Phenylpyruvic acid (PPA) is used as a food and feed additive and has a wide range of applications in the pharmaceutical, chemical and other fields. At present, PPA is mainly produced by chemical synthesis. With the green transformation of the manufacturing industry, biotransformation will be a good alternative for PPA production. The L-amino acid deaminase (PmiLAAD) from Proteus mirabilis has been widely studied for the production of PPA. However, the low yield limits its industrial production. To further enhance the production of PPA and better meet industrial demands, a more efficient synthesis method for PPA was established. In this study, PmiLAAD was heterologously expressed in Escherichia coli. Subsequently, a colorimetric reaction method was established to screen the strains with high PPA production. The semi-rational design of PmiLAAD was carried out, and the obtained triple-site mutant V18 (V437I/S93C/E417A) showed a 35% increase in catalytic activity compared with the wild type. Meanwhile, the effect of N-terminal truncation on the catalytic activity of the V18 mutant was investigated. After the optimization of the whole-cell conditions for the obtained mutant V18-N7, fed-batch conversion was carried out in a 5-L fermenter, and 44.13 g/L of PPA was synthesized with a conversion rate of 88%, which showed certain potential for industrial application. This study lays foundation for the industrial production of phenylpyruvic acid and also offers insights into the biosynthesis of other chemicals.
Escherichia coli/metabolism* ; Proteus mirabilis/genetics* ; Phenylpyruvic Acids/metabolism* ; Protein Engineering/methods* ; Recombinant Proteins/biosynthesis* ; Bacterial Proteins/metabolism*

Objective:To explore the challenges in the implementation of drug centralized volume-based procurement policies in hospitals and propose corresponding optimization suggestions.Methods:From August to December 2023, a purposive sampling was conducted to select 11 pharmaceutical experts from tertiary hospitals in China for Delphi method. The survey content included " policy recommendations for promoting the acceleration and expansion of national drug centralized procurement and retaining surplus medical insurance funds for centralized procurement" .Results:Survey participants gave feedback on a set of existing problems found in the implementation of drug centralized procurement policies and proposed corresponding optimization methods. Kendall′s W coefficient of the specialist consultation was 0.332( P<0.05), demonstrating good consistency and concentration of the expert opinions. Among the problems, the score of drug supply guarantee was the highest(mean value of importance was 4.45). At the same time, the recommendation of strengthening monitoring and early warning, coordination and dispatch, and effectively ensuring the supply of centralized drug procurement had the highest score and concentration(mean value of importance was 4.91, coefficient of variation was 0.06). Conclusions:Through Delphi method, this study revealed issues and optimization methods in the implementation of drug centralized procurement policies in hospitals. The findings could provide valuable insights for improvements in the pharmaceutical sector and future policy adjustments.

Objective:To summarize the construction and practice of the compulsory postgraduate course "reproductive genetics in gynecology and obstetrics" in Department of Gynecology and Obstetrics, Peking University, and to evaluate the teaching effect of this course.Methods:A total of 139 postgraduates who studied in Department of Obstetrics and Gynecology, Peking University, from 2019 to 2021 were enrolled as subjects, and a syllabus was constructed through a top-level design based on the "biological-psychological-social medicine pattern", with the teaching objectives of reproductive genetics theory, clinical translation, genetic counseling methods, and research advances. The teaching effect was evaluated by analyzing teaching assessment results, teaching evaluation feedback, and teaching achievements. SPSS 26.0 software was used to perform the t-test and the chi-square test. Results:The written test score, usual performance score, and total score of the postgraduate students in 2021 were higher than those in 2019 and 2020 [(73.50±8.19) vs. (70.94±14.90); (68.60±2.82) vs. (68.22±4.58); (90.58±4.18) vs. (89.49±7.60)], with significant differences in written test score and total score ( P<0.05). There was a high degree of satisfaction with the feedback of teaching, and 85.61% (119/139) of the students selected "great satisfaction"; in particular, there were increases in the degree of satisfaction with expanding research ideas and reflecting the advances in this discipline, but with no significant difference. There was a significant increase in the number of published articles. Conclusion:For the active implementation of the course of "Reproductive Genetics in Obstetrics and Gynecology", improving the teaching process in a planned and step-by-step way through a top-level design in advance can help to expand research ideas for future research work among postgraduates in obstetrics and gynecology and promote the sustainable development and improvement of the teaching of the emerging interdisciplinary discipline of reproductive genetics in obstetrics and gynecology.

【Objective】 To explore the correlation between red blood cell lifespan and adhesion molecules on the surface of red blood cell membrane, in order to establish a method to detect the duration of red blood cell storage. 【Methods】 10 samples(10 mL each) of fresh red blood cell, collectedf rom 10 healthy voluntary blood donors, were divided into 5 age groups (layers) by Percoll density gradient centrifugation. The expression of CD47, CD44 and CD147 on the surface of red blood cell membrane in each layer was detected using flow cytometry. The variance of protein expression in each layer of red blood cells was analyzed by SPSS statistical software. 【Results】 The expression levels (%) of 3 adhesion molecules on the surface of red blood cell membranes from young to old were CD47: 14.44±2.61, 9.30±1.75, 7.84±1.49, 6.54±1.32 and 5.53±1.12 (P<0.01); CD44: 25.01±1.94, 19.22±1.52, 17.10±1.28, 15.18±1.11 and 13.56±1.08 (P<0.01); CD147: 33.46±1.99, 28.31±2.95, 23.83±1.59, 20.40±1.56 and 18.03±1.65 (P<0.01). 【Conclusion】 The expression levels of CD47, CD44 and CD147 on the surface of red blood cell membranes have showed a downward trend as the storage extended. These three protein adhesion molecules have showed a correlation with red blood cells lifespan, and could be used as detection markers of cell age.

As a pattern recognition receptor, CD36 antigen participates in a series of pathophysiological processes, and has well been documented in transfusion medicine. This article reviews the discovery, structure and expression of CD36, the type and frequency of CD36 antigen deletion, as well as the relationship between anti-CD36 and transfusion-related immune diseases.

Objective:To stablish a transitional nursing model for patients with chronic diseases under the hierarchical diagnosis and treatment system and clarify the structure and functions, so as to provide a chronic disease management model with practical operation, promotion and reference.Methods:In April 2019, a research group was established. Based on a large amount of literature and policy analysis, guided by the three-dimensional quality theory model of structure-process-results, the transitional nursing model was constructed and the index system letter consultation questionnaire was initially prepared. After the preliminary survey, it was revised to form the first-round consultation questionnaire. Delphi method was used to conduct 2 rounds of letter consultation among 19 experts. The positive coefficient of experts, the authoritative coefficient of experts and Kendall 's coordination coefficient were calculated. Analytic hierarchy process (AHP) was used to determine the weight coefficient of each index and quantify the importance degree of the index. Results:The effective recovery rates of the two rounds of letter consultation questionnaires of expert were both 100%. The authoritative coefficients of experts were respectively 0.80 and 0.84, Kendall 's coordination coefficient was 0.137 to 0.253 ( P<0.001) , and the coefficients of variation were all less than 0.25. The index system for the transitional nursing model of patients with chronic diseases finally established included 6 first-level indicators, 38 second-level indicators and 32 third-level indicators of "transitional nursing procedures", and the structural and functional diagram was generated. Conclusions:The established transitional nursing model for patients with chronic diseases is scientifically reliable and comprehensive in content, which has practical significance for promoting the deep integration of mobile Internet and medical and health fields, guiding the subsidence of advantageous resources and promoting the implementation of hierarchical diagnosis and treatment.

Objective: To explore the effect of heme oxygenase-1 (HO-1) on level of reactive oxygen species (ROS) induced by zinc oxide nanoparticles (ZnO-NPs) in Human umbilical vein endothelial cells line EA.hy926. Methods: The EA.hy926 cells in logarithmic growth phase were incubated with 0.0, 2.5, 5.0, 10.0 and 15.0 mg/L ZnO-NPs respectively. The ROS level, reflected by mean fluorescence intensity (MFI), was examined by flow cytometer after 4 hours exposure, the protein expression of HO-1 which was determined by Western Blot after exposed to ZnO-NPs for 24 hours. Cells incubated with 15.0 mg/L were set as the ZnO-NPs group; a blank control group was set at the same time. Cells were pretreated with HO-1 inhibitor zinc protoporphyrin (ZnPPIx) and HO-1 activator cobalt protoporphyrin (CoPPIx), they were classified as ZnPPIx group and CoPPIx group. 15 mg/L ZnO-NPs was chosen to conduct the experiment of HO-1 activation and inhibition. Cells were classified as ZnPPIX+ ZnO-NPs group and CoPPIx+ ZnO-NPs group after pretreated with 10 μmol/L ZnPPIx or CoPPIx for 1 h, added 15 mg/L ZnO-NPs to cell culture medium. In all groups ROS levels were detected after exposed to ZnO-NPs for 4 hours, the protein expression of HO-1 was detected after exposed to ZnO-NPs for 24 hours. Results: With the increased dose of ZnO-NPs, levels of ROS and HO-1 in EA.hy926 cells were clearly elevated (the MFI of 0.0, 2.5, 5.0, 10.0 and 15.0 mg/L ZnO-NPs incubated groups was 22 627.22±718.27, 24 726.47±568.52, 31 141.75±1 312.24, 39 824.82±4 774.74, 50 569.03±1 497.63 respectively, and HO-1 relative expression were 0.16±0.01, 0.19±0.02, 0.16±0.01, 0.23±0.02, 0.92±0.06 respectively). HO-1 expression in ZnPPIx pretreatment group decreased compared with ZnO-NPs group (1.05±0.05 vs. 1.12±0.01, P<0.05), meanwhile ROS level enhanced (62 683.95±2 589.59 vs. 53 654.53±2 229.01, P<0.05). However, CoPPIx pretreatment had higher HO-1 level and lower level of ROS compared with ZnO-NPs group (HO-1: 1.74±0.11 vs. 0.22±0.03, P<0.05; ROS: 32 845.04±993.48 vs. 53 654.53±2 229.01, P<0.05). Conclusions Exposure to ZnO-NPs significantly induced ROS generation in EA.hy926 cells in a dose-dependent manner. HO-1 regulated ZnO-NPs-induced oxidative stress.

A 2-month-and-20-day-old girl was hospitalized because of slow weight gain for 20 days.The patient was diagnosed as anemia,pneumonia and neutrophils firstly,and showed no improvement during anti-infective therapy,blood transfusion,and leukopenia drugs.The urinary organic acid spectrum of the patient showed increased methylmalonic acid slightly,but the normal value was found by 4 tests,which was inconsistent with the typical methylmalonic aciduria.By analyzing clinical manifestations and plasma homocysteine,folic acid and vitamin B12 levels,she was considered to have intracellular cobalamin metabolic disorder.Then,normal diet,intramuscular injection of vitamin B12,and feeding calcium,betaine,and L-carnitine were given,and the girl's symptoms were improved significantly.Latter gene analysis further showed that the patient had methylmalonic aciduria CblC type.Clinical manifestations of methylmalonic aciduria are complex and individually various,different therapies showed be applied to different causes.The value of urinary methylmalonic acid level alone cannot judge etiology;diagnosis should be based on blood amino acids and carnitine spectrum,homocysteine level and vitamin B12 level.