Objective To investigate the efficacy of discontinuous density gradient combined with swim-up method for bacterial removal in frozen semen,and to provide experimental evidence for the safe application of frozen semen in assisted reproductive technology.Methods A total of 208 frozen semen samples containing non-pathogenic bacteria,cryopreserved in our human sperm bank from January 2019 to December 2023,were selected.The thawed semen samples were processed using discontinuous density gradient combined with swim-up method.To simulate in vitro fertilization procedures,the processed sperm suspensions were subjected to 5-day in vitro culture for bacterial residue monitoring.Prior to cryopreservation,all semen samples underwent routine bacterial culture to exclude pathogenic microorganisms.Post-thaw semen samples,processed sperm suspensions,and 5-day culture media were sent to an independent third-party laboratory for comprehensive bacterial identification.Results Among the 208 frozen semen samples containing non-pathogenic bacteria,41 bacterial species were identified,with 1～4 species per sample.After processing with discontinuous density gradient combined with swim-up method,the sterility rate of the sperm suspension reached 89.42%(186/208),with residual bacteria including Enterococcus faecalis,Corynebacterium glucuronolyticum,Group B Streptococcus,and Corynebacterium glycinophilum.After 5-day in vitro culture,the sterility rate increased to 93.75%(195/208),with residual bacteria limited to Entero-coccus faecalis and Group B Streptococcus.Conclusions The combined method of discontinuous density gradient and swim-up technique effectively eliminates most bacteria in frozen semen.However,the clearance rate for Entero-coccus faecalis and Group B Streptococcus remains below 90%.Optimization of pre-freezing bacterial screening and post-thaw processing techniques is necessary to enhance the safety of frozen semen in assisted reproductive technology.

Objective To investigate the efficacy of discontinuous density gradient combined with swim-up method for bacterial removal in frozen semen,and to provide experimental evidence for the safe application of frozen semen in assisted reproductive technology.Methods A total of 208 frozen semen samples containing non-pathogenic bacteria,cryopreserved in our human sperm bank from January 2019 to December 2023,were selected.The thawed semen samples were processed using discontinuous density gradient combined with swim-up method.To simulate in vitro fertilization procedures,the processed sperm suspensions were subjected to 5-day in vitro culture for bacterial residue monitoring.Prior to cryopreservation,all semen samples underwent routine bacterial culture to exclude pathogenic microorganisms.Post-thaw semen samples,processed sperm suspensions,and 5-day culture media were sent to an independent third-party laboratory for comprehensive bacterial identification.Results Among the 208 frozen semen samples containing non-pathogenic bacteria,41 bacterial species were identified,with 1～4 species per sample.After processing with discontinuous density gradient combined with swim-up method,the sterility rate of the sperm suspension reached 89.42%(186/208),with residual bacteria including Enterococcus faecalis,Corynebacterium glucuronolyticum,Group B Streptococcus,and Corynebacterium glycinophilum.After 5-day in vitro culture,the sterility rate increased to 93.75%(195/208),with residual bacteria limited to Entero-coccus faecalis and Group B Streptococcus.Conclusions The combined method of discontinuous density gradient and swim-up technique effectively eliminates most bacteria in frozen semen.However,the clearance rate for Entero-coccus faecalis and Group B Streptococcus remains below 90%.Optimization of pre-freezing bacterial screening and post-thaw processing techniques is necessary to enhance the safety of frozen semen in assisted reproductive technology.

Objective:To analyze the characteristics of transabdominal bowel ultrasound (TBUS) in immune checkpoint inhibitor-related colitis (IRC) and their correlation with endoscopic manifestations.Methods:A cross-sectional study was conducted. Clinical data from 10 patients with IRC treated at Peking Union Medical College Hospital from January 2022 to January 2024 were collected. The ulcerative colitis endoscopic index of severity (UCEIS) and Limberg classification were used to assess the severity of colonoscopy and TBUS examinations, respectively. Kendall's tau-b method was applied for correlation analysis between UCEIS scores and Limberg classification.Results:All the 10 patients were male with a median age of 65 years (59-74 years). The majority had lung cancer (8 patients) and all were in advanced stages, with 6 patients in stage Ⅲ and 4 in stage Ⅳ. They all received anti-programmed death 1 (PD-1) /anti-programmed death ligand 1 (PD-L1) combined with chemotherapy, among whom 2 patients were combined with anti-angiogenic drug treatment. The median time from the first immunotherapy to the onset of IRC was 1.50 (0.25-12.00) months; the median time from IRC treatment to clinical symptom relief to G1 was 2.45 (0.50-8.00) weeks. Nine patients were in the active phase, mainly G3 (8 patients) ; 1 was in the remission phase after treatment. TBUS showed that among the 9 active IRC patients, the entire colon was mainly involved (7 patients), with combined small intestine involvement (3 patients) ; the main manifestations were thickening of the bowel wall, with the thickest bowel wall being 7.0 (5.0-8.0) mm, mainly located in the sigmoid colon (3 patients) and descending colon (3 patients) ; increased bowel wall blood flow signals (Limberg classification 2-4) occurred in 7 patients; 3 active patients had perienteric fat wrapping, and 2 had blurred bowel wall stratification. The Kendall's tau-b correlation coefficient r between the entire colon UCEIS scores and Limberg classification was 0.891 ( P = 0.003), and the Kendall's tau-b correlation coefficient r between the colon segment UCEIS scores and Limberg classification was 0.690 ( P < 0.001) . Conclusion:During the active phase, the left colon of IRC is more severe in TBUS, which mainly manifests as the thickening bowel wall and increased blood flow signals, and the TBUS has good correlation with colonoscopy evaluation.

Objective:To investigate the relationship between alterations in dynamic functional connectivity (dFC) and spatial navigation abilities in individuals with subjective cognitive decline (SCD) across different Traditional Chinese Medicine (TCM) constitutions.Methods:Seventy-five participants with SCD, comprising 34 individuals with balanced constitutions and 41 individuals with biased constitutions, were recruited from the Affiliated Drum Tower Hospital of Nanjing University Medical School between August 2022 and January 2025. The participants underwent TCM constitution assessment, spatial navigation ability testing, and neuropsychological scale evaluation. Additionally, each participant was assessed using 3.0 T resting-state functional magnetic resonance imaging (rs-fMRI) and high-resolution T1-weighted imaging scans. Based on prior research, 20 spatial navigation-related regions of interest (ROIs) were defined. Afterwards, rs-fMRI time series were segmented using a sliding time window approach before calculating the dFC within the spatial navigation brain network.Results:Compared to the balanced constitution group, the biased constitution SCD group showed significantly lower scores on the Mini-Mental State Examination (MMSE) ( z=-3.05, P=0.002) and the Auditory Verbal Learning Test (AVLT) measures: immediate recall ( z=-2.12, P=0.035), short-delay recall ( z=-2.22, P=0.026), long-delay recall ( z=-2.88, P=0.004), cued recall ( z=-2.91, P=0.004), and recognition ( z=-2.20, P=0.028). They also exhibited significantly higher average error distances in ego-allocentric navigation ( z=-2.28, P=0.023), egocentric navigation ( z=-2.31, P=0.021), and delayed navigation ( z=-2.02, P=0.043). Participants with SCD who had a biased constitution also demonstrated significantly reduced dFC between the left parahippocampal gyrus (PHG) and left prefrontal cortex (PFC) ( t=2.43), right precuneus and right retrosplenial cortex (RSC) ( t=2.96), and left inferior parietal lobule (IPL) and left hippocampus ( t=2.42) (all P<0.05, Bonferroni-corrected). Conversely, the dFC was significantly increased between the right PHG and left PFC ( t=-2.29, P<0.05, Bonferroni-corrected). Significant correlations were also found in participants with SCD who had biased constitutions: the dFC between the left PHG and left PFC positively correlated with the egocentric navigation average total error ( r=0.34, P=0.030) and negatively correlated with the visuospatial memory cognitive domain ( r=-0.35, P=0.026); the dFC between the left IPL and left hippocampus negatively correlated with the egocentric navigation average total error ( r=-0.32, P=0.043); and the dFC between the right PHG and left PFC positively correlated with the delayed navigation average total error ( r=0.33, P=0.037). The area under the ROC curve for the combined differences in cognitive assessments, spatial navigation behavior, and navigation-related brain network dFC was 0.966 in predicting biased constitution versus balanced constitution in participants with SCD. Conclusions:Individuals with SCD and biased constitutions demonstrated poorer spatial navigation ability, possibly due to altered dFC within the spatial navigation brain network. Furthermore, the integrated model based on spatial navigation behaviors and dFC exhibited a high predictive value in distinguishing between individuals with SCD who had balanced and biased constitutions.

Lumbar disc (LD) herniation and aging are prevalent conditions that can result in substantial morbidity. This study aimed to clarify the mechanisms connecting the LD aging and herniation, particularly focusing on cellular senescence and molecular alterations in the nucleus pulposus (NP). We performed a detailed analysis of NP samples from a diverse cohort, including individuals of varying ages and those with diagnosed LD herniation. Our methodology combined histological assessments with single-nucleus RNA sequencing to identify phenotypic and molecular changes related to NP aging and herniation. We discovered that cellular senescence and a decrease in nucleus pulposus progenitor cells (NPPCs) are central to both processes. Additionally, we found an age-related increase in NFAT1 expression that promotes NPPC senescence and contributes to both aging and herniation of LD. This research offers fresh insights into LD aging and its associated pathologies, potentially guiding the development of new therapeutic strategies to target the root causes of LD herniation and aging.
Intervertebral Disc Displacement/metabolism* ; Humans ; Aging/pathology* ; Nucleus Pulposus/pathology* ; Male ; Female ; Transcriptome ; Middle Aged ; Lumbar Vertebrae/pathology* ; Adult ; Cellular Senescence ; Stem Cells/pathology* ; Aged ; Intervertebral Disc Degeneration/metabolism*

Objective:To analyze the characteristics of transabdominal bowel ultrasound (TBUS) in immune checkpoint inhibitor-related colitis (IRC) and their correlation with endoscopic manifestations.Methods:A cross-sectional study was conducted. Clinical data from 10 patients with IRC treated at Peking Union Medical College Hospital from January 2022 to January 2024 were collected. The ulcerative colitis endoscopic index of severity (UCEIS) and Limberg classification were used to assess the severity of colonoscopy and TBUS examinations, respectively. Kendall's tau-b method was applied for correlation analysis between UCEIS scores and Limberg classification.Results:All the 10 patients were male with a median age of 65 years (59-74 years). The majority had lung cancer (8 patients) and all were in advanced stages, with 6 patients in stage Ⅲ and 4 in stage Ⅳ. They all received anti-programmed death 1 (PD-1) /anti-programmed death ligand 1 (PD-L1) combined with chemotherapy, among whom 2 patients were combined with anti-angiogenic drug treatment. The median time from the first immunotherapy to the onset of IRC was 1.50 (0.25-12.00) months; the median time from IRC treatment to clinical symptom relief to G1 was 2.45 (0.50-8.00) weeks. Nine patients were in the active phase, mainly G3 (8 patients) ; 1 was in the remission phase after treatment. TBUS showed that among the 9 active IRC patients, the entire colon was mainly involved (7 patients), with combined small intestine involvement (3 patients) ; the main manifestations were thickening of the bowel wall, with the thickest bowel wall being 7.0 (5.0-8.0) mm, mainly located in the sigmoid colon (3 patients) and descending colon (3 patients) ; increased bowel wall blood flow signals (Limberg classification 2-4) occurred in 7 patients; 3 active patients had perienteric fat wrapping, and 2 had blurred bowel wall stratification. The Kendall's tau-b correlation coefficient r between the entire colon UCEIS scores and Limberg classification was 0.891 ( P = 0.003), and the Kendall's tau-b correlation coefficient r between the colon segment UCEIS scores and Limberg classification was 0.690 ( P < 0.001) . Conclusion:During the active phase, the left colon of IRC is more severe in TBUS, which mainly manifests as the thickening bowel wall and increased blood flow signals, and the TBUS has good correlation with colonoscopy evaluation.

Objective:To investigate the relationship between alterations in dynamic functional connectivity (dFC) and spatial navigation abilities in individuals with subjective cognitive decline (SCD) across different Traditional Chinese Medicine (TCM) constitutions.Methods:Seventy-five participants with SCD, comprising 34 individuals with balanced constitutions and 41 individuals with biased constitutions, were recruited from the Affiliated Drum Tower Hospital of Nanjing University Medical School between August 2022 and January 2025. The participants underwent TCM constitution assessment, spatial navigation ability testing, and neuropsychological scale evaluation. Additionally, each participant was assessed using 3.0 T resting-state functional magnetic resonance imaging (rs-fMRI) and high-resolution T1-weighted imaging scans. Based on prior research, 20 spatial navigation-related regions of interest (ROIs) were defined. Afterwards, rs-fMRI time series were segmented using a sliding time window approach before calculating the dFC within the spatial navigation brain network.Results:Compared to the balanced constitution group, the biased constitution SCD group showed significantly lower scores on the Mini-Mental State Examination (MMSE) ( z=-3.05, P=0.002) and the Auditory Verbal Learning Test (AVLT) measures: immediate recall ( z=-2.12, P=0.035), short-delay recall ( z=-2.22, P=0.026), long-delay recall ( z=-2.88, P=0.004), cued recall ( z=-2.91, P=0.004), and recognition ( z=-2.20, P=0.028). They also exhibited significantly higher average error distances in ego-allocentric navigation ( z=-2.28, P=0.023), egocentric navigation ( z=-2.31, P=0.021), and delayed navigation ( z=-2.02, P=0.043). Participants with SCD who had a biased constitution also demonstrated significantly reduced dFC between the left parahippocampal gyrus (PHG) and left prefrontal cortex (PFC) ( t=2.43), right precuneus and right retrosplenial cortex (RSC) ( t=2.96), and left inferior parietal lobule (IPL) and left hippocampus ( t=2.42) (all P<0.05, Bonferroni-corrected). Conversely, the dFC was significantly increased between the right PHG and left PFC ( t=-2.29, P<0.05, Bonferroni-corrected). Significant correlations were also found in participants with SCD who had biased constitutions: the dFC between the left PHG and left PFC positively correlated with the egocentric navigation average total error ( r=0.34, P=0.030) and negatively correlated with the visuospatial memory cognitive domain ( r=-0.35, P=0.026); the dFC between the left IPL and left hippocampus negatively correlated with the egocentric navigation average total error ( r=-0.32, P=0.043); and the dFC between the right PHG and left PFC positively correlated with the delayed navigation average total error ( r=0.33, P=0.037). The area under the ROC curve for the combined differences in cognitive assessments, spatial navigation behavior, and navigation-related brain network dFC was 0.966 in predicting biased constitution versus balanced constitution in participants with SCD. Conclusions:Individuals with SCD and biased constitutions demonstrated poorer spatial navigation ability, possibly due to altered dFC within the spatial navigation brain network. Furthermore, the integrated model based on spatial navigation behaviors and dFC exhibited a high predictive value in distinguishing between individuals with SCD who had balanced and biased constitutions.

Endometrial injury is the main fact leading to the damage of endometrial basal layer.It is usually caused by traumatic treatment,inflammatory injury,radiation,chemical drug-related injury,and delivery-related divertic-ulum.The use of hyaluronic acid,decellularized matrix,amniotic membrane,platelet-rich plasma,mesenchymal stem cells(especially scaffold-based therapeutic delivery)has been proved to reduce post-injury scar formation and promote endometrial growth and repair,which provides a new therapeutic strategy for repairing the damaged endo-metrium with improved tolerability and fertility outcomes.

Objective:To explore mechanism of Chaihu Longgu Muli Decoction and Ganmai Dazao Decoction in treatment of post-stroke depression(PSD)based on network pharmacology,molecular docking and animal experiment.Methods:TCMSP and other databases were used to predict active components and targets of Chaihu Longgu Muli Decoction and Ganmai Dazao Decoction.Targets of PSD were retrieved from PharmGKB and other databases,and"component-intersection target-disease"network was constructed by Cytoscape(v3.9.1)software.PPI network was constructed by String(v11.5)database,and GO enrichment and KEGG pathway analy-sis of intersection targets were performed by DAVID6.8 database.AutoDock vina(v1.1.2)software was used for molecular docking.Pymol(v 2.5)and other softwares were used to visualize optimal docking results.Animal experiments were setup in control group,model group,fluoxetine group,TCM group and TCM+fluoxetine group,neurobehavioral scores and expressions of neurotransmitters and inflammatory factors in brain tissues were detected.mRNA and protein expressions of key genes PPARG,MAPK3,AKT1,PIK3CA were detected by RT-qPCR and Western blot.Results:A total of 225 kinds of active ingredients of Chaihu Longgu Muli Decoction and Ganmai Dazao Decoction were obtained,which acted on 119 targets of PSD,among which key targets included MAPK3,AKT1,PIK3CA and PPARG,key pathways including MAPK signaling pathway,PI3K-Akt signaling pathway and etc.Compared with model group,MAPK3 mRNA and protein expressions were decreased,AKT1,PIK3CA,PPARG mRNA and protein expressions were increased in TCM group and TCM+fluoxetine group(P<0.05).Conclusion:Mechanism of Chaihu Longgu Muli Decoction and Ganmai Dazao Decoction in treatment of PSD may be related to inhibition of MAPK3 expression,promotion of AKT1,PIK3CA,PPARG expressions,alleviation of inflammatory response and oxidative stress in brain tissues.

Objective To investigate the effects of tanreqing injection(TRQ)on lipopolysaccharides(LPS)-induced colon injury and intestinal flora in mice.Methods BALB/c mice were randomly divided into normal group,LPS-induced group,low-dose TRQ group(LPS+TRQ 4mg/kg)and high-dose TRQ group(LPS+TRQ 8mg/kg)by completely randomalized design.Body weight and liv-ing status of mice were recorded daily.The serum inflammatory cytokines interleukin-1(IL-1),IL-6 and tumor necrosis factor-α(TNF-α)were detected by enzyme-linked immunosorbent assay(ELISA).Hematoxylin-eosin staining was used to observe the path-ological changes of colon.16S rDNA sequencing was used to analyze the changes of intestinal flora in mice.Results High-dose TRQ had significant therapeutic effect on LPS-induced weight loss in mice(P＜0.05).Both low-dose and high-dose TRQ can inhibit the LPS-induced increase in serum IL-1,IL-6 and TNF-α levels(P＜0.01).Both low-dose and high-dose TRQ groups could sig-nificantly improve the colonic shortening(P＜0.01,P＜0.05)and pathological changes induced by LPS.High-dose TRQ could signifi-cantly increase the Simpson index of intestinal flora(P＜0.05)and improve Beta diversity.The abundance of a variety of intestinal flora changed significantly in the high-dose TRQ group.Conclusion TRQ may improve the intestinal function by inhibiting the release of in-flammatory cytokines and regulating the structure of intestinal flora,and play a protective role in LPS-induced inflammatory response and colon injury in mice.