Objective To investigate the effect of tumor necrosis factor-α (TNF-α) on the levels of ezrin-radixin-moesin (ERM) proteins and the phosphorylated ERM proteins (p-ERM) in rat pulmonary microvascular endothelial cells (PMVEC),and to explore Rho kinase (ROCK) influencing on modulation of the ERM proteins phosphorylation.Methods Cultured rat pulmonary microvascular endothelial cells were randomly divided into dose-dependent and time-dependent groups.In dose-dependent group,cells were cultured with different doses of TNF-α (0,0.1,1,10 μg/LTNF-α) for 60 min.In time-dependent group,cells were cultured with TNF-α (10 μg/L) for 0,15,30,60,90,120,180 min.In ROCK inhibitor (Y27632) intervention group,cells were cultured with TNF-α (10 μg/L) or Y27632 (30 μmol/L) + TNF-α (10 μg/L) for 60 min respectively.The levels of ERM proteins and p-ERM were determined by western blot.One way analysis of variance (ANOVA) was employed for statistical analysis by using SPSS version 16.0 software to compare values among all groups.A significant difference was presumed as a P value ＜ 0.05.Results Western blot revealed that ERM and p -ERM proteins were present in rat PMVEC.Stimulation withTNF-α gradually up-regulated the level of pERM proteins in a dose-dependent manner [0 μg/LTNF-α group:(0.648 ± 0.102),0.1 μg/LTNF-αgroup:(0.728-±0.082),1 μg/LTNF-α group:(0.926±0.121),10 μg/LTNF-α group:(1.245 ±0.134),all P =0.000].In time-dependent group,the level of p-ERM proteins rose at 15 min (0.777 ±0.151),peaked at 90 min (1.295 ±0.176),then decreased gradually at 120 min (0.802 ±0.139),but stayed higher level at 180 min (0.669 ±0.128) than that in un-stimulated 0 min group (0.631 ±0.123,P=0.004,0.000,0.001,0.016,respectively).When PMVEC pre-incubated with ROCK inhibitor and TNF-t,the level of p-ERM proteins caused a marked attenuation of TNF-αstimulation [(0.634 ± 0.112) vs.(0.875 ± 0.164),P =0.002],however,there are no significant differences compared to ROCK inhibitor alone group (0.661 ± 0.108) and no intervention group (0.654 ± 0.125),P =0.973,P =0.900,respectively).Conclusions TNF-α could induce up-regulation of the level of the phosphorylated ERM proteins in rat PMVEC,and ROCK signal molecules might involve in modulation of the ERM proteins phosphorylation.

Objective To observe the long-term toxicity of sustained-release dropping pills of Rhynchophylla total alkaloids in rats.Methods 80 Wistar rats were randomly divided into high dose group, middle dose group, low dose group and normal group. The rats were consecutively drenched for 12 weeks. The general status of the animals was observed daily in medication duration and body weight, daily appetite, quantity were recorded every 2 weeks. All animals were sacrificed on drenched 12 weeks and 2 weeks after discontinuation, then the content of WBC, RBC, Hb, PLT, LYM, NEU and ALT, AST, ALP, BUN, Cr were detected. Ratio of viscera was measured and the major organs were examined by the pathology.Results Continuous intragastric administration for 12 weeks after high dose group rats, poor diet, weight growth was slow;no significant changes of Hematology was found;The ALT, AST and ALP of high dose group rats increased[respectively(77.5±11.9)U/L,(210.4±21.7)U/L,(220.6±19.8)U/L], compared with the blank control group[respectively(55.2±12.1)U/L,(180.4±21.3)U/L,(190.3±22.6)U/L], the difference was statistically significant(P<0.05). The ratio of liver body in high dose group(3.86±0.29)was higher than the control group(3.52±0.25), the difference was statistically significant(P<0.05). And liver degeneration, focal necrosis was found.Conclusion The main chronic toxic damage is liver damage caused by sustained-release dropping pills of Rhynchophylla total alkaloids of long-term large delivery.

Objective: To examine the effect of lifestyle behaviors on myopia among primary and middle school students in Yinzhou District, Ningbo City using latent class analysis, so as to provide insights into prevention of myopia among primary and middle school students. Methods: A total of 1 547 students were sampled from primary and middle schools in Yinzhou District using a stratified cluster sampling method. Myopia-associated lifestyle behaviors were collected using questionnaires, and visual acuity was tested. Myopia-associated lifestyle behaviors were subjected to latent class analysis among primary and middle school students, and the association between lifestyle behaviors and risk of myopia was examined using a logistic regression model. Results: A total of 1 547 questionnaires were allocated, and 1 494 valid questionnaires were recovered, with an effective recovery rate of 96.57%. There were 247 primary school students (16.53%), 452 junior high school students (30.25%), 532 senior high school students (35.61%) and 263 vocational high school students (17.60%), and 773 men (51.74%) and 721 women (48.26%). Latent class analysis showed that students' lifestyle behaviors were classified into three groups, including the healthy behavior group (37.68%), reading and writing fatigue group (42.64%), and video fatigue and poor diet group (19.68%), with myopia prevalence of 79.22%, 88.38% and 86.73%, respectively. Moderate myopia was predominant in the reading and writing fatigue group and video fatigue and poor diet group, and low myopia was predominant in the healthy behavior group. A higher risk of myopia was found in the reading and writing fatigue group (OR=1.996, 95%CI: 1.454-2.739) and video fatigue and poor diet group (OR=1.715, 95%CI: 1.157-2.543) than in the healthy behavior group. Conclusions Long reading and writing duration, insufficient exercise and sleep, long video watching duration, and high intake frequency of sugary drinks and fried foods may increase the risk of myopia in primary and middle school students in Yinzhou District. Targeted myopia interventions are required tailored to different categories of lifestyle behaviors.

Objective To investigate the correlation between vital capacity, pulmonary ventilation and morphological parameters among children living in different altitude areas, so as to provide a reference basis for the development of prevention and control strategies for high altitude illness. Methods From January 2019 to June 2020, primary and secondary school students aged 7 to 15 years old were randomly selected from three different altitude areas, which were Xining (2 260m, low altitude group), Haixi (2 900m, medium altitude group), and Yushu (4 493m, high altitude group), respectively. The vital capacity, pulmonary ventilation and morphological parameters of the selected children were recorded. Results The vital capacity, pulmonary ventilation and morphological parameters showed statistically significant difference among three groups (P<0.05). The vital capacity and pulmonary ventilation were positively correlated with lung volume, but negatively correlated with lung density and lung artery diameter (P<0.05). Logistic regression analysis showed that there were three factors affecting children's vital capacity and lung ventilation: mean lung density, total lung transverse diameter, and total lung volume (P<0.05). Conclusion The monitoring of lung morphological indexes, mean lung density, total lung transverse diameter, and total lung volume can effectively judge children's lung function, and have certain value in the prevention and treatment of related high-altitude illness.

Objective The aim of this study was to investigate the pulmonary function of healthy humans at middle and high altitudes using a 256-slice multidetector CT (MDCT) scanner.Methods We enrolled 40 healthy male volunteers aged 18－45 years in this study, 20 from the middle-altitude area (at a mean altitude of 2 260 m, the MA group) and the other 20 from the high-altitude area (at a mean altitude of 4 000 m, the HA group). Using 256-slice MDCT, we performed inspiratory and expiratory CT scanning of the lungs, analyzed the images with the GE (AW 4.6) Workstation software and collected such pulmonary function parameters as the mean lung density in the full inspiratory phase (MLDin) and expiratory phase (MLDex), lung volume in the full inspiratory phase (Vin) and expiratory phase (Vex), difference between Vin and Vex (Vin-Vex), and ratio of Vin to Vex (Vin/Vex), followed by comparison of the parameters between the two groups of subjects.Results Statistically significant differences were observed between the MA and HA groups in MLDex (\[-705.90±25.63] vs \[-745.50±12.76\] HU, P=0.000) but not in MLDin (\[-869.80±20.66\] vs \[-865.85±22.57\] HU, P=0.567). The Vex was markedly higher in the HA than in the MA group (\[2 279.59±520.25\] vs \[1 566.48±350.97\] mL, P<0.05) while both Vin-Vex and Vin/Vex were remarkably lower in the former than in the latter group (P<0.05).Conclusion CT quantitative technology may offer a deeper insight into human pulmonary function at a high altitude and provide some imaging evidence for the high-altitude medical explanation of the mechanisms underlying the adaptive capacity of human pulmonary function to hypoxic environment.

Objective To investigate the possibility of the involvement of RhoA/mDia1 pathway to cause the expression of phosphorylate ezrin-radixin-moesin (p-ERM) in rat pulmonary micro-vascular endothelial cell (PMVEC) after the stimulation of lipopolysaccharide (LPS).Methods The specimens of lung tissue were taken from healthy male SPF grade SD rat with 100-120 g body weight which was purchased from the laboratory animal center of Anhui province.After culture,the PMVECs were randomly divided into dose-dependent groups (0,0.1,1,10 μg/mL LPS added in PMVECs and cultured for30 min,n =8 in each),time-dependent groups (10 μg/mL LPS added to PMVECs cultured for 0,15,30,60,120 min,n =8 in each) and intervention group (n =8).In the intervention group,PMVECs were cultured with 1 μg/mL C3 transferase in serum free media for 240 min,followed by treatment with 10 μg/mL LPS for 30 min.Meanwhile,two control groups in serum-free DMEM medium were made by adding 10.μg/mL LPS to PMVECs and 1 μg/mL C3 transferase to PMVECs respectively cultured for 30 min (n =8 in each).Western blot was used to detect the level of p-ERM,ERM and mDia1.Data were analyzed with SPSS 16.0 software,while one way analysis of variance (ANOVA) was used to compare multiple sets of variables,the intergroup comparisons were analyzed by the least-significant-difference (LSD) tests,with P ＜0.05 for the statistically significant difference.Results ERM,p-ERM and mDia1 were presented in rat PMVEC.Stimulation with LPS up-regulated p-ERM,mDia1 in a dose-dependent manner:LPS [0 μg/mL LPS group:(0.520±0.101),0.1 μg/mL LPS group:(0.657 ±0.092),1 μg/mL LPS group:(0.891 ±0.167),10 μg/mL LPS group:(1.227 ±0.106);0 μg/mL group vs.0.1 μg/mL group,P ＞0.05;the rest P ＜0.01];and mDia1 [0 μg/mL LPS group:(0.200 ±0.102),0.1 μg/mL LPS group:(0.430 ±0.121),1 μg/mL LPS group:(0.603 ±0.154),10 μg/mL LPS group:(0.887 ±0.204);0.1 μg/mL group vs.1 μg/mL group,P ＞ 0.05;the rest P ＜ 0.05].In time-dependent group,the level of p-ERM protein increased at 15 min (0.670 ±0.149),peaked at 30 min (1.175 ±0.103),then decreased,at 60 min (0.959 ±0.189),90 min (0.842 ±0.129),but kept at higher level at 120 min (0.767 ±0.097) than that in control group (0.471 ±0.157,15 min group vs.120 min group,60 min group vs.90 min group and 90 min group vs.120 min group,P ＞ 0.05;the rest P ＜ 0.05);and the level of mDia1 increased at 15 min (0.779 ±0.035),peaked at 30 min (0.889 ±0.036) then decreased at 60 min (0.648 ±0.019),90 min (0.582 ±0.068),but kept at higher level at 120 min (0.526 ±0.059) than that in control group (0.284±0.118,all P ＜ 0.01).C3 transferase caused a marked attenuation of LPS induced p-ERM expression [control group:(0.339 ± 0.069);C3 + LPS group:(0.438 ± 0.07);C3 control group:(0.352 ± 0.071);LPS control group:(0.634 ± 0.191),C3 + LPS group vs.LPS control group,P =0.01],as the same in mDia1 [control group:(0.449 ±0.122);C3 + LPS group:(0.380 ±0.148);C3 control group:(0.404 ±0.164);LPS control group:(0.622 ±0.174),C3 + LPS group vs.LPS control group,P ＜ 0.01].Conclusions LPS could up-regulated the expression of p-ERM protein,and inhibition of RhoA/mDia1 signal pathway by C3 transferase could down-regulated the p-ERM levels.

Objective To study the role of src-suppressed C kinase substrate (SSeCKS) in the secretion of tumor necrosis factor (TNF-α) in rat pulmonary micro-vascular endothelial cells (PMVEC) induced by lipopolysaccharide (LPS).Methods Wistar rat PMVEC cultured in vitro were randomly (random number) divided into several groups (n =4) as per exposure to given dosage of LPS for different lengths of time and to different dosages of LPS for given length of time.After PMVEC exposed to 10 mg/L LPS for 1 hour (h),3 h,6 h,12 h and 24 h or 0.1 mg/L,1 mg/L and 10 mg/L LPS for 24 h,the levels of TNF-αin the supernatant of culture medium were examined by the method of enzyme linked immunosorbent assay (ELISA).Another PMVEC was pre-treated by protein kinase C (PKC) inhibitor bis-indolylmaleimide (BIM) for 0.5 h or had the transfection of SSeCKS-specific small interfering RNA (siRNA) for 48 h before 10 mg/L LPS challenge for 24 h,and subsequently the supernatant was also examined by ELISA.One-way analysis of variance (ANOVA) was employed for statistical analysis by SPSS version 10.0 to compare values among all groups.A significant difference was presumed as a probability value ＜ 0.05.Results After PMVEC incubated with 0.1 mg/L,1 mg/L and 10 mg/L LPS for 24 hours,the levels of TNF-αsecreted were (253.70 ± 23.55),(327.88 ± 37.25),(403.20 ± 36.22),respectively,which were higher than that in un-stimulated PMVEC (82.28 ± 22.56,all P =0.000).After 10 mg/L LPS challenge for one hour,the level of TNF-αin the supernatant of PMVEC raised substantially (170.11 ±49.22),peaked at the time of 6 h (404.82 ± 13.78),then persisted at a higher level until 24 h (395.67 ± 36.23) than that in un-stimulated PMVEC (84.60 ± 23.61,P =0.001,0.000,0.000,respectively).After PMVEC pre-incubated with BIM,the level of LPS-induced TNF-αdecreased obviously (200.44 ± 27.39 vs.402.28 ± 31.07,P =0.000).Compared with LPS challenged PMVEC (407.28 ± 32.64),depletion of endogenous SSeCKS in PMVEC after inhibited by SSeCKS-siRNA significantly attenuated increase in the level of LPS-induced TNF-α (195.20 ± 13.28,P =0.000).Conclusions Down-activation of SSeCKS and PKC can inhibit the secretion of TNF-αin PMVEC induced by LPS,relieving the inflammatory response of PMVEC.

Aim To investigate the role of cAMP re-sponse element binding protein (CREB)in the injury of rat pulmonary microvascular endothelial cell (RPM-VEC)induced by LPS.Methods RPMVECs were i-solated and cultured in vitro,Western-blot was used to assay phosphorylation levels of CREB.Endothelial per-meability was determined by measuring the influx of Evans blue-labeled albumin across endothelial mono-layer.Results LPS increased CREB phosphorylation at Ser 1 3 3 in RPMVEC in a time-dependent manner , peaked at 30 min,but still higher at 120 min compared with basal control group.Pretreatment of cells with PKA inhibitor V5681 nearly suppressed the CREB phosphorylation stimulated in the presence of LPS,and the monolayer permeability of PMVEC was significantly increased. Conclusions LPS rapidly induces the phosphorylation of CREB in RPMVEC,and PKA me-diates the process.During the process of LPS-stimula-ted injury of RPMVEC,phosphorylation of CREB may play a protective role.

Objective: To analyze the association between latent class of health-risk behaviors and depressive symptoms among middle school students, so as to provide the evidence for the prevention and intervention of depressive symptoms among middle school students. Methods: Students in two junior high schools, two senior high schools and one vocational high school in Yinzhou District, Ningbo City, Zhejiang Province, were selected using a stratified multi-stage cluster sampling method. Demography and health-risk behaviors were collected using questionnaire surveys, depressive symptoms were investigated using the Center for Epidemiological Studies Depression-10 Scale, and latent class analysis was conducted for health-risk behaviors. The association between different latent classes and depressive symptoms was analyzed using a multivariable logistic regression model. Results: A total of 1 247 students were surveyed, including 641 boys (51.40%) and 606 girls (48.60%). There were 452 junior high school students (36.25%), 532 high school students (42.66%) and 263 vocational high school students (21.09%). Latent class analysis showed that health-risk behaviors in students were classified into three groups, namely healthy behavior group (52.93%), poor diet group (39.94%) and high-risk behavior group (7.14%), and the detection rates of depressive symptoms were 7.12%, 18.88% and 52.81%, respectively, with a statistically significant difference between groups (P<0.05). Multivariable logistic regression analysis showed that after adjusting for age, gender, native place, only child and living on campus, the poor diet group (OR=3.107, 95%CI: 2.086-4.627) and high-risk behavior group (OR=15.401, 95%CI: 9.031-26.262) had higher risks of depressive symptoms compared with the healthy behavior group. Conclusion Having high-risk behaviors and poor diet may increase the risk of developing depressive symptoms among middle school students.

Objective To identify the differentially expressed proteins in different liver tissues in the mouse model of cystic echinococcosis (CE), so as to provide insights into the research and development of therapeutic drugs targeting CE. Methods Female Kunming mice at ages of 6 to 8 weeks were randomly assigned into the CE group and the control group. Mice in the CE group were intraperitoneally infected with 2 000 Echinococcus multilocularis protoscoleces, while mice in the control group were injected with the same volume of physiological saline. All mice in both groups were sacrificed after breeding for 350 d, and the lesions (the lesion group) and peri-lesion specimens (the peri-lesion group) were sampled from the liver of mice in the CE group and the normal liver specimens (the normal group) were sampled from mice in the control group for data independent acquisition (DIA) proteomics analysis, and the differentially expressed proteins were subjected to Gene Ontology (GO) term enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Results A total of 26 differentially expressed proteins were identified between the lesion group and the normal group and between the peri-lesion group and the normal group, including 8 up-regulated proteins and 18 down-regulated proteins. GO term enrichment analysis showed that these differentially expressed proteins were predominantly enriched in endoplasmic reticulum membrane (biological components), oxidoreductase activity (molecular function) and oxoacid metabolic process and monocarboxylic acid metabolic process (biological processes). KEGG pathway enrichment analysis revealed that the differentially expressed protein Acyl-CoA oxidase 1 (Acox1), which contributed to primary bile acid biosynthesis during the fatty acid oxidation, was involved in peroxisome signaling pathway, and the differentially expressed protein fatty acid binding protein 1 (Fabp1), which contributed to fatty acid transport, was involved in the peroxisome proliferator-activated receptor (PPAR) signaling pathway. Conclusion Differentially expressed proteins are identified in the liver specimens between mouse models of CE and normal mice, and some differentially expressed proteins may serve as potential drug targets for CE.