ObjectiveTo establish an in vitro culture model of rat corpora cavernous smooth muscle cells （CCSMCs） using a modified tissue block adherence method. MethodsCorpus cavernosum smooth muscle tissue was digested with collagenase type I and subsequently cultured using an adherent method. Cells were purified via differential adhesion and identified through immunofluorescence and Western blotting. ResultsCCSMCs began to emerge from the tissue block after 3 days， increased significantly by day 7， and converged by day 12. Post-passage， CCSMCs exhibited strong proliferation and a “peak-to-valley” phenomenon. After purification， the cells tested positive for α-smooth muscle actin （α-SMA）， confirming the successful establishment of the in vitro culture model. ConclusionThe modified tissue block adherence method is a cost-effective and efficient way to obtain high-purity CCSMCs.

Andrological diseases have become an important public health problem threatening men's health worldwide， which significantly affects the quality of life of patients and brings a heavy disease burden. Western medicine often faces the dilemma of obvious side effects and limited efficacy. Traditional Chinese medicine has unique advantages in the prevention and treatment of andrological diseases and has accumulated rich clinical experience. Epimedii Folium， as a traditional Chinese medicine for strengthening kidney and Yang， exerts a key therapeutic effect on andrology diseases through multi-component synergy， multi-target regulation， and multi-pathway intervention. Recent studies have found that the main active components of Epimedii Folium， such as icariin， icariside， and icaritin， are the key material basis for the treatment of andrological diseases. The active components of Epimedii Folium can play a role in common andrological diseases such as erectile dysfunction， male infertility， and prostate cancer by regulating the activity of the nitric oxide/cyclic guanosine monophosphate （NO/cGMP） pathway， participating in oxidative stress response， regulating the secretion of hypothalamic-pituitary-gonadal axis hormones， improving spermatogenic dysfunction， and inhibiting the proliferation of cancer cells. However， the systematic action network and molecular mechanisms of the active components of Epimedii Folium have not been fully elucidated， thereby limiting its potential for clinical translation and application. In the future， it is necessary to combine cutting-edge technologies such as metabolomics， single-cell sequencing， and targeted nanoscale drug delivery systems， strengthening the research on the compatibility rules of active components and organ-specific delivery， providing a scientific basis for the development of innovative andrology traditional Chinese medicine formulas with international competitiveness， and promoting the innovation and breakthrough of andrology disease treatment modes.

Oligoasthenospermia （OAS） is the common cause of male infertility，bringing fertility problems to many families. Its pathogenesis is complex，involving endocrine disorders，reproductive system infection，oxidative stress injury，immune dysfunction and other aspects. At present，the Western medicine treatment of OAS is mainly based on drugs，surgery，and assisted reproductive technology. However，drug therapy has certain side effects，and long-term use is easy to produce drug resistance. Surgical treatment has certain trauma and risk. Assisted reproductive technology is not widely available because of its high cost. Therefore，there is an urgent need to develop safer，economical，and effective treatment options. Traditional Chinese medicine （TCM） has obvious advantages in the treatment of OAS. It can play a role in the treatment of OAS by reducing oxidative stress，inhibiting inflammatory response，reducing apoptosis，and regulating cell autophagy through multiple pathways and targets. Recent studies have shown that TCM compound prescriptions，single medicinal materials，medicinal pairs，and their active components can ameliorate OAS by regulating several key signaling pathways such as the phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） pathway，Janus kinase 2/signal transducer and activator of transcription 3（JAK2/STAT3） pathway，nuclear factor E₂-related factor 2/heme oxygenase-1 （Nrf2/HO-1） pathway，stem cell factor/tyrosine kinase receptor （SCF/c-kit） pathway，nuclear factor-κB （NF-κB） pathway，transforming growth factor-β （TGF-β）/Smads pathway，and extracellular regulated protein kinase （ERK） pathway. This article summarizes the role of the above signaling pathways in the occurrence and development of OAS and the therapeutic mechanism of TCM，aiming to provide a scientific basis for the clinical treatment of OAS and the research and development of drugs.

Objective To analyze the expression levels and the clinical significance of serum interleukin(IL)-4,IL-5,IL-6,IL-13 and IL-17 in children with mycoplasma pneumoniae(MP)infection accompanied by airway hyperresponsiveness(AHR).Methods A total of 120 children diagnosed with MP infection pneumonia and admitted to Nantong First People's Hospital between June 2022 and April 2024 were enrolled in the study group.According to whether their MP infection pneumonia was accompanied by AHR,the participants were divided into an AHR group(n=41)and a non-AHR group(n=79).An additional 90 healthy children undergoing physical examination were included in the control group.Baseline data,fractional exhaled nitric oxide(FeNO),tidal breathing lung function—including the ratio of volume at peak tidal expiratory flow to total expiratory volume(VPTEF/VE),the ratio of time to peak tidal expiratory flow to total expiratory time(TPTEF/TE),and inspiratory-to-expiratory time ratio(TI/TE)—as well as the levels of serum IL-4,IL-5,IL-6,IL-13 and IL-17 were compared among the 3 groups.The correlation between serum IL-4,IL-5,IL-6,IL-13 and IL-17 and clinical data of children with MP infection pneumonia accompanied by AHR was analyzed.Logistic regression analysis was used to identify the independent influencing factors of AHR in children with MP infection pneumonia.Receiver operating characteristic(ROC)curves were plotted to assess the diagnostic value of serum IL-4,IL-5,IL-6,IL-13 and IL-17 for MP infection pneumonia accompanied by AHR in children.Results Significant differences were observed among the AHR,non-AHR,and control groups in white blood cell(WBC)count,neutrophil percentage,eosinophil(EOS)count,platelet(PLT)count,C-reactive protein(CRP),FeNO,VPTEF/VE,TPTEF/TE,TI/TE,forced vital capacity(FVC),and forced expiratory volume in one second(FEV?)(all P<0.05).Serum IL-4,IL-5,IL-6,IL-13 and IL-17 levels showed significant differences among the 3 groups(P<0.05).Correlation analysis showed that these cytokines were positively correlated with WBC count,neutrophil percentage,EOS count,CRP,and FeNO(P<0.05),and negatively correlated with VPTEF/VE,TPTEF/TE,TI/TE,FVC,and FEV1(P<0.05).Binary logistic regression analysis identified neutrophil percentage(odds ratio[OR]=1.923;95%CI,1.496-2.472),EOS count(OR=3.074;95%CI,1.228-7.693),CRP(OR=2.382;95%CI,1.854-3.061),FeNO(OR=1.931;95%CI,1.635-2.281),VPTEF/VE(OR=0.294;95%CI,0.200-0.432),TPTEF/TE(OR=0.358;95%CI,0.177-0.722),TI/TE(OR=0.399;95%CI,0.221-0.722),IL-4(OR=1.064;95%CI,1.019-1.111),IL-5(OR=1.234;95%CI,1.095-1.390),IL-6(OR=1.013;95%CI,1.001-1.025),IL-13(OR=1.058;95%CI,1.005-1.113),IL-17(OR=1.759;95%CI,1.293-2.393),and allergy history(OR=2.989;95%CI,1.058-8.447)as independent factors associated with with AHR in children with MP infection pneumonia(P<0.05).ROC curves revealed that the areas under the ROC curves(AUC)of serum IL-4,IL-5,IL-6,IL-13 and IL-17 for predicting AHR in children with MP infection pneumonia were 0.815,0.769,0.782,0.793,and 0.815,respectively,with the sensitivities being 85.37%,75.61%,87.80%,75.61%,and 80.49%,and 95%CI being 0.739-0.892,0.677-0.861,0.679-0.867,0.715-0.882 and 0.732-0.899,respectively.Conclusion The levels of serum IL-4,IL-5,IL-6,IL-13,and IL-17 in children with MP infection pneumonia and AHR are abnormally elevated.These cytokines may serve as valuable biomarkers for assessing the risk of AHR in children with MP infection pneumonia.

Objective To predict and preliminarily validate potential shared key genes involved in cardiac complications caused by influenza A virus(IAV)and severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infections.Methods Differentially expressed genes(DEGs)associated with cardiac complications were obtained from the Gene Expression Omnibus(GEO)database.A hierarchical intersection strategy was applied.First,cardiac complication related DEGs were overlapped with 2 independent virus related gene sets:3 454 human genes linked to IAV infection in GeneCards and 333 human protein-coding genes interacting with SARS-CoV-2 in the Human Protein Atlas.The 2 overlap results were then intersected to yield 22 hub genes.Lasso regression,random forest(RF)and support vector machine algorithms(SVM)were employed to refine this list.Predicted genes were validated in vitro in H1N1-infected human cardiomyocyte AC16 cells and in vivo in IFITM3 knockout mice challenged with H1N1,assessing transcriptional changes.Results A total of 22 hub genes were identified through integrative bioinformatics analysis.Application of the 3 machine learning algorithms resulted in 5 common key genes:ACE2,TBK1,NUP210,PUSL1,and MEPCE.In vitro infection of AC16 cells with H1N1 revealed dynamic transcriptional changes in all 5 genes post-infection(P<0.05).In vivo experiments using H1N1-infected IFITM3 knockout mice confirmed the dynamic mRNA expression changes of these 5 genes,consistent with the in vitro results(P<0.05).Conclusion By combining multilayered bioinformatics analysis with 3 machine learning approaches,5 common key genes are identified:ACE2,TBK1,NUP210,PUSL1 and MEPCE.Validation in H1N1 infection models confirms their relevance to IAV-induced cardiac complications.

Objective To explore the effects of graphene quantum dots(GQDs)on the reprogramming of Müller cells.Methods Müller cells were randomly divided into the control,dedifferentiation,dedifferentiation+GQD,and GQD groups.The cells in the dedifferentiation group and the dedifferentiation+GQD group were dedifferentiated using the serum-free medium containing DMEM/F12(1∶1),20 μg·L-1 EGF,10 μg·L-1 bFGF,2 × B27,and 1 × N2 for 5 d.Then,the cells in the dedifferentiation+GQD group and the GQD group were treated with 50 mg·L-1 GQDs for 72 h.The Müller cells in the control group were subjected to normal cell culture.Immunofluorescence staining was used to identify the ex-pression of Müller cell markers,including glial fibrillary acidic protein(GFAP),glutamate-aspartate transporter(GLAST),and glutamine synthetase(GS).Phalloidin staining was performed to observe whether Müller cells could take up GQDs.The effect of different concentrations of GQDs on the proliferation of Müller cells was assessed using the cell counting kit-8(CCK-8)assay.The effect of GQDs on the expression of neural progenitor/stem cell markers(SRY-box transcription factor 2[SOX-2]and Nestin)and the astrocyte marker GFAP in normal and dedifferentiated Müller cells was examined using im-munofluorescence staining and Western blotting.Results The immunofluorescence staining results showed that the fluo-rescence of GFAP was extremely weak and almost invisible in Müller cells,while the fluorescence of GLAST and GS was extremely strong and predominantly appeared in the cytoplasm of Müller cells.After 24 h of GQD treatment,trace amounts of GQD fluorescence were visible in Müller cells.The amount of GQD fluorescence in the cytoplasm of Müller cells gradual-ly increased with time.The CCK-8 assay results showed that the activity of Müller cells tended to decrease with an increase in the treatment time and concentration of GQD.The statistical analysis results showed that the mean fluorescence intensity of GFAP,Nestin,and SOX-2 in Müller cells of the dedifferentiation,dedifferentiation+GQD,and GQD groups was higher than that of the control group,and the differences were statistically significant(all P＜0.05).The mean fluorescence inten-sity of GFAP in the dedifferentiation+GQD group was higher than that in the dedifferentiation group,the mean fluores-cence intensity of Nestin and SOX-2 was lower than that in the dedifferentiation group,and the differences were all statisti-cally significant(P＜0.05).The relative protein expression of GFAP,Nestin,and SOX-2 in Müller cells in the dedifferentia-tion,dedifferentiation+GQD,and GQD groups was higher than that in the control group,and the differences were statis-tically significant(all P＜0.05).The relative protein expression of GFAP in the dedifferentiation+GQD group was higher than that in the dedifferentiation group,the relative protein expression of Nestin and SOX-2 was lower than that in the dedi-fferentiation group,and the differences were statistically significant(all P＜0.05).Conclusion GQDs facilitate the re-programming of Müller cells into astrocytes.

Objective To investigate the rehabilitation outcome in hemiplegic stroke patients with upper limb dysfunction treated with electromyographic(EMG)biofeedback therapy in combination with an intelligent Tongluo therapeutic apparatus.Methods A total of 132 stroke patients with hemiplegia were chosen and divided into observation group and control group according to the rehabilitation strategies,with 66 cases in each group.Control group was treated with EMG biofeedback therapy,while observation group was given the integrated therapy of EMG biofeedback therapy and intelligent Tongluo therapeutic apparatus.The two groups were compared for motor function of the hemiplegic upper limb(shoulder,hand and wrist muscle),integrated electromyography(iEMG)in the affected limb flexion and extension(biceps brachii and triceps brachii),range of motion of the shoulder joint(abduction,external rotation,internal rotation)and Barthel index of activities of daily living(ADL).Results The differences in the iEMG of biceps brachii and triceps brachii of the elbow joint,motor function of the hemiplegic upper limb,range of motion of the shoulder joint and ADL between two groups before treatment were trivial(P>0.05).The treatment in observation group increased the iEMG of biceps brachii and triceps brachii in elbow flexion(P<0.05),decreased the iEMG of biceps brachii in elbow extension but increased the iEMG of triceps brachii in elbow extension(P<0.05),and these improvements were more significant than those in control group(P<0.05).After treatment,the scores for shoulder function,hand function,and wrist muscle function in observation group increased from(12.23±2.76)to(26.77±4.28),(6.54±1.34)to(15.35±2.46),and(2.36±0.46)to(8.13±2.52),and the scores of the above indexes were higher than those in control group(P<0.05).The shoulder abduction,external rotation and internal rotation ranges of motion in observation group after treatment were greater than those before treatment[(90.77±8.35)° vs(61.23±6.35)°,(45.36±5.33)° vs(26.33±4.65)°,(52.55±5.86)° vs(32.44±3.87)°],and the observation group had greater shoulder ranges of motion than control group(P<0.05).The ADL score in observation group after treatment was higher than that before treatment[(62.45±5.25)vs(33.35±4.85),P<0.05],and it was also higher than that in control group(P<0.05).Conclusion EMG biofeedback therapy combined with intelligent Tongluo therapeutic apparatus can effectively improve the iEMG in the affected limb flexion and extension,enhance the ranges of motion of the shoulder joint,and ultimately improve the upper limb motor function and life quality of hemiplegic stroke patients.

A comprehensive phytochemical investigation of the leaves and twigs of Physalis angulata. var. villosa resulted in the isolation of 23 withanolide derivatives, including one novel 13,20-γ-lactone withanolide derivative (1) and three new withanolide derivatives (2-4). Architecturally, physalinin A (1) represents the first identified type B withanolide featuring a 13,20-γ-lactone moiety. The molecular structures of all isolates were elucidated using an integrated approach combining nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (MS), infrared (IR) spectroscopy, and quantum chemical calculations to confirm structural assignments. The antiproliferative activities of all isolated withanolides were evaluated against four human cancer cell lines (HEL, HCT-116, Colo320DM, and MDA-MB-231). Among them, eight derivatives (2, 5-8, 14, 15, and 23) exhibited significant inhibitory effects, with half-maximal inhibitory concentration (IC₅₀) values of 0.18 ± 0.03 to 17.02 ± 0.21 μmol·L^-1. Structure-activity relationship (SAR) analysis suggested that the presence of an epoxide ring enhances anticancer activity, potentially through increased reactivity or specific interactions with molecular targets involved in cancer progression. These findings underscore the pharmacological potential of withanolides as promising lead compounds for the development of novel anticancer therapeutics.
Withanolides/isolation & purification* ; Physalis/chemistry* ; Humans ; Molecular Structure ; Cell Line, Tumor ; Antineoplastic Agents, Phytogenic/isolation & purification* ; Cell Proliferation/drug effects* ; Plant Leaves/chemistry* ; Plant Extracts/pharmacology*

AIM:The study aims to investigate the effects of Huayu-Qutan formula(HYQT)-medicated serum on oleic acid-induced lipid damage and endoplasmic reticulum stress(ERS)in HepG2 cells,and to explore the mecha-nism underlying the prevention and treatment of lipid metabolism disorders by HYQT.METHODS:The HepG2 cells were treated with various concentrations of oleic acid,and CCK8 assay,oil red O staining and ELISA were used to identify the optimal treatment concentration and time of oleic acid and HYQT-medicated serum.Moreover,the cells were divided into control,model,thapsigargin(Tg),and Tg+HYQT groups.Cellular lipid deposition was measured using oil red O staining,while triglyceride(TG)and free fatty acid(FFA)levels were assessed by ELISA.Transmission electron micros-copy was used to examine endoplasmic reticulum structures,and RT-qPCR and Wes fully automated protein quantification analysis system were used to measure the mRNA and protein expression of glucose-regulated protein 78(GRP78),sterol regulatory element binding protein 1c(SREBP1c),acetyl coenzyme A carboxylase(ACC1),fatty acid synthase(FAS)and stearoyl coenzyme A desaturase 1(SCD1)in different groups.RESULTS:Significant lipid deposition was induced in HepG2 cells after treatment with 1 000 μmol/L oleic acid,while treatment with serum containing 10%HYQT for 48 h was found to be optimal.Compared with control group,the cells in model group showed significant deposition of oil red O-stained lipid droplets in the cytoplasm,associated with endoplasmic reticulum expansion,ERS,and nuclear condensa-tion.The TG and FFA levels,and the mRNA and protein expression levels of GRP78,SREBP1c,ACC1,FAS and SCD1 were increased significantly(P<0.01).Compared with model group,the cells treated with HYQT-mediated serum showed marked decrease in the number of lipid droplets in the cytoplasm,with restored endoplasmic reticulum morphology.The TG and FFA levels were significantly reduced(P<0.01),and the mRNA and protein levels of factors related to ERS and de novo fatty acid synthesis were markedly decreased(P<0.01).Treatment with Tg,an ERS agonist,led to greater accu-mulation of cytoplasmic lipid droplets and increased endoplasmic reticulum expansion.Marked variations in the morpholo-gy and size of the endoplasmic reticulum were observed,with fusion and clustering of vacuoles.The TG and FFA levels,and the expression of ERS-and fatty acid synthesis-related factors were increased(P<0.01).Compared with Tg group,the cells treated with Tg+HYQT showed reduced number of cytoplasmic lipid droplets,attenuated endoplasmic reticulum dilation,and decreased number and volume of vacuoles,while the TG and FFA levels and the expression of ERS-and fatty acid synthesis-associated factors were significantly decreased(P<0.01).CONCLUSION:Serum containing HYQT alle-viates oleic acid-induced lipid damage in HepG2 cells by inhibiting ERS-induced de novo synthesis of fatty acids.

Objective To investigate the early diagnostic value of combined detection of serum TIR-domain-containing adapter-inducing interferon-β(TRIF),cyclophilin A(CyPA),ubiquitin carboxyl-terminal hydrolase L1(UCH-L1),and hepcidin(Hepc)in brain injury among premature infants(BIPI).Methods Clinical data from 105 infants with brain injury following perinatal asphyxia(BIPI)admitted to our hospital between January 2022 and December 2024 were retrospectively collected and assigned to the case group,which was further subdivided into a severe subgroup(n=31)and a mild subgroup(n=74)based on the extent of brain injury.As controls,clinical data from 105 healthy preterm infants born in our hospital during the same period were retrospectively enrolled in a 1∶1 ratio.Serum levels of TRIF,CyPA,UCH-L1,and Hepc,along with clinical characteristics,were compared between the groups.Pearson correlation analysis was performed to evaluate the association between serum biomarker levels and neonatal behavioral neurological assessment(NBNA)scores.The diagnostic performance of these bio-markers for early detection of BIPI was assessed by constructing receiver operating characteristic(ROC)curves and calculating the area under the curve(AUC).Results Serum levels of TRIF,CyPA,UCH-L1,and Hepc were significantly higher in the case group than in the control group(P<0.05),while NBNA scores were significantly lower in the case group(P<0.05).Similarly,serum levels of these biomarkers were significantly elevated in the severe group compared to the mild group(P<0.05),accompanied by significantly lower NBNA scores in the severe group(P<0.05).Pearson correlation analysis revealed that serum levels of TRIF,CyPA,UCH-L1,and Hepc were negatively correlated with NBNA scores(r=-0.579,-0.514,-0.609,-0.588;all P<0.05).ROC analysis demon-strated that the combined detection of these four markers yielded an AUC of 0.927 for early diagnosis of BIPI,which was significantly higher than the AUCs of individual markers(0.819,0.803,0.776,and 0.767,respectively;P<0.05).Conclusions The serum levels of TRIF,CyPA,UCH-L1,and Hepc in children with BIPI were significantly elevated.These biomarkers were closely associated with disease progression and neurological development,and their combined measurement demonstrated superior performance in the early diagnosis of BIPI.