Objective To investigate the role of RNA-binding motif protein X-linked(RBMX)in regulating the proliferation,migration,invasion and glycolysis in human bladder cancer cells.Methods A lentivirus vectors system and RNA interference technique were used to construct bladder cancer 1376 and UC-3 cell models with RBMX overexpression and knockdown,respectively,and successful cell modeling was verified using RT-qPCR and Western blotting.Proliferation and colony forming ability of the cells were evaluated using EdU assay and colony-forming assay,and cell migration and invasion abilities were determined using Transwell experiment.The expressions of glycolysis-related proteins M1 pyruvate kinase(PKM1)and M2 pyruvate kinase(PKM2)were detected using Western blotting.The effects of RBMX overexpression and knockdown on glycolysis in the bladder cancer cells were assessed using glucose and lactic acid detection kits.Results RT-qPCR and Western blotting confirmed successful construction of 1376 and UC-3 cell models with RBMX overexpression and knockdown.RBMX overexpression significantly inhibited the proliferation,clone formation,migration and invasion of bladder cancer cells,while RBMX knockdown produced the opposite effects.Western blotting results showed that RBMX overexpression increased the expression of PKM1 and decreased the expression of PKM2,while RBMX knockdown produced the opposite effects.Glucose consumption and lactate production levels were significantly lowered in the cells with RBMX overexpression(P<0.05)but increased significantly following RBMX knockdown(P<0.05).Conclusion RBMX overexpression inhibits bladder cancer progression and lowers glycolysis level in bladder cancer cells by downregulating PKM2 expression,suggesting the potential of RBMX as a molecular target for diagnosis and treatment of bladder cancer.

Objective To investigate the role of RNA-binding motif protein X-linked(RBMX)in regulating the proliferation,migration,invasion and glycolysis in human bladder cancer cells.Methods A lentivirus vectors system and RNA interference technique were used to construct bladder cancer 1376 and UC-3 cell models with RBMX overexpression and knockdown,respectively,and successful cell modeling was verified using RT-qPCR and Western blotting.Proliferation and colony forming ability of the cells were evaluated using EdU assay and colony-forming assay,and cell migration and invasion abilities were determined using Transwell experiment.The expressions of glycolysis-related proteins M1 pyruvate kinase(PKM1)and M2 pyruvate kinase(PKM2)were detected using Western blotting.The effects of RBMX overexpression and knockdown on glycolysis in the bladder cancer cells were assessed using glucose and lactic acid detection kits.Results RT-qPCR and Western blotting confirmed successful construction of 1376 and UC-3 cell models with RBMX overexpression and knockdown.RBMX overexpression significantly inhibited the proliferation,clone formation,migration and invasion of bladder cancer cells,while RBMX knockdown produced the opposite effects.Western blotting results showed that RBMX overexpression increased the expression of PKM1 and decreased the expression of PKM2,while RBMX knockdown produced the opposite effects.Glucose consumption and lactate production levels were significantly lowered in the cells with RBMX overexpression(P<0.05)but increased significantly following RBMX knockdown(P<0.05).Conclusion RBMX overexpression inhibits bladder cancer progression and lowers glycolysis level in bladder cancer cells by downregulating PKM2 expression,suggesting the potential of RBMX as a molecular target for diagnosis and treatment of bladder cancer.

Objective To investigate the expression levels of selenoprotein genes in the patients with coronavirus disease 2019(COVID-19)and the possible regulatory mechanisms.Methods The dataset GSE177477 was obtained from the Gene Expression Omnibus,consisting of a symptomatic group(n=11),an asymptomatic group(n=18),and a healthy control group(n=18).The dataset was preprocessed to screen the differentially expressed genes(DEG)related to COVID-19,and gene ontology functional annotation and Kyoto encyclopedia of genes and genomes enrichment analysis were performed for the DEGs.The protein-protein interaction network of DEGs was established,and multivariate Logistic regression was employed to analyze the effects of selenoprotein genes on the presence/absence of symptoms in the patients with COVID-19.Results Compared with the healthy control,the symptomatic COVID-19 patients presented up-regulated expression of GPX1,GPX4,GPX6,DIO2,TXNRD1,SELENOF,SELENOK,SELENOS,SELENOT,and SELENOW and down-regulated ex-pression of TXNRD2 and SELENON(all P＜0.05).The asymptomatic patients showcased up-regulated expres-sion of GPX2,SELENOI,SELENOO,SELENOS,SELENOT,and SELENOW and down-regulated expression of SELP(all P＜0.05).The results of multivariate Logistic regression analysis showed that the abnormally high expression of GPX1(OR=0.067,95％CI=0.005-0.904,P=0.042)and SELENON(OR=56.663,95％CI=3.114-856.999,P=0.006)was the risk factor for symptomatic COVID-19,and the abnormally high expres-sion of SELP was a risk factor for asymptomatic COVID-19(OR=15.000,95％CI=2.537-88.701,P=0.003).Conclusions Selenoprotein genes with differential expression are involved in the regulation of COVID-19 development.The findings provide a new reference for the prevention and treatment of COVID-19.

Objective:To preliminarily evaluate the immunogenicity and efficacy of two novel tuberculosis vaccine candidates (a fusion multicomponent protein EPDPA015f and a mixed multicomponent protein EPDPA015m) and to provide a new antigen combination for the development of tuberculosis vaccines.Methods:Recombinant plasmids for the expression of EPDPA015f and EPDPA015m proteins were constructed. Six-week-old BALB/c mice were immunized with EPDPA015f or EPDPA015m in combination with aluminium adjuvant (50 μg/mouse) for three times with an interval of 10 d. The mice were sacrificed 10 d after the last immunization to collect blood and spleen samples. Serum antibody titers and cytokine levels were measured by ELISA, Luminex technique and enzyme-linked immunospot assay (ELISPOT). Mycobacterial growth inhibition assay (MGIA) was used to detect the ability of mouse splenocytes to inhibit the growth of Mtb in vitro. One-way analysis of variance and t-test were used for statistical analysis. Results:Both EPDPA015f and EPDPA015m could induce the production of various cytokines and IgG antibodies at a high level. The levels of cytokines related to Th1 (IL-2, TNF-α, IFN-γ), Th2 (IL-4, IL-6, IL-10) and Th17 (IL-17) as well as other proinflammatory cytokines (GM-CSF, IL-12) were higher in the EPDPA015f group than in the adjuvant group ( P<0.05). The titer of IgG antibody induced by EPDPA015f was as high as 1∶4×10 6. The results of MGIA showed that the numbers of Mtb (lgCFU) in the PBS, adjuvant, EPDPA015f and EPDPA015m groups were 3.46±0.11, 3.51±0.06, 2.98±0.09 and 3.19±0.08, respectively. The number of colonies in the EPDPA015f group was the least as compared with that in the other three groups ( P<0.001, P<0.001, P<0.01). Conclusions:The vaccine candidate EPDPA015f could elicit more comprehensive and high-level cellular and humoral immune responses, and exhibited superior in vitro inhibitory activity against the growth of Mtb. EPDPA015f had the potential to be used as a preventive vaccine or a booster vaccine

ObjectiveTo investigate the individual factors of postural adjustment reaction time and movement time during adaptive equilibrium. MethodsFrom March to December, 2021, 126 healthy adults aged 18 to 80 years were recruited at the First Medical Center of the General Hospital of the Chinese PLA. The DE-A somatosensory balance detection system was used to detect their postural adjustment reaction time (RT) and movement time (MT) as the platform tilting in multiple directions during standing (static) or walking (dynamic). The ages, genders, body mass index (BMI) and physical activity level of them were investigated. ResultsThe age was the only factor independently associated with dynamic RT and MT in all the directions (β > 0.632, P < 0.05). For static MT, as the platform tilting forward, physical activity level (β = -0.143, P < 0.05), BMI (β = 0.154, P < 0.05) and age (β = 0.663, P < 0.05) were the independently associated factors; as the platform tilting leftward, gender (β = -0.173, P < 0.05) and age (β = 0.647, P < 0.05) were the independently associated factors; and age was the only independently associated factor for other directions (β > 0.571, P < 0.05). For the static RT, age was the only independently associated factor for all the directions (β > 0.615, P < 0.05). ConclusionAge is the most important independently factor related to postural adjustment during adaptive equilibrium, and aging may delay the postural adjustment after instability.

Objective:To evaluate the immunogenicity and protective effect of a multicomponent recombinant protein vaccine EPRHP014 constructed independently and provide a scientific basis for developing new tuberculosis (TB) vaccine and effective prevention and control of TB.Methods:Three full-length Mycobacterium ( M.) tuberculosis protein antigens (EsxH, Rv2628, and HspX) and two epitope-predicted and optimized epitope-dominant protein antigens (nPPE18 and nPstS1) were selected, from which five protein antigens were used to construct a protein antigen composition EPRHP014, including a fusion expression multi-component protein antigen (EPRHP014f) and a multi-component mixed protein antigen (EPRHP014m) formed with the five single protein using clone, purification, and purification respectively. Multicomponent protein vaccines EPRHP014f and EPRHP014m were prepared with aluminum adjuvant, and the BCG vaccine was used as a control. ELISA detected the titer of serum-specific antibodies, the secretion of various cytokines was detected by ELISpot and Luminex, and immune protection was observed by the M.tuberculosis growth inhibition test in vitro. The results were statistically analyzed by t-test or rank sum test, and P<0.05 was considered a statistically significant difference. Results:Mice Immunized with EPRHP014m and EPRHP014f could produce highly effective IgG antibodies and their subtypes IgG1 and IgG2a, and the antibody titers were similar to those of mice immunized with BCG, with no statistical significance ( P>0.05). The number of spot-forming cells (SFC) secreting IFN-γ and IL-4 induced by EPRHP014f group was significantly higher than those by EPRHP014m group and BCG group ( P<0.05), but there was no significant difference in the number of SFC for IFN-γ and IL-4 induced between EPRHP014m group and BCG group ( P>0.05). The secretion levels of GM-CSF and IL-12p70 induced by the EPRHP014m group were higher than those of the BCG group ( P<0.05), but there was no significant difference in the levels of IL-6 and IL-10 induced between EPRHP014m group and BCG group ( P>0.05). There was no significant difference in the secretions of IL-6, IL-10, IL-12, and GM-CSF between the EPRHP014f and BCG groups ( P>0.05). EPRHP014m group, EPRHP014f group, and BCG group had obvious antibacterial effects in vitro, and the difference was insignificant ( P>0.05). Conclusion:Both EPRHP014f and EPRHP014m can induce strong humoral and cellular immune responses in mice after immunization, and have a strong ability to inhibit the growth of M. tuberculosis in vitro, indicating that the antigen composition EPRHP014 has good potential in the development and application of TB vaccine.

Objective: To examine the association between the cross-resistance to ethionamide (Eto) and isoniazid (INH) and mutations of drug resistant genes in Mycobacterium tuberculosis (MTB), so as to provide the evidence for clinical diagnosis and treatment for multidrug-resistant (MDR) tuberculosis. Methods: Totally 126 MTB clinical isolates were selected, including 88 MDR-MTB clinical isolates and 38 INH- and rifampicin (RFP)-sensitive isolates. The resistance to INH and Eto was tested in MTB clinical isolates using the drug susceptibility test, and the mutations in the spacer region of INH and Eto resistance-related katG, inhA, ethA, mshA, ndh, spacer region of oxyR-ahpC and inhA promoter were detected using PCR assay. The phenotypic resistance served as a gold standard, and the sensitivity, specificity and accuracy of gene mutation tests were calculated for detection of MTB clinical isolates cross-resistant to INH and Eto. Results: Of the 126 MTB clinical isolates, there were 37 isolates cross-resistant to INH and Eto (29.37%), 51 isolates with resistance to INH and susceptibility to Eto (40.48%), 4 isolates with susceptibility to INH and resistance to Eto (3.17%) and 34 isolates with susceptibility to INH and Eto (26.98%). Among the 41 Eto-resistant MTB clinical isolates, there were 37 isolates with resistance to INH (90.24%). There were 64 MTB clinical isolates detected with katG mutations (50.79%), 4 isolates with mutation in the spacer region of oxyR-ahpC (3.17%), 2 isolates with inhA mutations (1.59%), and these isolates were all resistant to INH. There were 11 MTB clinical isolates detected with mutation in the inhA promoter (8.73%) and one isolate with ndh mutation, and all these isolates were cross-resistant to INH and Eto. There were 23 MTB clinical isolates detected with ethA mutations (18.25%) and 40 isolates with mshA mutations (31.75%), in which Eto-susceptible and -resistant isolates were detected. The diagnostic sensitivity, specificity and accuracy of inhA promoter tests for detection of cross-resistance to INH and Eto were 29.73% (95%CI: 16.44%-47.17%), 100.00% (95%CI: 87.36%-100.00%) and 63.38% (95%CI: 51.76%-73.63%) in MTB clinical isolates. Conclusions The prevalence of INH resistance is high in Eto-resistant MTB clinical isolates. Mutation in the inhA promoter region correlates with the cross-resistance to INH and Eto in MTB clinical isolates, and detection of mutation in the inhA promoter may be feasible to detect the cross-resistance to INH and Eto in MTB clinical isolates.

Objective:To understand the knowledge and skills of professional technicians in prevention and control of parasitic diseases in Shandong Province, and to provide a basis for further capacity building.Methods:On October 12 and 13, 2017, five professional technicians from each of 17 prefectures and cities in Shandong Province participated in the "Shandong Province Parasitic Diseases Prevention and Control Job Skills Competition in 2017". This competition included theoretical knowledge examination (written test) and operation skill examination (operation test). The written test included basic knowledge of parasites life history, pathogen, diagnosis, epidemic and prevention; the operation test included blood smear making, Kato-Katz thick smear making, microscopic examination of Plasmodium and Helminth eggs. The competition results of the competitors were statistically analyzed, and the pass rate, correct answer rate and accuracy of microscopic examination were calculated. According to the report of malaria cases from 2010 to 2017, 17 prefectures and cities in Shandong Province were divided into malaria classⅠreport area (≥100 cases) and malaria classⅡreport area (< 100 cases), and the competition results of the two types of areas were compared. Results:A total of 85 competitors in Shandong Province participated in the competition, including 19 males (22.35%) and 66 females (77.65%); the age was (34.67±6.04) years old, the youngest was 25 years old, and the oldest was 51 years old. The scores of written test and operation test were (67.06±12.73) and (59.31±14.23) points, respectively, and the difference between them was statistically significant ( t=4.949, P < 0.01). The pass rate of written test was 74.12% (63/85); the correct answer rates of morphology, life history, pathogenicity, diagnosis, epidemic and prevention were 71.76% (366/510), 71.61% (913/1 275), 67.94% (462/680), 71.18% (847/1 190), 66.91% (455/680) and 65.76% (1 062/1 615), respectively, there was statistically significant difference in the correct answer rates of different knowledge points (χ 2=18.185, P < 0.01). The pass rate of operation test was 55.29% (47/85); among them, the pass rates of blood smear making, Kato-Katz thick smear making, microscopic examination of Plasmodium and Helminth eggs were 98.82% (84/85), 98.82% (84/85), 70.59% (60/85) and 31.76% (27/85), respectively. Four Plasmodium species were examined, and the overall accuracy of microscopic examination was 61.41% (261/425), there was no statistically significant difference in the accuracy of microscopic examination between different Plasmodium species (χ 2=4.791, P > 0.05). Nine common Helminth eggs were examined, and the overall accuracy of microscopic examination was 47.29% (402/850), there was statistically significant difference in the accuracy of microscopic examination between different Helminth eggs (χ 2=180.064, P < 0.01). The scores [(28.27±3.74) and (23.20±3.39) points, n=30] of microscopic examination of Plasmodium and Helminth eggs in the malaria classⅠ report area were higher than those in the malaria classⅡreport area [(22.40±5.81) and (18.25±3.41) points, n=55], and the differences were statistically significant ( t=2.217, 2.860, P < 0.05). Conclusions:For professional technicians, the mastery of theoretical knowledge in prevention and control of parasitic diseases is better than operation skills in Shandong Province. So the training and assessment of prevention and control skills of parasitic diseases should be strengthened in areas with weak abilities.

Objective:To explore the effect of recombinant human syntaxin-4(STX4) on lipopolysaccharide(LPS)-induced injury in islet β-cells(INS-1).Methods:Pancreatic islet β-cells(INS-1) were divided into Control (blank control), LPS (LPS treatment), LPS+ NC (transfection of negative control vector, LPS treatment), and LPS+ STX4 (transfection of pcDNA-STX4, LPS treatment) groups. RT-qPCR and Western blot were used to detect STX4 mRNA and protein expression, flow cytometry to detect apoptosis, DCFHDA method to detect reactive oxygen species(ROS) level, xanthine oxidation method to detect superoxide orgotein dismutase(SOD) level, colorimetric method to detect glutathione peroxidase(GSH-Px) level, ammonium molybdate colorimetric method to detect catalase(CAT) level, thiobarbituric acid method to detect malonaldehyde(MDA) level, ELISA method to detect the level of interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), and insulin secretion levels under glucose conditions secreted by cells, Western blot method to detect Cleared Caspase-3, Bcl-2 Associated X Protein(Bax), p65 protein expression. After treatment with NF-κB signaling pathway activator, STX4 up-regulated islet β-cell INS-1 was given LPS stimulation, and the same method was used to measure apoptosis, ROS, SOD, GSH-Px, CAT, MDA levels and secreted IL-1β, TNF-α, insulin levels and Cleaved Caspase-3, Bax, p65 protein levels.Results:Compared with the Control group, the expression of STX4 mRNA and protein in islet β cells of the LPS group decreased, the apoptosis rate, ROS level, and MDA levels increased, and the levels of SOD, GSH-Px, and CAT decreased, the levels of IL-1β, TNF-α increased, the level of insulin secreted by the cells decreased, and the expression levels of Cleaved Caspase-3, Bax, and p65 also increased. NF-κB pathway activator treatment reversed the effect of up-regulated STX4 on islet β-cell apoptosis, ROS, SOD, GSH-Px, CAT, MDA levels and secreted IL-1β, TNF-α levels, and Cleaved Caspase-3 , Bax and p65 protein levels.Conclusion:Up-regulation of STX4 alleviated LPS-induced islet β cell oxidative damage, apoptosis and inflammatory factor release. The underlying mechanism might be related to the inhibition of activated NF-κB signaling pathway.

Objective:To analyze the application of Cho peak value and color doppler ultrasound blood flow score in the early diagnosis of breast cancer, and to evaluate the relationship between Cho peak value, blood flow score, TNM stage and prognosis quality.Methods:A total of 82 patients with breast cyst admitted from Jan. 2015 and Dec. 2019 were selected as subjects for the study. ROC curve was used to compare the ability of color doppler flow score and functional magnetic resonance imaging (fmri) in the diagnosis of breast cancer when used alone or in combination. Logistic regression model was used to analyze the factors affecting the prognosis quality and TNM staging of patients.Results:The breast cancer group’s Cho value and blood flow signal score were significantly higher than the benign breast lesion group, and the difference was statistically significant (Cho value: t=43.977, P<0.001; blood flow signal score: t=22.071, P<0.001) ; The sensitivity, specificity and AUC of MRS combined with Doppler ultrasound for differential diagnosis of breast cancer are significantly higher than MRS or Doppler ultrasound alone, and the difference was statistically significant (sensitivity: χ2=4.514, P=0.016; specificity: χ2=4.858, P=0.013; AUC: Z=5.251, P<0.001) ; Cho value of patients with good prognosis group ( t=3.984, P<0.001) and blood flow signal score ( t=4.213, P<0.001) were significantly lower than those in the poor prognosis group; Cho value ( t=3.612, P<0.001) and blood flow signal score ( t=3.835, P<0.001) of TNM stage 0-Ⅱ patients were significantly lower than those of stage Ⅲ-Ⅳ group, the difference was statistically significant; the Cho value of the MRS scan and the patient’s prognosis quality ( OR=1.837, 95% CI=1.210-2.788, P=0.004) and TNM stage ( OR=1.818, 95% CI=1.224~2.702, P=0.003) was significantly positively correlated. The blood flow signal and the patient’s prognostic quality ( OR=1.906, 95% CI=1.105~3.287, P=0.020) and TNM stages ( OR=1.799, 95% CI=1.232-2.626, P=0.002) also showed a significantly positive correlation. Conclusion:The combination of Cho peak value and color doppler ultrasound blood flow score can significantly improve the early diagnosis efficiency of breast cancer, and Cho peak value and blood flow score are independent factors affecting TNM staging and prognosis.