A novel analytical method was developed in this study by combining online solid phase extraction with ultra performance liquid chromatography-tandem mass spectrometry(Online SPE-UPLC-MS/MS)for simultaneous determination of 50 kinds of steroid hormones in surface water.Specifically,after high-speed centrifugation of 4 mL water samples,the supernatant was directly injected into an Oasis HLB online SPE column for enrichment and purification.Subsequently,the target compounds were transferred to the analytical column via valve switching for separation and analysis.The chromatographic separation was performed on a Thermo Acclaim RSLC C18 column(100 mm×2.1 mm,2.2 μm),using a mobile phase composed of 5 mmol/L ammonium fluoride aqueous solution and acetonitrile.Mass spectrometric detection was conducted in positive ion mode,utilizing multiple reaction monitoring(MRM)with quantification achieved by the internal standard method.The method validation demonstrated that the limits of detection(LOD)for the 50 kinds of steroid hormones ranged from 0.02 to 0.50 ng/L,while the limits of quantification(LOQ)were between 0.08 and 1.67 ng/L.The average recoveries in surface water samples at spiked concentrations of 5,20 and 200 ng/L were between 74.1％and 119％,with relative standard deviations(RSDs)of 0.2％to 9.9％.This method was applied to analyze 11 surface water samples collected from sites surrounding a pharmaceutical and chemical industrial park.A total of 44 kinds of steroid hormones were detected,with concentrations ranging from 0.11 to 88.6 ng/L,revealing the presence of hormone contamination in the environmental waters surrounding industrial areas.Compared with the traditional offline SPE methods,the proposed online SPE technique significantly reduced sample volume requirements and pretreatment time,while minimizing the loss of target compounds during the pretreatment process.Moreover,compared to reported online SPE techniques,this method achieved high-throughput analysis of multiple classes of steroid hormones,with lower detection limits and higher recoveries.Overall,this method provided rapid sample preparation,high sensitivity,and excellent stability,making it suitable for the direct analysis of trace steroid hormones in surface water.

Stem cells play a crucial role in maintaining tissue regenerative capacity and homeostasis. However, mechanisms associated with stem cell senescence require further investigation. In this study, we conducted a proteomic analysis of human dental pulp stem cells (HDPSCs) obtained from individuals of various ages. Our findings showed that the expression of NUP62 was decreased in aged HDPSCs. We discovered that NUP62 alleviated senescence-associated phenotypes and enhanced differentiation potential both in vitro and in vivo. Conversely, the knocking down of NUP62 expression aggravated the senescence-associated phenotypes and impaired the proliferation and migration capacity of HDPSCs. Through RNA-sequence and decoding the epigenomic landscapes remodeled induced by NUP62 overexpression, we found that NUP62 helps alleviate senescence in HDPSCs by enhancing the nuclear transport of the transcription factor E2F1. This, in turn, stimulates the transcription of the epigenetic enzyme NSD2. Finally, the overexpression of NUP62 influences the H3K36me2 and H3K36me3 modifications of anti-aging genes (HMGA1, HMGA2, and SIRT6). Our results demonstrated that NUP62 regulates the fate of HDPSCs via NSD2-dependent epigenetic reprogramming.
Humans ; Dental Pulp/cytology* ; Nuclear Pore Complex Proteins/genetics* ; Cellular Senescence/genetics* ; Stem Cells/metabolism* ; Epigenesis, Genetic ; Cell Proliferation ; Cell Differentiation ; Histone-Lysine N-Methyltransferase/metabolism* ; Cells, Cultured ; Cellular Reprogramming ; Cell Movement ; Proteomics

The objective of this study was to develop a rapid and precise detection technique for por-cine Senecavirus A(SVA)employing reverse transcription recombinase polymerase amplification(RT-RAA)in conjunction with CRISPR Cas13a technology.Additionally,the study aimed to opti-mize the assay's reaction conditions to enhance amplification efficiency.Eight RT-RAA primer sets were designed based on the conserved gene sequence of porcine SVA,and a series of reaction condi-tions were evaluated to refine the RT-RAA reaction system.Subsequently,CRISPR-derived RNA(crRNA)sequences were developed and selected to construct the RT-RAA-CRISPR reaction sys-tem.The method's specificity was determined by examining six prevalent porcine pathogenic nucleic acids,while its sensitivity was assessed using SVA cRNA standards quantified by digital PCR.The method's stability and the consistency of clinical sample analysis were also evaluated.The findings revealed that the optimized RT-RAA and CRISPR reaction systems exhibited the highest amplifi-cation efficiency at a reaction temperature of 37 ℃.Among the eight crRNAs,five were identified as exhibiting the strongest detection signals.The formulated RT-RAA-CRISPR Cas13a method demonstrated exceptional specificity,showing no cross-reactivity with other common porcine disea-ses,including ASFV,PRRSV,PEDV,PCV2,CSFV,and PRV.The method achieved high sensitivi-ty,detecting as low as 0.86 copies/μL of SVA,and exhibited stable fluorescence output,robust re-producibility,and the ability to complete clinical sample analysis within 50 minutes.Consistency e-valuation with six positive and 58 negative samples indicated 100％agreement in outcomes.These results substantiate that the study successfully established a rapid and specific RT-RAA-CRISPR Cas13a detection method for the on-site identification of porcine Senecavirus A,demonstrating high specificity and sensitivity,and holds promise for application in SVA monitoring and control initia-tives.

The objective of this study was to develop a rapid and precise detection technique for por-cine Senecavirus A(SVA)employing reverse transcription recombinase polymerase amplification(RT-RAA)in conjunction with CRISPR Cas13a technology.Additionally,the study aimed to opti-mize the assay's reaction conditions to enhance amplification efficiency.Eight RT-RAA primer sets were designed based on the conserved gene sequence of porcine SVA,and a series of reaction condi-tions were evaluated to refine the RT-RAA reaction system.Subsequently,CRISPR-derived RNA(crRNA)sequences were developed and selected to construct the RT-RAA-CRISPR reaction sys-tem.The method's specificity was determined by examining six prevalent porcine pathogenic nucleic acids,while its sensitivity was assessed using SVA cRNA standards quantified by digital PCR.The method's stability and the consistency of clinical sample analysis were also evaluated.The findings revealed that the optimized RT-RAA and CRISPR reaction systems exhibited the highest amplifi-cation efficiency at a reaction temperature of 37 ℃.Among the eight crRNAs,five were identified as exhibiting the strongest detection signals.The formulated RT-RAA-CRISPR Cas13a method demonstrated exceptional specificity,showing no cross-reactivity with other common porcine disea-ses,including ASFV,PRRSV,PEDV,PCV2,CSFV,and PRV.The method achieved high sensitivi-ty,detecting as low as 0.86 copies/μL of SVA,and exhibited stable fluorescence output,robust re-producibility,and the ability to complete clinical sample analysis within 50 minutes.Consistency e-valuation with six positive and 58 negative samples indicated 100％agreement in outcomes.These results substantiate that the study successfully established a rapid and specific RT-RAA-CRISPR Cas13a detection method for the on-site identification of porcine Senecavirus A,demonstrating high specificity and sensitivity,and holds promise for application in SVA monitoring and control initia-tives.

Objective To explore the regulatory mechanism of Macelignan on oxidative stress-mediated neuronal injury in autophagy and apoptosis.Methods Murine hippocampal neuronal HT22 cells were treated with 2.5 mmol·L-1 glutamic acid(Glu)to establish an oxidative stress cell model.The cells were divided into normal group(normal cultured cells),model group(2.5 mmol·L-1 Glu)and experimental-L,-M,-H groups(2.5,5,10 μmol·L-1Macelignan treatment),inhibitor group(2.5 mmol·L-1 Glu+10 μmol·L-1 Macelignan+10 μmol·L-1 LY294002).Aoptosis rate was detected by flow cytometry;the protein expression level of autophagy-related protein LC3B(LC3B),anti-SQSTM1/p62(p62),p21,B-cell lymphoma-2(Bcl-2)and Bcl-2 associated X protein(Bax)was detected by Western blot.Results The apoptosis rates in the normal group,model group and experimental-L,-M,-H groups were(4.58±1.25)％,(8.75±0.55)％,(6.30±1.71)％,(5.97±2.27)％and(5.49±1.71)％.The difference between model group and normal group was statistically significant(P＜0.01).The difference between experimental-L,-M,-H groups and model group was statistically significant(all P＜0.01).The levels of LC3B in normal group,model group,experimental-L,experimental-M,experimental-H groups and inhibitor group were 0.28±0.02,0.74±0.02,1.02±0.04,0.70±0.03,0.26±0.02 and 0.21±0.01;p62 levels were 0.49±0.08,0.33±0.03,0.50±0.07,0.59±0.01,0.64±0.13 and 0.65±0.06;p21 levels were 0.87±0.02,1.18±0.03,0.98±0.03,0.88±0.03,0.72±0.06 and 0.81±0.02;Bcl-2/Bax levels were 1.74±0.23,1.11±0.10,1.38±0.05,1.66±0.26,1.58±0.29 and 1.53±0.09,respectively.The differences between model group and normal group,between model group and experimental-H group,between model group and inhibitor group,were also statistically significant(all P＜0.01).Conclusion Macelignan can reduce the damage of hippocampal neurons induced by glutamate acid by regulating the process of autophagy and apoptosis,and has obvious neuroprotective effect.

Objective To investigate the effect of exosomal miR-146a-5p expression on gray matter volume in patients with major depressive disorder(MDD).Methods A total of 113 MDD patients(MDD group)and 107 healthy controls(HC)(HC group)were selected.Peripheral blood was collected and exosomes were isolated to quantify miR-146a-5p expression.Brain high-resolution T1 WI images of MDD and HC were obtained via MR,and gray matter volume was computed via SPM12 software.The interaction effect of"Depression×miR-146a-5p expression"on gray matter volume was analyzed using SPM's Flexible factorial design,and the between-group difference was assessed by extracting the mean value,thus to analyze whether MDD-related gray matter volume abnormalities were dependent on miR-146a-5p expression.Results Exosomal miR-146a-5p expression was significantly elevated in MDD group compared to HC group.Voxel-based factorial analysis revealed a relationship between high miR-146a-5p expression in MDD group and reduced gray matter volume in the anterior and posterior cingulate cortices(independent voxel threshold P＜0.001,AlphaSim corrected),and a significantly reduced gray matter volume as compared with HC group was detected in the two regions.Conclusion The exosomal miR-146a-5p is overexpressed in patients with MDD and may be associated with specific cortical atrophy in patients with MDD.

Objective To evaluate and compare the success rate and short-term clinical prognosis of transfemoral transcatheter aortic valve replacement(TF-TAVR)for patients with pure aortic regurgitation(PAR)of different annulus sizes.Methods This study is a single center retrospective study,selecting symptomatic PAR patients who received TF-TAVR treatment at Zhongshan Hospital Fudan University from September 2019 to September 2023.Based on preoperative CT results,all patients were divided into three groups:Group A(aortic annulus circumference＜80 mm),Group B(80 mm≤aortic annulus circumference＜85 mm),and Group C(aortic annulus circumference≥ 85 mm).The primary endpoint was success rate and 30d all-cause mortality,while the secondary endpoint was TAVR related complications.Results A total of 61 PAR patients were included in this study,including 27 in Group A,21 in Group B,and 13 in Group C.The overall success rate is 82.0％,and the 30 d all-cause mortality rate is 3.3％.The success rate of Group C patients was significantly lower(P=0.012),with higher rates of conversion to surgery and valve-in-valve implantation(P=0.022 and P=0.040).In terms of secondary endpoint events,there were no significant differences among the three groups in major bleeding events,major vascular complications,stroke,myocardial infarction,newly developed atrial fibrillation,implantation of new pacemakers,coronary artery occlusion,and postoperative moderate to severe perivalvular leakage(all P＞0.05).Conclusions The circumference of the aortic valve annulus is a key factor affecting the success rate of TF-TAVR in PAR,and PAR patients with an aortic valve annulus circumference less than 85mm may be more suitable for TF-TAVR.

Humans ; Tricuspid Valve/surgery* ; Heart Valve Prosthesis Implantation ; Surgical Instruments ; Treatment Outcome ; Cardiac Catheterization ; Tricuspid Valve Insufficiency/surgery*

Objective: To explore the efficacy of chemotherapy re-challenge in the third-line setting for patients with metastatic colorectal cancer (mCRC) in the real world. Methods: The clinicopathological data, treatment information, recent treatment efficacy, adverse events and survival data of mCRC patients who had disease progression after treatment with oxaliplatin-based and/or irinotecan-based chemotherapy and received third-line chemotherapy re-challenge from January 2013 to December 2020 at Tianjin Medical University Cancer Institute and Hospital were retrospectively collected. Survival curves were plotted with the Kaplan-Meier method, and the Cox proportional hazard model was used to analyze the prognostic factors. Results: A total of 95 mCRC patients were included. Among them, 32 patients (33.7%) received chemotherapy alone and 63 patients (66.3%) received chemotherapy combined with targeted drugs. Eighty-three patients were treated with dual-drug chemotherapy (87.4%), including oxaliplatin re-challenge in 35 patients and irinotecan re-challenge in 48 patients. The remaining 12 patients were treated with triplet chemotherapy regimens (12.6%). Among them, as 5 patients had sequential application of oxaliplatin and irinotecan in front-line treatments, their third-line therapy re-challenged both oxaliplatin and irinotecan; 7 patients only had oxaliplatin prescription before, and these patients re-challenged oxaliplatin in the third-line treatment. The overall response rate (ORR) and disease control rate (DCR) reached 8.6% (8/93) and 61.3% (57/93), respectively. The median progression free survival (mPFS) and median overall survival (mOS) were 4.9 months and 13.0 months, respectively. The most common adverse events were leukopenia (34.7%) and neutropenia (34.7%), followed by gastrointestinal adverse reactions such as nausea (32.6%) and vomiting (31.6%). Grade 3-4 adverse events were mostly hematological toxicity. Cox multivariate analysis showed that gender (HR=1.609, 95% CI: 1.016-2.548) and the PFS of front-line treatments (HR=0.598, 95% CI: 0.378-0.947) were independent prognostic factors. Conclusion: The results suggested that it is safe and effective for mCRC patients to choose third-line chemotherapy re-challenge, especially for patients with a PFS of more than one year in front-line treatments.
Humans ; Irinotecan/therapeutic use* ; Oxaliplatin/therapeutic use* ; Colorectal Neoplasms/pathology* ; Retrospective Studies ; Fluorouracil ; Colonic Neoplasms/chemically induced* ; Rectal Neoplasms/drug therapy* ; Antineoplastic Combined Chemotherapy Protocols/adverse effects* ; Camptothecin/adverse effects*

Objective: To explore the efficacy of chemotherapy re-challenge in the third-line setting for patients with metastatic colorectal cancer (mCRC) in the real world. Methods: The clinicopathological data, treatment information, recent treatment efficacy, adverse events and survival data of mCRC patients who had disease progression after treatment with oxaliplatin-based and/or irinotecan-based chemotherapy and received third-line chemotherapy re-challenge from January 2013 to December 2020 at Tianjin Medical University Cancer Institute and Hospital were retrospectively collected. Survival curves were plotted with the Kaplan-Meier method, and the Cox proportional hazard model was used to analyze the prognostic factors. Results: A total of 95 mCRC patients were included. Among them, 32 patients (33.7%) received chemotherapy alone and 63 patients (66.3%) received chemotherapy combined with targeted drugs. Eighty-three patients were treated with dual-drug chemotherapy (87.4%), including oxaliplatin re-challenge in 35 patients and irinotecan re-challenge in 48 patients. The remaining 12 patients were treated with triplet chemotherapy regimens (12.6%). Among them, as 5 patients had sequential application of oxaliplatin and irinotecan in front-line treatments, their third-line therapy re-challenged both oxaliplatin and irinotecan; 7 patients only had oxaliplatin prescription before, and these patients re-challenged oxaliplatin in the third-line treatment. The overall response rate (ORR) and disease control rate (DCR) reached 8.6% (8/93) and 61.3% (57/93), respectively. The median progression free survival (mPFS) and median overall survival (mOS) were 4.9 months and 13.0 months, respectively. The most common adverse events were leukopenia (34.7%) and neutropenia (34.7%), followed by gastrointestinal adverse reactions such as nausea (32.6%) and vomiting (31.6%). Grade 3-4 adverse events were mostly hematological toxicity. Cox multivariate analysis showed that gender (HR=1.609, 95% CI: 1.016-2.548) and the PFS of front-line treatments (HR=0.598, 95% CI: 0.378-0.947) were independent prognostic factors. Conclusion: The results suggested that it is safe and effective for mCRC patients to choose third-line chemotherapy re-challenge, especially for patients with a PFS of more than one year in front-line treatments.
Humans ; Irinotecan/therapeutic use* ; Oxaliplatin/therapeutic use* ; Colorectal Neoplasms/pathology* ; Retrospective Studies ; Fluorouracil ; Colonic Neoplasms/chemically induced* ; Rectal Neoplasms/drug therapy* ; Antineoplastic Combined Chemotherapy Protocols/adverse effects* ; Camptothecin/adverse effects*