BACKGROUND:The cryopreservation technology enables tissues/cells to be stored for a long time in a low-temperature environment while maintaining the integrity of their activity and function,which is of great significance for the construction of cell therapy,tissue engineering and biological sample banks.Cryoprotective agents often contain dimethyl sulfoxide and serum.To avoid the toxic side effects of dimethyl sulfoxide,the complexity of serum components and immune responses,although some finished cryoprotective agents have been marketed,they are faced with many difficulties such as high cost and limited application.Therefore,there is an urgent need to develop a cryoprotective agent with clear components and the ability to solve the above problems.OBJECTIVE:To evaluate the effects of a novel cryoprotectant on cryopreservation efficiency of different tissue and cell sources.METHODS:By applying the novel cryoprotectant as an experimental group with the commercially available and widely used cryoprotectant(control group)to umbilical cord Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,NK and CIK cells,comparative analyses were conducted in terms of cell morphology,number,viability,surface markers,differentiation potential,and cell-killing toxicity assay before cryopreservation and after resuscitation thawing.We confirmed the cryopreservation effect of the new cryoprotectant and its potential application value.RESULTS AND CONCLUSION:(1)The novel cryoprotectant facilitated the normal growth of cryopreserved Wharton's jelly tissue upon recovery,exhibiting mesenchymal stem cell morphology.No significant differences were observed between the experimental and control groups in terms of cell recovery rate,surface markers,and differentiation potential.(2)There was no significant difference in the number and viability of cells between the experimental group and the control group after cryopreservation of cord blood/peripheral blood mononuclear cells,and the cryo-resuscitated cell numbers and viability of derived NK cells/CIK cells did not show significant difference between the experimental and control groups.(3)For NK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportion of CD56+CD16+cell subpopulations between the experimental group and the control group.For CIK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportions of CD3+CD8+and CD3+CD56+cell subpopulations between the experimental group and the control group.(4)In terms of cytotoxicity testing,when the effective-target ratio of immune cells and melanoma cell line Mel624 was 20:1,whether it was NK cells/CIK cells derived from cord blood or peripheral blood mononuclear cells,there was no significant difference in the tumoricidal activity of cells between the experimental group and the control group.These findings suggest that the novel cryoprotectant can replace existing commercially available and widely used cryoprotectants,and is applicable to Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,as well as NK and CIK cells,providing a solid technical foundation for the scaling,standardization,and commercialization of universal cryoprotectants.

BACKGROUND:Long non-coding RNA(LncRNA)plays an important role in nervous system development and neurological diseases.Previous studies by the research team have demonstrated that human umbilical cord mesenchymal stem cells overexpressing erythropoietin(EPO-MSCs)under ischemic and hypoxic conditions have better neuroprotective functions and significantly activate the expression of LncRNA XIST.Research suggests that XIST is related to the pathogenesis of hypoxic-ischemic encephalopathy,but the role and mechanism of its regulation by EPO-MSCs in protecting ischemic-hypoxic neurons remain unclear.OBJECTIVE:To explore the new mechanism by which LncRNA XIST,in response to EPO-MSC signaling,affects the apoptosis of ischemic-hypoxic SH-SY5Y cells.METHODS:(1)SH-SY5Y cell lines with knockdown of LncRNA XIST(sh-XIST)and negative control(NC-XIST)were constructed through lentiviral transfection.Oxygen-glucose deprivation was used to induce ischemic-hypoxic injury in the cells.Transwell chambers were used to create a non-contact co-culture system with EPO-MSCs,sh-XIST,and NC-XIST ischemic-hypoxic SH-SY5Y cells.Cell proliferation ability was detected using the CCK-8 assay.Cell migration ability was assessed using the scratch assay,and cell apoptosis was measured by flow cytometry.(2)RNA-seq bioinformatics analysis was performed to screen for differentially expressed genes and pathways between sh-XIST and NC-XIST cell lines.Dual-luciferase experiments were used to verify the relationship between miR-124-3p and the target genes XIST and GRIN1.qRT-PCR was conducted to validate the expression levels of downstream miR-124-3p and GRIN1 genes.(3)miR-124-3p inhibitors and mimics were added to verify phenotypic changes in SH-SY5Y cells after ischemic-hypoxic injury and co-culture with EPO-MSCs.RESULTS AND CONCLUSION:(1)Compared with the NC-XIST group,SH-SY5Y cells in the sh-XIST group showed reduced proliferation and migration abilities and increased apoptosis after ischemic-hypoxic injury and co-culture with EPO-MSCs.(2)Dual-luciferase experiments showed that miR-124-3p interacted with the target gene XIST.SH-SY5Y cells transfected with miR-124-3p mimics and co-cultured with EPO-MSCs showed decreased apoptosis after ischemic-hypoxic injury,while SH-SY5Y cells transfected with miR-124-3p inhibitors showed increased apoptosis after co-culture with EPO-MSCs.(3)Transcriptomic sequencing and bioinformatics analysis of sh-XIST revealed significant downregulation of the neuroactive ligand-receptor pathway and the key receptor gene GRIN1 for central nervous system development.(4)Dual-luciferase experiments showed that miR-124-3p interacted with GRIN1.GRIN1 expression was significantly downregulated in the sh-XIST group after ischemic-hypoxic injury compared with the NC-XIST group.These findings indicate that LncRNA XIST promotes GRIN1 expression by upregulating miR-124-3p,thereby reducing cell apoptosis after ischemic-hypoxic injury and co-culture with EPO-MSCs and enhancing proliferation and migration.sh-XIST can block this protective function.

Objective:To explore the predictive factors for extubation in the operating room after single lung transplantation without the assistance of extracorporeal membrane oxygenation (ECMO) in adult patients with end-stage interstitial lung disease (ILD), as well as their short-term (1-year postoperative) prognosis.Methods:A retrospective analysis was conducted on the clinical data of 78 adult ILD recipients who underwent single lung transplantation without ECMO assistance at the First Affiliated Hospital of Guangzhou Medical University from June 2018 to June 2023. Based on whether extubation was completed in the operating room (OR), patients were divided into the OR group (19 cases) and ICU group (59 cases). Baseline characteristics of donors and recipients, as well as intraoperative events, were compared between the two groups. Univariate logistic regression analysis was used to identify potential predictors, and variables with P<0.2 were included in multivariate logistic regression to determine independent predictors for OR extubation. Receiver operating characteristic (ROC) curves were plotted to evaluate predictive performance. The Kaplan-Meier method was used to analyze survival, and short-term prognosis between groups was compared. Results:The rate of OR extubation after single lung transplantation in ILD recipients was 24%(19/78). Compared with the ICU group, the OR group had shorter operation times, lower fluid volumes, reduced transfusions of red blood cells and plasma, less intraoperative bleeding, and lower lactate levels 15 minutes after pulmonary artery reperfusion (all P<0.05). Univariate logistic regression analysis identified the following factors as significantly associated with OR extubation: recipient age ( P=0.100), operative time ( P=0.001), fluid infusion volume ( P=0.005), red blood cell transfusion volume ( P=0.037), plasma transfusion volume ( P=0.039), blood loss ( P=0.004), oxygenation index at 15 minutes after reperfusion ( P=0.174), and blood lactate at 15 minutes after reperfusion ( P=0.041). Multivariate analysis revealed that intraoperative blood loss was an independent predictor of OR extubation ( OR=0.993, 95% CI: 0.986 - 0.999, P=0.026). ROC curve analysis showed that blood loss had an area under the curve (AUC) of 0.822 in predicting OR extubation, with a sensitivity of 64.4% and specificity of 89.5%. Postoperatively, patients in the OR group had significantly shorter durations of mechanical ventilation [0 vs 5 (3,11) days, P<0.001], ICU stay [7(4,8) vs 9(6,20) days, P=0.012], and overall postoperative hospitalization [19 (15,23) vs 25 (19,39) days, P=0.015]. Within one year after surgery, 2 patients (11%) in the OR group and 19 patients (32%) in the ICU group had died, but the difference in 1-year survival rates between the two groups was not statistically significant. Conclusions:Intraoperative blood loss is an independent predictor of extubation in the operating room. Early extubation in non-ECMO-assisted single lung transplantation for ILD patients is associated with improved short-term outcomes.

OBJECTIVE: Humans are exposed to complex mixtures of environmental chemicals and other factors that can affect their health. Analysis of these mixture exposures presents several key challenges for environmental epidemiology and risk assessment, including high dimensionality, correlated exposure, and subtle individual effects. METHODS: We proposed a novel statistical approach, the generalized functional linear model (GFLM), to analyze the health effects of exposure mixtures. GFLM treats the effect of mixture exposures as a smooth function by reordering exposures based on specific mechanisms and capturing internal correlations to provide a meaningful estimation and interpretation. The robustness and efficiency was evaluated under various scenarios through extensive simulation studies. RESULTS: We applied the GFLM to two datasets from the National Health and Nutrition Examination Survey (NHANES). In the first application, we examined the effects of 37 nutrients on BMI (2011-2016 cycles). The GFLM identified a significant mixture effect, with fiber and fat emerging as the nutrients with the greatest negative and positive effects on BMI, respectively. For the second application, we investigated the association between four pre- and perfluoroalkyl substances (PFAS) and gout risk (2007-2018 cycles). Unlike traditional methods, the GFLM indicated no significant association, demonstrating its robustness to multicollinearity. CONCLUSION GFLM framework is a powerful tool for mixture exposure analysis, offering improved handling of correlated exposures and interpretable results. It demonstrates robust performance across various scenarios and real-world applications, advancing our understanding of complex environmental exposures and their health impacts on environmental epidemiology and toxicology.
Humans ; Environmental Exposure/analysis* ; Linear Models ; Nutrition Surveys ; Environmental Pollutants ; Body Mass Index

Objective To investigate the occurrence of hemodialysis events in patients who received maintenance hemodialysis in outpatient setting in Liangshan Yi Autonomous Prefecture of Sichuan Province,and provide basis for the formulation of infection prevention and control strategies in this area.Methods Based on the standards from Center for Disease Control and Prevention-National Healthcare Safety Network(CDC-NHSN)as well as the WS/T 312-2023 standards,four public hospitals(two secondary and two tertiary hospitals)in three counties and one city of Liangshan Yi Autonomous Prefecture were selected,and prospective surveillance method was adopted.A total of 826 patients(n=8 675 cases)who received maintenance hemodialysis on the first two working day of each month from March 2024 to March 2025 were surveilled.Data were collected with standardized form by infection control teams in dialysis centers.Statistical analysis was conducted.Results During the surveillance period,the incidence of hemodialysis events was 5.15％(447/8 675).The incidence(16.67％,36/216)and mortality(10.00％,4/40)of hemodialysis events in patients with non-tunneled catheters were the highest among patients with all types of ac-cess.Patients with tunneled-catheters had the highest hospitalization rate(42.86％).Systemic use of antimicrobial agents accounted for 82.33％(368/447).Upper respiratory tract infection was the main infection(38.32％).In-fection at the puncture site of vascular access accounted for 16.78％(75/447).Four cases(0.89％,4/447)had positive blood culture.Conclusion Systemic antimicrobial use and infection at vascular access puncture site is the most frequent events in patients who receive maintenance hemodialysis in outpatient setting in Liangshan Yi Autono-mous Prefecture of Sichuan Province.Patients with non-tunneled catheters have the highest rate of hemodialysis events.

BACKGROUND:The cryopreservation technology enables tissues/cells to be stored for a long time in a low-temperature environment while maintaining the integrity of their activity and function,which is of great significance for the construction of cell therapy,tissue engineering and biological sample banks.Cryoprotective agents often contain dimethyl sulfoxide and serum.To avoid the toxic side effects of dimethyl sulfoxide,the complexity of serum components and immune responses,although some finished cryoprotective agents have been marketed,they are faced with many difficulties such as high cost and limited application.Therefore,there is an urgent need to develop a cryoprotective agent with clear components and the ability to solve the above problems.OBJECTIVE:To evaluate the effects of a novel cryoprotectant on cryopreservation efficiency of different tissue and cell sources.METHODS:By applying the novel cryoprotectant as an experimental group with the commercially available and widely used cryoprotectant(control group)to umbilical cord Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,NK and CIK cells,comparative analyses were conducted in terms of cell morphology,number,viability,surface markers,differentiation potential,and cell-killing toxicity assay before cryopreservation and after resuscitation thawing.We confirmed the cryopreservation effect of the new cryoprotectant and its potential application value.RESULTS AND CONCLUSION:(1)The novel cryoprotectant facilitated the normal growth of cryopreserved Wharton's jelly tissue upon recovery,exhibiting mesenchymal stem cell morphology.No significant differences were observed between the experimental and control groups in terms of cell recovery rate,surface markers,and differentiation potential.(2)There was no significant difference in the number and viability of cells between the experimental group and the control group after cryopreservation of cord blood/peripheral blood mononuclear cells,and the cryo-resuscitated cell numbers and viability of derived NK cells/CIK cells did not show significant difference between the experimental and control groups.(3)For NK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportion of CD56+CD16+cell subpopulations between the experimental group and the control group.For CIK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportions of CD3+CD8+and CD3+CD56+cell subpopulations between the experimental group and the control group.(4)In terms of cytotoxicity testing,when the effective-target ratio of immune cells and melanoma cell line Mel624 was 20:1,whether it was NK cells/CIK cells derived from cord blood or peripheral blood mononuclear cells,there was no significant difference in the tumoricidal activity of cells between the experimental group and the control group.These findings suggest that the novel cryoprotectant can replace existing commercially available and widely used cryoprotectants,and is applicable to Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,as well as NK and CIK cells,providing a solid technical foundation for the scaling,standardization,and commercialization of universal cryoprotectants.

BACKGROUND:Long non-coding RNA(LncRNA)plays an important role in nervous system development and neurological diseases.Previous studies by the research team have demonstrated that human umbilical cord mesenchymal stem cells overexpressing erythropoietin(EPO-MSCs)under ischemic and hypoxic conditions have better neuroprotective functions and significantly activate the expression of LncRNA XIST.Research suggests that XIST is related to the pathogenesis of hypoxic-ischemic encephalopathy,but the role and mechanism of its regulation by EPO-MSCs in protecting ischemic-hypoxic neurons remain unclear.OBJECTIVE:To explore the new mechanism by which LncRNA XIST,in response to EPO-MSC signaling,affects the apoptosis of ischemic-hypoxic SH-SY5Y cells.METHODS:(1)SH-SY5Y cell lines with knockdown of LncRNA XIST(sh-XIST)and negative control(NC-XIST)were constructed through lentiviral transfection.Oxygen-glucose deprivation was used to induce ischemic-hypoxic injury in the cells.Transwell chambers were used to create a non-contact co-culture system with EPO-MSCs,sh-XIST,and NC-XIST ischemic-hypoxic SH-SY5Y cells.Cell proliferation ability was detected using the CCK-8 assay.Cell migration ability was assessed using the scratch assay,and cell apoptosis was measured by flow cytometry.(2)RNA-seq bioinformatics analysis was performed to screen for differentially expressed genes and pathways between sh-XIST and NC-XIST cell lines.Dual-luciferase experiments were used to verify the relationship between miR-124-3p and the target genes XIST and GRIN1.qRT-PCR was conducted to validate the expression levels of downstream miR-124-3p and GRIN1 genes.(3)miR-124-3p inhibitors and mimics were added to verify phenotypic changes in SH-SY5Y cells after ischemic-hypoxic injury and co-culture with EPO-MSCs.RESULTS AND CONCLUSION:(1)Compared with the NC-XIST group,SH-SY5Y cells in the sh-XIST group showed reduced proliferation and migration abilities and increased apoptosis after ischemic-hypoxic injury and co-culture with EPO-MSCs.(2)Dual-luciferase experiments showed that miR-124-3p interacted with the target gene XIST.SH-SY5Y cells transfected with miR-124-3p mimics and co-cultured with EPO-MSCs showed decreased apoptosis after ischemic-hypoxic injury,while SH-SY5Y cells transfected with miR-124-3p inhibitors showed increased apoptosis after co-culture with EPO-MSCs.(3)Transcriptomic sequencing and bioinformatics analysis of sh-XIST revealed significant downregulation of the neuroactive ligand-receptor pathway and the key receptor gene GRIN1 for central nervous system development.(4)Dual-luciferase experiments showed that miR-124-3p interacted with GRIN1.GRIN1 expression was significantly downregulated in the sh-XIST group after ischemic-hypoxic injury compared with the NC-XIST group.These findings indicate that LncRNA XIST promotes GRIN1 expression by upregulating miR-124-3p,thereby reducing cell apoptosis after ischemic-hypoxic injury and co-culture with EPO-MSCs and enhancing proliferation and migration.sh-XIST can block this protective function.

Objective:To explore the predictive factors for extubation in the operating room after single lung transplantation without the assistance of extracorporeal membrane oxygenation (ECMO) in adult patients with end-stage interstitial lung disease (ILD), as well as their short-term (1-year postoperative) prognosis.Methods:A retrospective analysis was conducted on the clinical data of 78 adult ILD recipients who underwent single lung transplantation without ECMO assistance at the First Affiliated Hospital of Guangzhou Medical University from June 2018 to June 2023. Based on whether extubation was completed in the operating room (OR), patients were divided into the OR group (19 cases) and ICU group (59 cases). Baseline characteristics of donors and recipients, as well as intraoperative events, were compared between the two groups. Univariate logistic regression analysis was used to identify potential predictors, and variables with P<0.2 were included in multivariate logistic regression to determine independent predictors for OR extubation. Receiver operating characteristic (ROC) curves were plotted to evaluate predictive performance. The Kaplan-Meier method was used to analyze survival, and short-term prognosis between groups was compared. Results:The rate of OR extubation after single lung transplantation in ILD recipients was 24%(19/78). Compared with the ICU group, the OR group had shorter operation times, lower fluid volumes, reduced transfusions of red blood cells and plasma, less intraoperative bleeding, and lower lactate levels 15 minutes after pulmonary artery reperfusion (all P<0.05). Univariate logistic regression analysis identified the following factors as significantly associated with OR extubation: recipient age ( P=0.100), operative time ( P=0.001), fluid infusion volume ( P=0.005), red blood cell transfusion volume ( P=0.037), plasma transfusion volume ( P=0.039), blood loss ( P=0.004), oxygenation index at 15 minutes after reperfusion ( P=0.174), and blood lactate at 15 minutes after reperfusion ( P=0.041). Multivariate analysis revealed that intraoperative blood loss was an independent predictor of OR extubation ( OR=0.993, 95% CI: 0.986 - 0.999, P=0.026). ROC curve analysis showed that blood loss had an area under the curve (AUC) of 0.822 in predicting OR extubation, with a sensitivity of 64.4% and specificity of 89.5%. Postoperatively, patients in the OR group had significantly shorter durations of mechanical ventilation [0 vs 5 (3,11) days, P<0.001], ICU stay [7(4,8) vs 9(6,20) days, P=0.012], and overall postoperative hospitalization [19 (15,23) vs 25 (19,39) days, P=0.015]. Within one year after surgery, 2 patients (11%) in the OR group and 19 patients (32%) in the ICU group had died, but the difference in 1-year survival rates between the two groups was not statistically significant. Conclusions:Intraoperative blood loss is an independent predictor of extubation in the operating room. Early extubation in non-ECMO-assisted single lung transplantation for ILD patients is associated with improved short-term outcomes.

The success of"New Medical Sciences"in higher education requires effective tools in evalua-ting students'performance in courses.Previously,we reported a teamwork(T),critique(C)and ap-preciation(A)(TCA)ideological and political model,a teaching model widely applied in Basic Medical courses.TCA is an abbreviation derived from Tricarboxylic Acid Cycle in Biochemistry as an analogy for nurturing the abilities of thinking and teamwork(T),critique(C)and appreciation(A),which hope-fully could provide students with moral norms for cognition,science and life.This paper further explores the tools to assess the educational outcomes of the TCA model,by which teachers can collect feedback and reflect on teaching quality and effectiveness.Addressing the challenges of individual differences in large classes,fragmented learning feedback,and the difficulty of measuring meta-cognition in educational evaluation,this study employs strategies of value-added assessment,matrix assessment and norm-transfer-able assessment to evaluate the TCA abilities from the aspects of thinking quality,thinking creativity,co-operation ability,iterative thinking,dialectical thinking and job responsibility.By modifying/using 18 e-valuation tools in Education and Psychology,we have established a rubric system composed of 30 primary indicators(with 11 newly designed,10 partly modified and 9 directly adopted),along with 49 secondary indicators and 98 tertiary indicators to enhance the feasibility of the evaluation process.This rubric sys-tem was applied to Biochemistry teaching among the five-year-program undergraduates at Air Force Medi-cal University.Specifically,thinking and teamwork are evaluated by creative works from"the magic bio-chemical-circle",while critiques are assessed in large classes under the guidance of basic and clinical teachers,coupled with appreciation measured by job responsibility in a task-driven virtual reality(VR)project.The results indicate that Biochemistry teaching not only accumulates knowledge in students,but also achieves the goals in nurturing values and cognition.The inclusion of creative performance evalua-tion,cooperative learning and clinical case studies,can enhance students'interpersonal skills,coopera-tion,quality of thinking,creative thinking,iterative thinking and dialectical thinking to varying degrees.TCA-based Biochemistry teaching has a long-lasting impact on character education,and is capable of in-ducing positive long-term changes in students'cognition and lifestyle.Taken together,with the help of this rubric system,teachers can promptly acknowledge the effectiveness of their teaching,thereby facilita-ting their teaching strategies.

Previous studies have shown that the diversity and composition of intestinal microbiota are related to the effect of tumor immunotherapy,but the mechanism of intestinal microbiota affecting tumor immunotherapy resistance has rarely been sum-marized.This article not only expounds the current clinical sta-tus of tumor immunotherapy resistance,but also summarizes the correlation and regulatory mechanism between the composition and homeostasis of intestinal microbiota and drug resistance to different types of tumor immunotherapy,so as to provide a refer-ence for the study of potential targets for improving tumor immu-notherapy resistance based on intestinal microbiota.