Objective:To develop a novel ovarian biological age prediction model based on DNA methylation for ovarian aging assessment.Methods:A prospective cohort study method was used. From March 2024 to January 2025, we collected 96 ovarian granulosa cell samples of infertility patients due to fallopian tube factors or male factors from the Reproductive Medicine Center of the Second Affiliated Hospital of Naval Medical University. Then we analyzed DNA methylation patterns across five age-associated gene regions ( ELOVL2, miR29B2C, TRIM59, KLF14 and FHL2) in a discovery cohort comprising 63 human ovarian granulosa cell samples. Targeted bisulfite sequencing was performed through PCR amplification followed by next-generation DNA sequencing. Leveraging elastic net regression analysis, we developed a predictive model incorporating 29 methylation sites that demonstrated strong age correlation. The model was subsequently validated using an independent cohort comprising 33 human ovarian granulosa cell samples. Results:The DNA methylation age prediction model based on ovarian granulosa cells showed the following results in the discovery cohort as follows: median absolute error (MAE) was 2.534 ( R=0.742, P<0.001). In the independent validation cohort, MAE was 3.019 ( R=0.729, P<0.001). Conclusion:In this study, we utilized human ovarian granulosa cells as experimental samples to develop a novel DNA methylation-based model for predicting ovarian biological age. By integrating multiple methylation sites across five age-related gene regions, this model serves as a robust indicator of ovarian aging status.

OBJECTIVE To investigate the distribution,drug resistance and molecular characteristics of the Staph-ylococcus aureus strains isolated from intensive care unit(ICU)environment in some hospitals of Shanghai Pudong New Area so as to provide bases for prevention and control of infections.METHODS The specimens were collected from ICU environment of 8 medical institutions in Pudong District in 2019 and 2023,the S.aureus strains were i-solated and identified.The drug susceptibility testing was performed by microbroth dilution method.The drug re-sistance genes and virulence genes were predicted based on virulence factors of bacterial pathogens,the compre-hensive antibiotic resistance database(mcmaster.ca)and PubMLST,multilocus sequence typing(MLST)and phylogenetic analysis were performed.RESULTS Totally 3408 samples were collected,and the isolated S.aureus strains accounted for 1.29％(44/3408);the isolated strains form both the shared articles and the articles that health care workers directly contact accounted for 20.45％.The drug resistance rate to penicillin was highest(72.73％,32/44);the strains were completely susceptible to vancomycin,rifampicin,sulfamethoxazole-trime-thoprim,daptomycin and nitrofurantoin.The methicillin-resistant Staphylococcus aureus(MRSA)strains accoun-ted for 25.00％.The multi-drug resistance rate was 40.91％.The strains carried 26 types of drug resistance genes in total,covering 9 categories;the carrying rate of fluroquinolones resistance gene nor A was 100.00％.The strains carried 12 types of enterotoxin genes,the carrying rate of sea gene was highest(50.00％,22/44),some of the strains carried lukS-PV,lukF-PV and tsst gene.There were 14 types of sequence type(ST),dominated by ST7(13/44,29.55％),and novel ST type ST9342 was discovered.The phylogenetic analysis showed that all of the strains were divided into 2 major branches,and the strains with the same ST type were phylogenetically close on the evolutionary tree.CONCLUSIONS The S.aureus strains are prevalent in ICU environment and are highly re-sistant to antibiotics.The strains exhibit various ST types,which,to a certain degree,is associated with the path-ogenicity.

Objective To investigate the association of spleen volume with the risk of non-alcoholic fatty liver disease(NAFLD)as well as their causal relationship.Methods We included 90 NAFLD cases and 47 healthy controls who had received contrast-enhanced computed tomography(CT)scan of the abdomen at the Second Affiliated Hospital of Xi'an Jiaotong University from November 2022 to November 2023.We conducted three-dimensional reconstruction of the spleen through a deep learning network model using a two-stage coarse-to-fine segmentation approach.We compared the two groups using the two-sample t test or Mann-Whitney U test for continuous data and using the chi-square test for categorical data;evaluated the correlation between spleen volume and liver function indicators through Pearson correlation or Spearman rank correlation analyses;determined the factors influencing the development of NAFLD through multivariable Logistic regression analysis;and further assessed the casual relationship between spleen volume and NAFLD using the inverse variance-weighted two-sample Mendelian randomization(IVW-MR)method.Results Spleen volume was significantly larger in NAFLD cases than in controls(272.93±104.16 vs 204.37±81.20 cm3,P<0.001).The Spearman rank correlation analysis showed that spleen volume was positively correlated with the hepatic steatosis index(rs=0.422,P<0.001)and gamma-glutamyl transferase levels(rs=0.211,P=0.047)in patients with NAFLD.The multivariable Logistic regression analysis indicated that spleen volume was an independent risk factor for the development of NAFLD(odds ratio[OR]=1.01,95%confidence interval[CI]:1.00-1.02,P=0.049).The IVW-MR analysis detected a causal relationship between spleen volume and NAFLD(OR=1.16,95%CI:1.05-1.28,P=0.005).Conclusion Increased spleen volume may be a risk factor for the development and progression of NAFLD.Further studies are still needed to investigate the specific mechanism.

OBJECTIVE To establish a preparation system for megakaryocytes(MKs)derived from human embryonic stem cells(hESCs)and MK microparticles(MKMPs),and to assess the pro-angio-genic efficiency of these microparticles.METHODS ①hESCs were induced to mesodermal progenitor cells via monolayer culture with the first-stage induction medium for 2 days before the cells were induced to hemogenic endothelial/hematopoietic progenitor cells by culturing with the second-stage induction medium for another 3 days.Then,the cells were dissociated into single cells,seeded into the third-stage induction medium,and cultured using the suspension method for 8 days to obtain MKs.The specific characters of differentiated cells were identified through morphological observation and flow cytometry before stage-specific marker proteins in different periods were analyzed[hESCs:TRA-1-60,sialyl glycolipid stage-specific embryonic antigen4(SSEA4)];mesodermal progenitor cells:brachyury;hemogenic endothelial/hematopoietic progenitor cells:CD34,CD43;MKs:CD41a,CD42b),and immu-nofluorescence staining[β1-tubulin,von Willebrand factor(VWF)],[friend leukemia integration 1(FLI1),CD42].② MKMP collection and verification:MKMPs were collected via differential centrifugation.The concentration and size of these MKMPs were determined by nanoparticle tracking analysis(NTA),and both the morphology and ultrastructure were examined by transmission electron microscopy(TEM).Besides,the MKMPs-specific proteins[CD41,tumor susceptibility gene 101(TSG101)and CD9]were detected by Western blotting analysis.③ Biological function of MKMPs:MKMPs were stained with CD41a-PE antibodies and co-cultured with human umbilical veinvascular endothelial cells(HUVECs)labeled by CD34-APC for 3 h.Live-cell immunofluorescence was employed to find out whether HUVECs could absorb MKMPs.To find out whether MKMPs could affect the role of HUVECs in angio-genesis and cell migration,platelet microvesicles(PMPs)were used as positive controls.The experi-mental groups were added with different concentrations of microparticles(1,5,10 and 20 mg·L-1)while the control group was given no microparticles(0 mg·L-1).The number of nodes that formed the lumen after 5 h of incubation in Matrigel was counted,and the size of healing of the scratch area was analyzed after 6 h.To elucidate the mechanism through which MKMPs impacted angiogenesis,ELISA was used out to quantitatively detect the concentration of proteins in microparticles.RESULTS ① A three-stage differentiation cultural system was established to develop hESCs into MKs.Flow cytometry revealed progressive loss of pluripotency markers SSEA4 and TRA-1-60,while the mesodermal progenitor marker brachyury peaked at d 2.Subsequently,hemogenic endothelial/hematopoietic progenitor markers CD34 and CD43 emerged at d 5,followed by megakaryocytic markers CD41a and CD42b at d 13.Immunofluorescent images further demonstrated that MKs expressed specific proteins CD42,β1-tubulin,von VWF and FLI1 at d 13.②Microparticles were collected via differential centrifuga-tion.Transmission electron microscopy revealed that their substructure exhibited a typical double-layered membrane.Nanoparticle tracking analysis indicated that the size was(164.3±14.0)nm.The result of WB demonstrated that the microparticles expressed specific markers,including TSG101,CD9 and CD41.③ MKMPs were absorbed after being co-cultured with HUVECs for 3 h and enhanced the ability of HUVECs to form tubes and migrate.Notably,the treatment of 5 mg·L-1 MKMPs was more effective than 5 mg·L-1 PMPs treatment.The results of ELISA showed that the content of VEGF from MKMPs was higher than from PMPs,which may be the key factor in regulating endothelial biological function.CONCLUSION MKs derived from hESCs can generate functional microparticles which can promote angiogenesis.

Objective:To develop a novel ovarian biological age prediction model based on DNA methylation for ovarian aging assessment.Methods:A prospective cohort study method was used. From March 2024 to January 2025, we collected 96 ovarian granulosa cell samples of infertility patients due to fallopian tube factors or male factors from the Reproductive Medicine Center of the Second Affiliated Hospital of Naval Medical University. Then we analyzed DNA methylation patterns across five age-associated gene regions ( ELOVL2, miR29B2C, TRIM59, KLF14 and FHL2) in a discovery cohort comprising 63 human ovarian granulosa cell samples. Targeted bisulfite sequencing was performed through PCR amplification followed by next-generation DNA sequencing. Leveraging elastic net regression analysis, we developed a predictive model incorporating 29 methylation sites that demonstrated strong age correlation. The model was subsequently validated using an independent cohort comprising 33 human ovarian granulosa cell samples. Results:The DNA methylation age prediction model based on ovarian granulosa cells showed the following results in the discovery cohort as follows: median absolute error (MAE) was 2.534 ( R=0.742, P<0.001). In the independent validation cohort, MAE was 3.019 ( R=0.729, P<0.001). Conclusion:In this study, we utilized human ovarian granulosa cells as experimental samples to develop a novel DNA methylation-based model for predicting ovarian biological age. By integrating multiple methylation sites across five age-related gene regions, this model serves as a robust indicator of ovarian aging status.

Objective To explore the efficacy of a model combining clinicopathological characteristics and multiparametric MRI features for predicting BRCA-mutated breast cancer(BC).Methods A total of 256 BC patients were retrospectively selected and divided into BRCA mutation group(116 cases)and BRCA wild group(140 cases)based on the BRCA results.Chi-square tests or independ-ent sample t-tests were used to compare the differences in clinicopathological characteristics and multiparametric MRI features between the BRCA mutation group and the wild group.Risk factors for BRCA-mutated BC were identified through univariate and multivariate logistic regression ananlyses,and a combined predictive model was constructed.Receiver operating characteristic(ROC)curve was used to ana-lyze the diagnostic efficacy of the model.Results There were statistically significant differences in T stage,human epidermal growth factor receptor 2(HER-2),Ki-67,non-mass enhancement,enhancement pattern,time-signal intensity curve(TIC)type,and apparent diffusion coefficient(ADC)values between the BRCA mutation group and the wild group.Univariate logistic regression analysis showed that T stage,HER-2,Ki-67,non-mass enhancement,enhancement pattern,TIC type,and ADC values were risk factors for BRCA-mutated BC(P<0.05).Multivariate logistic regression analysis revealed that T stage,HER-2,Ki-67,enhancement pattern,and TIC type were independent risk factors for BRCA-mutated BC(P<0.05).The combined model incorporating T stage,HER-2,Ki-67,enhancement pattern,and TIC type had the best diagnostic efficacy in predicting BRCA-mutated BC,with an area under the curve(AUC)of 0.751.Conclusion The combined model integrating T stage,HER-2,Ki-67,enhancement pattern,and TIC type has good efficacy in predicting BRCA-mutated BC.

OBJECTIVE To establish a preparation system for megakaryocytes(MKs)derived from human embryonic stem cells(hESCs)and MK microparticles(MKMPs),and to assess the pro-angio-genic efficiency of these microparticles.METHODS ①hESCs were induced to mesodermal progenitor cells via monolayer culture with the first-stage induction medium for 2 days before the cells were induced to hemogenic endothelial/hematopoietic progenitor cells by culturing with the second-stage induction medium for another 3 days.Then,the cells were dissociated into single cells,seeded into the third-stage induction medium,and cultured using the suspension method for 8 days to obtain MKs.The specific characters of differentiated cells were identified through morphological observation and flow cytometry before stage-specific marker proteins in different periods were analyzed[hESCs:TRA-1-60,sialyl glycolipid stage-specific embryonic antigen4(SSEA4)];mesodermal progenitor cells:brachyury;hemogenic endothelial/hematopoietic progenitor cells:CD34,CD43;MKs:CD41a,CD42b),and immu-nofluorescence staining[β1-tubulin,von Willebrand factor(VWF)],[friend leukemia integration 1(FLI1),CD42].② MKMP collection and verification:MKMPs were collected via differential centrifugation.The concentration and size of these MKMPs were determined by nanoparticle tracking analysis(NTA),and both the morphology and ultrastructure were examined by transmission electron microscopy(TEM).Besides,the MKMPs-specific proteins[CD41,tumor susceptibility gene 101(TSG101)and CD9]were detected by Western blotting analysis.③ Biological function of MKMPs:MKMPs were stained with CD41a-PE antibodies and co-cultured with human umbilical veinvascular endothelial cells(HUVECs)labeled by CD34-APC for 3 h.Live-cell immunofluorescence was employed to find out whether HUVECs could absorb MKMPs.To find out whether MKMPs could affect the role of HUVECs in angio-genesis and cell migration,platelet microvesicles(PMPs)were used as positive controls.The experi-mental groups were added with different concentrations of microparticles(1,5,10 and 20 mg·L-1)while the control group was given no microparticles(0 mg·L-1).The number of nodes that formed the lumen after 5 h of incubation in Matrigel was counted,and the size of healing of the scratch area was analyzed after 6 h.To elucidate the mechanism through which MKMPs impacted angiogenesis,ELISA was used out to quantitatively detect the concentration of proteins in microparticles.RESULTS ① A three-stage differentiation cultural system was established to develop hESCs into MKs.Flow cytometry revealed progressive loss of pluripotency markers SSEA4 and TRA-1-60,while the mesodermal progenitor marker brachyury peaked at d 2.Subsequently,hemogenic endothelial/hematopoietic progenitor markers CD34 and CD43 emerged at d 5,followed by megakaryocytic markers CD41a and CD42b at d 13.Immunofluorescent images further demonstrated that MKs expressed specific proteins CD42,β1-tubulin,von VWF and FLI1 at d 13.②Microparticles were collected via differential centrifuga-tion.Transmission electron microscopy revealed that their substructure exhibited a typical double-layered membrane.Nanoparticle tracking analysis indicated that the size was(164.3±14.0)nm.The result of WB demonstrated that the microparticles expressed specific markers,including TSG101,CD9 and CD41.③ MKMPs were absorbed after being co-cultured with HUVECs for 3 h and enhanced the ability of HUVECs to form tubes and migrate.Notably,the treatment of 5 mg·L-1 MKMPs was more effective than 5 mg·L-1 PMPs treatment.The results of ELISA showed that the content of VEGF from MKMPs was higher than from PMPs,which may be the key factor in regulating endothelial biological function.CONCLUSION MKs derived from hESCs can generate functional microparticles which can promote angiogenesis.

Objective To explore the efficacy of a model combining clinicopathological characteristics and multiparametric MRI features for predicting BRCA-mutated breast cancer(BC).Methods A total of 256 BC patients were retrospectively selected and divided into BRCA mutation group(116 cases)and BRCA wild group(140 cases)based on the BRCA results.Chi-square tests or independ-ent sample t-tests were used to compare the differences in clinicopathological characteristics and multiparametric MRI features between the BRCA mutation group and the wild group.Risk factors for BRCA-mutated BC were identified through univariate and multivariate logistic regression ananlyses,and a combined predictive model was constructed.Receiver operating characteristic(ROC)curve was used to ana-lyze the diagnostic efficacy of the model.Results There were statistically significant differences in T stage,human epidermal growth factor receptor 2(HER-2),Ki-67,non-mass enhancement,enhancement pattern,time-signal intensity curve(TIC)type,and apparent diffusion coefficient(ADC)values between the BRCA mutation group and the wild group.Univariate logistic regression analysis showed that T stage,HER-2,Ki-67,non-mass enhancement,enhancement pattern,TIC type,and ADC values were risk factors for BRCA-mutated BC(P<0.05).Multivariate logistic regression analysis revealed that T stage,HER-2,Ki-67,enhancement pattern,and TIC type were independent risk factors for BRCA-mutated BC(P<0.05).The combined model incorporating T stage,HER-2,Ki-67,enhancement pattern,and TIC type had the best diagnostic efficacy in predicting BRCA-mutated BC,with an area under the curve(AUC)of 0.751.Conclusion The combined model integrating T stage,HER-2,Ki-67,enhancement pattern,and TIC type has good efficacy in predicting BRCA-mutated BC.

Objective To investigate the association of spleen volume with the risk of non-alcoholic fatty liver disease(NAFLD)as well as their causal relationship.Methods We included 90 NAFLD cases and 47 healthy controls who had received contrast-enhanced computed tomography(CT)scan of the abdomen at the Second Affiliated Hospital of Xi'an Jiaotong University from November 2022 to November 2023.We conducted three-dimensional reconstruction of the spleen through a deep learning network model using a two-stage coarse-to-fine segmentation approach.We compared the two groups using the two-sample t test or Mann-Whitney U test for continuous data and using the chi-square test for categorical data;evaluated the correlation between spleen volume and liver function indicators through Pearson correlation or Spearman rank correlation analyses;determined the factors influencing the development of NAFLD through multivariable Logistic regression analysis;and further assessed the casual relationship between spleen volume and NAFLD using the inverse variance-weighted two-sample Mendelian randomization(IVW-MR)method.Results Spleen volume was significantly larger in NAFLD cases than in controls(272.93±104.16 vs 204.37±81.20 cm3,P<0.001).The Spearman rank correlation analysis showed that spleen volume was positively correlated with the hepatic steatosis index(rs=0.422,P<0.001)and gamma-glutamyl transferase levels(rs=0.211,P=0.047)in patients with NAFLD.The multivariable Logistic regression analysis indicated that spleen volume was an independent risk factor for the development of NAFLD(odds ratio[OR]=1.01,95%confidence interval[CI]:1.00-1.02,P=0.049).The IVW-MR analysis detected a causal relationship between spleen volume and NAFLD(OR=1.16,95%CI:1.05-1.28,P=0.005).Conclusion Increased spleen volume may be a risk factor for the development and progression of NAFLD.Further studies are still needed to investigate the specific mechanism.

Lung cancer, particularly non-small cell lung cancer (NSCLC), is a leading cause of cancer-related mortality worldwide. In recent years, metabolomics has emerged as a key systems biology approach for analyzing small-molecule metabolites in cells, tissues and organisms. It provides new strategies for early diagnosis and metabolic profiling. Additionally, metabolomics plays a crucial role in studying resistance mechanisms in lung cancer. Tumor cell metabolic reprogramming is a key driving factor in the initiation and progression of lung cancer. Metabolomics studies have revealed how lung cancer cells regulate critical pathways such as energy metabolism, lipid metabolism, and amino acid metabolism to adapt to the demands of rapid proliferation and invasive metastasis. This review summarizes the latest advances in metabolomics research in lung cancer, focusing on the characteristics of metabolic reprogramming, the identification of potential metabolic biomarkers, and the prospects of metabolomics in early diagnosis and the elucidation of resistance mechanisms in lung cancer.
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Humans ; Metabolomics/methods* ; Lung Neoplasms/pathology* ; Animals ; Biomarkers, Tumor/metabolism*