Objective: To examine the causal association and potential mechanisms between bone mineral density in different age groups and primary malignant bone tumor based on two sample Mendelian randomization (MR), so as to provide a reference for the prevention and treatment of primary malignant bone tumor. Methods: The genome-wide association study (GWAS) of bone mineral density was obtained from the GEFOS database,which included 66 628 subjects divided into five age groups (0-15, 15-30, 30-45, 45-60, and >60 years) based on the phases of human bone development. The GWAS of primary malignant bone tumor was sourced from the FinnGen database, including 648 cases and 378 749 controls. Using bone mineral density of five age groups as the exposure and primary malignant bone tumor as the outcome, an MR analysis was performed with the inverse-variance weighted (IVW) method. Sensitivity analysis were conducted using Cochran's Q test, MR-Egger regression, MR-PRESSO test and MR Steiger test. The potential mechanisms underlying the causal association between bone density and primary malignant bone tumors were explored using Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Results: The MR analysis results showed that there was a negative causal association between bone density and primary malignant bone tumors in the 30-45 age group (OR=0.301, 95%CI: 0.126-0.721). No statistically significant associations between bone density and primary malignant bone tumors were found in the 0-15, 15-30, 45-60, and >60 age groups (all P>0.05). Sensitivity analysis did not detect heterogeneity, pleiotropy (all P>0.05) and reverse causality. KEGG enrichment analysis revealed that genes highly associated with bone density and primary malignant bone tumors were enriched in the mTOR signaling pathway and the Wnt signaling pathway, among which Low Density lipoprotein Receptor Related protein 5 and Wnt Family Member 16 are key regulatory genes. Conclusion The decrease in bone mineral density among individuals aged 30-45 may increase the risk of primary malignant bone tumors through the mTOR signaling pathway and the Wnt signaling pathway.

Objective:To study effects of different pretreatment methods on the detection of pesticide residues in Angelicae Sinensis Radix,ChrysanthemI Flos,Lych Fructus,Astragali Radix and Lonicerae Japonicae Flos.Methods:The samples were treated with QuEChERS method and high-speed homogenization combined with hydro-phile-lipophile balance(HLB)solid-phase extraction method,and the residual amounts of carbofuran,3-carboxyl-carbofuran,phorate,phorate sulfone,phorate sulfoxide,and methyl isoflurophos were simultaneously determined using UHPLC-MS/MS.With a gradient elutionof 0.1％formic acid solution containing 5 mmol·L-1 ammonium formate and t 0.1％formic acid solution containing 5 mmol·L-1 ammonium formate-acetonitrile(volume ratio of 5:95),Ultra-performance liquid chromatography column Agilent Poroshell 120 SB-C18(2.1 mm × 100 mm,2.7μm)was used at 35 ℃ and the electrospray ion source was scanned in the positive ion mode and detected in the multiple reaction monitoring mode in mass spectrometry.Results:The deviation of the results measured by QuECh-ERS method and HLB solid-phase extraction method was between 9.09％-55.56％.Conclusion:In the selection of the pretreatment method for the detection of pesticide residues in traditional Chinese medicine,it is recommen-ded to take the measurement data of positive samples as the evaluation index and basis,and choose the method with higher measurement value and high extraction efficiency.

Objective:To explore the training needs for clinical core competence of master of nursing specialist (MNS) from the perspective of supervisors, providing reference for the development of future MNS clinical practice training programs.Methods:Using phenomenological research methods from qualitative research, purposive sampling was used to select 10 MNS supervisors from Jilin Province, Heilongjiang Province, Sichuan Province, and Zhejiang Province as research subjects for semi-structured interviews from May to July 2023. Colaizzi 7-step analysis method was used to extract themes.Results:Six themes were extracted, including the need to strengthen MNS ideological and political education, differences in clinical training needs and ability goals between fresh and non-fresh students, the need to enhance MNS clinical practice ability, clinical research should be a key training content, thinking ability training should be integrated throughout the entire clinical training process, and achievement transformation.Conclusions:Relevant training institutions should attach importance to the cultivation of MNS ideological and political education, specialized practical abilities, thinking abilities, clinical research, and achievement transformation abilities, distinguish the tendency of cultivating fresh and non-fresh students, and actively set up relevant courses to improve students' core competence and job competitiveness, and cultivate nursing expert talents that truly meet the needs of clinical development.

Objective:To develop a joint clinical practice teaching and assessment program tailored to the clinical training needs of master of nursing specialist (MNS) postgraduates which focuses on core competency requirements.Methods:Totally 10 MNS postgraduate supervisors were selected by convenience sampling for semi-structured interviews between May and July 2023. Subsequently, a Delphi method was employed with 22 MNS postgraduate supervisors over two rounds of consultations from October to December 2023.Results:A total of 22 experts participated in the Delphi consultations, with an effective response rate of 100.00% (22/22) in both rounds. The expert authority coefficients were 0.822 and 0.833, respectively, for the two rounds. The Kendall's W for various levels of indicators ranged from 0.097 to 0.243 and 0.159 to 0.256, respectively ( P<0.01). The final training program included five primary indicators, 10 secondary indicators, and 26 tertiary indicators. Conclusions:The development process for the joint clinical practice teaching and assessment program for MNS postgraduates is scientific and reliable. The program can serve as a reference for the clinical practice training of MNS postgraduates.

Objective To explore the effect of 12-week simplified Tai Chi training on rehabilitation and neuromuscular control in individuals with chronic ankle instability(CAI).Methods Thirty-four partic-ipants with CAI were randomly divided into an experimental group and a control group,each of 17.The control group received 12-week health education,while the experimental group underwent simpli-fied Tai Chi training for the same length.Before and after the intervention,both groups were evaluat-ed their self-reported instability feeling,ankle muscle strength,proprioception and dynamic postural stability.Results After intervention,a significant increase was observed in the average Cumberland An-kle Instability Tool(CAIT),the maximum extension distance in the posteromedial and posterolateral di-rections of the modified Star Excursion Balance Test(mSEBT)in,as well as the ankle peak torque in plantarflexion,inversion and eversion(P<0.05)in the experimental.Moreover,the ankle passive motion proprioceptive threshold in plantarflexion and inversion decreased significantly in the experimental group after intervention(P<0.05).Meanwhile,after intervention,the average CAIT score,maximum ex-tension distances in the posteromedial and posterolateral directions in the mSEBT,and the ankle peak torques in plantarflexion,inversion and eversion of the experimental group were significantly higher than the control group(P<0.05),while the ankle passive motion proprioceptive threshold in plantarflex-ion was significantly lower than the latter(P<0.05).Conclusion Twelve-week simplified Tai Chi train-ing can improve the clinical subjective instability of CAI participants,maybe related to the improve-ment of ankle muscle strength,proprioception and dynamic postural stability.Therefore,it is suggest-ed that simplified Tai Chi should be one of the rehabilitation methods for CAI patients.

Acylcarnitines are metabolic intermediates of fatty acids and branched-chain amino acids having vital biofunctions and pathophysiological significances.Here,we developed a high-throughput method for quantifying hundreds of acylcarnitines in one run using ultrahigh performance liquid chromatography and tandem mass spectrometry(UPLC-MS/MS).This enabled simultaneous quantification of 1136 acyl-carnitines(C0-C26)within 10-min with good sensitivity(limit of detection＜0.7 fmol),linearity(cor-relation coefficient＞0.992),accuracy(relative error＜20％),precision(coefficient of variation(CV),CV＜15％),stability(CV＜15％),and inter-technician consistency(CV＜20％,n=6).We also established a quantitative structure-retention relationship(goodness of fit＞0.998)for predicting retention time(tR)of acylcarnitines with no standards and built a database of their multiple reaction monitoring parameters(tR,ion-pairs,and collision energy).Furthermore,we quantified 514 acylcarnitines in human plasma and urine,mouse kidney,liver,heart,lung,and muscle.This provides a rapid method for quantifying acyl-carnitines in multiple biological matrices.

[Objective] To explore the expression of Nectin-4 in invasive bladder urothelial carcinoma (BUC) tissue and its clinical significance, so as to provide reference for clinical diagnosis and treatment of BUC. [Methods] Nectin-4 expression in 60 cases of invasive BUC and 40 cases of chronic inflammation of bladder mucosa was detected with immunohistochemical staining (IHC) and RNAscope.The results of the two methods were analyzed and compared, and the relationship between the two methods and the clinicopathological characteristics of invasive BUC was discussed.The correlation between the protein expression of Nectin-4 in BUC tissues, human epidermal growth factor receptor 2 (Her-2) and programmed death factor ligand 1 (PD-L1) was analyzed. [Results] The positive protein expression rates of Nectin-4 detected by IHC were 78.33%(47/60) and 17.50% (7/40) in the invasive BUC group and inflammatory group, respectively, while the positive mRNA expression rates of Nectin-4 detected by RNAscope were 83.33% (50/60) and 12.50% (5/40), respectively.The Kappa values of Nectin-4 in the invasive BUC group and inflammatory group were 0.732 and 0.610, respectively, with general consistency.The protein expression of Nectin-4 in invasive BUC was correlated with muscular invasion, histological grade, vascular thrombus, lymph node metastasis and clinical stage (P<0.05). The mRNA expression of Nectin-4 in invasive BUC was correlated with max tumor diameter, muscular invasion, histological grade, vascular thrombus, lymph node metastasis and clinical stage (P<0.05). The high expression of Nectin-4 in invasive BUC was positively correlated with the expression of Her-2 (P=0.002), but not with the expression of PD-L1 (P>0.05). [Conclusion] Nectin-4 is highly expressed in invasive BUC, and is usually associated with the pathological parameters of poor prognosis.Detection of Nectin-4 expression will help to guide clinical diagnosis and treatment.

Objective To construct myeloid-specific Spi1 gene knockout mice and analyze their genotypes,so as to provide animal model basis for the study of pathological mechanism of diseases and drug targets.Methods Ac-cording to the principle of CRISPR/Cas9 technology and Cre/LoxP system,sgRNA and Donor vectors were de-signed and constructed.The transcript of Exon 2(Exon 2)was used as the knockout region,and Loxp elements were placed on both sides of Exon 2.Cas9 protein,sgRNA and Donor vector were mixed and microinjected into the fertilized eggs of C57BL/6J mice,the fertilized eggs were transplanted into the uterus of C57BL/6J pregnant female mice,and F0 generation was obtained after 19～20 days.Positive F0 mice were mated with C57BL/6J mice to ob-tain stable F1 Spi1flox/+mice.Spi1flox/+mice of F1 generation were selfed to obtain Spi1flox/flox mice.Spi1flox/flox mated with Lyz2-Cre+mice to obtain Spi1flox/+/Lyz2-Cre+mice,and then mated with Spi1flox/flox,the Spi1flox/flox/Lyz2-Cre+mice were myeloid-specific Spi1 gene knockout(KO)mice.Spi1flox/flox/Lyz2-cre-mice were used as wild-type(WT)mice.DNA of WT and KO mice was extracted,and the genotypes were identified by agarose gel electro-phoresis after PCR amplification.Western blot was used to detect the expression of spleen focus forming virus provi-ral integration oncogene,Spi-1/purine rich box-1(PU.1)in immune cells of WT and KO mice.Results The results of PCR identification showed that the genotype of mice with only 220 bp amplified by flox primer was Spi1flox/flox homozygote,and the genotype of mice with 700 bp amplified by Lyz2-Cre primer was Lyz2-Cre+.Western blot showed that compared with WT group,the protein PU.1 was not expressed in bone marrow-derived macropha-ges(BMDMs)and peritoneal macrophages(PM)in KO group(P<0.01).There was no significant difference of statistics in the expression level of PU.1 in T cells between KO mice and WT mice.The results of PCR and West-ern blot showed that myeloid-specific Spi1 KO mice were successfully constructed.Conclusion The myeloid-spe-cific Spi1 gene KO mice are successfully constructed and identified,which provides animal model basis for further revealing the potential mechanism of PU.1 inimmune regulation.

Objective To breed and identify the T lymphocyte-conditional Spi1 knockout mice for the further in-vestgation of the specific role of Spi1-encoded protein PU.1.Methods The Lck-Cre mice were mated with Spi1flox/flox mice to obtain Lck-Cre×Spi1flox/flox mice(T lymphocyte-specific Spi1 knockout mice),and the genotype was determined by polymerase chain reaction(PCR)and agarose gel electrophoresis.Magnetic beads were used to sort out the splenic T lymphocytes,and the knockdown efficiency of PU.1 in T cells was detected by Western blot,quantitative real-time PCR(qPCR)and flow cytometry.Results The Lck-Cre×Spi1flox/flox mouse genotype was stably inherited.Compared with Spi1flox/flox mice,the expression level of PU.1 was significantly reduced in splenic T cells of Lck-Cre×Spi1flox/flox mice.Conclusion In this study,the T lymphocyte-specific Spi1 knockout mice was successfully constructed by applying Cre/LoxP system and CRISPR/Cas9 technology,which provided a reliable an-imal model for the subsequent experiments of the specific role of PU.1 in T cell-related diseases.

Objective To construct Csf1r-CreERT2 R26REYFP reporter gene mice and assess the efficacy of Csf1r-CreERT2-mediated enhancement of CSF1R in CD45+cells labeled with yellow fluorescein protein EYFP.Methods Csf1r-CreERT2 mice were crossbred with R26REYFP homozygous mice,and Csf1r-CreERT2R26REYFP mice were identified through PCR and Western Blot analyses.Flow cytometry was employed to evaluate CSF1R tag-efficiency in CD45+cells across different mouse tissues following tamoxifen induction.Results Csf1r-CreERT2 R26REYFP reporter gene mice were acquired.In addition,it was found that Csf1r-CreERT2-mediated EYFP could effectively mark CSF1R in various tissues of mice and CD45+cells in different locations.Compared to the R26REYF P group,the highest labeling efficiency was observed in the brain tissue(P＜0.001),the lowest in the thymus tissue(P＜0.05),and no sig-nificant difference was observed in the spleen tissue.Conclusion Adult Csf1r-CreERT2 mice and R26REYFP mice are effective ways to obtain Csf1r-CreERT2 R26REYFP induced conditional fluorescence mice.Csf1r-CreERT2 can mediate EYFP to effectively trace CSF1R in CD45+cells in different parts of mice.