prepared by mixing MHSP65 with TCL derived from B16 melanoma cell lysate by repeated freezing and thawing. The MHSP65-TCL vaccine was administered in mice bearing B16 melanoma, and the changes in melanoma growth was observed. To investigate the influence of TCL in MHSP65-TCL on the activity of the immunocytes, we co-cultured TCL and mouse spleen cells in vitro, and analyzed CD69 expression on the cells, cell apoptosis, and levels of IL-10 and IFN-γ in the cell culture supernatant.

RESULTSThe MHSP65-TCL vaccine showed an anti-melanoma effect in the tumor-bearing mice. In the in vitro experiment, TCL in MHSP65-TCL strongly stimulated the activation of mouse spleen cells while causing apoptosis in some of the immunocytes and promoting cellular IL-10 secretion, but not IFN-γ.

CONCLUSIONSMHSP65-TCL derived from B16 melanoma cells has an anti-melanoma effect mediated by the activation of immunocytes. TCL in MHSP65-TCL also has immunosuppressive effect on immunocytes possibly due to the presence of suppressive components in TCL, and identifying and eliminating these components may potentially improve the anti-tumor actovoty of MSHP65-TCL vaccine.

Animals ; Antigens, CD ; metabolism ; Antigens, Differentiation, T-Lymphocyte ; metabolism ; Apoptosis ; Bacterial Proteins ; administration & dosage ; immunology ; Cancer Vaccines ; Cell Extracts ; administration & dosage ; immunology ; Cell Line, Tumor ; Chaperonin 60 ; administration & dosage ; immunology ; Female ; Interferon-gamma ; metabolism ; Interleukin-10 ; metabolism ; Lectins, C-Type ; metabolism ; Melanoma, Experimental ; immunology ; pathology ; Mice ; Mice, Inbred C57BL ; Random Allocation ; Spleen ; cytology ; immunology ; metabolism ; Tumor Burden ; immunology

Objective To analyze the related risk factors affecting the prognosis of elderly patients with severe community acquired pneumonia (SCAP). Methods A retrospective study method was conducted; the elderly (≥ 75 years old) patients with SCAP treated in the First Affiliated Hospital of Hainan Medical College from January 2015 to January 2019 were enrolled. The general data of patients were collected, including sex, age, oxygenation index (PaO2/FiO2), involved organs, presence or absence of following diseases or treatment: damage in multiple lung lobes, septic shock, basic diseases (cardiovascular disease, chronic lung disease, diabetes, hypertension, and cerebrovascular disease), invasive mechanical ventilation, ventilator-associated pneumonia (VAP), misinhalation event, hyponatremia, respiratory acidosis, hypoproteinemia, intubation times, total mechanical ventilation time, etc. According to the prognosis, the patients were divided into a death group and a survival group. The general data were compared between the two groups with different prognoses. Single factor analysis was carried out by selecting variables. The indicators with statistical significant differences in the results of univariate analysis were introduced into the multivariate Logistic regression analysis to analyze the related risk factors affecting the prognosis of elderly patients with SCAP. The receiver operating characteristic (ROC) curve was drawn to analyze the predictive values of risk factors in the patients with SCAP. Results A total of 112 patients were included, 33 died, and the mortality rate was 29.46%. Univariate analysis showed that the following factors were higher in the death group than those in the survival group: organ involvement >2 [69.70% (23/33) vs. 35.44% (28/79)], lung lobe damage ≥ 3 [75.76% (25/33) vs. 51.90% (41/79)], invasive mechanical ventilation [72.73% (24/33) vs. 32.91% (26/79)], diabetes [30.30% (10/33) vs. 12.66% (10/79)], intubation times ≥2 [57.58% (19/33) vs. 48.10% (38/79)], hypoproteinemia [75.76% (25/33) vs. 41.77% (33/79)], hyponatremia [72.73% (24/33) vs. 48.10% (38/79)], respiratory acidosis [66.67% (22/33) vs. 44.30 %(35/79)] and total mechanical ventilation time ≥ 15 days [69.70% (23/33) vs. 40.51 (32/79)]; the factors in the death group lower than those in the survival group were: septic shock [3.03% (1/33) vs. 17.72% (14/79)], chronic lung disease [6.06% (2/33) vs. 25.32% (20/79)] and PaO2/FiO2 [mmHg (1 mmHg = 0.133 kPa): 102.89±14.78 vs. 109.56±14.08],the differences were statistically significant (all P < 0.05); there were no significant differences in gender, age, cardiovascular disease, hypertension, VAP, misinhalation events and cerebrovascular disease between the two groups (all P > 0.05). Multivariate Logistic regression analysis showed that diabetes mellitus [odds ratio (OR) = 1.074, 95% confidence interval (95%CI) = 1.017-1.287, P =0.045], septic shock (OR = 2.765, 95%CI = 1.083-3.411, P = 0.047), hyponatremia (OR = 1.792, 95%CI = 1.128-1.417, P = 0.006), hypoalbuminemia (OR = 2.187, 95%CI = 1.872-5.462, P = 0.046), invasive mechanical ventilation (OR = 5.870, 95%CI = 2.324-23.796, P = 0.001), respiratory acid poisoning (OR = 2.934, 95%CI = 2.454-7.275, P = 0.043), time of mechanical ventilation (OR= 1.986, 95%CI = 2.467-3.483, P = 0.034), number of intubation (OR = 6.760, 95%CI = 2.116-24.696, P = 0.001), PaO2/FiO2 (OR = 1.981, 95%CI = 1.006-1.417, P = 0.007), organ involvement > 2 (OR = 2.924, 95%CI = 2.534-6.285, P = 0.048), chronic lung disease (OR = 2.887, 95%CI = 1.487-3.483, P = 0.039), and lung lobe damage≥3 (OR = 2.754, 95%CI = 1.131-1.798, P = 0.045) were independent risk factors affecting the prognosis of elderly patients with SCAP. ROC analysis showed that hyponatremia, hypoalbuminemia, invasive mechanical ventilation, total mechanical ventilation time, PaO2/FiO2, organ involvement > 2, damage of lung lobes ≥ 3, had predictive values for the prognosis of SCAP [the areas under ROC curve (AUC) were 0.377, 0.267, 0.301, 0.646, 0.650, 0.329, and 0.381, respectively, all P < 0.05]. Conclusions Underlying disease, invasive mechanical ventilation, respiratory acidosis, total mechanical ventilation time, PaO2/FiO2, intubation times ≥ 2, chronic lung disease and lung damage≥ 3 lobes are the independent risk factors for the prognosis of elderly patients with severe community acquired pneumonia. Clinical treatment should focus on the above aspects to minimize the mortality of patients.

Objective To evaluate the therapeutic effect of MHSP65-TCL on melanoma and its effect on the activity of the immunocytes. Methods MHSP65-TCL was prepared by mixing MHSP65 with TCL derived from B16 melanoma cell lysate by repeated freezing and thawing. The MHSP65-TCL vaccine was administered in mice bearing B16 melanoma, and the changes in melanoma growth was observed. To investigate the influence of TCL in MHSP65-TCL on the activity of the immunocytes, we co-cultured TCL and mouse spleen cells in vitro, and analyzed CD69 expression on the cells, cell apoptosis, and levels of IL-10 and IFN-γin the cell culture supernatant. Results The MHSP65-TCL vaccine showed an anti-melanoma effect in the tumor-bearing mice. In the in vitro experiment, TCL in MHSP65-TCL strongly stimulated the activation of mouse spleen cells while causing apoptosis in some of the immunocytes and promoting cellular IL-10 secretion, but not IFN-γ. Conclusions MHSP65-TCL derived from B16 melanoma cells has an anti-melanoma effect mediated by the activation of immunocytes. TCL in MHSP65-TCL also has immunosuppressive effect on immunocytes possibly due to the presence of suppressive components in TCL, and identifying and eliminating these components may potentially improve the anti-tumor actovoty of MSHP65-TCL vaccine.

Objective To evaluate the therapeutic effect of MHSP65-TCL on melanoma and its effect on the activity of the immunocytes. Methods MHSP65-TCL was prepared by mixing MHSP65 with TCL derived from B16 melanoma cell lysate by repeated freezing and thawing. The MHSP65-TCL vaccine was administered in mice bearing B16 melanoma, and the changes in melanoma growth was observed. To investigate the influence of TCL in MHSP65-TCL on the activity of the immunocytes, we co-cultured TCL and mouse spleen cells in vitro, and analyzed CD69 expression on the cells, cell apoptosis, and levels of IL-10 and IFN-γin the cell culture supernatant. Results The MHSP65-TCL vaccine showed an anti-melanoma effect in the tumor-bearing mice. In the in vitro experiment, TCL in MHSP65-TCL strongly stimulated the activation of mouse spleen cells while causing apoptosis in some of the immunocytes and promoting cellular IL-10 secretion, but not IFN-γ. Conclusions MHSP65-TCL derived from B16 melanoma cells has an anti-melanoma effect mediated by the activation of immunocytes. TCL in MHSP65-TCL also has immunosuppressive effect on immunocytes possibly due to the presence of suppressive components in TCL, and identifying and eliminating these components may potentially improve the anti-tumor actovoty of MSHP65-TCL vaccine.

Purpose/Significance To expound the content features of online medical reviews and its impact on the utilization of pa-tients'online consultation services,and to provide references for the information management of online medical platforms.Method/Process The LDA topic model based on TF-IDF is constructed,the semantic analysis method based on sentiment dictionary is adopted to mine the content features of online medical reviews,and the principal component analysis(PC A)is applied to classify and explore them,and the regression analysis is carried out by binary logistic.Result/Conclusion The content features of online medical reviews are divided into 3 categories,including 9 indicators.Among them,the attributes of reviewers,timeliness,consultation process,service atti-tude and platform feedback have a significant impact on the utilization of efficient online consultation services by patients.It is suggested to improve the utilization level of online consultation services by optimizing related services of online medical platform review section.

Objective To analyze the clinical values of super early enteral nutrition combined with microecopharmaceutics and delayed enteral nutrition on patients with severe acute pancreatitis. Methods Clinical data of thirty patients diagnosed as severe acute pancreatitis in our emergency department during January 2013 and December 2017 were reviewed retrospectively. Patients were divided into the treatment group (n=15, patients given enteral nutrition combined with microecopharmaceutics within 24 h after admission) and the control group (n=15, patients given delayed enteral nutrition after 48 h of admission). Two weeks after the treatment, the serum variables of C-reactive protein, total protein, albumin, recovery time of urine and blood amylase, length of hospital stay and APACHE Ⅱ score were compared between the two groups by using paired samples t test. Results The C-reactive protein [(46.7±13.1) mg/L vs. (190.72±19.3) mg/L, t=10.4, P<0.01] and APACHE Ⅱ score [(7.2±1.9) vs.(9.3±2.4),t=2.7,P<0.05] of the treatment group were significantly lower than those in the control group. The total protein [(58.1±6.3)g/L vs.(52.6±5.4)g/L, t=2.5, P<0.05] and albumin [(29.9±3.2)g/L vs.(22.0±2.8)g/L, t=7.12, P<0.01] of the treatment group were significantly higher than those in the control group. The recovery time of urine amylase [(13.2±2.1)d vs.(18.7±3.9)d, t=4.9, P<0.01] and blood amylase [(7.5±3.0)d vs.(11.1±3.4)d, t=3.1, P<0.01], and length of hospital stay[(14.9±4.5)d vs.(27.1±5.3)d, t=6.9, P<0.01] were significantly shorter in the treatment group compared with those in the control group. Conclusions Ultra-early enteral nutrition combined with microecopharmaceutics can shorten the length of hospital stay of patients with severe acute pancreatitis, and is safe and effective.

Objective To explore the molecular mechanism of resveratrol(RES)in the treatment of oral squamous cell carcinoma(OSCC)through the use of biological information methods such as network pharmacology and molecular docking and to provide a theoretical reference for the clinical application of RES in the treatment of OSCC.Methods The Swiss Target Prediction(http://www.swisstargetprediction.ch),SEA(http://sea.bkslab.org)database,and Pharm map-per database(http://lilab-ecust.cn)were used to retrieve RES-related targets,and the DISGENET(www.disgenet.org),OMIM(https://omim.org)and GeneCards(https://www.genecards.org)databases were used to screen OSCC disease tar-gets.The intersection of drugs and disease targets was determined,and Cytoscape 3.7.2 software was used to construct a"drug-diseasetarget pathway"network.The Search Tool for the Retrieval of Interacting Genes/Proteins(STRING)data-base was used to construct a target protein interaction network,and the DAVID database was used for enrichment analy-sis of key proteins.Finally,molecular docking validation of key proteins was performed using AutoDock and PyMOL.The enrichment analysis and molecular docking results were integrated to predict the possible molecular mechanisms of RES treatment in OSCC;western blot was used to determine the effect of resveratrol at different concentrations(50,100)μmol/L on the expression of Src tyrosine kinase(SRC),epidermal growth factor receptor(EGFR),estrogen re-ceptor gene 1(ESR1),and phosphatidylinositol 3 kinase/protein kinase B(PI3K/AKT)signaling pathway proteins in OSCC HSC-3 cells.Results A total of 243 targets of RES drugs and 6 094 targets of OSCC were identified.A total of 116 potential common targets were obtained by intersecting drugs with disease targets.These potential targets mainly participate in biological processes such as in vivo protein self-phosphorylation,peptide tyrosine phosphorylation,trans-membrane receptor protein tyrosine kinase signaling pathway,and positive regulation of RNA polymerase Ⅱ promot-er transcription,and they interfere with the PI3K/AKT signaling pathway to exert anti-OSCC effects.The docking results of resveratrol with OSCC molecules indicated that key targets,such as EGFR,ESR1,and SRC,have good binding activi-ty.The results of cell-based experiments showed that resveratrol inhibited the protein expression of SRC,EGFR,ESR1,p-PI3K,and p-AKT in HSC-3 cells in a dose-dependent manner.Conclusion RES can inhibit the expres-sion of its targets EGFR,ESR1,SRC,p-PI3K,and p-AKT in OSCC cells.