Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.

Aim To establish a reliable in vitro evaluation sys-tem to assess human liver UGTs activities and the inhibitory effect of drugs on human liver UGTs and validate its accuracy and stability.Methods Six probe substrate of the six major subtypes of human liver microsomal UGT enzymes were select-ed.The activity of human liver microsomal UGT enzymes was determined under optimized incubation conditions using a metab-olite generation method.The concentrations of metabolites were determined using a validated liquid chromatography-tandem mass spectrometry(LC-MS/MS)method.Known inhibitors were used to verify the established incubation system.Results The estab-lished LC-MS/MS analysis method complied with the require-ments of bioanalysis.In the incubation system of this experi-ment,known UGT inhibitors exhibited significant inhibitory effects.Conclusions This study successfully establishes an ac-curate and reliable in vitro evaluation system,which can be used to assess the in vitro inhibition of drugs on UGT enzymes,provi-ding important guidance for drug development and rational clini-cal medication.

This study was aimed at investigating the effect of lymphocyte activation gene-3(LAG3)on liver B lymphocyte subsets and their functions in WT and LAG3-KO mice infected with Echinococcus multilocularis(E.multilocularis).In a mouse model of E.multilocularis infection,the expression and localization of CD19 and α-SMA in liver were detected by immu nohistochemistry.CD80,CD86 and MHC-Ⅱ molecules expressed on B cells and their subsets in mice liver were detected by flow cytometry.After 12 weeks of infection,the area and percentage of CD19 in LAG3-KO group was slightly higher than that in WT group,but the difference was not statistically(t=-1.241、-1.237,P＞0.05).The area and percentage of a-SMA in LAG3-KO group was higher than that in WT group(t=-3.224、-3.227,P＜0.05).The proportion of CD80 and MHC-Ⅱ molecules expressed on liver B cells in LAG3-KO group was up-regulated(t=-2.379,-3.321,P＜0.05).The percentage of liver B2 cells in LAG3-KO group was higher than that in WT group(t=-2.695,P＜0.05).The expression of CD80 on Blb cells in LAG3-KO group was significantly up-regulated(t=-5.315,P＜0.001).The proportion of CD80 of B2 cells in LAG3-KO group was lower than that in WT group(t=2.806,P＜0.05).The expression of MHC-Ⅱ molecule in B2 cells in LAG3-KO group was up-regulated(t=-4.227,P＜0.01).It is suggested that LAG3 molecules affected the B cell subsets and func-tion of mouse liver in the middle stage of E.multilocularis infection,especially B2 lymphocytes.LAG3 molecule exerted an in-hibitory effect on the activation of B cells and the expression of MHC-class Ⅱ molecules,suggesting that it may be involved in B cell exhaustion caused by E.multilocularis.

AIM To study the chemical constituents form the leaves of Castanopsis orthacantha Fance and their α-glucosidase inhibitory activities.METHODS The methanol extract form the leaves of C.orthacanth was isolated and purified by various column chromatography methods,such as MCI gel CHP 20P,Sephadex LH-20,Diaion HP20SS,then the structures of obtained compounds were identified by physicochemical properties and spectral data.The α-glucosidase inhibitory activities were determined by PNPG method.RESULTS Eighteen compounds were isolated and identified as protocatechuic acid(1),gallic acid(2),3-O-α-L-arabininopyranosyl-4-hydroxybenzoic acid(3),3-O-galloyl shikimic acid(4),methyl 4-epi-shikimate-3-O-gallate(5),5-O-galloyl shikimic acid(6),5-O-caffeoylshikimic acid(7),6-O-galloyl-glucoside(8),1,6-di-O-galloyl-β-D-pyranogluloside(9),1,3-di-O-galloyl-α-D-glucoside(10),2,3-di-O-galloyl-D-glucoside(11),β-O-methylgluco-2,3-digalloyl esters(12),(3R,1'S)-[1'-(6"-O-galloyl-β-D-gluco-pyranosyl)oxyethyl]-3-hydroxy-dihydrofuran-2(3H)-one(13),4-O-D-(6'-O-galloyl)glucopyranyl-(E)-p-coumaryl acid(14),chestanin(15),1-desgalloyl eugeniin(16),picrorhiza acid(17),11-methyl chebulate(18).The IC50 values of compounds 2 and 16 were(0.12±0.059),(0.00089±0.00025)mmol/L,respectively.CONCLUSION All compounds are isolated from the leaves of C.orthacantha for the first time.Compounds 2 and 16 have strong α-glucosidase inhibitory activities.

Objective To investigate the biomechanical effects of the backside design of tibial trays on the bone-prosthesis fixation interfaces in unicompartmental knee arthroplasty(UKA).Methods Finite element models of medial knee arthroplasty were constructed using a fixed UKA prosthesis.The knee joint load and joint motion under walking motion were considered as boundary conditions,and the differences in tibial von Mises stress,contact stress,and micromotion of the bone-prosthesis fixation interface of the UKA tibial trays with big keel,small keel,two-peg with fin,three-oblique peg,and three-upright peg types were compared.Results At the maximum medical knee force moment,compared to the two-peg with fin type,the tibial von Mises stress,contact stress,and micromotion of the bone-prosthesis fixation interface decreased by 8%and 15.9%and increased by 9.9%for the big keel type;decreased by 12.3%and increased by 7.5%and 0.9%for the small keel type;decreased by 10%,10.5%,and increased by 1.2%for the three-oblique peg type;and decreased by 7.7%,14.7%,and 1.6%for the three-upright peg type,respectively.However,the maximum micromotion of the bone-prosthesis fixation interface occurred at 21%of the gait cycle.Compared to the two-peg with fin type,the micromotion of the bone-prosthesis fixation interface increased by 11.6%for the big keel type,increased by 1.6%for the small keel type,decreased by 0.4%for the three-oblique peg type,and decreased by 2.3%for the three-upright peg type.Conclusions To improve the long-term fixation effects of tibial prostheses,it is recommended to focus on a two-upright peg with fin or small keel designs when UKA tibial trays are designed,which can effectively balance the stress transfer and interface micromotion,thereby ensuring prosthesis stability and reducing the risk of aseptic loosening.

Objective To understand the respiratory protection competency of staff in hospitals.Methods Staff from six hospitals of different levels and characteristics in Beijing were selected,including doctors,nurses,medical technicians,and servicers,to conduct knowledge assessment on respiratory protection competency.According to exposure risks of respiratory infectious diseases,based on actual cases and daily work scenarios,content of respira-tory protection competency assessment was designed from three aspects:identification of respiratory infectious di-seases,transmission routes and corresponding protection requirements,as well as correct selection and use of masks.The assessment included 6,6,and 8 knowledge points respectively,with 20 knowledge points in total,all of which were choice questions.For multiple-choice questions,full marks,partial marks,and no mark were given respective-ly if all options were correct,partial options were correct and without incorrect options,and partial options were correct but with incorrect options.Difficulty and discrimination analyses on question of each knowledge point was conducted based on classical test theory.Results The respiratory protection competency knowledge assessment for 326 staff members at different risk levels in 6 hospitals showed that concerning the 20 knowledge points,more than 60％participants got full marks for 6 points,while the proportion of full marks for other questions was relatively low.Less than 10％participants got full marks for the following 5 knowledge points:types of airborne diseases,types of droplet-borne diseases,conventional measures for the prevention and control of healthcare-associated infec-tion with respiratory infectious diseases,indications for wearing respirators,and indications for wearing medical protective masks.Among the 20 knowledge questions,5,1,and 14 questions were relatively easy,medium,and difficult,respectively;6,1,4,and 9 questions were with discrimination levels of ≥0.4,0.30-0.39,0.20-0.29,and ≤0.19,respectively.Conclusion There is still much room for hospital staff to improve their respiratory protection competency,especially in the recognition of diseases with different transmission routes and the indications for wearing different types of masks.

Objective To investigate the implementation of surveillance,prevention and control measures for healthcare-associated infection(HAI)in maternal and child healthcare(MCH)institutions,and provide policy evi-dence for optimizing HAI prevention and control in MCH institutions.Methods Stratified sampling was conducted among the MCH institutions at provincial,municipal and county levels in 8 provinces/autonomous regions.A uni-fied questionnaire was designed and the online survey was conducted through"Questionnaire Star".Results The data from 123 MCH institutions were included in the analysis.90.24％of the MCH institutions carried out compre-hensive surveillance on HAI.The ratios of MCH institutions which implemented targeted surveillance on HAI in neonatal intensive care unit(NICU),surgical site infection,multidrug-resistant organisms(MDROs)and HAI in intensive care units(non-NICU excluded)were 89.66％,85.96％,80.77％,and 74.19％,respectively.51.22％MCH institutions adopted information surveillance system on HAI cases.94.31％MCH institutions carried out surveillance on hand hygiene compliance.Over 90％MCH institutions carried out surveillance on environment hy-giene in high-risk departments.71.54％MCH institutions conducted centralized cleaning,disinfection,sterilization and supply for reusable medical instruments in the central sterile supply department(CSSD).Over 90％MCH insti-tutions established three-level pre-examination triage systems.86.18％set up transitional wards.MCH institutions generally adopted a management model with established effective communication,full appointment visits,and sepa-rate visits for special medical groups,such as registered pregnant women,high-risk newborns,healthcare groups,and long-term rehabilitation patients.However,the ratio of institutions conducting on-line follow-up visits was less than 50％.Conclusion MCH institutions have generally carried out comprehensive and targeted surveillance on HAI.Information surveillance need to be facilitated.Hand hygiene and environmental hygiene surveillance has been popularized to a certain extent at all levels of MCH institutions.The cleaning,disinfection,sterilization,and supply processes of reusable medical devices in a few MCH institutions are not standardized.Special medical populations get effective management.On-line healthcare is to be further promoted.

Objective To investigate the impact of silymarin(SM)on the malignant growth of glioma cells and the regulatory mechanism on the miR-124-3p/WEE1 axis.Methods Glioma U87 cells were grouped into control,SM low,medium,and high concentration groups,and SM high concentration + miR-124-3p inhibitor group(SM high + miR-124-3p inhibitor group).CCK-8 was used to measure the proli-feration rate of cells;Transwell? assay was applied to assay the migration and invasion of cells;cell cycle progression was detected by flow cytometry;Western blotting was applied to measure the expression of cyclin D1 and apoptosis-related proteins;the levels of miR-124-3p and WEE1 mRNA were determined by qRT-PCR;and a luciferase activity test was applied to verify the targeting relationship between miR-124-3p and WEE1;in addition,the establishment,administration,and analysis of a NOD/SCID mouse model of intracranial trans-planted tumor were conducted.Results Compared with the control group,the cell proliferation,the numbers of migrating and invading cells,the expression of cyclin D1,and the level of WEE1 mRNA in the various SM treatment groups decreased,the number of cells in G0/G1 phase,the expression of cleaved caspase-8,cleaved caspase-9,cleaved caspase-3 and miR-124-3p increased(P＜0.05);furthermore,transfection of miR-124-3p inhibitor reversed the inhibitory effect of SM on the malignant behavior of glioma cells.In vivo experiments with mice showed that the weights and volumes of tumors in the SM treatment group were lower than those in the model group(P＜0.05),and there was no discernible change in the weight of the mice(P＞0.05).Conclusion SM can inhibit the malignant growth of glioma cells by upregulating miR-124-3p and downregulating WEE1.

Objective: To determine the inhibitory effects of pachymic acid on lung adenocarcinoma (LUAD) cells and elucidate its underlying mechanism. Methods: CCK-8, wound healing, Transwell, Western blot, tube formation, and immunofluorescence assays were carried out to measure the effects of various concentrations of pachymic acid on LUAD cell proliferation, metastasis, angiogenesis as well as autophagy. Subsequently, molecular docking technology was used to detect the potential targeted binding association between pachymic acid and protein tyrosine phosphatase 1B (PTP1B). Moreover, PTP1B was overexpressed in A549 cells to detect the specific mechanisms of pachymic acid. Results: Pachymic acid suppressed LUAD cell viability, metastasis as well as angiogenesis while inducing cell autophagy. It also targeted PTP1B and lowered PTP1B expression. However, PTP1B overexpression reversed the effects of pachymic acid on metastasis, angiogenesis, and autophagy as well as the expression of Wnt3a and β-catenin in LUAD cells. Conclusions: Pachymic acid inhibits metastasis and angiogenesis, and promotes autophagy in LUAD cells by modulating the Wnt/ β-catenin signaling pathway via targeting PTP1B.

Objective To investigate the effect and mechanism of endothelin-1(ET-1)on atrial fibrosis in Atrial fibrillation(AF)rats.Methods Fourteen adult male SD rats were randomly divided into normal control(NC)group and Atrial fibrillation(AF)group.The rat model of Atrial fibrillation was established by injecting 0.1 ml/100g CaCl2-Ach mixture into the tail vein once a day for one week.The control group was injected with the same dose of normal saline.An electrocardiogram of normal or atrial fibrillation was recorded on the first day and the eighth day in each group,and echocardiography was used to monitor atrial size and cardiac function.The fibrosis of atrial was observed using Masson and HE staining.The expression of endothelin-1(ET-1),collagen-I(Col-I),transforming growth factor-β(TGF-β)and the store operated calcium channel(SOCC)protein Orai1,stromal in-teraction molecule 1(STIM1)in atrial tissue were detected by Western blot.HL-1 cells were cultured and treated with gradient concentration of ET-1 for 24 hours.Western blot was used to observe changes in the expression of TGF-β,Orai1 and STIM1 proteins in ET-1/SOCC/TGF-β signaling pathway of HL-1 cells.Small interfering RNA(siRNA)transfection method was used to knock down the expression of Orai1 in HL-1 cells,then the cells were treated with appropriate concentrations of ET-1 for 24 hours,and the expression of TGF-β protein in HL-1 cells was detected by Western blot.Results Compared with the control group,echocardiography showed a significant in-crease in left atrial diameter(LAD)of the heart in atrial fibrillation rats(P<0.05).The HE and Masson staining results showed significant fibrosis in the myocardial tissue of AF group rats(P<0.05),and the Western blot re-sults indicated the expression of ET-1,Orai1,STIM1,TGF-β and COL-Ⅰ in the myocardial tissue of AF group significantly increased compared to the NC group(P<0.05).After ET-1 treatment of HL-1 cells,the protein ex-pression of Orai1,STIM1and TGF-β increased(P<0.05),while knocking down Orai1 in HL-1 cells,ET-1 treat-ment no longer caused the expression of TGF-β a significant upregulation.Conclusion AF caused by atrial fibril-lation results in a significant increase in ET-1 expression in atrial tissue,and ET-1/SOCC/TGF-β signal pathway promotes atrial fibrillation and fibrosis.