Objective To investigate the effect and mechanism of zanthoxylum bungeanum seed oil on autophagy and apoptosis of laryngeal cancer cells.Methods Human bronchial epithelial cells BEAS-2B and human laryngeal cancer cells Hep-2 were treated with different volume fractions(v/v)of Zanthoxylum bungeanum seed oil(0,0.02%,0.04%,0.06%,0.08%,0.10%),respectively,cell viability was detected by cell counting kit-8 method(CCK-8).Hep-2 cells in logarithmic growth phase were randomly divided into control group,Zanthoxylum bungeanum seed oil low,medium and high dose groups and Zanthoxylum bungeanum seed oil high dose+insulin-like growth factor 1(IGF-1)(PI3K agonist)group,cell apoptosis and cell cycle progression were detected by flow cytometry(FCM)assay,the formation of autophagic vesicles was detected by monodansyl cadaverine(MDC)staining,the expression of apoptosis,autophagy and phosphatidylinositide 3-kinases(PI3K)/protein kinase B(AKT)/mechanistic target of rapamycin(mTOR)pathway-related proteins were detected by Western blotting(WB).Results Compared with the control group Hep-2(99.03%±0.82%),treatment with zanthoxylum bungeanum seed oil(0.02%,0.04%,0.06%,0.08%,0.10%)(v/v)could reduce cell survival rate(84.63%±0.73%,57.34%±0.84%,19.76%±0.62%,17.22%±0.72%,12.19%±0.81%),and the differences were statistically significant(t=22.718～133.559,all P<0.001),while it has no inhibitory effect on BEAS-2B activity(t=0.283～1.980,all P>0.05).Compared with the control group,the Hep-2 apoptosis rate,G1/G0 phase cell proportion,autophagic vesicle integrated optical density(IOD)value,Cleaved-caspase-3,Bcl-2 associated X protein(Bax),microtubule-associated protein 1A/1B 1ight chain 3I/Ⅱ(LC 3I/Ⅱ)and Beclin-1 expression were all increased in the low,medium,and high-close groups of zantheoxylum bungeanum seed oil(t=4.270～58.425);the proportion of G2/M phase cells,ubiquitin-binding protein P62,Bcl-2 expression and p-PI3K/PI3K,p-AKT/AKT and p-mTOR/mTOR expression were all decreased(t=3.041～58.765),and the differences were statistically significant(all P<0.05).Compared with the high-dose zanthoxylum bungeanum seed oil group,the apoptosis rate of Hep-2,the proportion of G1/G0 phase cells,the expression of Cleaved-caspase-3,Bax,LC3Ⅱ/I and Beclin-1,the IOD value of autophagic vesicles(t=4.931～39.507),the expression of Bcl-2 and ubiquitin-binding protein P62,the proportion of G2/M phase cells,the ratio of p-PI3K/PI3K,p-AKT/AKT and p-mTOR/mTOR in the high-dose zanthoxylum bungeanum seed oil+IGF-1 group were decreased(t=3.402～14.207),and the differences were statistically significant(all P<0.05).Conclusion Zanthoxylum bungeanum seed oil can promote autophagy and apoptosis of Hep-2 cells,and its mechanism may be related to the inhibition of PI3K/AKT/mTOR pathway activation.

Objective To investigate the effect and mechanism of zanthoxylum bungeanum seed oil on autophagy and apoptosis of laryngeal cancer cells.Methods Human bronchial epithelial cells BEAS-2B and human laryngeal cancer cells Hep-2 were treated with different volume fractions(v/v)of Zanthoxylum bungeanum seed oil(0,0.02%,0.04%,0.06%,0.08%,0.10%),respectively,cell viability was detected by cell counting kit-8 method(CCK-8).Hep-2 cells in logarithmic growth phase were randomly divided into control group,Zanthoxylum bungeanum seed oil low,medium and high dose groups and Zanthoxylum bungeanum seed oil high dose+insulin-like growth factor 1(IGF-1)(PI3K agonist)group,cell apoptosis and cell cycle progression were detected by flow cytometry(FCM)assay,the formation of autophagic vesicles was detected by monodansyl cadaverine(MDC)staining,the expression of apoptosis,autophagy and phosphatidylinositide 3-kinases(PI3K)/protein kinase B(AKT)/mechanistic target of rapamycin(mTOR)pathway-related proteins were detected by Western blotting(WB).Results Compared with the control group Hep-2(99.03%±0.82%),treatment with zanthoxylum bungeanum seed oil(0.02%,0.04%,0.06%,0.08%,0.10%)(v/v)could reduce cell survival rate(84.63%±0.73%,57.34%±0.84%,19.76%±0.62%,17.22%±0.72%,12.19%±0.81%),and the differences were statistically significant(t=22.718～133.559,all P<0.001),while it has no inhibitory effect on BEAS-2B activity(t=0.283～1.980,all P>0.05).Compared with the control group,the Hep-2 apoptosis rate,G1/G0 phase cell proportion,autophagic vesicle integrated optical density(IOD)value,Cleaved-caspase-3,Bcl-2 associated X protein(Bax),microtubule-associated protein 1A/1B 1ight chain 3I/Ⅱ(LC 3I/Ⅱ)and Beclin-1 expression were all increased in the low,medium,and high-close groups of zantheoxylum bungeanum seed oil(t=4.270～58.425);the proportion of G2/M phase cells,ubiquitin-binding protein P62,Bcl-2 expression and p-PI3K/PI3K,p-AKT/AKT and p-mTOR/mTOR expression were all decreased(t=3.041～58.765),and the differences were statistically significant(all P<0.05).Compared with the high-dose zanthoxylum bungeanum seed oil group,the apoptosis rate of Hep-2,the proportion of G1/G0 phase cells,the expression of Cleaved-caspase-3,Bax,LC3Ⅱ/I and Beclin-1,the IOD value of autophagic vesicles(t=4.931～39.507),the expression of Bcl-2 and ubiquitin-binding protein P62,the proportion of G2/M phase cells,the ratio of p-PI3K/PI3K,p-AKT/AKT and p-mTOR/mTOR in the high-dose zanthoxylum bungeanum seed oil+IGF-1 group were decreased(t=3.402～14.207),and the differences were statistically significant(all P<0.05).Conclusion Zanthoxylum bungeanum seed oil can promote autophagy and apoptosis of Hep-2 cells,and its mechanism may be related to the inhibition of PI3K/AKT/mTOR pathway activation.

Objective:To analyze the clinical characteristics, treatment, and outcomes of children with complete left bundle branch block (CLBBB) mediated by maternal autoantibodies.Methods:A retrospective analysis was conducted on nine children diagnosed with maternal autoantibody-mediated CLBBB, treated at Beijing Anzhen Hospital and Fujian Provincial Hospital from March 2015 to August 2023. Their clinical characteristics, electrocardiographic and echocardiographic findings before and after treatment were reviewed. Paired sample t-test was used for inter-group comparison. Results:Among the mothers, 6 had positive antinuclear antibodies (ANA), 5 had anti-Sjogren syndrome antigen A antibodies, and 3 had anti-Ro-52 antibodies. The cohort included one female and eight male children, diagnosed with CLBBB at the age of 1 (2, 13) months. The positive autoantibodies in the infants, consisted with maternal antibodies, were detected within the first 3 months of life among 3 cases. Treatments included anti-heart failure therapy, myocardial nutritional support, intravenous immunoglobulin (IVIG) and glucocorticoids. Before treatment, the levels of troponin I (0.175 (0.060, 10.270) μg/L) and N-terminal pro-B-type natriuretic peptide (420 (327, 12 865) ng/L) were elevated, which normalized in most cases after treatment. Post-treatment, the QRS duration significantly shortened compared to pre-treatment ((137±15) vs.(169±25) ms, t=3.76, P<0.001), and the QTc interval significantly decreased ((433±41) vs. (514±27) ms, t=4.95, P=0.001). Before treatment, varying degrees of mitral and tricuspid regurgitation and marked interventricular septal dyskinesia were observed in echocardiography. After treatment, valve regurgitation and ventricular septum motion significantly improved, with a marked increase in left ventricular ejection fraction ((51±13)% vs. (27±6)%, t=-6.66, P<0.001). Conclusions:Maternal autoantibody-mediated CLBBB in children presents with chronic heart failure in infancy. Early treatment with anti-heart failure medications, IVIG and glucocorticoids can improve clinical symptoms.

Objective     To investigate the risk factors for postoperative complications Clavien-Dindo classification≥grade Ⅱ after lung cancer surgery. Methods     The patients who underwent lung cancer surgery in a multicenter observational study from November 2017 to January 2020 were included. The Clavien-Dindo classification of postoperative complications was analyzed. Logistic regression was used to identify the risk factors for complications≥ gradeⅡ. Results     A total of 388 patients were enrolled, including 203 males and 185 females with a mean age of 56.14±10.36 years. The incidence of postoperative complications was 25.52% (99/388) after lung cancer surgery and the incidence of complications≥gradeⅡ was 20.10% (78/388). The five most common postoperative complications were pneumonia (6.96%), prolonged pulmonary air leak (>7 days, 5.67%), incision dehiscence (4.64%), arrhythmia (3.87%), and postoperative pleural effusion (3.35%). Multivariate analysis showed that open surgery [reference: uniportal thoracoscopic surgery, OR=2.18, 95%CI (1.01, 4.70), P=0.047], extended resection [reference: sublobar resection, OR=2.86, 95%CI (1.11, 7.19), P=0.030; reference: lobectomy, OR=2.20, 95%CI (1.10, 4.40), P=0.026] and operative time≥3 h [OR=2.07, 95%CI (1.12, 3.85), P=0.021] were independent risk factors for postoperative complications≥gradeⅡ after lung cancer surgery. Conclusion     Surgical approach, extent of resection and operative time are independent influencing factors for postoperative complications≥gradeⅡ after lung cancer surgery.

Objective:To investigate the diagnostic value of neutrophil CD64 index (nCD64) in disseminated nontuberculous mycobacteria (NTM) infection.Methods:Thirty-six patients with NTM infection from January 2020 to June 2021 in Huashan Hospital, Fudan University were included. Patients were classified into groups of disseminated infection and focal infection according to their medical history and discharge diagnosis. The expressions of nCD64 in patients with focal infection and disseminated infection before treatment were collected and analyzed. Statistical analysis was performed using the Mann-Whitney U test, and the diagnostic value of nCD64 for disseminated NTM infection was analyzed using the receiver operator characteristic curve (ROC curve). Results:Among the 36 patients with NTM infection, 18 cases were focal infection (due to the low white blood cell count of the patient with myelodysplastic syndrome, the detection results were biased, which were excluded from the subsequent analysis) and 18 cases were disseminated infection. The expression of nCD64 in focal infection was 0.72(0.50, 1.55), and that in disseminated infection was 13.63(6.77, 32.31). The difference was statistically significant ( U=15.50, P<0.001). Using focal infection as a control, the area under the ROC curve for the operational characteristics of the subjects was 0.949 3 for disseminated NTM infection. The diagnostic cut-off value of nCD64 was 3.06, with the sensitivity and specificity of the disseminated NTM infection were 88.89% and 100.00%, respectively. Conclusions:In patients with NTM infection before effective treatment, the diagnostic cut-off value of nCD64 of 3.06 has high sensitivity and specificity, which is useful for the aided diagnosis of disseminated NTM infection.

Objective: To establish the norm of the Physical Activity afterschool Questionnaire for Preschooler(P-PAQ) in urban areas of China, so as to provide a basis for graded guidance from the family perspective and to improve children s physical activity levels. Methods: From October 2020 to January 2021, 6 267 children aged 3-6 years old were recruited from 40 kindergartens in eight cities across six major administrative regions by stratified cluster sampling, and the P-PAQ initially developed by the researchers of this study were completed by the primary caregivers. The questionnaire was administered to collect data relating to the amount of physical activity undertaken by the preschoolers, and the norm was determined by quartiles. Data relating to parental concepts of sports and parental behavior were assessed by calculating mean scores in order to establish the norm. Results: Among preschoolers in urban areas, the M(P 25 ，P 75 ) of total physical activity time (min/day), moderate-to-vigorous physical activity time (min/day), outdoor time (min/day) and screen time (min/day) on school days outside kindergarten and on weekends were 84 (54,120), 22 (8,40), 12 (0,24) and 18 (6,30), and 170 (115,240), 60 (30,95), 90 (35,120) and 30 (20,60), respectively. When the score of parents sports concept and behavior (total score of 40) were≥34, 29-<34, 24-<29, <24, it was defined as four levels about above medium, medium, lower medium and lower, respectively. And for two dimensions，when the score of parental sports concept were ≥19, 17-<19, 15-<17, <15，and the score of parental behaviors were ≥16, 12-<16, 8-<12, <8, it was defined as four levels about upper medium, medium, lower medium and lower, respectively. Conclusion The norm of extracurricular activities among preschool children in Chinese cities has good representativeness and appropriate threshold values, which could provide a valuable reference for early assessment, as well as guidance in relation to out-of-school physical activity behaviors among children aged 3-6 years old.

Objective:To explore the expression and clinical significance of immunosuppressive receptor T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT) on the peripheral blood mononuclear cells (PBMC) in silicosis patients with Mycobacterium tuberculosis infection. Methods:August 2018, a total of 78 patients with silicosis (all were quarry workers in Sanmen County, Zhejiang Province) were enrolled and divided into silicosis combined with active pulmonary tuberculosis group (APTB group), silicosis combined with latent tuberculosis infection group (LTBI group), and simple silicosis with non-tuberculosis infection group (non-TB group). Flow cytometry was used to analyze the expressions of TIGIT, programmed death-1 (PD-1) and transcription factor T-bet on PBMC from patients. Mann-Whitney U test and Pearson correlations analysis were used for statistical analysis. Results:Among the 78 patients, eight were in the APTB group, 24 in the LTBI group, and 46 in the non-TB group. The expressions of PD-1 and TIGIT on CD8 + T cells in the APTB group (29.45%(16.78%) and 65.40%(12.12%), respectively) were significantly higher than those in the LTBI group (17.40%(11.17%) and 48.30%(28.75%), respectively; U=23.500 and 43.500, respectively, P=0.000 8 and 0.020 5, respectively) and non-TB group (15.95%(12.46%) and 45.30%(19.75%), respectively; U=64.000 and 69.000, respectively, P=0.002 3 and 0.003 8, respectively), and the differences were all statistically significant. The expression of TIGIT was positively correlated with PD-1 on CD8 + T cells in silicosis patients ( r=0.434 3, P<0.01). The proportion of PD-1 + TIGIT + CD8 + T cells in the APTB group (19.90%(22.67%)) was significantly higher than those in the non-TB group (11.55%(11.29%), U=76.500, P=0.007 1) and LTBI group (11.55%(10.53%), U=41.000, P=0.015 4), while the proportion of PD-1 -TIGIT -CD8 + T cells in the APTB group (30.60%(12.90%)) was significantly lower than non-TB group (48.90%(18.98%), U=58.000, P=0.001 3) and LTBI group (47.20%(24.59%), U=41.000, P=0.015 4). The differences were all statistically significant. The expression of T-bet on the peripheral blood CD8 + T cells in the APTB group (29.45%(16.78%)) was higher than that in the non-TB group (15.95%(12.46%)) and the LTBI group (17.40%(11.17%)), and the differences were both statistically significant ( U=46.500 and 46.000, respectively, P=0.000 3 and 0.028 3, respectively). The expression of T-bet on CD8 + T cells was positively correlated with TIGIT on CD8 + T cells ( r=0.456 7, P<0.01). The expression of T-bet on PD-1 + TIGIT + CD8 + T cells in the APTB group (65.40%(12.12%)) was higher than those in the LTBI group (48.30%(28.75%), U=23.500, P=0.000 8) and non-TB group (45.30%(19.75%), U=65.000, P=0.002 6), and the differences were both statistically significant. Conclusion:The immunosuppressive receptor PD-1 and TIGIT are highly expressed on CD8 + T cells in silicosis patients with active pulmonary tuberculosis, which indicates CD8 + T cells exhaustion in these population, while the highly co-expression of T-bet suggests the exhausted subsets may have reversed potentiality.

In order to explore the effect of extract (SNE) on skin wound healing in mice and its mechanism, hemostasis effect of SNE was measured, the mouse skin wound model was established by full-thickness excision. The morphological changes of the wound were observed after the treatment with SNE and the healing rate was measured. The changes of wound histology were observed by hematoxylin eosin (HE) staining, Masson staining and transmission electron microscope (TEM). The expression of cell factors and related proteins was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Results showed that the SNE possessed hemostatic function. SNE could obviously improve the healing rate of wound in mouse and shorten time of scab removal compared with the none-treatment (NT) group ( < 0.05).The pathological histology analysis results showed complete epidermal regeneration, with remarkable capillary and collagen fiber observed in the SNE group. The expression level of tumor necrosis factor-α (TNF -α), interleukin-1β (IL-1β) and transforming growth factor-β1 (TGF-β1) in SNE group was significantly lower than that of the NT group on 7 d ( < 0.05). Moreover, compared with the NT group, the gene expressions level of Smad7 was significantly increased and the level of type II TGF-β receptors (TGF-βRII), collagen I (COL1A1) and α-smooth muscle actin (α-SMA) were significantly reduced in the SNE group on 28 d ( < 0.05), but the difference was not statistically significant compared to Yunnanbaiyao group (PC group) ( > 0.05). These results indicated that SNE possessed obvious activity of accelerating wound healing and inhibiting scar formation, and its mechanism was closely related to hemostatic function, regulation of inflammatory factors, collagen deposition, collagen fiber remodeling and intervening TGF-β/Smads signal pathway. Therefore, SNE may have promising clinical applications in skin wound repair and scar inhibition.

Objective:To analyze the differences of peripheral blood transcriptome between mild and severe influenza A (H1N1) patients, and to find indicators for the assessment of disease severity.Methods:A total of ten patients (five patients with mild disease and five patients with severe disease) diagnosed with H1N1 infection from January to May 2018 at Huashan Hospital, Fudan University in Shanghai were enrolled, and five healthy people were also enrolled as controls. The peripheral blood of patients was collected for transcriptome sequencing at the time when they were first diagnosed. Measurement data were compared using t test or Mann-Whitney U test. The count data were compared using Fisher exact test when appropriate. Data analysis of transcriptome predictions was performed using bioinformatics methods. Results:The platelet counts were significantly different between mild and severe groups ((163.4±21.5 )×10 9/L vs (255.6±52.5)×10 9/L, t=3.636, P=0.007). There were no differences between the two groups in gender, age, white blood cell counts, neutrophil percentage, lymphocyte percentage and hemoglobin levels (all P>0.05). However, the average expression levels of matrix metalloproteinase (MMP) 8 and MMP9 in severe group (18.41 and 174.00, respectively) were both higher than those in mild group (2.33 and 22.91, respectively) and healthy control (1.43 and 34.65, respectively; all P<0.01). Conclusion:MMP8 and MMP9 could be expected to serve as the molecular biological markers for predicting the disease severity in patients with influenza A (H1N1) infection.

Objective: To compare the diagnostic efficacy of transient elastography (TE) FibroScan and acoustic radiation force impulse imaging (ARFI) combined with serological models including aspartate aminotransferase-to-platelet ratio (APRI) and fibrosis-4 (FIB-4) in hepatitis B virus-related fibrosis. Methods: Sixty-seven patients with chronic HBV infection from October 2014 to May 2017 in Department of Infectious Diseases, Putuo Hospital were enrolled. Both FibroScan and ARFI were conducted in all patients together with serological tests. According to the golden standard of pathology results, the diagnosis values of FibroScan, ARFI combined with APRI or FIB-4 were compared as noninvasive assessment for liver fibrosis. Data with homogeneity of variance were tested by t test, and data with heterogeneity of variance were tested by Mann-Whitney U test. Results: Based on the pathology results, the receiver operating characteristic (ROC) areas under the curve (AUC) of APRI, FIB-4, FibroScan and ARFI in diagnosis of hepatic fibrosis ≥S2 were 0.752, 0.612, 0.885, and 0.850, respectively. The AUC of ROC curve in diagnosis of hepatic fibrosis ≥S3 were 0.746, 0.733, 0.851, and 0.863, respectively. The AUC of ROC curve in diagnosis of hepatic fibrosis ≥S4 were 0.782, 0.705, 0.962 and 0.981, respectively. Combined liver imaging technique and serological tests, such as APRI with FibroScan, APRI with ARFI, FIB-4 with FibroScan or FIB-4 with ARFI, the AUC of ROC curve in the 4 groups in diagnosis of hepatic fibrosis ≥S2 were 0.887, 0.861, 0.893, and 0.853, respectively; in the diagnosis of hepatic fibrosis ≥S3 were 0.873, 0.871, 0.900, and 0.875, respectively; and in diagnosis of hepatic fibrosis ≥S4 were 0.952, 0.981, 0.969, and 0.981, respectively. FibroScan and ARFI were positively correlated with liver inflammation (r=0.467, P=0.000; r=0.371, P=0.002) and jaundice (r=0.424, P=0.000; r=0.0.312, P=0.01), while negatively correlated with platelet (r=-0.331, P=0.006; r=-0.312, P=0.01). The AUC of ROC curve of FibroScan, ARFI and their combination with serological model were significantly increased compared with the single serological model (all P<0.05). Conclusions Serological models such as APRI and FIB-4 as well as liver imaging techniques such as FibroScan and ARFI are all valuable in assessment of hepatic fibrosis, while FibroScan and ARFI have better diagnostic value. ARFI is convenient to application for its integration with the ordinary ultrasound system. The sensitivity and specificity for diagnosis of hepatic fibrosis could be improved by combining serological model with FibroScan or ARFI. Combination of APRI and ARFI show the highest accuracy in diagnosis of hepatic fibrosis. Combination of serological models and transient elastic liver imaging is recommended for assessment and follow-up of HBV-related fibrosis.