ObjectiveTo investigate whether Huanglian Jiedutang （HLJD） and its major active constituents （geniposide， baicalin， and berberine） can inhibit the inflammatory response of BV2 cells under lipopolysaccharide （LPS） stimulation via the high-mobility group protein B1 （HMGB1）/Toll-like receptor 4 （TLR4）/nuclear factor-κB （NF-κB） signaling pathway， and to explore differences in therapeutic efficacy among the three monomers， their combined formula， and HLJD under equal content ratios. MethodsBV2 microglial cells were used as the primary experimental model. Cell viability was assessed using the cell counting kit-8 （CCK-8） method to examine the effects of different concentrations of dimethyl sulfoxide （DMSO， 0.8%， 0.4%， 0.2%， 0.1%， and 0.05%） on cell viability. IncuCyte was employed to monitor the growth of cells under different concentrations of HLJD （200， 100， 50， 25， 12.5， 6.25 mg·L^-1）. Nitric oxide （NO） assay was used to screen the optimal HLJD concentration. High-performance liquid chromatography （HPLC） determined the content of geniposide， baicalin， and berberine in HLJD， and experimental groups were subsequently established according to the relative proportions of these constituents. CCK-8 assay evaluated cell viability under different treatments. Enzyme-linked immunosorbent assay （ELISA） measured levels of inflammatory factors （TNF-α， IL-1β， IL-6， IL-10） in the supernatant. Flow cytometry assessed the effects of treatments on M1-type polarization of BV2 cells. Western blot determined the expression levels of HMGB1， TLR4， and NF-κB-related proteins. ResultsCompared with the blank group， DMSO at concentrations ≤0.2% did not affect cell viability within 48 h. BV2 cell growth plateaued at 24 h after treatment with 200 mg·L^-1 HLJD. Under stimulation with 2 mg·L^-1 LPS， this concentration of HLJD effectively reduced NO release， and 6 h pre-treatment had a stronger inhibitory effect on NO than direct administration. HPLC results showed that 1 mg of HLJD freeze-dried powder contained approximately 24 μg of geniposide， 15 μg of baicalin， and 30 μg of berberine. Based on these ratios， experimental groups were blank， LPS （2 mg·L^-1）， HLJD （200 mg·L^-1）， monomer combination， geniposide （4.8 mg·L^-1）， baicalin （3 mg·L^-1）， and berberine （6 mg·L^-1）. The monomer combination group consisted of all three active constituents dissolved together. LPS and HLJD or its active constituents did not affect cell viability compared with the blank group. LPS significantly increased TNF-α， IL-1β， IL-6， and IL-10 in the supernatant （P<0.01）. HLJD and its active constituents significantly reduced pro-inflammatory factors TNF-α， IL-1β， and IL-6 （P<0.05， P<0.01） while upregulating anti-inflammatory IL-10 （P<0.01）， with the monomer combination showing the strongest effect （P<0.05， P<0.01）. Compared with the blank group， LPS significantly increased the proportion of CD80⁺CD86⁺ （M1-type） BV2 cells （P<0.01）. HLJD and its constituents partially inhibited M1 polarization （P<0.05， P<0.01）， with the monomer combination exhibiting the most pronounced effect （P<0.05， P<0.01）. Compared with the blank group， LPS upregulated HMGB1， TLR4， and NF-κB-related proteins （P<0.01）， whereas HLJD and its active constituents significantly reduced their expression （P<0.05， P<0.01）， with the monomer combination having the strongest regulatory effect （P<0.05， P<0.01）. ConclusionHLJD and its major active constituents （geniposide， baicalin， berberine） can inhibit LPS-induced inflammatory responses in BV2 cells. The combination of the three active constituents demonstrates the most potent anti-inflammatory effect， significantly attenuating M1-type polarization of BV2 cells via the HMGB1/TLR4/NF-κB signaling pathway.

Gene regulation relies on the precise binding of transcription factors (TFs) at regulatory elements, but simultaneously detecting hundreds of TFs on chromatin is challenging. We developed cFOOT-seq, a cytosine deaminase-based TF footprinting assay, for high-resolution, quantitative genome-wide assessment of TF binding in both open and closed chromatin regions, even with small cell numbers. By utilizing the dsDNA deaminase SsdAtox, cFOOT-seq converts accessible cytosines to uracil while preserving genomic integrity, making it compatible with techniques like ATAC-seq for sensitive and cost-effective detection of TF occupancy at the single-molecule and single-cell level. Our approach enables the delineation of TF footprints, quantification of occupancy, and examination of chromatin influences on TF binding. Notably, cFOOT-seq, combined with FootTrack analysis, enables de novo prediction of TF binding sites and tracking of TF occupancy dynamics. We demonstrate its application in capturing cell type-specific TFs, analyzing TF dynamics during reprogramming, and revealing TF dependencies on chromatin remodelers. Overall, cFOOT-seq represents a robust approach for investigating the genome-wide dynamics of TF occupancy and elucidating the cis-regulatory architecture underlying gene regulation.
Transcription Factors/genetics* ; Humans ; Chromatin/genetics* ; Animals ; Binding Sites ; Mice ; DNA Footprinting/methods*

Surgical methods for varicocele remain controversial. This study intends to evaluate the efficacy and safety of different surgical approaches for treating varicocele through a network meta-analysis (NMA). PubMed, Embase, Cochrane, and Web of Science databases were thoroughly searched. In total, 13 randomized controlled trials (RCTs) and 24 cohort studies were included, covering 9 different surgical methods. Pairwise meta-analysis and NMA were performed by means of random-effects models, and interventions were ranked based on the surface under the cumulative ranking curve (SUCRA). According to the SUCRA, microsurgical subinguinal varicocelectomy (MSV; 91.6%), microsurgical retroperitoneal varicocelectomy (MRV; 78.2%), and microsurgical inguinal varicocelectomy (MIV; 76.7%) demonstrated the highest effectiveness in reducing postoperative recurrence rates. In this study, sclerotherapy embolization (SE; 87.2%), MSV (77.9%), and MIV (67.7%) showed the best results in lowering the risk of hydrocele occurrence. MIV (82.9%), MSV (75.9%), and coil embolization (CE; 58.7%) were notably effective in increasing sperm motility. Moreover, CE (76.7%), subinguinal approach varicocelectomy (SV; 69.2%), and SE (55.7%) were the most effective in increasing sperm count. SE (82.5%), transabdominal laparoscopic varicocelectomy (TLV; 76.5%), and MRV (52.7%) were superior in shortening the length of hospital stay. The incidence rates of adverse events for MRV (0), SE (3.3%), and MIV (4.1%) were notably low. Cluster analyses indicated that MSV was the most effective in the treatment of varicocele. Based on the existing evidence, MSV may represent the optimal choice for varicocele surgery. However, selecting clinical surgical strategies requires consideration of various factors, including patient needs, surgeon experience, and the learning curve.
Humans ; Male ; Embolization, Therapeutic/methods* ; Microsurgery/methods* ; Randomized Controlled Trials as Topic ; Sclerotherapy/methods* ; Treatment Outcome ; Urologic Surgical Procedures, Male/methods* ; Varicocele/surgery*

OBJECTIVE: To investigate the application value of bronchoalveolar lavage fluid (BALF) metagenomic next-generation sequencing (mNGS) in etiological diagnosis of children with hematological malignancies complicated with pneumonia. METHODS: We retrospectively analyzed the clinical data of children with hematological malignancies who underwent BALF mNGS pathogenic detection due to pneumonia. All patients underwent mNGS detection of bronchoalveolar lavage fluid as well as traditional methods(including sputum culture, bronchoalveolar lavage fluid culture, blood culture, serological detection of pathogens, etc.). By analyzing the results of mNGS and traditional methods, we compared key indicators such as the positive rate, etiological distribution. RESULTS: A total of 26 children with hematological malignancies enrolled in the study, including 12 males and 14 females, with a median age of 4.9 (1.8-14.9) years, underwent bronchoalveolar lavage (BAL) 35 times. A total of 17 pathogenic microorganisms were detected in BALF mNGS, including 9 cases of bacterial infection, 10 cases of viral infection, 3 cases of fungal infection, 2 cases of mycoplasma infection and 8 cases of mixed infection, and the most commonly detected bacteria, viruses and fungi were streptococcus pneumoniae, cytomegalovirus and pneumocystis jirovecii, respectively. The positive rate of mNGS detection (91.43%) was significantly higher than that of traditional methods detection (20%, P <0.001). A total of 25 cases were adjusted according to BALF mNGS results. CONCLUSION The application of BALF mNGS technology can improve the detection rate of the pathogens in children with hematological malignancies complicated with pneumonia, initially revealed the pathogen spectrum of pulmonary infection in this group, and effectively guide clinical medication, improve treatment outcomes.
Humans ; Bronchoalveolar Lavage Fluid/microbiology* ; Hematologic Neoplasms/complications* ; Child ; Child, Preschool ; Infant ; Retrospective Studies ; Male ; Female ; Adolescent ; High-Throughput Nucleotide Sequencing ; Pneumonia/microbiology*

OBJECTIVES: This study aims to analyze the biomechanics of three kinds of rigid internal fixation methods for condylar head fractures. METHODS: A three dimensional finite element model of the normal mandible was constructed. It was then used to prepare condylar head fracture finite element model and three kinds of rigid internal fixation finite element model (unilateral tension screw, bilateral tension screw, tension screw+titanium plate). The mechanical characteristics and changes of the mandible condyle under the same mechanical conditions were compared among the three different rigid internal fixation methods. RESULTS: The maximum equivalent stress and displacement of the non-free end of condyle under the rigid internal fixation method of unilateral tension screw were 71.03 MPa and 4.72 mm, respectively. The maximum equivalent stress and displacement of the free end of condyle were 78.45 MPa and 4.50 mm, respectively. The maximum stress of fracture suture was 3.27 MPa. The maximum equivalent stress and displacement of the non-free end of condyle under the rigid internal fixation method of bilateral tension screw were 70.52 MPa and 4.00 mm, respectively. The maximum equivalent stress and displacement of the free end of condyle were 72.49 MPa and 3.85 mm, respectively. The maximum stress of fracture suture was 2.33 MPa. The maximum equivalent stress and maximum displacement of the non-free end of condyle under the rigid internal fixation method of tension screw+titanium plate were 67.26 MPa and 2.66 mm, respectively. The maximum equivalent stress and maximum displacement of the free end of condyle were 69.66 MPa and 2.50 mm, respectively. The maximum stress of fracture suture was 2.18 MPa. CONCLUSIONS The tension screw+titanium plate rigid internal fixation method is the most conducive to biomechanical distribution for condylar head fractures.
Fracture Fixation, Internal/instrumentation* ; Mandibular Condyle/surgery* ; Biomechanical Phenomena ; Bone Screws ; Finite Element Analysis ; Humans ; Mandibular Fractures/surgery* ; Bone Plates ; Titanium ; Stress, Mechanical

This study aims to investigate the therapeutic effects of the Coptidis Rhizoma-Scutellariae Radix on cytokine storm secondary lung injury(CSSLI) induced by CpG1826 in mice, and to elucidate the potential molecular mechanisms by which its major active components, i.e., coptisine and wogonin, alleviate CSSLI by inhibiting the mitochondrial permeability transition pore(mPTP)/nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3) inflammasome pyroptosis pathway. In vivo, a mouse model of CSSLI was established by CpG1826 induction. Pulmonary edema was assessed by lung wet-to-dry weight ratio(W/D), lung injury was evaluated by hematoxylin-eosin(HE) staining, and ultrastructural changes in lung tissue were observed by transmission electron microscopy(TEM). The levels of interleukin(IL)-1β, high mobility group box 1 protein(HMGB1), IL-18, and IL-1α in bronchoalveolar lavage fluid were measured by enzyme-linked immunosorbent assay(ELISA). The results showed that the decoction of the Coptidis Rhizoma-Scutellariae Radix significantly reduced pulmonary edema, alleviated lung injury, and decreased the concentrations of related cytokines in BALF more effectively than either single herb alone, thereby improving CSSLI. In vitro, a CpG1826-induced CSSLI model was established in mouse alveolar macrophage MH-S cells. Calcein-AM quenching was used to screen for the most effective monomer components from the herb pair in inhibiting mPTP opening. Coptisine(5, 10, 20 μmol·L~(-1)) and wogonin(10, 20, 40 μmol·L~(-1)) markedly inhibited mPTP opening, with optimal effects and a clear dose-dependent pattern. These components suppressed mPTP opening, thereby reducing the release of mitochondrial DNA(mtDNA) and the accumulation of reactive oxygen species(ROS), effectively reversing the CpG1826-induced decrease in mitochondrial membrane potential(MMP). Further studies revealed that both coptisine and wogonin inhibited pyroptosis and downregulated the expression of key proteins in the NLRP3/Caspase-1/gasdermin D(GSDMD) pathway. In conclusion, the Coptidis Rhizoma-Scutellariae Radix improves CpG1826-induced CSSLI in mice, and this effect is associated with the inhibition of the mPTP/NLRP3 pyroptosis pathway, providing scientific evidence for its clinical application and further development.
Animals ; Mice ; Drugs, Chinese Herbal/administration & dosage* ; Pyroptosis/drug effects* ; NLR Family, Pyrin Domain-Containing 3 Protein/immunology* ; Male ; Lung Injury/immunology* ; Cytokines/immunology* ; Scutellaria baicalensis/chemistry* ; Oligodeoxyribonucleotides/adverse effects* ; Mice, Inbred C57BL ; Coptis chinensis

Sepsis is a systemic inflammatory response caused by severe infection or trauma, and is one of the common causes of acute lung injury(ALI) and acute respiratory distress syndrome(ARDS). Sepsis-acute lung injury(SALI) is a critical clinical condition with high morbidity and mortality. Its pathogenesis is complex and not yet fully understood, and there is currently a lack of targeted and effective treatment options. Pyroptosis, a novel form of programmed cell death, plays a key role in the pathological process of SALI by activating inflammasomes and releasing inflammatory factors, making it a potential therapeutic target. In recent years, the role of traditional Chinese medicine(TCM) in regulating signaling pathways related to pyroptosis through multi-components and multi-targets has attracted increasing attention. TCM may intervene in pyroptosis by inhibiting the activation of NLRP3 inflammasomes and regulating the expression of Caspase family proteins, thus alleviating inflammatory damage in lung tissues. This paper systematically reviews the molecular regulatory network of pyroptosis in SALI and explores the potential mechanisms and research progress on TCM intervention in cellular pyroptosis. The aim is to provide new ideas and theoretical support for basic research and clinical treatment strategies of TCM in SALI.
Pyroptosis/drug effects* ; Humans ; Sepsis/genetics* ; Acute Lung Injury/physiopathology* ; Animals ; Drugs, Chinese Herbal/therapeutic use* ; Medicine, Chinese Traditional ; Inflammasomes/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics*

OBJECTIVE T o explore the action mechanisms of Ganghuo Qingwen granules(GHQWG)in treatment of influenza based on network pharmacology and molecular docking technique.METHODS The TCMSP database was logged in to search for active compounds,targets of GHQWG and targets of influenza so as to screen out the intersected targets between HGQWG and influenza and establish a drug-ingredient-target network.The poten-tial mechanisms were analyzed by means of protein-protein interaction analysis,Gene Ontology(GO)function an-notation and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment,and the activity of binding the key targets with ingredients was validated by molecular docking.RESULTS GHQWG contained 90 types of po-tential active ingredients,312 active ingredient targets,1996 influenza targets and 134 intersection targets.The core ingredients included quercetin,kaempferol and luteolin;the key targets involved V-Rel reticuloendotheliosis viral oncogene homolog A(RELA),tumor protein 53(TP53),mitogen-activated protein kinase 3(MAPK3),tumor necrosis factor(TNF),serine/threonine kinase proteins 1(AKT1)and mitogen-activated protein kinase 1(MAPK1).The function enrichment showed that GHQWG could regulate the processes of gene expression,in-flammatory reaction and apoptosis.The result of KEGG indicated that GHQWG played therapeutic effect on influ-enza mainly through C type agglutinin receptor,TNF and in terleukin-17(IL-17)signaling pathway.The molecu-lar docking analysis showed that RELA,TP53,M APK3,TNF,AKT1 and M APK1 had strong activities of bind-ing with quercetin,kaempferol,luteolin,β-sitosterol,wogonin,stigmasterol,caffeic acid and isorhamnetin.CONCLUSION GHQWG may regulate the key targets of signaling pathways of C type agglutinin receptor,TNF and IL-17 through multiple ingredients-multiple targes-multiple pathway mechanisms so as to play the therapeutic effect on influenza.

Objective:To investigate the relationship between serum insulin-like growth factor-1 (IGF-1) levels and intracranial or extracranial atherosclerosis in patients with cerebral small vessel disease (CSVD).Methods:A total of 407 patients with CSVD admitted to Xiangya Hospital of Central South University between July 2021 and September 2023 were enrolled in the study. Carotid duplex ultrasound was used to measure the internal diameter, intima-media thickness (IMT), vascular wall thickness, plaque property score, stenosis index, and stenosis ratio of the bilateral common carotid arteries, internal carotid arteries, external carotid arteries, and vertebral arteries. Magnetic resonance angiography was used to assess the degree of stenosis in intracranial arteries. Patients were divided into 4 groups based on the serum IGF-1 levels (low level group:≤5.21 ng/ml, medium level group:>5.21 ng/ml and ≤10.73 ng/ml, high level group:>10.73 ng/ml and ≤24.26 ng/ml, extremely high level group:>24.26 ng/ml). The IMT of the common carotid artery, carotid plaques, diameters of various cervical vascular lumens, carotid artery diameter stenosis, and intracranial artery stenosis in 4 groups of the patients were compared. The relationship between IGF-1 and intracranial and extracranial atherosclerosis was analyzed by univariate Logistic regression analysis and multivariate Logistic regression analysis.Results:There were inter group differences among the 4 groups in internal carotid artery diameter [low level group 5.45 (0.50) mm vs medium level group 5.32 (0.55) mm vs high level group 5.30 (0.55) mm vs extremely high level group 5.30 (0.50) mm; H=8.210, P=0.042]. The carotid IMT [low level group 0.80 (0.05) mm vs medium level group 0.80 (0.05) mm vs high level group 0.83 (0.03) mm vs extremely high level group 0.83 (0.09) mm; H=8.107, P=0.044], the proportion of carotid artery vascular wall thickening [low level group 52.9%(54/102) vs medium level group 48.0%(49/102) vs high level group 68.3%(69/101) vs extremely high level group 60.8%(62/102); χ2=9.889, P=0.020], the carotid artery plaque property score [low level group 1 (2) vs medium level group 2 (2) vs high level group 2 (2) vs extremely high level group 2 (2); H=8.913, P=0.030] and the proportion of anterior cerebral artery stenosis [low level group 2.9%(3/102) vs medium level group 2.0%(2/102) vs high level group 4.0%(4/101) vs extremely high level group 10.8%(11/102); χ2=10.473, P=0.014] had inter group differences among the 4 groups, and the differences were statistically significant. Univariate Logistic regression analysis indicated that carotid artery vascular wall thickening ( OR=1.197, 95% CI 1.003-1.429, P=0.046), anterior cerebral artery stenosis ( OR=1.814, 95% CI 1.148-2.867, P=0.011), and basilar artery stenosis ( OR=1.530, 95% CI 1.084-2.159, P=0.015) were correlated with IGF-1 levels. Multivariate Logistic regression analysis revealed that after adjusting for age, gender, low-density lipoprotein cholesterol (LDL-C), and C-reactive protein, IGF-1 was positively correlated with the carotid artery vascular wall thickening ( OR=1.311, 95% CI 1.014-1.696, P=0.039); after adjusting for age, IGF-1 was positively correlated with the anterior cerebral artery stenosis ( OR=2.130, 95% CI 1.201-3.776, P=0.010); after adjusting for gender, low-density lipoprotein cholesterol, and cholesterol levels, IGF-1 was positively correlated with basilar artery stenosis ( OR=1.688, 95% CI 1.063-2.681, P=0.027). Conclusions:There is an association between IGF-1 levels and intracranial and extracranial atherosclerosis in patients with CSVD. IGF-1 may play a role in the development and progression of atherosclerosis in CSVD.

AIM To study the effect of Zuogui Pills on improving ovarian function in rats with premature ovarian failure(POF)by regulating autophagy.METHODS Seven rats were randomly selected as the blank group,and the remaining rats were injected with cisplatin(4.0 mg/kg)intraperitoneally to establish POF model.The success of the model was evaluated by observing the changes of estrous cycle.Twenty-one successful model of rats were randomly divided into model group and estradiol group(0.01 mg/mL)and Zuogui Pills group(1.85 g/kg),with 7 rats in each group,the drug was administered continuously for 21 days.Serum E2,FSH and LH levels were detected by ELISA;HE staining was used to observe the pathological morphology of ovarian tissue;immunohistochemical(IHC)method was used to detect the protein expressions of LC3B,SIRT1 and FoxO1 in ovarian tissues;RT-qPCR method was used to detect the mRNA expressions of LC3B,Beclin-1 and Atg5 in ovarian tissues;the protein expressions of SIRT1,FoxO1 and Ac-FoxO1 in ovarian tissue was detected by Western blot.RESULTS Compared with the blank group,the ovarian index of the model group decreased(P＜0.01);serum FSH and LH levels increased(P＜0.01)and E2 level decreased(P＜0.01);the structure of ovary is disordered,especially atresia follicle;the mRNA expressions of LC3B,Beclin-1 and Atg5 in ovarian tissue increased(P＜0.01),while the protein expressions of LC3B and Ac-FoxO1 increased(P＜0.01),while the protein expressions of SIRT1 and FoxO1 decreased(P＜0.01).Compared with the model group,the ovarian index of rats in estradiol group and Zuogui Pills group increased(P＜0.01);serum FSH and LH levels decreased(P＜0.01)and E2 level increased(P＜0.01);the number of primordial follicles in ovary increased and the number of atresia follicles decreased;the mRNA expressions of LC3B,Beclin-1 and Atg5 in ovarian tissue decreased(P＜0.01),while the protein expressions of LC3B and Ac-FoxO1 decreased(P＜0.01),while the protein expressions of SIRT1 and FoxO1 increased(P＜0.01).CONCLUSION Zuogui Pills may inhibit autophagy by activating SIRT1/FoxO1 signaling pathway,thus improving the pathological state of POF rats,regulating the level of sex hormones in rats,restoring endocrine balance,enhancing ovarian reserve function,promoting the normal development of follicles and delaying the progress of POF.