BACKGROUND:The mesenchymal stem cell secretome contains bioactive substances,cytokines,and growth factors.Three-dimensional cell culture can regulate the secretion of these components,potentially enhancing the ability to promote injury repair.OBJECTIVE:To investigate the repair effect of three-dimensional cultured human umbilical cord mesenchymal stem cell secretome on skin injuries in mice.METHODS:Human umbilical cord mesenchymal stem cells were cultured in conventional two-dimensional culture dishes and 96-well U-bottom cell culture plates,from which their secretory components were subsequently collected.The expression of skin damage repair related secretory factors in umbilical cord mesenchymal stem cells was analyzed using RT-qPCR.The protein expression level of skin damage repair related factors in umbilical cord mesenchymal stem cell secretome was detected using enzyme-linked immunosorbent assay.The potential of human umbilical cord mesenchymal stem cell secretome to repair vascular injuries was evaluated using an immortalized human umbilical vein endothelial cell migration model.A mouse skin injury model was established,and the human umbilical cord mesenchymal stem cell secretome was injected subcutaneously.Repair effects on skin injury were assessed through wound healing rates and histopathological analysis.RESULTS AND CONCLUSION:(1)After three days of cultivation,human umbilical cord mesenchymal stem cells cultured in two dimensions exhibited a fibroblast-like,swirling growth pattern,whereas three-dimensional culture led to the formation of uniform microspheres.(2)Compared with two-dimensional culture,three-dimensional culture significantly increased the mRNA expression of transforming growth factor β and basic fibroblast growth factor in human umbilical cord mesenchymal stem cells.(3)Compared with two-dimensional culture,three-dimensional cultured human umbilical cord mesenchymal stem cell secretome significantly enhanced the protein expression of vascular endothelial growth factor,interleukin-10,and granulocyte-macrophage colony-stimulating factor in the human umbilical cord mesenchymal stem cell secretome.(4)Compared with two-dimensional culture,three-dimensional cultured human umbilical cord mesenchymal stem cell secretome significantly promoted the migration of immortalized human umbilical cord mesenchymal stem cells.(5)Compared with the untreated control group and the two-dimensional cultured human umbilical cord mesenchymal stem cell secretome,the three-dimensional cultured human umbilical cord mesenchymal stem cell secretome can significantly accelerate the skin wound healing rate and wound skin structure remodeling in mice.These results indicate that three-dimensional culture can enhance the expression of paracrine factors of human umbilical cord mesenchymal stem cells,and their secretome can significantly promote the repair of mouse skin damage.

BACKGROUND:The mesenchymal stem cell secretome contains bioactive substances,cytokines,and growth factors.Three-dimensional cell culture can regulate the secretion of these components,potentially enhancing the ability to promote injury repair.OBJECTIVE:To investigate the repair effect of three-dimensional cultured human umbilical cord mesenchymal stem cell secretome on skin injuries in mice.METHODS:Human umbilical cord mesenchymal stem cells were cultured in conventional two-dimensional culture dishes and 96-well U-bottom cell culture plates,from which their secretory components were subsequently collected.The expression of skin damage repair related secretory factors in umbilical cord mesenchymal stem cells was analyzed using RT-qPCR.The protein expression level of skin damage repair related factors in umbilical cord mesenchymal stem cell secretome was detected using enzyme-linked immunosorbent assay.The potential of human umbilical cord mesenchymal stem cell secretome to repair vascular injuries was evaluated using an immortalized human umbilical vein endothelial cell migration model.A mouse skin injury model was established,and the human umbilical cord mesenchymal stem cell secretome was injected subcutaneously.Repair effects on skin injury were assessed through wound healing rates and histopathological analysis.RESULTS AND CONCLUSION:(1)After three days of cultivation,human umbilical cord mesenchymal stem cells cultured in two dimensions exhibited a fibroblast-like,swirling growth pattern,whereas three-dimensional culture led to the formation of uniform microspheres.(2)Compared with two-dimensional culture,three-dimensional culture significantly increased the mRNA expression of transforming growth factor β and basic fibroblast growth factor in human umbilical cord mesenchymal stem cells.(3)Compared with two-dimensional culture,three-dimensional cultured human umbilical cord mesenchymal stem cell secretome significantly enhanced the protein expression of vascular endothelial growth factor,interleukin-10,and granulocyte-macrophage colony-stimulating factor in the human umbilical cord mesenchymal stem cell secretome.(4)Compared with two-dimensional culture,three-dimensional cultured human umbilical cord mesenchymal stem cell secretome significantly promoted the migration of immortalized human umbilical cord mesenchymal stem cells.(5)Compared with the untreated control group and the two-dimensional cultured human umbilical cord mesenchymal stem cell secretome,the three-dimensional cultured human umbilical cord mesenchymal stem cell secretome can significantly accelerate the skin wound healing rate and wound skin structure remodeling in mice.These results indicate that three-dimensional culture can enhance the expression of paracrine factors of human umbilical cord mesenchymal stem cells,and their secretome can significantly promote the repair of mouse skin damage.

ObjectiveTo investigate the molecular mechanism by which Yifei Xuanfei Jiangzhuo prescription regulates the nuclear factor-κB （NF-κB）/NOD-like receptor protein 3 （NLRP3） signaling pathway to improve neuronal function in vascular dementia （VaD） rats. MethodsA VaD model was established by intermittently clamping the bilateral common carotid arteries （CCA） combined with bilateral vascular occlusion （2-VO）. Eighty-four SD rats were randomly divided into a blank group， sham group， model group， piracetam group （0.2 g·kg^-1）， and low-， medium-， and high-dose Yifei Xuanfei Jiangzhuo prescription groups （6.09， 12.18， and 24.36 g·kg^-1）. Drug administration began on day 7 after surgery， once daily for 28 consecutive days. Behavioral experiments were used to evaluate learning and spatial memory. Hematoxylin-eosin （HE） staining was applied to observe pathological morphological changes in the CA1 region of the hippocampus. Transmission electron microscopy was used to examine the ultrastructure of hippocampal neurons. Terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） was used to detect neuronal apoptosis in the CA1 region. Immunohistochemistry was performed to determine the positive expression rate of neuronal nuclear antigen （NeuN）. Immunofluorescence single staining was used to assess nuclear expression of NF-κB p65 in brain tissue. Western blot was used to detect the protein expression levels of inhibitor of κB kinase （IKK）， NF-κB p65， NLRP3， Caspase-1， apoptosis-associated speck-like protein （ASC）， and interleukin-1β （IL-1β）. ResultsCompared with the blank group， the model group showed a significant reduction in platform-crossing frequency （P0.01）， aggravated hippocampal injury， a significant increase in neuronal apoptosis （P0.05）， decreased NeuN positivity in the CA1 region （P0.05）， increased nuclear expression of NF-κB p65 （P0.05）， and significantly elevated expression of p-IKK， p-NF-κB p65， NLRP3， cleaved Caspase-1， ASC， and cleaved IL-1β （P0.05）. Compared with the model group， all drug-treated groups improved learning and spatial memory in VaD rats， alleviated hippocampal pathological injury and neuronal apoptosis， and protected neuronal ultrastructure. Yifei Xuanfei Jiangzhuo prescription at doses of 12.18 and 24.36 g·kg^-1 reduced hippocampal expression levels of p-IKK， p-NF-κB p65， NLRP3， Caspase-1， ASC， and cleaved IL-1β in VaD rats （P0.05）， showing dose-dependent inhibition of the NF-κB/NLRP3 signaling pathway. ConclusionYifei Xuanfei Jiangzhuo prescription may exert neuroprotective effects by regulating the NF-κB/NLRP3 signaling pathway， thereby reducing neuroinflammation and inhibiting hippocampal neuronal apoptosis.

ObjectiveTo investigate the molecular mechanism by which Yifei Xuanfei Jiangzhuo prescription regulates the nuclear factor-κB （NF-κB）/NOD-like receptor protein 3 （NLRP3） signaling pathway to improve neuronal function in vascular dementia （VaD） rats. MethodsA VaD model was established by intermittently clamping the bilateral common carotid arteries （CCA） combined with bilateral vascular occlusion （2-VO）. Eighty-four SD rats were randomly divided into a blank group， sham group， model group， piracetam group （0.2 g·kg^-1）， and low-， medium-， and high-dose Yifei Xuanfei Jiangzhuo prescription groups （6.09， 12.18， and 24.36 g·kg^-1）. Drug administration began on day 7 after surgery， once daily for 28 consecutive days. Behavioral experiments were used to evaluate learning and spatial memory. Hematoxylin-eosin （HE） staining was applied to observe pathological morphological changes in the CA1 region of the hippocampus. Transmission electron microscopy was used to examine the ultrastructure of hippocampal neurons. Terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） was used to detect neuronal apoptosis in the CA1 region. Immunohistochemistry was performed to determine the positive expression rate of neuronal nuclear antigen （NeuN）. Immunofluorescence single staining was used to assess nuclear expression of NF-κB p65 in brain tissue. Western blot was used to detect the protein expression levels of inhibitor of κB kinase （IKK）， NF-κB p65， NLRP3， Caspase-1， apoptosis-associated speck-like protein （ASC）， and interleukin-1β （IL-1β）. ResultsCompared with the blank group， the model group showed a significant reduction in platform-crossing frequency （P0.01）， aggravated hippocampal injury， a significant increase in neuronal apoptosis （P0.05）， decreased NeuN positivity in the CA1 region （P0.05）， increased nuclear expression of NF-κB p65 （P0.05）， and significantly elevated expression of p-IKK， p-NF-κB p65， NLRP3， cleaved Caspase-1， ASC， and cleaved IL-1β （P0.05）. Compared with the model group， all drug-treated groups improved learning and spatial memory in VaD rats， alleviated hippocampal pathological injury and neuronal apoptosis， and protected neuronal ultrastructure. Yifei Xuanfei Jiangzhuo prescription at doses of 12.18 and 24.36 g·kg^-1 reduced hippocampal expression levels of p-IKK， p-NF-κB p65， NLRP3， Caspase-1， ASC， and cleaved IL-1β in VaD rats （P0.05）， showing dose-dependent inhibition of the NF-κB/NLRP3 signaling pathway. ConclusionYifei Xuanfei Jiangzhuo prescription may exert neuroprotective effects by regulating the NF-κB/NLRP3 signaling pathway， thereby reducing neuroinflammation and inhibiting hippocampal neuronal apoptosis.

ObjectiveIn the context of the digital transformation of education, this study aims to construct a triadic interactive teaching model for surgical animal surgery in clinical medicine using modern information technology. It explores the effectiveness of different teaching methods in improving students' practical skills, aseptic awareness, and teamwork abilities, providing a reference for the reform of clinical practice education. MethodsA quasi-experimental research design was adopted. A total of 80 students from the eight-year clinical medicine program at Nanjing University were selected, including the Class of 2020 (control group, n=40) and the Class of 2021 (experimental group, n=40). The control group received traditional teaching methods, while the experimental group implemented the "Teacher-Student-AI" triadic interactive teaching model. This model utilized a smart teaching platform for personalized pre-class preparation , as well as data-driven post-class review and feedback throughout the entire teaching process. The "assessment indicators and scoring criteria for the surgical animal surgery course" were used to evaluate teaching effectiveness, with independent samples t-tests used for statistical analysis. ResultsPre-course assessments revealed no statistically significant differences in baseline theoretical knowledge or practical skills between the two groups (P>0.05). Upon completion of the course, the experimental group achieved higher scores than the control group across three key dimensions: practical skills (47.98±1.34 vs 46.92±2.51, P=0.022), aseptic awareness (17.84±1.16 vs 16.94±2.29, P=0.029), and teamwork (16.82±1.44 vs 15.95±1.22, P=0.004). However, no statistically significant difference was observed in the scores for humane care awareness between the two groups (8.24±0.70 vs 8.16±0.53, P=0.589). ConclusionThe AI-based triadic interactive teaching model can, to some extent, address the limitations of traditional surgical animal surgery education. It plays a positive role in enhancing medical students' surgical skills, aseptic awareness, and collaborative abilities. This model facilitates the transition from traditional to personalized teaching and offers a practical framework for the digital reform of clinical practice education.

Objective:To study genotype analysis and influence factors analysis of hepatitis B virus(HBV)deoxyribonucleic acid(DNA)positive samples of negative hepatitis B surface antigen(HBsAg)donors in blood donors without compensation in Nan'an area.Methods:A total of 62,000 HBsAg fast-screened blood donor samples from January 2021 to June 2024 were selected as the study objects.HBV infection,viral load and genotype of HBV DNA positive samples,single factor and influencing factor of HBV DNA positive samples were detected and analyzed.Results:Among the 62000 HBsAg fast-screened blood donor samples,the results were negative by HBsAg screening,moreover,285 cases(0.46％)were HBsAg positive and 61715 cases(99.54％)were negative.NAT positive 180 cases and 118 cases were positive for HBV DNA.The HBV DNA positive rate of HBsAg negative donors was 0.19％(118/62000).HBV DNA of the viral load of positive specimens with＜20 IU/mL was higher than those with 20 to 99 IU/mL and ≥ 100 IU/mL(P＜0.05).HBV DNA of C style of the genotype of positive specimens was higher than B style and unsamples(P＜0.05).Univariate analysis found that HBV DNA positivity was mainly related to the history of intravenous drug use and risky sexual behavior,as well as the history of hepatitis B vaccination and blood transfusion(P＜0.05).According to the Logistic regression analysis,history of intravenous drug use and risky sexual behavior,and blood transfusion were risk factors for positive HBV DNA(OR＞1,P＜0.05),vaccination of hepatitis B vaccine was a protective factor for HBV DNA positivity(OR＜1,P＜0.05).Conclusion:The proportion of HBsAg in Nan'an area is relatively high,but there are still a small number of positive samples,the main genotype was type C,and history of intravenous drug use and risky risk behaviors,as well as history of blood transfusion were all risk factors for positive HBV DNA,vaccination of hepatitis B vaccine is protective factor for HBV DNA positivity.

Objective:To investigate the MRI characteristics of ductal lesions with microcalcifications detected by mammography,and to evaluate the value of MRI in the differential diagnosis of benign and malignant ductal lesions.Methods:The clinical data of 112 patients with pathologically confirmed ductal lesions with microcalcifications detected by mammography in our hospital from January 2022 to June 2024 were retrospectively analyzed.All patients underwent mammography and MRI examinations.MRI morphological features,dynamic enhancement characteristics,diffusion-weighted imaging(DWI)findings,and apparent diffusion coefficient(ADC)values were analyzed and compared with pathological results.The diagnostic performance of various MRI features was analyzed using ROC curves.Results:Among the 112 patients,there were 47 benign lesions and 65 malignant lesions.Malignant lesions were more likely to present with irregular morphology,heterogeneous T2 signal,lobulation,and spiculation(P＜0.05),while benign lesions were more likely to present with round or oval shapes and homogeneous T2 signal(P＜0.05).Malignant lesions predominantly showed fast rise-fast washout(Washout type)enhancement curves(58.5％),while benign lesions predominantly showed persistent rise(Progressive type)enhancement curves(55.3％).The ADC values of malignant lesions(0.92±0.21× 10-3 mm2/s)were significantly lower than those of benign lesions(1.45±0.28× 10-3 mm2/s)(P＜0.001).The combined application of morphological features,dynamic enhancement curves,and ADC values had the highest diagnostic performance,with a sensitivity of 92.3％,specificity of 89.4％,and accuracy of 91.1％.Conclusion:Multi-parameter MRI can comprehensively evaluate ductal lesions with microcalcifications detected by mammography.The combined application of morphological features,dynamic enhancement curve types,and ADC values can effectively improve the accuracy of differential diagnosis between benign and malignant lesions,providing important basis for clinical treatment decisions.

Objective:To investigate the relationship between Ig class switch recombination(CSR)and mismatch repair(MMR)protein,microsatellite phenotype in extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue(MALT lymphoma).Methods:Forty cases of MALT lymphoma archived in the Department of Pathology,Jiading District Central Hospital,Shanghai University of Medicine & Health Sciences were selected as the observation group,and twenty cases of benign lymphoid tissue hyperplasia were as the control group.The expressions of IgG,IgM,IgD,and IgA in both groups were detected by immunohistochemical double staining,and MMR proteins including MLH1,MSH2,MSH6,and PMS2 in both groups were detected by immunohistochemistry.Multiplex fluorescence PCR capillary electrophoresis was used to detect microsatellite phenotype in tumor and adjacent tissues of the experimental group.Results:In the observation group,the proportions of single Ig heavy chain expression(mode Ⅰ),negative expression(mode Ⅱ),and multiple expression(mode Ⅲ)were 65％(26/40),27.5％(11/40),and 7.5％(3/40),respectively,while in the control group were 0(0/20),5％(1/20),and 95％(19/20).The proportion of Ig heavy chain expression mode Ⅰ+Ⅱ in the observation group was 92.5％,which was significantly higher than 5％in the control group(P＜0.0 1).In the observation group,partial deletion of MMR protein was observed in 3 cases(7.5％),including 2 cases of MSH6 deletion and 1 case of both MSH6 and PMS2 deletion.In the control group,there was 1 case(5％)with PMS2 deletion.There was no significant difference in the deletion rate of MMR protein between the two groups(P＞0.05).A total of 5 cases of microsatellite instability(MSI)were detected in the observation group,including 1 case of low-frequency MSI(MSI-L),4 cases of high-frequency MSI(MSI-H),and 2 cases of MSI-H with MSH6 deletion.When the loss expression of MSI-H or MMR protein was counted as a positive result,the MSI-H rate detected by PCR capillary electrophoresis was 10％(4/40),which was slightly higher than the MMR protein deletion rate detected by immunohistochemistry(7.5％,3/40),but there was no statistically significant difference between the two groups(P＞0.05).The MMR protein deletion rates among the Ig heavy chain protein expression mode Ⅰ,mode Ⅱ,and mode Ⅲ groups were 0(0/26),18.2％(2/11),and 33.3％(1/3),respectively.There was a statistically significant difference in the constituent ratios among the three groups(P＜0.05).The MMR protein deletion rates among the MSS,MSI-L,and MSI-H groups were 2.9％(1/35),0(0/1),and 50％(2/4),respectively.There was a statistically significant difference in the constituent ratios among the three groups(P＜0.05).MMR protein deficiency was positively correlated with Ig heavy chain expression pattern and MSI(r=0.41,P＜0.05;r=0.48,P＜0.05),but Ig heavy chain expression pattern was not correlated with MSI(r=0.02,P＞0.05).Conclusion:Ig heavy chain CSR detection is helpful for the differential diagnosis of MALT lymphoma.Low frequency MMR protein deletion and MSI-H phenotype exist in MALT lymphoma,which may be of certain value for the study of its occurrence,development and clinical treatment.

Objective To explore the effect of curcumin(CUR)on oxidative damage of keratinocytes induced by ultraviolet B(UVB)irradiation through Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB)/nucleotide-binding oligomerization domain-containing protein 3(NLRP3)signaling pathway.Methods Human keratinocytes of HaCaT were cultured normally in vitro,and the keratinocyte oxidative damage model was established by the irradiation of 57 mJ/cm2 UVB.The cells with normal culture were as the control group,the cells treated after modeling were as the UVB group,the cells treated with 5 μmol/L CUR after modeling were as the CUR group,the cells treated with 100 μg/L TLR4 inhibitor of TAK-242 after modeling were as the TAK-242 group,and the cells treated with 5 μmol/L CUR and 100 nmol/L TLR4 activator of lipopolysaccharide(LPS)were as the CUR+LPS group.qRT-PCR was applied to detect the relative expression levels of TLR4,NF-κB,and NLRP3 mRNAs of cells in each group.CCK-8 was applied to detect the cell proliferation in each group.The relative content of reactive oxygen species(ROS),the viabilities of superoxide dismutase(SOD)and catalase(CAT),and the concentrations of glutathione(MDA)and glutathione(GSH)of cells in each group were detected by fluorescence assay according to the kit instruction.ELISA kit was used to detect the expression of inflammatory factors of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)of cells in each group.Flow cytometry was applied to detect the cell apoptosis in each group.Western blot was applied to detect the expression of proliferation related protein of proliferating cell nuclear antigen(PCNA),apoptosis related proteins[B-cell lymphoma-2(Bcl-2)and Bcl-2-associated X protein(Bax)],and TLR4/NF-κB/NLRP3 signaling pathway related proteins(TLR4,NF-κB and NLRP3)of cells in each group.Results Compared with the Control group,the cell survival rate,the expression levels of PCNA and Bcl-2 proteins,the viabilities of SOD and CAT,and the GSH concentration in the UVB group decreased,while the apoptosis rate,the level of Bax protein,the relative content of ROS,the concentration of MDA,the levels of TNF-α and IL-1β,and the mRNA and protein levels of TLR4,NF-κB and NLRP3 increased(P<0.05).Compared with the UVB group,the cell survival rate,the expression levels of PCNA and Bcl-2 proteins,the viabilities of SOD and CAT,and the GSH concentration in the TAK-242 group and CUR group increased,while the apoptosis rate,the level of Bax protein,the relative content of ROS,the concentration of MDA,the levels of TNF-α and IL-1β,and the mRNA and protein levels of TLR4,NF-κB and NLRP3 decreased(P<0.05).Compared with the CUR group,the cell survival rate,the expression levels of PCNA and Bcl-2 proteins,the viabilities of SOD and CAT,and the GSH concentration in the CUR+LPS group decreased,while the apoptosis rate,the level of Bax protein,the relative content of ROS,the concentration of MDA,the levels of TNF-α and IL-1β,and the mRNA and protein levels of TLR4,NF-κB and NLRP3 increased(P<0.05).Conclusion CUR can increase the antioxidant stress level of keratinocytes,alleviate inflammatory response,promote cell proliferation,and improve cell damage caused by UVB irradiation,which may be related to the inhibition of TLR4/NF-κB/NLRP3 signaling pathway.

BACKGROUND:Allogeneic hematopoietic stem cell transplantation is currently the most effective method for the radical treatment of thalassemia major,but only half of patients can find compatible bone marrow or peripheral blood stem cells.Sib-derived umbilical cord blood stem cells have different characteristics from bone marrow and peripheral blood stem cells,and are a potential alternative source of hematopoietic stem cells for transplantation in patients with thalassemia major.OBJECTIVE:To investigate the therapeutic effect of human leukocyte antigen matched sibling fresh umbilical cord blood transplantation in the treatment of β-thalassemia major in children.METHODS:Forty-eight children with β-thalassemia major,including 28 males and 20 females,with a median age of 4 years old,were selected from Nanfang Hospital of Southern Medical University from June 2010 to June 2020.All of them received fresh cord blood transplantation from human leukocyte antigen matched sibling.Transplantation conditioning adopted a myeloablative regiment without anti-thymocyte globulin.A combination of cyclosporine A and mycophenolate mofetil with or without short-range methotrexate was administered for graft-versus-host disease.RESULTS AND CONCLUSION:(1)The median infused doses of total nucleated cells and CD34+cells were 8.17×107/kg and 2.40×105/kg,respectively in 48 children.The median follow-up time after cord blood transplantation was 98 months,and 44 cases were successfully engrafted.The median time to neutrophil and platelet engraftment was 28 and 31 days,respectively.Among them,37 cases were found to be donor-type complete chimerism detected as evidence of implantation after transplantation,7 cases were found to be stable mixed chimerism.(2)Among the 44 children with successful implantation,four patients developed acute graft-versus-host disease,and were scored as grade Ⅰ(n=2)and grade Ⅱ(n=2).All the affected organs were skin,and no chronic graft-versus-host disease occurred.(3)After umbilical cord blood transplantation,cytomegalovirus infection and activation occurred in 5 of the 48 cases,sepsis in 12 cases,invasive fungal disease in 3 cases,stomatitis in 21 cases,hemorrhagic cystitis in 8 cases,and hepatic vein occlusion in 1 case.(4)Among 48 children,47 patients survived;1 died of severe pneumonia combined with acute heart failure 28 days after transplantation;43 survived without disease;3 had primary implantation failure,and 1 had pancytopenia after transplantation.The 5-year probabilities of overall survival and disease-free survival were 98％and 89％,respectively.The cumulative incidence of transplant-related deaths at 1 year was 2.1％.(5)The above results indicate that human leukocyte antigen matched sibling fresh umbilical cord blood transplantation is effective in the treatment of β-thalassemia major in children with a low incidence of graft-versus-host disease.