Ultrasound technology has been applied in the biomedical field,particularly in drug delivery and cell processing.In this study,the effects of different ultrasound power levels(40 W to 100 W)and time durations(1 min,5 min,or 5 min discontinuously)on the morphology of human red blood cells(hRBCs)membranes were systematically investigated using cryo-electron tomography(Cryo-ET).The hRBCs membranes were firstly subjected to ultrasound at power levels of 40 W and 60 W for 5 min each.Cryo-ET observations revealed minimal morphological changes in the hRBCs membranes following the 40 W treatment,with the membrane structure remaining relatively intact and only minor undulations appearing on the membrane surface.These undulations might result from the mild mechanical stress induced by ultrasound,which was insufficient to disrupt the overall membrane structure.At power of 60 W,the hRBCs membranes largely preserved their structural integrity.When the ultrasonic power was increased to 80 W,the structural damage to the hRBCs membranes became more severe.Cryo-ET images showed irregular ruptures and larger pores on the membrane surface,indicating a significant compromise in membrane integrity.At ultrasound power of 100 W,the hRBCs membranes were completely disrupted,resulting in the formation of numerous membrane fragments,and a complete loss of membrane continuity.To further explore the effects of ultrasound duration on erythrocyte membrane morphology,the ultrasonic power was fixed at 100 W and the impacts of varying treatment durations(1 min,5 min,and intermittent ultrasound)on the membrane structure were systematically investigated.After 1 min of ultrasonic treatment,Cryo-ET images showed minimal changes in erythrocyte membrane morphology.Although some small pores and undulations appeared on the membrane surface,the overall structure remained relatively intact.As the ultrasound duration extended to 5 min,the degree of membrane damage increased significantly.Cryo-ET images revealed extensive rupture and detachment of the membrane,with continuity being severely compromised.As to treatment alternating 1 min of ultrasound with 1 min of rest,for a total of 5 min of ultrasound exposure,Cryo-ET observations showed the integrity of the membrane-cytoskeleton attachment remained.Under intermittent ultrasound treatment,although some pores and ruptures were observed on the membrane surface,the overall structure remained more intact compared to continuous ultrasonic treatment.This preservation might be due to the intermittent treatment providing buffer periods for the membrane,allowing partial recovery after mechanical stress,thereby reducing the cumulative damage caused by continuous ultrasound.This work provided experimental basis for further understanding of mechanism of ultrasound induced change of cell membrane and cytoskeleton.

Aim To investigate the effect of chrysoeriol on pulmonary vascular remodeling in pulmonary hyper-tension by animal experiments combined with cell ex-periments,and to explore its potential therapeutic tar-gets by network pharmacology.Methods The target of chrysoeriol was collected in Targetnet,SEA and SwissTargetPrediction database.Pulmonary arterial hy-pertension(PAH)targets were collected in the Dis-GeNET and GeneCards databases,and PPI network map was drawn in the STRING database,and key tar-gets were screened.The GO and KEGG pathway en-richment analysis was carried out through DAVID data-base and Weishengxing platform.AutoDock software was used for molecular docking of key core targets.The PAH model of rats was constructed,and the pulmo-nary hemodynamics and vascular remodeling were de-tected by echocardiography,HE and Masson staining.Primary pulmonary smooth muscle cells were extracted,and the effects of drugs on pathway proteins were de-tected in vitro.Results The results of network phar-macology showed that chrysoeriol exerted therapeutic effects on pulmonary hypertension by affecting key tar-gets such as AKT1,SRC,EGFR,MMP9 and gsk3 β,and signaling pathways such as EGFR and PI3K-AKT.Molecular docking showed that chrysoeriol had good binding ability with 5 key target genes.Animal experi-ments showed that the pulmonary hemodynamic func-tion of PAH rats was significantly improved after ad-ministration of chrysoeriol.The remodeling of small pulmonary arteries was significantly reduced.Cell ex-periments showed that chrysoeriol could inhibit the ex-pression of proliferation,migration and phenotypic transformation genes.Conclusion Chrysoeriol may play a role in the treatment of pulmonary hypertension through multiple targets.

AIM:This study investigated the potential mechanism of acupuncture regulating adenosine A1 re-ceptor(ADORA1)in the mouse caudate putamen(CPu)to improve neuroplasticity and alleviate inflammatory pain.METHODS:Male C57BL/6 mice were randomly divided into five groups,namely,saline,complete Freund's adjuvant(CFA),acupuncture(MA),acupuncture+ADORA1 shRNA(MA+shRNA),and acupuncture+negative shRNA(MA+NCshRNA)groups.Twenty-one days before modeling,the mice in the MA+shRNA and MA+NCshRNA groups were in-jected with ADORA1 shRNA and control virus into the CPu.Modeling was performed 21 d later by injection of CFA into the right plantar skin of mice in the model and acupuncture groups to induce adjuvant-mediated arthritis.On day 2 after modeling,mice in the acupuncture groups received acupuncture at bilateral"Zusanli"points,30 min per session,once a day,for a total of 7 days.The paw thermal radiation pain threshold was used as an indicator of the effects on pain in the mice.Changes in the protein levels of ADORA1,synaptophysin(SYP),synapsin(SYN)I,SYN II,and brain-derived neurotrophic factor(BDNF)in the CPu were assessed by Western blot,and transmission electron microscopy was used to examine the dendritic structure and synaptic ultrastructure of neurons within the CPu.RESULTS:(1)Compared with the saline group,CFA modeling significantly reduced the thermal radiation pain threshold in mice(P<0.01),as well as the protein levels of ADORA1(P<0.01).Compared with the CFA group,the thermal pain threshold in the MA group in-creased between days 1 and 7 of acupuncture(P<0.05 or P<0.01),and ADORA1 protein levels increased(P<0.01).(2)Compared with the saline group,SYN I,SYN II,and BDNF protein levels were increased in the CFA group(P<0.05 or P<0.01),while relative to the CFA group,the levels of SYN I,SYN II,and BDNF were reduced in the acupuncture group(P<0.05 or P<0.01).No significant changes in SYP protein levels were observed in any of the groups.(3)Golgi staining and Sholl analysis showed that compared with the saline group,there were reductions in the total dendritic length and number of intersection points in the CFA group,while the dendritic spine density increased(P<0.05).Relative to the CFA group,the total dendritic length and the number of intersection points were increased in the MA group,while the dendritic spine density decreased(P<0.05).(4)Transmission electron microscopy revealed that compared with the Sa-line group,the synaptic clefts were narrower in the CFA group,while the density of synaptic vesicles in the presynaptic membrane was increased.Compared with the CFA group,synaptic clefts in the MA group were wider,and synaptic vesi-cle densities in the presynaptic membrane were reduced.(5)Twenty-one days after viral transfection,the thermal pain threshold in the MA and MA+NCshRNA groups increased from day 1 to day 7 of acupuncture relative to the CFA and MA+shRNA groups(P<0.05 or P<0.01),while the expression of ADORA1 was increased in both the MA and MA+NCshRNA groups(P<0.05 or P<0.01).(6)Compared with the CFA and MA+shRNA groups,the protein expression of SYN II and BDNF in the MA and MA+NCshRNA groups was reduced(P<0.05 or P<0.01).No significant changes in SYN I protein were observed in any of the groups.CONCLUSION:Acupuncture on Zusanli point can alleviate chronic pain in CFA-treated mice,potentially mediated by up-regulation of ADORA1 expression in the CPu brain region,thereby improving neuroplasticity.

Aim To investigate the effect of chrysoeriol on pulmonary vascular remodeling in pulmonary hyper-tension by animal experiments combined with cell ex-periments,and to explore its potential therapeutic tar-gets by network pharmacology.Methods The target of chrysoeriol was collected in Targetnet,SEA and SwissTargetPrediction database.Pulmonary arterial hy-pertension(PAH)targets were collected in the Dis-GeNET and GeneCards databases,and PPI network map was drawn in the STRING database,and key tar-gets were screened.The GO and KEGG pathway en-richment analysis was carried out through DAVID data-base and Weishengxing platform.AutoDock software was used for molecular docking of key core targets.The PAH model of rats was constructed,and the pulmo-nary hemodynamics and vascular remodeling were de-tected by echocardiography,HE and Masson staining.Primary pulmonary smooth muscle cells were extracted,and the effects of drugs on pathway proteins were de-tected in vitro.Results The results of network phar-macology showed that chrysoeriol exerted therapeutic effects on pulmonary hypertension by affecting key tar-gets such as AKT1,SRC,EGFR,MMP9 and gsk3 β,and signaling pathways such as EGFR and PI3K-AKT.Molecular docking showed that chrysoeriol had good binding ability with 5 key target genes.Animal experi-ments showed that the pulmonary hemodynamic func-tion of PAH rats was significantly improved after ad-ministration of chrysoeriol.The remodeling of small pulmonary arteries was significantly reduced.Cell ex-periments showed that chrysoeriol could inhibit the ex-pression of proliferation,migration and phenotypic transformation genes.Conclusion Chrysoeriol may play a role in the treatment of pulmonary hypertension through multiple targets.

AIM:This study investigated the potential mechanism of acupuncture regulating adenosine A1 re-ceptor(ADORA1)in the mouse caudate putamen(CPu)to improve neuroplasticity and alleviate inflammatory pain.METHODS:Male C57BL/6 mice were randomly divided into five groups,namely,saline,complete Freund's adjuvant(CFA),acupuncture(MA),acupuncture+ADORA1 shRNA(MA+shRNA),and acupuncture+negative shRNA(MA+NCshRNA)groups.Twenty-one days before modeling,the mice in the MA+shRNA and MA+NCshRNA groups were in-jected with ADORA1 shRNA and control virus into the CPu.Modeling was performed 21 d later by injection of CFA into the right plantar skin of mice in the model and acupuncture groups to induce adjuvant-mediated arthritis.On day 2 after modeling,mice in the acupuncture groups received acupuncture at bilateral"Zusanli"points,30 min per session,once a day,for a total of 7 days.The paw thermal radiation pain threshold was used as an indicator of the effects on pain in the mice.Changes in the protein levels of ADORA1,synaptophysin(SYP),synapsin(SYN)I,SYN II,and brain-derived neurotrophic factor(BDNF)in the CPu were assessed by Western blot,and transmission electron microscopy was used to examine the dendritic structure and synaptic ultrastructure of neurons within the CPu.RESULTS:(1)Compared with the saline group,CFA modeling significantly reduced the thermal radiation pain threshold in mice(P<0.01),as well as the protein levels of ADORA1(P<0.01).Compared with the CFA group,the thermal pain threshold in the MA group in-creased between days 1 and 7 of acupuncture(P<0.05 or P<0.01),and ADORA1 protein levels increased(P<0.01).(2)Compared with the saline group,SYN I,SYN II,and BDNF protein levels were increased in the CFA group(P<0.05 or P<0.01),while relative to the CFA group,the levels of SYN I,SYN II,and BDNF were reduced in the acupuncture group(P<0.05 or P<0.01).No significant changes in SYP protein levels were observed in any of the groups.(3)Golgi staining and Sholl analysis showed that compared with the saline group,there were reductions in the total dendritic length and number of intersection points in the CFA group,while the dendritic spine density increased(P<0.05).Relative to the CFA group,the total dendritic length and the number of intersection points were increased in the MA group,while the dendritic spine density decreased(P<0.05).(4)Transmission electron microscopy revealed that compared with the Sa-line group,the synaptic clefts were narrower in the CFA group,while the density of synaptic vesicles in the presynaptic membrane was increased.Compared with the CFA group,synaptic clefts in the MA group were wider,and synaptic vesi-cle densities in the presynaptic membrane were reduced.(5)Twenty-one days after viral transfection,the thermal pain threshold in the MA and MA+NCshRNA groups increased from day 1 to day 7 of acupuncture relative to the CFA and MA+shRNA groups(P<0.05 or P<0.01),while the expression of ADORA1 was increased in both the MA and MA+NCshRNA groups(P<0.05 or P<0.01).(6)Compared with the CFA and MA+shRNA groups,the protein expression of SYN II and BDNF in the MA and MA+NCshRNA groups was reduced(P<0.05 or P<0.01).No significant changes in SYN I protein were observed in any of the groups.CONCLUSION:Acupuncture on Zusanli point can alleviate chronic pain in CFA-treated mice,potentially mediated by up-regulation of ADORA1 expression in the CPu brain region,thereby improving neuroplasticity.

Objective To investigate the differences in demographic characteristics,reproductive health status,and the distribution of pregnancy-related diseases between couples conceived via assisted reproductive technology(ART)and naturally conceived couples,and to analyze the impact of ART treatment on the incidence of preterm birth(PTB)in singleton and twin and multiple pregnancies.Methods We conducted a retrospective analysis of the maternal and infant cohort data of Jing'an District from 2013 to 2020.Based on the conception method,the subjects were categorized into two groups:the ART group and the natural conception group.Chi-square test was applied to compare baseline characteristics and disease distributions differences between the two groups,and logistic regression models were used to evaluate the association between ART and the PTB risks.A causal mediation model was used to evaluate the mediating effect of twin and multiple pregnancy in the relationship between ART and PTB.Results A total of 117 717 parturients were included,6 265 in the ART group and 111 452 in the natural conception group.Compared with the natural conception group,couples in the ART group were significantly older and had a higher prevalence of reproductive system diseases.The incidences of diabetes and hypertensive disorders during pregnancy in ART parturient were 13.76%and 9.99%,respectively,which were significantly higher than 7.88%and 4.75%in the natural conception group(both P<0.001).The overall PTB rate in the ART group was 14.81%,higher than 5.35%in the natural conceptions group(P<0.001).The PTB rate in ART for singleton pregnancies in the ART group was 6.40%,higher than 4.83%in the natural conception group(P<0.001),while the PTB rate in ART for twin and multiple pregnancies in the ART group was 53.97%,lower than 60.42%in the natural conception group(P<0.05).Mediation analysis showed that 97.99%of the effect of ART on PTB was mediated by twin and multiple pregnancy,with ART increasing the PTB risk by 3.44 times through multiple pregnancy.Conclusion The overall PTB rate of ART recipients is higher than that of natural recipients,but ART does not increase the PTB risk in singleton and twin and multiple pregnancies.Twin and multiple pregnancy is the key mediating factor contributing to PTB in ART-conceived recipients.Compared with naturally conceived couples,ART conception couples own more advanced maternal age,and have higher risks of suffering gestational diabetes,gestational hypertension,and PTB.

Chronic pain has emerged as a prevalent medical challenge in contemporary society.Patients suffering from chronic pain frequently develop comorbid psychological disorders,including anxiety,depression,post-traumatic stress disorder,and various psychiatric syndromes.These psychological complications not only affect patients' pain perception and responses,but may also constitute critical obstacles during pain management interventions.Acupuncture is a long-established clinical practice that has demonstrated remarkable efficacy in alleviating diverse pain types and has shown favorable therapeutic outcomes in ameliorating emotional disturbances such as anxiety and depression.The precise mechanisms underlying acupuncture-induced analgesia and anxiolytic effects,however,remain to be fully elucidated.In this context,it is essential to establish suitable and stable animal models to allow in-depth investigations into the pathogenesis of pain-related emotional disorders and the mechanistic foundations of acupuncture.This article presents a comprehensive review of recent literature regarding the selection of experimental animals,model-establishment method ologies,and behavioral-assessment paradigms pertaining to animal model platforms of chronic pain with comorbid anxiety.We also provide an in-depth discussion of research advancements regarding acupuncture intervention parameters,including needling techniques,acupoint selection,treatment duration,and efficacy evaluation within these animal models.This review proposes comprehensive and reference strategies for constructing preclinical animal models to investigate the mechanisms of acupuncture in managing chronic pain with comorbid anxiety,thus supporting scientific advancements in related research fields.

Objective To investigate the mechanism by which Senegenin(SEN)alleviates microglial inflammatory response through the nuclear factor erythroid 2-related factor 2(Nrf2)/NOD-like receptor protein 3(NLRP3)pathway.Methods BV2 mouse microglia cells were randomly divided into control group,model group,SEN group and MCC950 group.Cells in control group were not treated,and cells in model group were added with 1 μg/ml lipopolysaccharide(LPS);Cells in SEN group were added with 1 μg/ml LPS+4 μmol/L SEN,and cells in MCC950 group were added with 1 μg/ml LPS+10 μmol/L MCC950 for 24 hours.CCK-8 method was used to detect the effect of different concentrations of SEN on the viability of BV2 cells.Griess method was used to determine the release amount of nitric oxide(NO)in the supernatant.Real-time fluorescent quantitative PCR was used to determine the mRNA expression levels of NLRP3,lymphocyte apoptosis-associated spect-like protein containing a CARD(ASC),caspase-1,interleukin(IL)-1β and IL-18 mRNA.Immunofluorescence staining was used to detect the expression levels of ASC,IL-1β,Nrf2 and heme oxygenase-1(HO-1).Western blotting was used to detect the expression levels of NLRP3,caspase-1,ASC,IL-1β,IL-18,Nrf2,HO-1,nuclear factor kappa B(NF-κB)and inducible nitric oxide synthase(iNOS).Results The results of CCK-8 method showed that there was no significant difference in the viability of BV2 cells treated with 2～20 μmol/L SEN compared with control group(P>0.05).Compared with control group,the viability of BV2 cells in model group decreased significantly(P<0.05).Compared with model group,the viability of BV2 cells in 4 μmol/L SEN group was significantly restored(P<0.05).Compared with control group,the results of Griess method showed that the release amount of NO in cells of model group increased significantly(P<0.05);the results of real-time PCR showed that the expression levels of NLRP3,ASC,caspase-1,IL-1β and IL-18 mRNA in cells of model group increased significantly(P<0.05);the results of Western blotting showed that the protein expression levels of NLRP3,ASC,caspase-1,IL-1β and IL-18 proteins in cells of model group increased significantly(P<0.05),and the immunofluorescence staining results showed that the expression levels of iNOS and NF-κB protein in cells of model group increased,and the expression levels of Nrf2 and HO-1 decreased,with statistically significant differences(P<0.05).Compared with model group,the release amount of NO in cells of SEN group and MCC950 group decreased,and the expression levels of NLRP3,ASC,caspase-1,IL-1β and IL-18 mRNA and proteins decreased,with statistically significant differences(P<0.05);in the SEN group,the expression levels of iNOS and NF-κB decreased,and immunofluorescence staining showed that Nrf2 was translocated into the nucleus,and the expression levels of Nrf2 and HO-1 proteins increased significantly,with statistically significant differences(P<0.05).Conclusions SEN could alleviate the inflammatory response of mouse microglia cells induced by LPS and inhibit the activation and expression of NLRP3 inflammasome,with an effect comparable to that of the inflammasome inhibitor MCC950.The mechanism may be related to the regulation of the expression of upstream factors Nrf2 and HO-1.

Objective: To establish a machine learning-based precision blood donor recruitment model at the county level and assess its generalizability and applicability. Methods: A retrospective study was conducted using blood donation and SMS recruitment data from the Taicang Branch of the Suzhou Blood Center between 2019 and 2024. Multiple machine learning algorithms were employed, including extreme gradient boosting, support vector machine, k-nearest neighbor, logistic regression, decision tree, random forest, and multilayer perceptron. These were combined with techniques such as synthetic minority oversampling, undersampling, and cost-sensitive learning (using MFE and MSFE loss functions). Model parameters were optimized through grid search to identify the best-performing model. Results: In a prospective comparative study against conventional methods, the machine learning models increased the recruitment success rate among high-willingness donors by an average of 129.15%, and the recruitment efficiency per SMS improved by 125.02% compared with the traditional method. Under full-scale SMS sending, the recruitment rate per SMS increased by 42.61%, and SMS sending efficiency improved by 31.77%, significantly enhancing recruitment performance. Conclusion: This study represents the first application of a machine learning-based precision donor recruitment model at the county-level in China. The precise recruitment framework not only improves recruitment efficiency and reduces recruitment costs but also demonstrates strong scalability and generalizability. It provides a scientific and feasible intelligent pathway to ensure the safety and sustainability of the blood supply.

OBJECTIVE: To explore a predictive model that can better predict the prognosis of patients with diffuse large B-cell lymphoma (DLBCL), and validate its clinical value. METHODS: Clinical data of 134 newly treated DLBCL patients were collected from Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine from January 2015 to January 2020. Several risk factors of the patients were screened and analyzed, a novel prognostic model were then established based on this, and its clinical application potential was validated. RESULTS: In the novel model, predicting progression-free survival (PFS) based on the age at initial treatment, albumin level, Hans classification, Ann Arbor stage, and BCL2 expression showed better predictive performance than International Prognostic Index (IPI) score (AUC: 0.788 vs 0.620，P <0.001). Predicting overall survival (OS) based on the age at initial treatment, albumin level, lactate dehydrogenase (LDH) level, and expressions of BCL2 and MUM1 proteins also showed better predictive performance for mortality risk than IPI score (AUC: 0.817 vs 0.624，P <0.001). CONCLUSION This novel prognostic model can better predict the survival prognosis of DLBCL patients compared to the IPI scoring system.
Humans ; Lymphoma, Large B-Cell, Diffuse/diagnosis* ; Prognosis ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Risk Factors ; Male ; Female ; Middle Aged