ObjectiveTo investigate the mechanism by which the Zishen Huoxue prescription （ZSHXP） ameliorates cognitive dysfunction in rats with vascular dementia （VD） induced by the bilateral common carotid artery ligation （2-VO model rats） through regulating the glucose-regulated protein 78 （GRP78）/protein kinase R-like endoplasmic reticulum kinase （PERK）/activating transcription factor 4 （ATF4） signaling pathway. MethodsA VD rat model was established via the 2-VO method. A total of 72 male Sprague-Dawley （SD） rats were randomly divided into six groups： Sham group， Model group， donepezil hydrochloride group （0.45 mg·kg^-1）， and ZSHXP groups at low （8.90 g·kg^-1）， medium （17.80 g·kg^-1）， and high （35.60 g·kg^-1） doses，with 12 rats in each group. The Morris Water Maze test was utilized to assess spatial learning and memory abilities of rats， and the Novel Object Recognition test was used to evaluate cognitive performance. Hematoxylin-eosin （HE） and Nissl staining were applied to observe the histological and morphological changes in hippocampal tissues. Transmission electron microscopy （TEM） was used to observe the morphological changes of endoplasmic reticulum in rat hippocampal neurons. Immunofluorescence staining was adopted to detect the colocalization of neuronal nuclei antigen （NeuN） with GRP78 and βⅢ Tubulin with gasdermin D （GSDMD） in hippocampal neurons. Western blot was used to detect the expression levels of endoplasmic reticulum stress （ERS）-related proteins including GRP78， PERK， ATF4， phosphorylated protein kinase R-like endoplasmic reticulum kinase （p-PERK）， C/EBP homologous protein （CHOP）， NOD-like receptor protein 3 （NLRP3）， Caspase-1 and GSDMD. ResultsCompared with the sham operation group， the model group showed a significantly prolonged escape latency （P<0.01）， a significant decrease in the number of platform crossings and the residence time in the target quadrant （P<0.01）， and a markedly reduced recognition index （P<0.01）. Histological observations revealed that the hippocampal neurons in the model group were disorderly arranged with reduced quantity， deformed and shrunken cell bodies， and pyknotic and hyperchromatic nuclei. The number of Nissl bodies decreased significantly. The number of endoplasmic reticula reduced obviously， accompanied by abnormal dilation and swelling， and the loss of normal folding structure. The fluorescence colocalization of NeuN with GRP78 and βⅢ Tubulin with GSDMD in the hippocampus was significantly increased in the model group. The protein expression levels of GRP78， p-PERK/PERK， ATF4， CHOP， NLRP3， GSDMD and Caspase-1 in the model group were significantly elevated （P<0.01）. Compared with the model group， the donepezil hydrochloride group and the ZSHXP medium- and high-dose groups had a significantly shortened escape latency （P<0.01） and an increased number of platform crossings （P<0.05， P<0.01）. The residence time in the target quadrant was increased in the donepezil hydrochloride group and all ZSHXP groups （P<0.05， P<0.01）， with a significantly improved recognition index （P<0.01）. In the donepezil hydrochloride group and all ZSHXP groups， the number of hippocampal neurons increased with a more compact arrangement and reduced nuclear hyperchromasia. The number of Nissl bodies increased with morphological structures tending to be normal. In the ZSHXP high-dose group， the number of endoplasmic reticula increased and the folding structure was restored. The fluorescence colocalization of NeuN with GRP78 and βⅢ Tubulin with GSDMD in the hippocampus was significantly weakened in the treatment groups. In the donepezil hydrochloride group， the protein expressions of GRP78， ATF4 and CHOP were increased （P<0.01）， while the expression of p-PERK/PERK was decreased （P<0.05）. In the ZSHXP low-dose group， the expressions of GRP78， p-PERK/PERK and CHOP were elevated （P<0.05， P<0.01）. The ZSHXP medium- and high-dose groups showed a significant decrease in the protein expressions of p-PERK/PERK， ATF4 and CHOP （P<0.01）， and the high-dose group had a markedly reduced GRP78 protein expression （P<0.01）. In the donepezil hydrochloride group， the Caspase-1 protein expression was increased （P<0.01） and the NLRP3 protein expression was decreased （P<0.01）. In the ZSHXP low-dose group， the GSDMD expression was elevated （P<0.01） while the NLRP3 protein expression was reduced （P<0.01）. After treatment with medium and high doses of ZSHXP， the protein expression levels of NLRP3， GSDMD and Caspase-1 were significantly decreased （P<0.01）. ConclusionThe ameliorative effect of ZSHXP on cognitive function in 2-VO model rats may be associated with its regulation of the GRP78/PERK/ATF4 signaling pathway， which ameliorates ERS and inhibits neuronal pyroptosis.

ObjectiveThis paper aims to explore the in vitro anti-psoriasis activity and potential mechanism of Kangfuxin liquid （KFX liquid）， providing experimental evidence for the anti-psoriasis effect of KFX liquid. MethodsFirstly， the uninduced human immortalized keratinocyte cells （HaCaT cells） were divided into seven groups， namely the control group and KFX liquid groups with different doses （5， 10， 20， 40， 80， 160 g·L^-1）. After being treated with different concentrations of KFX liquid， the effect of KFX liquid on the normal cell proliferation was detected by using the cell counting kit-8 （CCK-8） method. Secondly， the uninduced HaCaT cells were divided into six groups， namely the control group and recombinant human interleukin-7A （rh-IL-7A） groups with different doses （5， 10， 50， 100， 120 g·L^-1）. After being treated with different concentrations of recombinant human interleukin-17A （rh IL-17A） liquid， the effect of rh IL-17A on cell proliferation was detected. The optimal induction concentration was screened. Then， normal HaCaT cells were divided into a control group and KFX liquid groups with different doses （5， 10， 20， 40， 80， 160 g·L^-1）. Except for the control group， the other groups established psoriasis cell models with the optimal induction concentration of rh IL-17A. After being treated with different concentrations of KFX liquid， the effects of KFX liquid on the psoriasis-like HaCaT cell proliferation were investigated. Finally， the uninduced HaCaT cells were divided into six groups， namely the control group， rh IL-17A group， methotrexate （MTX） group， and KFX liquid groups with different doses （20， 40， 80 g·L^-1）. Except for the control group， the other groups used the optimal induction concentration of rh IL-17A to establish psoriasis cell models. After being treated with different drugs， the cell migration levels were detected through scratch assays， and real-time quantitative polymerase chain reaction （Real-time PCR） was used to detect the relative mRNA expression levels of Ki-67 antigen （Ki67）， S100 calcium-binding protein A7 （S100A7）， S100 calcium-binding protein A8 （S100A8）， and S100 calcium-binding protein A9 （S100A9）， thereby comprehensively evaluating the in vitro anti-psoriasis activity of KFX liquid. By detecting the relative mRNA expression levels of interleukin-1β （IL-1β）， interleukin-6 （IL-6）， and chemokine-20 （CXCL-20） inflammatory-related factors in psoriasis-like HaCaT cells and the protein expression levels of Janus kinase 3 （JAK3）， phosphorylated Janus kinase 3 （p-JAK3）， signal transducer and activator of transcription 3 （STAT3）， and phosphorylated signal transducer and activator of transcription 3 （p-STAT3）， the mechanism was explored. ResultsCompared with that of control group， when treated with 80 g·L^-1 KFX liquid for 72 h （P<0.05） and at different times with 160 g·L^-1 KFX liquid， the HaCaT cell proliferation activity was significantly affected （P<0.01）， while the other concentrations of KFX liquid had no significant differences in cell morphology and cell proliferation activity at different times， indicating that the KFX liquid is relatively safe for HaCaT cells and has no obvious toxic side effects. Compared with that of control group， when treated with different concentrations of rh IL-17A for 24 h， the HaCaT cell proliferation activity was significantly enhanced， and the cell activity was the strongest when the concentration was 100 μg·L^-1 （P<0.05）， with a density close to 100% and intact cell morphology， indicating that 100 μg·L^-1 is the optimal concentration for inducing HaCaT cell proliferation. The results of the KFX liquid treatment on rh IL-17A-induced psoriasis-like cells show that the KFX liquid not only effectively inhibits the rh IL-17A-induced psoriasis-like HaCaT cell proliferation activity （P<0.01）， but also significantly reduces the migration ability of rh IL-17A-induced psoriasis-like HaCaT cells （P<0.01）， and the relative mRNA expression levels of Ki67， S100A7， S100A8， and S100A9 （P<0.01）. Moreover， the KFX liquid can significantly reduce the relative mRNA expression levels of IL-1β， IL-6， and CXCL-20 in rh IL-17A-induced psoriasis-like cells （P<0.01）， and significantly inhibit the phosphorylation levels of JAK3 and STAT3 proteins （P<0.05， P<0.01）. ConclusionThe KFX liquid has no obvious toxicity to uninduced HaCaT cells. It can inhibit rh IL-17A-induced psoriasis-like HaCaT cell proliferation， reduce the cell migration ability， and has good in vitro anti-psoriasis activity. Its action mechanism may be related to downregulating the expression levels of inflammation-related cytokines in the JAK3/STAT3 signaling pathway and inhibiting the phosphorylation levels of JAK3 and STAT3 proteins.

ObjectiveTo investigate the mechanism by which the Zishen Huoxue prescription （ZSHXP） ameliorates cognitive dysfunction in rats with vascular dementia （VD） induced by the bilateral common carotid artery ligation （2-VO model rats） through regulating the glucose-regulated protein 78 （GRP78）/protein kinase R-like endoplasmic reticulum kinase （PERK）/activating transcription factor 4 （ATF4） signaling pathway. MethodsA VD rat model was established via the 2-VO method. A total of 72 male Sprague-Dawley （SD） rats were randomly divided into six groups： Sham group， Model group， donepezil hydrochloride group （0.45 mg·kg^-1）， and ZSHXP groups at low （8.90 g·kg^-1）， medium （17.80 g·kg^-1）， and high （35.60 g·kg^-1） doses，with 12 rats in each group. The Morris Water Maze test was utilized to assess spatial learning and memory abilities of rats， and the Novel Object Recognition test was used to evaluate cognitive performance. Hematoxylin-eosin （HE） and Nissl staining were applied to observe the histological and morphological changes in hippocampal tissues. Transmission electron microscopy （TEM） was used to observe the morphological changes of endoplasmic reticulum in rat hippocampal neurons. Immunofluorescence staining was adopted to detect the colocalization of neuronal nuclei antigen （NeuN） with GRP78 and βⅢ Tubulin with gasdermin D （GSDMD） in hippocampal neurons. Western blot was used to detect the expression levels of endoplasmic reticulum stress （ERS）-related proteins including GRP78， PERK， ATF4， phosphorylated protein kinase R-like endoplasmic reticulum kinase （p-PERK）， C/EBP homologous protein （CHOP）， NOD-like receptor protein 3 （NLRP3）， Caspase-1 and GSDMD. ResultsCompared with the sham operation group， the model group showed a significantly prolonged escape latency （P<0.01）， a significant decrease in the number of platform crossings and the residence time in the target quadrant （P<0.01）， and a markedly reduced recognition index （P<0.01）. Histological observations revealed that the hippocampal neurons in the model group were disorderly arranged with reduced quantity， deformed and shrunken cell bodies， and pyknotic and hyperchromatic nuclei. The number of Nissl bodies decreased significantly. The number of endoplasmic reticula reduced obviously， accompanied by abnormal dilation and swelling， and the loss of normal folding structure. The fluorescence colocalization of NeuN with GRP78 and βⅢ Tubulin with GSDMD in the hippocampus was significantly increased in the model group. The protein expression levels of GRP78， p-PERK/PERK， ATF4， CHOP， NLRP3， GSDMD and Caspase-1 in the model group were significantly elevated （P<0.01）. Compared with the model group， the donepezil hydrochloride group and the ZSHXP medium- and high-dose groups had a significantly shortened escape latency （P<0.01） and an increased number of platform crossings （P<0.05， P<0.01）. The residence time in the target quadrant was increased in the donepezil hydrochloride group and all ZSHXP groups （P<0.05， P<0.01）， with a significantly improved recognition index （P<0.01）. In the donepezil hydrochloride group and all ZSHXP groups， the number of hippocampal neurons increased with a more compact arrangement and reduced nuclear hyperchromasia. The number of Nissl bodies increased with morphological structures tending to be normal. In the ZSHXP high-dose group， the number of endoplasmic reticula increased and the folding structure was restored. The fluorescence colocalization of NeuN with GRP78 and βⅢ Tubulin with GSDMD in the hippocampus was significantly weakened in the treatment groups. In the donepezil hydrochloride group， the protein expressions of GRP78， ATF4 and CHOP were increased （P<0.01）， while the expression of p-PERK/PERK was decreased （P<0.05）. In the ZSHXP low-dose group， the expressions of GRP78， p-PERK/PERK and CHOP were elevated （P<0.05， P<0.01）. The ZSHXP medium- and high-dose groups showed a significant decrease in the protein expressions of p-PERK/PERK， ATF4 and CHOP （P<0.01）， and the high-dose group had a markedly reduced GRP78 protein expression （P<0.01）. In the donepezil hydrochloride group， the Caspase-1 protein expression was increased （P<0.01） and the NLRP3 protein expression was decreased （P<0.01）. In the ZSHXP low-dose group， the GSDMD expression was elevated （P<0.01） while the NLRP3 protein expression was reduced （P<0.01）. After treatment with medium and high doses of ZSHXP， the protein expression levels of NLRP3， GSDMD and Caspase-1 were significantly decreased （P<0.01）. ConclusionThe ameliorative effect of ZSHXP on cognitive function in 2-VO model rats may be associated with its regulation of the GRP78/PERK/ATF4 signaling pathway， which ameliorates ERS and inhibits neuronal pyroptosis.

ObjectiveThis paper aims to explore the in vitro anti-psoriasis activity and potential mechanism of Kangfuxin liquid （KFX liquid）， providing experimental evidence for the anti-psoriasis effect of KFX liquid. MethodsFirstly， the uninduced human immortalized keratinocyte cells （HaCaT cells） were divided into seven groups， namely the control group and KFX liquid groups with different doses （5， 10， 20， 40， 80， 160 g·L^-1）. After being treated with different concentrations of KFX liquid， the effect of KFX liquid on the normal cell proliferation was detected by using the cell counting kit-8 （CCK-8） method. Secondly， the uninduced HaCaT cells were divided into six groups， namely the control group and recombinant human interleukin-7A （rh-IL-7A） groups with different doses （5， 10， 50， 100， 120 g·L^-1）. After being treated with different concentrations of recombinant human interleukin-17A （rh IL-17A） liquid， the effect of rh IL-17A on cell proliferation was detected. The optimal induction concentration was screened. Then， normal HaCaT cells were divided into a control group and KFX liquid groups with different doses （5， 10， 20， 40， 80， 160 g·L^-1）. Except for the control group， the other groups established psoriasis cell models with the optimal induction concentration of rh IL-17A. After being treated with different concentrations of KFX liquid， the effects of KFX liquid on the psoriasis-like HaCaT cell proliferation were investigated. Finally， the uninduced HaCaT cells were divided into six groups， namely the control group， rh IL-17A group， methotrexate （MTX） group， and KFX liquid groups with different doses （20， 40， 80 g·L^-1）. Except for the control group， the other groups used the optimal induction concentration of rh IL-17A to establish psoriasis cell models. After being treated with different drugs， the cell migration levels were detected through scratch assays， and real-time quantitative polymerase chain reaction （Real-time PCR） was used to detect the relative mRNA expression levels of Ki-67 antigen （Ki67）， S100 calcium-binding protein A7 （S100A7）， S100 calcium-binding protein A8 （S100A8）， and S100 calcium-binding protein A9 （S100A9）， thereby comprehensively evaluating the in vitro anti-psoriasis activity of KFX liquid. By detecting the relative mRNA expression levels of interleukin-1β （IL-1β）， interleukin-6 （IL-6）， and chemokine-20 （CXCL-20） inflammatory-related factors in psoriasis-like HaCaT cells and the protein expression levels of Janus kinase 3 （JAK3）， phosphorylated Janus kinase 3 （p-JAK3）， signal transducer and activator of transcription 3 （STAT3）， and phosphorylated signal transducer and activator of transcription 3 （p-STAT3）， the mechanism was explored. ResultsCompared with that of control group， when treated with 80 g·L^-1 KFX liquid for 72 h （P<0.05） and at different times with 160 g·L^-1 KFX liquid， the HaCaT cell proliferation activity was significantly affected （P<0.01）， while the other concentrations of KFX liquid had no significant differences in cell morphology and cell proliferation activity at different times， indicating that the KFX liquid is relatively safe for HaCaT cells and has no obvious toxic side effects. Compared with that of control group， when treated with different concentrations of rh IL-17A for 24 h， the HaCaT cell proliferation activity was significantly enhanced， and the cell activity was the strongest when the concentration was 100 μg·L^-1 （P<0.05）， with a density close to 100% and intact cell morphology， indicating that 100 μg·L^-1 is the optimal concentration for inducing HaCaT cell proliferation. The results of the KFX liquid treatment on rh IL-17A-induced psoriasis-like cells show that the KFX liquid not only effectively inhibits the rh IL-17A-induced psoriasis-like HaCaT cell proliferation activity （P<0.01）， but also significantly reduces the migration ability of rh IL-17A-induced psoriasis-like HaCaT cells （P<0.01）， and the relative mRNA expression levels of Ki67， S100A7， S100A8， and S100A9 （P<0.01）. Moreover， the KFX liquid can significantly reduce the relative mRNA expression levels of IL-1β， IL-6， and CXCL-20 in rh IL-17A-induced psoriasis-like cells （P<0.01）， and significantly inhibit the phosphorylation levels of JAK3 and STAT3 proteins （P<0.05， P<0.01）. ConclusionThe KFX liquid has no obvious toxicity to uninduced HaCaT cells. It can inhibit rh IL-17A-induced psoriasis-like HaCaT cell proliferation， reduce the cell migration ability， and has good in vitro anti-psoriasis activity. Its action mechanism may be related to downregulating the expression levels of inflammation-related cytokines in the JAK3/STAT3 signaling pathway and inhibiting the phosphorylation levels of JAK3 and STAT3 proteins.

The engineering application of microbially induced carbonate precipitation(MICP)is limited by pH-dependent conformational dynamics of urease.Focusing on the α-subunit urease from Sporosarcina pasteurii,this study integrated conductivity experiments and constant-pH molecular dynamics simulations to analyze active site conformational dynamics and catalytic function across pH 3-11.Results showed that under neutral conditions(pH 7-8),key histidine residues(HIS139/HIS249)exhibited minimal dis-placement(＜0.5 ?),the longest hydrogen bond lifetime(＞8 ps),highest conformational stability(root mean square deviation,RMSD:0.15-0.18 nm),and optimal catalytic activity(conductivity change rate:0.03 mS/cm·min-1,CaCO3 precipitation:3.84 g).Extreme pH(pH 3/11)induced structural collapse(displacement up to 1.8 ?)and complete activity loss.Simulations revealed that neutral pH sta-bilizes a protonation-dependent cooperative allosteric network by maintaining active site cavity volume(～120 ?3)and moderate conformational coherence(correlation coefficient～0.8).This work deciphers the molecular mechanism of pH-regulated urease dynamics through protonation states,providing theoreti-cal support for MICP applications in acidic mine tailing remediation and alkaline soil stabilization.

Chronic prostatitis,a common condition in andrology,is clinically characterized by a prolonged course,resistance to treatment,and frequent recurrence.In traditional Chinese medicine,it is classified under the categories of"turbid sperm,""overstrain strangury,"and"vaginal pain."Based on the"host-guest interaction-collateral disease"theory,we believe that healthy qi deficiency,latent pathogenic factors,and collateral obstruction are the primary pathological factors of this disease,which run through the entire process of chronic prostatitis occurrence and development.Accordingly,we propose that obstruction of collaterals and apathesia of the semen chamber are the core pathogenesis.The disease progression can be divided into three pathological stages:"deficiency,depression,and blood stasis."Spleen and kidney deficiency and malnutrition of collaterals form the pathological foundation.In the deficiency stage,treatment strategies involve reinforcing qi and nourishing the collaterals,using Fuzheng Yangrong Decoction.During disease progression,dampness and heat invasion,as well as collateral stagnation qi,are key contributors to disease progression.Thus,treatment focuses on clearing heat and dampness,promoting qi flow,and smoothing the collaterals,achieved with a modified Qiantongding Decoction.In the final stage,blood stasis and collateral obstruction dominate,warranting therapeutic strategies aimed at tonifying and removing blood stasis,addressing both the body and the collaterals simultaneously using the modified Guiling Huayu Decoction.Overall,the clinical treatment generally focuses on the concept of function through free flow,combination of unblocking and tonifying.This study provides a novel perspective and reference for clinical differentiation and treatment of chronic prostatitis.

Oral squamous cell carcinoma(OSCC)is a common malignant tumor.Interleukin-8(IL-8)is an important biomarker of OSCC,and its level can reflect the occurrence and development of OSCC.It is of great significance to detect IL-8 rapidly and sensitively for the purpose of early diagnosis of OSCC.In this study,gold nanoparticles(AuNPs)with uniform particle size were synthesized by reduction of chloroauric acid with trisodium citrate,and the probe(AuNPs@mAb1)was prepared by coupling AuNPs with the murine anti-IL-8 monoclonal antibody mAb1.Cy5-NHS and murine anti-IL-8 monoclonal antibody mAb2 complex(Cy5-mAb2)and sheep anti-mouse IgG antibody were sprayed on nitrocellulose membrane to form test line(T line)and control line(C line)respectively,and reverse fluorescence-enhanced test strips were thus constructed,based on which an immunochromatographic method was established for highly sensitive detection of IL-8 in saliva samples.The experimental results showed that the test strip had good stability,high specificity and high sensitivity.The linear range for fluorescence detection of IL-8 was 0.01-100 ng/mL,and the limit of detection(3σ)was 7.93 pg/mL.The linear range for visualization detection was 6-100 ng/mL,with limit of detection(3σ)of 0.85 ng/mL.The fabricated test trip had good preparation reproducibility,with inter-and intra-batch assay precision of less than 5.5%.The test strip was used for detection of IL-8 in healthy human saliva samples,with spiked recoveries of 93.7% -102.4%,and relative standard deviations of 2.1% -4.3%.The fabricated test strip could be used for early screening of OSCC.

Objective To propose an in vitro cytotoxicity evaluation method for novel iron-based biodegradable mateirals focusing on their degrading characteristics.Methods Half-finished scaffolds of nitrided iron tubes with a higher iron content of over 99％and a nitrogen content of 0.03％to 0.20％were selected as test samples,and based on the treatment mode were divided into an anaerobic treatment group,a non-anaerobic treatment group,a non-anaerobic treatment with removed iron particle group and a non-anaerobic treatment with plate-washing group.The anaerobic treatment group was processed in an anaerobic workstation;the conventional treatment group underwent standard handling in a biosafety cabinet;the conventional treatment with removed iron particle group was subjected to iron particle elimination using a Midimacs Starting Kit manual sorter after conventional treatment;and the conventional treatment with plate-washing group was rinsed with 0.9％sodium chloride injection before tetrazolium salt reagent treatment.Different extraction media(simulated body fluid,cell culture medium,phosphate buffer and 0.9％sodium chloride injection)were used for the immersion treatment of the test samples in each group to compare the effects of the degradation products on the survival rate of L929 cells.Results The anaerobic treatment group and the conventional treatment with removed iron particle group exhibited no detectable cytotoxicity.Trypan blue staining revealed significant cytotoxicity in the conventional treatment group,while false-negative results emerged due to interactions between the tetrazolium salt reagent and degradation products.In the non-anaerobic treatment with washing plate group,the false negative from iron particles was eliminated while potential cytotoxicity was showen,and the result accuracy was affected because some cells were washed out.Conclusion The proposed method can be used for the in vitro cytotoxicity evaluation,and facilitates the safety evaluation of the application of such materials.[Chinese Medical Equipment Journal,2025,46(3):35-41]

Objective:To establish a complete system for the isolation, purification, identification, and culture of Kaposiform hemangioendothelioma-derived endothelial cells (KHE-ECs), to analyze the phenotype of KHE-ECs, and to explore the possibility of establishing a KHE-EC bank.Methods:A novel digestion solution for KHE tumors (patent number: CN202410500224.2) was formulated using collection fluid, Liberase TM and dispase stock solutions, and was used to process tumor tissues to obtain cells. High-purity KHE-ECs were purified using CD31 + immunomagnetic beads. The EGM-2 complete medium containing 10% fetal bovine serum and 2% penicillin-streptomycin solution was employed for cell culture. To verify the characteristics of KHE-ECs, immunofluorescence assay was conducted to determine the expression of endothelial cell-specific markers CD31 and CD34, KHE disease markers podoplanin (D2-40), prospero-related homeobox 1 (Prox-1), and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1), as well as an infantile hemangioma-specific diagnostic marker glucose transporter 1 (GLUT-1). Human umbilical vein endothelial cells (HUVECs) served as controls for the phenotype analysis of KHE-ECs, including cell viability, cytoskeleton, proliferation, migration, invasion, tube formation, and sprouting ability. Results:Primary cells were successfully isolated from KHE tumor tissues, and high-purity KHE-ECs were obtained by using CD31 + immunomagnetic beads. The cells exhibited typical spindle-shaped morphology and an adherent growth pattern. Immunofluorescence assay showed that KHE-ECs expressed CD31, CD34, D2-40, Prox-1, and LYVE1, but did not express GLUT-1. There were significant differences in cell morphology, cell viability, and cytoskeletal structures between KHE-ECs and HUVECs. Additionally, the KHE-EC group showed significantly increased percentages of proliferative cells (29.1% ± 2.5%), numbers of migratory cells (114.3 ± 9.4) and invasive cells (110.0 ± 6.1), tube length (32 121.0 ± 892.0 μm), and number of sprouting cells (25.0 ± 3.6) compared with the HUVEC group (13.0% ± 2.2%, 38.0 ± 3.6, 35.3 ± 2.3, 25 345.0 ± 448.1 μm, 5.0 ± 1.0, respectively, all P ≤ 0.001) . Conclusion:An innovative digestion solution specifically for KHE tumors was formulated for the first time, and high-purity and well-growing KHE-EC strains were successfully isolated and purified by using the novel digestion solution in combination with CD31 + immunomagnetic beads, providing a stable and reliable cell source for subsequent experimental studies on KHE and laying the foundation for establishing a KHE-EC bank.

Objective:To investigate the predictive value of interventricular septal thickness(IVST),the ratio of mitral ear-ly diastolic peak flow velocity to late diastolic peak flow velocity(E/A)combined with aortic valve flow velocity(AV)for hypertensive heart disease(HHD)and its association with blood pressure grade.Methods:A total of 202 patients with hy-pertension suspected of HHD admitted to Qinhuangdao Hospital of Dongfang Hospital,Beijing University of Chinese Medi-cine between May 2020 and May 2023 were enrolled and divided into HHD group(n=102)and non-HHD group(n=100)according to color Doppler energy imaging and blood pressure diagnosis.Blood pressure,IVST,E/A and AV were measured in two groups.The predictive value of IVST,E/A and AV for HHD were evaluated by receiver operating charac-teristic(ROC)curve.According to hypertension grading,patients in HHD group was divided into grade 1～2 group(n=59)and grade 3 group(n=41).The association of IVST,E/A,AV with blood pressure grade in HHD patients were ana-lyzed by Spearman correlation analysis.Results:Compared with patients in non-HHD group,those in HHD group had significant higher IVST,and significant lower E/A and AV(P＜0.001 all);Compared with patients in grade 1～2 group,those in grade 3 group had significant higher IVST,and significant lower E/A and AV(P＜0.001 all).ROC curve showed that the AUC of IVST,E/A and AV alone and their combination in HHD diagnosis was 0.798(95％CI 0.736～0.851),0.682(95％CI 0.613～0.746),0.810(95％CI 0.748～0.862)and 0.927(95％CI 0.881～0.959)respectively,and the diagnostic value of combined detection was significantly higher than those of single detections(Z=4.431,5.669,3.948,P=0.001 all).Spearman correlation analysis showed that E/A and AV were significant negatively correlated with blood pressure grade(r=-0.725,-0.781),while I VST was significant positively correlated with blood pressure grade(r=0.714),and combined detection possessed a stronger correlation with it in HHD patients(r=0.875,P＜0.001 all).Con-clusion:The combined detection of I VST,E/A and AV demonstrates a good predictive value for HHD and exhibits a sig-nificant correlation with blood pressure grade in HHD patients.