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.

ObjectiveTo analyze the differences in color， odor and volatile components of Citri Reticulatae Pericarpium（CRP） before and after being stir-fried with Halloysitum Rubrum， and to explore the material basis of enhancing the effect of strengthening spleen after processing and the scientific connotation of decoction pieces processed with Halloysitum Rubrum as the auxiliary material. MethodsThe volatile components of the samples before and after processing were identified and relatively quantified by headspace gas chromatography-mass spectrometry（HS-GC-MS）， and the volatile components were analyzed by principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA）. According to the principle of variable importance in the projection（VIP） value>1.5， volatile differential components before and after processing were screened. And combined with intelligent sensory technologies such as colorimeter and electronic nose， the chroma and odor information of CRP before and after being stir-fried with Halloysitum Rubrum were identified. Pearson correlation analysis was used to explore the correlation between volatile differential components and chroma values. ResultsA total of 112 volatile components were identified from CRP and CRP stir-fried with Halloysitum Rubrum， of which 84 were from CRP and 97 were from CRP stir-fried with Halloysitum Rubrum. And 7 differential components were selected， including α-pinene， β-myrcene， linalool， sabinene， ocimene isomer mixture， A-ocimene， and δ-elemene. After being processed with Halloysitum Rubrum， the brightness value（L^*）， yellow-blue value（b^*） and total chromatic value（E^*_ab） of CRP were decreased（P<0.01）， and red-green value（a^*） was increased（P<0.01）， the response values of S4， S5， S10 and S13 sensors were significantly increased（P<0.05）， and the response values of S3 and S8 sensors were significantly decreased（P<0.05）. Correlation analysis showed that α-pinene and β-myrcene were negatively correlated with L^* and E^*_ab， but positively correlated with a^*. Sabinene was positively correlated with L^* and E^*_ab. Linalool was positively correlated with L^* and E^*_ab， and negatively correlated with a^*. The ocimene isomer mixture was positively correlated with the L^*. ConclusionAfter being processed with Halloysitum Rubrum， the appearance color， odor and volatile components of CRP change significantly， and α-pinene， β-myrcene， sabinene， linalool and A-ocimene are the characteristic volatile components before and after processing， which can provide references for the quality evaluation and clinical application of CRP and its processed products.

ObjectiveTo analyze the differences in color， odor and volatile components of Citri Reticulatae Pericarpium（CRP） before and after being stir-fried with Halloysitum Rubrum， and to explore the material basis of enhancing the effect of strengthening spleen after processing and the scientific connotation of decoction pieces processed with Halloysitum Rubrum as the auxiliary material. MethodsThe volatile components of the samples before and after processing were identified and relatively quantified by headspace gas chromatography-mass spectrometry（HS-GC-MS）， and the volatile components were analyzed by principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA）. According to the principle of variable importance in the projection（VIP） value>1.5， volatile differential components before and after processing were screened. And combined with intelligent sensory technologies such as colorimeter and electronic nose， the chroma and odor information of CRP before and after being stir-fried with Halloysitum Rubrum were identified. Pearson correlation analysis was used to explore the correlation between volatile differential components and chroma values. ResultsA total of 112 volatile components were identified from CRP and CRP stir-fried with Halloysitum Rubrum， of which 84 were from CRP and 97 were from CRP stir-fried with Halloysitum Rubrum. And 7 differential components were selected， including α-pinene， β-myrcene， linalool， sabinene， ocimene isomer mixture， A-ocimene， and δ-elemene. After being processed with Halloysitum Rubrum， the brightness value（L^*）， yellow-blue value（b^*） and total chromatic value（E^*_ab） of CRP were decreased（P<0.01）， and red-green value（a^*） was increased（P<0.01）， the response values of S4， S5， S10 and S13 sensors were significantly increased（P<0.05）， and the response values of S3 and S8 sensors were significantly decreased（P<0.05）. Correlation analysis showed that α-pinene and β-myrcene were negatively correlated with L^* and E^*_ab， but positively correlated with a^*. Sabinene was positively correlated with L^* and E^*_ab. Linalool was positively correlated with L^* and E^*_ab， and negatively correlated with a^*. The ocimene isomer mixture was positively correlated with the L^*. ConclusionAfter being processed with Halloysitum Rubrum， the appearance color， odor and volatile components of CRP change significantly， and α-pinene， β-myrcene， sabinene， linalool and A-ocimene are the characteristic volatile components before and after processing， which can provide references for the quality evaluation and clinical application of CRP and its processed products.

ObjectiveTo learn the inner experience of decisions to seek medical attention among patients with ischemic stroke and to explore the factors that affect patients’ early decisions to seek medical attention， providing a basis for formulating intervention measures to reduce patient delays. MethodsUsing qualitative research methods， semi-structured in-depth interviews were conducted with 24 patients with ischemic stroke who had delayed seeking medical attention through purposive sampling， and the data were analyzed by category analysis method. ResultsThe five major themes that affected ischemic stroke patients’ decisions to seek medical attention were symptom experience （insufficient attention to disease and the impact of symptoms on the patients）， symptom assessment （symptom perception， symptom interpretation， and symptom recognition）， symptom response （psychological denial and self-relief）， prior knowledge of the diseases （deficient knowledge and stroke experience）， as well as environmental-social factors （morbidity context， external information support， and medical insurance）. ConclusionUnder the guidance of cognitive， psychosocial， and environmental frameworks， targeted intervention strategies should be proposed to reduce the incidence of delayed decisions to seek medical attention among patients with ischemic stroke.

Objective:To explore the effects of participatory interactive teaching in the clinical internship course of pediatric infectious diseases.Methods:217 undergraduate students of grade 2018 majoring in pediatrics were selected as the experimental group, and 208 undergraduate students of grade 2017 majoring in pediatrics were selected as the control group. The experimental group used the teaching method of BOPPPS combined with case-based learning and team-based learning, while the control group adopted the traditional teaching method. Evaluate the learning effectiveness by comparing the formative evaluation and theoretical exam scores of the two groups , and a questionnaire survey was conducted for the students in the experimental group to investigate teaching feedback. SPSS 19.0 was used to perform the t test for data analysis. Results:The experimental group had significantly better scores than the control group in final theoretical examination [(75.04±9.12) vs. (71.03±9.51), P<0.05] and formative evaluation [(81.03±6.13) vs. (70.02±10.32), P<0.05]. According to the results of the questionnaire survey on teaching satisfaction, the students were satisfied with the interactive participatory teaching method in the four aspects of improving learning interest, improving learning ability, classroom satisfaction level, and course acceptance level. Conclusions:The pragmatic teaching reform in the internship of pediatric infectious diseases is highly accepted by students and beneficial to improving students' clinical thinking ability and comprehensive quality, which is worth further application and promotion.

Objective To identify small molecule inhibitors of APC-mutant colon cancer and provide lead compounds for targeted therapy of colon cancer.Methods APC-mutant colon cancer cell lines that stably express 7*Tcf-GFP/SV40-Cherry(7TGC)dual fluorescence reporter system was constructed for small-molecule inhibitor screening.Cell viability,colony formation,EdU incorporation,and xenograft tumor assay were used to evaluate the inhibitory effect of these inhibitors on APC-mutant colon cancer in vitro and in vivo.Western blot and co-immunoprecipitation assays were used to explore the molecular mechanism.Results Four small molecules that inhibited Wnt activity in APC-mutant colon cancer cells were discovered.Shikonin exhibited significant inhibition of cell viability and proliferation while inducing apoptosis of APC-mutant colon cancer cells.Xenograft tumor assay demonstrated that shikonin significantly reduced tumor growth in vivo.Furthermore,Western blot and co-immunoprecipitation assays revealed that shikonin markedly decreased β-catenin levels.Conclusion Shikonin effectively inhibits Wnt activity and suppresses tumor growth in APC-mutant colon cancer.

Objective:To investigate the recognition of the post competency index system among rural general practice assistant physicians and its influencing factors.Methods:This study was a cross-sectional survey. A questionnaire survey on the recognition of post competency index system was conducted from October 2020 to September 2021 among rural general practice assistant physicians from 10 provinces/municipalities selected by stratified cluster sampling method. The recognition of rural general practice assistant physicians at all levels of indexs and the factors influencing recognition were analyzed.Results:A total of 1 123 questionnaires were distributed and 1 024 valid ones were collected with a recovery rate of 91.18%. Of the 1 024 respondents, 529 were male(51.7%) and 435 were aged 40-49 years(42.5%), which was the highest proportion by age group. The average overall recognition score of the index system was 4.41, and the scores of the primary indexes were 4.32-4.45. Three primary indexes had the highest recognition scores: professional competence, basic health care services, and interpersonal communication and teamwork. The recognition scores on the second level index were 4.18-4.61, and the proportion of recognition scores greater than 4 was over 80%. There were significant differences in the recognition scores of the index system among assistant physicians with different working years, educational background, professional title and work unit ( F/H=6.41, 14.83, 12.45, 7.53, P<0.01). Educational background(associate degree: B=0.091, P=0.015; bachelor degree and above: B=0.196, P<0.001) and professional title(intermediate professional title and above: B=-0.234, P<0.001) were the independent factors influencing the recognition degree of the index system for rural general practice assistant physicians. Conclusions:The post competency index system is generally recognized by rural general practice assistant physician, and academic qualifications and professional title status may influence its recognition.

Objective:To evaluate the application feasibility of the post competency index system of rural general practice assistant physicians, and to analyze and compare the post competency of rural general practice assistant physicians with different characteristics through a survey among rural general practice assistant physicians in China.Methods:This study was a cross-sectional study. A questionnaire survey was conducted among rural general practice assistant physicians in 10 provinces/municipalities selected by stratified cluster sampling method from October 2020 to September 2021. The post competency scores were self-evaluated based on the post competency index system of rural general practice assistant physicians. The results of the survey were analyzed and the application feasibility of the index system was evaluated.Results:A total of 1 123 questionnaires were distributed and 1 024 valid questionnaires were returned with a recovery rate of 91.2%. Of the 1 024 respondents, 529 were males (51.7%), 435 were aged 40-49 years (42.5%), 434 had a secondary school education or less (42.4%), and 531 were junior practitioners (51.9%). The Cronbach′s α coefficient of the overall questionnaire was 0.987, and the Cronbach′s α coefficient of the first level index ranged from 0.897 to 0.974. The cumulative variance contribution rate of exploratory factor analysis was 72.012%. The confirmatory factor analysis showed χ2/ df=3.926, RMSEA=0.076, CFI=0.858, IFI=0.859, indicating that the model fit was basically good. The average self-evaluation scores of the first level index ranged from 3.95 to 4.25, and the average self-evaluation scores of the second level index ranged from 3.74 to 4.36. There were significant differences in self-evaluation scores of post competency among rural general practice assistant physicians with different working years, professional titles, working units and economic regions( F=4.67, 10.54, 22.16 and 20.90,all P<0.05). People with low self-evaluation scores of post competency had the following characteristics: working 10-19 years, intermediate or above titles, working in community health service centers, located in the eastern region.People with high self-evaluation scores of post competency had the following characteristics: primary professional title, working in the village clinic. Conclusion:The competency index system of rural general practice assistant physicians has good application feasibility,and it can be used to evaluate post competency for the education and training of rural general practice assistant physicians.

Objective:Evaluate the occurrence of repeated hospitalizations during the DRG payment reform of a tertiary general hospital and attempt to establish an operational method for evaluating decomposed hospitalization.Methods:The relevant data of inpa-tients in a tertiary public hospital in Beijing from November 1,2020,to August 31,2023,were collected,the occurrence of repeated hospitalization before and after the DRG reform were compared,and the suspicious decomposed hospitalization groups were screened and analyzed.Results:After the DRG reform,the incidence of 14-day,7-day,3-day,and same-day repeated hospitalization increased significantly compared with that before the reform(P＜0.05).Malignancies of the digestive system,a disease with a high readmission rate in the four categories,had a consistency of 6.02％in the ADRG group for two adjacent repeated hospitalizations,and the subsequent admissions of this disease may be assigned to different DRG groups for payment settlement.Conclusion:It is crucial to further enhance the medical insurance payment system,clarify criteria for"decomposed hospitalization",reduce the supervision cost of decomposed hospitalization,and prioritize management strategies targeting avoidable unplanned readmissions.

Objective: To investigate the incidence trend of diabetes among children and adolescents in Ningbo City, Zhejiang Province from 2011 to 2021, so as to provide the basis for the prevention and control of diabetes among children and adolescents. Methods: Data on diabetes incidence among children and adolescents aged 0 to 18 years in Ningbo City were collected through the Diabetes Monitoring Platform of the Ningbo Chronic Disease Collaborative Management System from 2011 to 2021. Crude incidence rates were calculated and standardized using data from the Sixth National Population Census in 2010. The trend of incidence rates were analyzed by average annual percent change (AAPC). Results: A total of 701 cases of diabetes among children and adolescents were reported in Ningbo City from 2011 to 2021. The crude and standardized incidence rates were 6.86/105 and 7.27/105, respectively, showing upward trends (AAPC=5.886%, 7.147%, both P<0.05). The crude and standardized incidence rates of type 1 diabetes mellitus were 3.36/105 and 3.35/105, respectively, with no significant trend observed (AAPC=1.229%, 1.449%, both P>0.05). The crude incidence rate was higher in children and adolescents aged 10 to <15 years (4.56/105) than in other age groups (all P<0.05). The standardized incidence rate was higher in females than in males (3.49/105 vs. 3.04/105, P<0.05). The standardized incidence rate was higher in urban areas than in rural areas (3.60/105 vs. 3.15/105, P<0.05). The crude and standardized incidence rates of type 2 diabetes mellitus were 3.43/105 and 3.87/105, respectively, showing upward trends (AAPC=4.904%, 7.579%, both P<0.05). The crude incidence rate was higher in children and adolescents aged 15 to 18 years (10.53/105) than in other groups (all P<0.05). The crude incidence rates in children and adolescents who aged 10 to <15 years and 15 to 18 years showed upward trends (AAPC=15.030%, 6.637%, both P<0.05). The standardized incidence rate was higher in males than in females (4.01/105 vs. 3.57/105, P<0.05). The standardized incidence rate was higher in urban areas than in rural areas (4.57/105 vs. 3.34/105, P<0.05). Conclusions From 2011 to 2021, the incidence of type 2 diabetes mellitus showed an upward trend, with cases mainly concentrated in children and adolescents aged 15 to 18 years, males, and those living in urban areas. The incidence of type 1 diabetes mellitus remained stable, with cases mainly concentrated in children and adolescents aged 10 to <15 years, females, and those living in urban areas.