Network pharmacology and molecular docking were employed to predict the mechanism of Zhizhu Decoction in regulating macrophage polarization to reduce adipose tissue inflammation in obese children, and animal experiments were then carried out to validate the prediction results. The active ingredients and targets of Zhizhu Decoction were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP). The inflammation related targets in the adipose tissue of obese children were searched against GeneCards, OMIM, and DisGeNET, and a drug-disease-target network was established. STRING was used to construct a protein-protein interaction(PPI) network and screen for core targets. R language was used to carry out Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analyses. AutoDock was used for the molecular docking between core targets and active ingredients. 24 SPF grade 6-week C57B/6J male mice were adaptively fed for 1 week, and 8 mice were randomly selected as the blank group. The remaining 16 mice were fed with high-fat diet for 8 weeks to onstruct a high-fat diet induced mouse obesity model. After successful modeling, the 16 mice were randomly divided into model group and Zhizhu Decoction group, with 8 mice in each group. Zhizhu Decoction group was intervened by gavage for 14 days, once a day. Blank group and model group were given an equal amount of sterile double distilled water(ddH_2O) by gavage daily. After the last gavage, serum and inguinal adipose tissue were collected from mice for testing. The morphology of inguinal adipose tissue was observed by hematoxylin-eosin(HE) staining, the levels of inflammatory factors interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α)were detected by enzyme-linked immunosorbent assay(ELISA), and the protein expression of macrophage marker molecule nitric oxide synthase(iNOS) and epidermal growth factor like hormone receptor 1(F4/80) was detected by immunofluorescence staining. Network pharmacology predicted luteolin, naringenin, and nobiletin as the main active ingredients in Zhizhu Decoction and 15 core targets. KEGG pathway enrichment analysis revealed involvement in the key signaling pathway of nuclear factor κB(NF-κB). Molecular docking showed that the active ingredients of Zhizhu Decoction bound well to the core targets. Animal experiment showed that compared with the model group, Zhizhu Decoction reduced the distribution of inflammatory cytokines in the inguinal adipose tissue of mice, lowered the levels of TNF-α and IL-6 in the serum(P<0.05, P<0.01), and down-regulated the expression of iNOS and F4/80(P<0.05). The results showed that the active ingredients in Zhizhu Decoction, such as luteolin, naringenin, and nobiletin, inhibit the aggregation of macrophages in adipose tissue, downregulate their classic activated macrophage(M1) polarization, reduce the expression of inflammatory factors IL-6 and TNF-α, and thus improve adipose tissue inflammation in obese mice.
Animals ; Drugs, Chinese Herbal/pharmacology* ; Molecular Docking Simulation ; Adipose Tissue/immunology* ; Mice ; Male ; Humans ; Network Pharmacology ; Macrophages/immunology* ; Mice, Inbred C57BL ; Child ; Protein Interaction Maps/drug effects* ; Obesity/genetics* ; Inflammation/drug therapy*

Buyang Huanwu Decoction(BYHWD), as one of the classic formulas in traditional Chinese medicine(TCM) for the treatment of cerebral ischemic stroke(CIS), has demonstrated definite effects in clinical practice. However, the material basis and mechanism of treatment have not been systematically elucidated. This study employed network pharmacology and molecular docking to analyze the potential targets and mechanisms of blood-and brain-penetrating active components of BYHWD in reducing cell apoptosis in CIS. Cell experiments were then carried out to validate the prediction results. In the experiments, five active components including hydroxysafflor yellow A( HSYA), tetramethylpyrazine( TMP), astragaloside Ⅳ( AS-Ⅳ), amygdalin( AMY), and paeoniflorin(PF) were selected to explore the pharmacological effects of BYHWD. HT22 cells were treated with BYHWD, and the cell counting kit-8(CCK-8) method was employed to examine the toxic and side effects of BYHWD. A cell model of oxygen-glucose deprivation/reoxygenation( OGD/R) was constructed, with apoptosis and pyroptosis as the main screening indicators. The levels of lactate dehydrogenase(LDH) and glutathione(GSH) were measured to assess the cell membrane integrity. Flow cytometry was employed to detect apoptosis, and the activities of caspase-3 and caspase-1 were measured to clarify the status of apoptosis and pyroptosis. ELISA was employed to determine the levels of interleukin(IL)-1β and IL-18 to confirm pyroptosis. HSYA and AMY were identified in this study as the active components regulating apoptosis and pyroptosis. TUNEL was employed to detect the apoptosis rate, and Western blot was employed to determine the expression levels of apoptosis-related proteins B-cell lymphoma-2(Bcl-2), Bcl-2-associated X protein(Bax), and caspase-3, which confirmed that the anti-apoptotic effect of the combined component group was superior to that of the single component groups. The molecular docking results revealed strong binding affinity of HSYA and AMY with SDF-1α and CXCR4.AMD3100, a selective antagonist of CXCR4, was then used for intervention. The results of Western blot showed alterations in the expression levels of apoptosis-associated proteins, SDF-1α, and CXCR4. In conclusion, HSYA and AMY influence cellular apoptosis by modulating the SDF-1α/CXCR4 signaling cascade.
Drugs, Chinese Herbal/chemistry* ; Apoptosis/drug effects* ; Animals ; Neurons/cytology* ; Mice ; Molecular Docking Simulation ; Cell Line ; Glucose/metabolism* ; Humans ; Neuroprotective Agents/pharmacology*

This study aims to explore the effects and action mechanisms of the active ingredients in Buyang Huanwu Decoction(BYHWD), namely tetramethylpyrazine(TMP) and hydroxy-safflor yellow A(HSYA), on oxygen-glucose deprivation/reglucose-reoxygenation(OGD/R)-induced inflammation and oxidative stress of microglia(MG). Network pharmacology was used to screen the effective monomer ingredients of BYHWD and determine the safe concentration range for each component. Inflammation and oxidative stress models were established to further screen the best ingredient combination and optimal concentration ratio with the most effective anti-inflammatory and antioxidant effects. OGD/R BV2 cell models were constructed, and BV2 cells in the logarithmic growth phase were divided into a normal group, a model group, an HSYA group, a TMP group, and an HSYA + TMP group. Enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of inflammatory cytokines such as interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), and interleukin-6(IL-6). Oxidative stress markers, including superoxide dismutase(SOD), nitric oxide(NO), and malondialdehyde(MDA), were also measured. Western blot was used to analyze the protein expression of both inflammation-related pathway [Toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB)] and oxidative stress-related pathway [nuclear factor erythroid 2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)]. Immunofluorescence was used to assess the expression of proteins such as inducible nitric oxide synthase(iNOS) and arginase-1(Arg-1). The most effective ingredients for anti-inflammatory and antioxidant effects in BYHWD were TMP and HSYA. Compared to the normal group, the model group showed significantly increased levels of IL-1β, TNF-α, IL-6, NO, and MDA, along with significantly higher protein expression of NF-κB, TLR4, Nrf2, and HO-1 and significantly lower SOD levels. The differences between the two groups were statistically significant. Compared to the model group, both the HSYA group and the TMP group showed significantly reduced levels of IL-1β, TNF-α, IL-6, NO, and MDA, lower expression of NF-κB and TLR4 proteins, higher levels of SOD, and significantly increased protein expression of Nrf2 and HO-1. Additionally, the expression of the M1-type MG marker iNOS was significantly reduced, while the expression of the M2-type MG marker Arg-1 was significantly increased. The results of the HSYA group and the TMP group had statistically significant differences from those of the model group. Compared to the HSYA group and the TMP group, the HSYA + TMP group showed further significant reductions in IL-1β, TNF-α, IL-6, NO, and MDA levels, along with significant reductions in NF-κB and TLR4 protein expression, an increase in SOD levels, and elevated Nrf2 and HO-1 protein expression. Additionally, the expression of the M1-type MG marker iNOS was reduced, while the M2-type MG marker Arg-1 expression increased significantly in the HSYA + TMP group compared to the TMP or HSYA group. The differences in the results were statistically significant between the HSYA + TMP group and the TMP or HSYA group. The findings indicated that the combined use of HSYA and TMP, the active ingredients of BYHWD, can effectively inhibit OGD/R-induced inflammation and oxidative stress of MG, showing superior effects compared to the individual use of either component.
Oxidative Stress/drug effects* ; Drugs, Chinese Herbal/pharmacology* ; Animals ; Mice ; Glucose/metabolism* ; Cell Line ; Inflammation/genetics* ; Oxygen/metabolism* ; Pyrazines/pharmacology* ; Microglia/metabolism* ; NF-E2-Related Factor 2/immunology* ; NF-kappa B/immunology* ; Toll-Like Receptor 4/immunology* ; Anti-Inflammatory Agents/pharmacology* ; Humans

This article reports the clinical features and gene mutation types of a large family with Nascimento form of syndromic X-linked intellectual developmental disorder (MRXSN), involving 9 individuals across 3 generations, and a literature review was conducted. In this family, 9 individuals had similar manifestations including mental retardation and unusual facies, and 4 of them had passed away. Genetic testing showed that the proband had the deletion of exons 2-3 of the UBE2A gene, which was inherited from the mother. Fluorescent quantitative polymerase chain reaction showed that the proband and his uncle had the deletion of exons 2-3 of the UBE2A gene; the proband's mother, grandmother, and great-aunt had a heterozygous deletion of exons 2-3 of the UBE2A gene; the proband's father, sister, and aunt had a normal copy number of exons 2-3 of the UBE2A gene. The 34 patients reported in the literature had diverse clinical phenotypes, and UBE2A gene mutations (22/34, 65%) and large fragment deletions (12/34, 35%) were the main mutation types. Moderate to severe mental retardation (34/34, 100%), speech and language impairment (33/34, 97%), and unusual facies (32/34, 94%) were the main clinical manifestations of MRXSN patients. The disease has obvious phenotypic heterogeneity, and early diagnosis facilitates optimal prenatal and postnatal management to improve reproductive outcomes.
Humans ; Male ; Ubiquitin-Conjugating Enzymes/genetics* ; Female ; X-Linked Intellectual Disability/genetics* ; Gene Deletion ; Child ; Pedigree ; Child, Preschool ; Adult

Objective:To report the nationwide surveillance results of pathogenic profiles and antimicrobial resistance patterns of Gram-positive bloodstream infections in China in 2023.Methods:The clinical isolates of Gram-posttive bacteria from blood cultures were collected in member hospitals of National Bloodstream Infection Bacterial Resistant Investigation Collaborative System（BRICS）during January to December 2023. Antimicrobial susceptibility testing was performed using the dilution method recommended by the Clinical and Laboratory Standards Institute（CLSI）. Statistical analyses were conducted using WHONET 5.6 and SPSS 25.0 software.Results:A total of 4 385 Gram-positive bacterial isolates were obtained from 60 participating center. The top five pathogens were Staphylococcus aureus（ n=1 544，35.2%），coagulase-negative Staphylococci（ n=1 441，32.9%）， Enterococcus faecium（ n=574，13.1%）， Enterococcus faecalis（ n=385，8.8%），and α-hemolytic Streptococci（ n=187，4.3%）. The prevalence of methicillin-resistant Staphylococcus aureus（MRSA）and methicillin-resistant coagulase-negative Staphylococci（MRCNS）was 26.2%（405/1 544）and 69.8%（1 006/1 441），respectively. Notably，all Staphylococci remained susceptible to glycopeptide or daptomycin. Staphylococcus aureus demonstrated excellent susceptibility（>97.0%）to cephalobiol，rifampicin，trimethoprim-sulfamethoxazole，linezolid，minocycline，tigecycline，and eravacycline. No Enterococcus exhibiting resistance to linezolid were detected. Glycopeptide resistance was uncommon but more frequent in Enterococcus faecium（resistance to vancomycin and teicoplanin：both 1.7%）compared to Enterococcus faecalis（both 0.3%）. The detection rates of MRSA and MRCNS exhibited significant regional variations across the country（ χ2=17.674 and 148.650，respectively，both P<0.001）. No vancomycin-resistant Enterococci were detected in central China. Institutional comparison demonstrated higher prevalence of MRSA（ χ2=14.111， P<0.001）and MRCNS（ χ2=4.828， P=0.028）in provincial hospitals than that in municipal hospitals. Socioeconomic analysis identified elevated detection rates of both MRSA（ χ2=18.986， P<0.001）and MRCNS（ χ2=4.477， P=0.034）in less developed regions（per capita GDP

Objective:To report the results of bacterial resistant investigation collaborative system（BRICS）on the distribution and antimicrobial resistance profile of clinical Gram-negative bacteria isolates from bloodstream infections in China in 2023，and provide reference for clinical tretment of bloodstream infections and prevention and control of bacterial resistance.Methods:The clinical isolates of Gram-negative bacteria from blood cultures in member hospitals of BRICS were collected during January 2023 to December 2023. Antibiotic susceptibility tests were conducted by agar dilution or broth dilution methods recommended by Clinical and Laboratory Standards Institute（CLSI）. WHONET 5.6 and SPSS 25.0 were used to analyze the data.Results:During the study period，11 492 strains of Gram-negative bacteria were collected from 60 hospitals，of which 10 098（87.9%）were Enterobacterales and 1 394（12.1%）were non-fermentative bacteria. The top 5 bacterial species were Escherichia coli（50.0%）， Klebsiella pneumoniae（26.1%）， Pseudomonas aeruginosa（5.1%）， Acinetobacter baumannii complex（5.0%）and Enterobacter cloacae complex（4.1%）. The ESBL-producing rates in Escherichia coli， Klebsiella pneumoniae and Proteus mirablilis were 46.8%（2 685/5 741），18.3%（549/2 999）and 44.0%（77/175），respectively. The prevalence of carbapenem-resistant Escherichia coli（CREC）and carbapenem-resistant Klebsiella pneumoniae（CRKP）were 1.3%（76/5 741）and 15.0%（450/2 999）；32.9%（25/76）and 78.0%（351/450）of CREC and CRKP were sensitive to ceftazidime/avibactam combination，respectively. 94.7%（72/76）and 90.2%（406/450）of CREC and CRKP were sensitive to aztreonam/avibactam combination. Furthermore，57.9%（44/76）and 79.1%（356/450）were sensitive to imipenem/relebactam combination. The prevalence of carbapenem-resistant Acinetobacter baumannii（CRAB）complex was 64.6%（370/573），while more than 80.0% of CRAB complex was sensitive to tigecycline，eravacycline and polymyxin B. The prevalence of carbapenem-resistant Pseudomonas aeruginosa（CRPA）was 17.0%（99/581）. There were differences in the composition ratio of Gram-negative bacteria in bloodstream infections and the prevalence of important Gram-negative bacteria resistance among different regions in China，with statistically significant differences in the prevalence of CREC，CRKP，CRPA and CRAB complex（ χ2=10.6，28.6，10.8 and 19.3， P<0.05）. The prevalence of ESBL-producing Escherichia coli， CREC，CRAB complex and CRKP were higher in provincial hospitals than those in municipal hospitals（ χ2=12.5，9.8，12.7 and 57.8，all P<0.01）. Conclusions:Gram-negative bacteria are the main pathogens causing bloodstream infections in China，and Escherichia coli is ranked in the top，while the trend of Klebsiella pneumoniae increases continuously with time. CRKP infection shows a slow upward trend，CREC infecton maintains a low prevalence level，and CRAB complex infection continues to exhibit a high prevalence rate. The composition and resistance patterns of pathogens causing bloodstream infections vary to some extent across different regions and levels of hospitals in China.

Gastric cancer remains one of the most prevalent and lethal malignancies of the digestive tract worldwide,underscoring the urgent need for more effective targeted therapeutic strategies.Poly(ADP-ri-bose)polymerase(PARP)inhibitors have demonstrated remarkable efficacy in tumors with homologous recombination repair(HRR)deficiency;however,their clinical application in gastric cancer remains limited.Clinical evidence suggests that patients harboring Helicobacter pylori infection in combination with HRR gene mutations exhibit a significantly elevated risk of developing gastric cancer,thereby supporting the potential benefit of PARP inhibition in this setting.In this study,a kinase inhibitor library was screened in combination with the PARP inhibitor olaparib in gastric cancer cells.And we identify the cy-clin-dependent kinase 8/19(CDK8/19)inhibitor Senexin A as a compound that synergistically enhances the cytotoxic effect of PARP inhibition(P<0.05).Phenotypic validation using CCK-8 and colony for-mation assays demonstrated that the combination treatment significantly suppressed cellular proliferation and clonogenic potential compared to either monotherapy(P<0.0001).Mechanistically,alkaline comet assays revealed a significant increase in DNA damage in the combination treatment group relative to either single-agent group(P<0.0001),suggesting that the synergistic effect results from the exacerbation of DNA damage via impaired DNA repair mechanisms.In addition,treatment with CDK8/19 inhibitors a-lone markedly increased the formation of γH2AX and 53BP1 foci in irradiated gastric cancer cells(P<0.0001),indicating inhibition of DNA damage repair pathways.Transcriptome sequencing further re-vealed that CDK8/19 inhibition impacts critical cellular pathways,including DNA repair,cell cycle reg-ulation,and RNA splicing.Co-immunoprecipitation assays confirmed that inhibition of CDK8/19 kinase activity significantly reduces the phosphorylation level of PARP1,suggesting a potential regulatory inter-action.Immunohistochemical analysis of tumor and adjacent non-tumor tissues from gastric cancer pa-tients demonstrated that CDK8 is significantly overexpressed in tumor tissues,supporting its potential as both a prognostic biomarker and a therapeutic target.Collectively,this study elucidates a mechanistic ba-sis by which CDK8/19 inhibition enhances the sensitivity of gastric cancer cells to PARP inhibitors.These findings provide a strong rationale for the combined use of CDK8/19 and PARP inhibitors as a tar-geted therapeutic strategy and offer promising translational implications for advancing personalized medi-cine in gastric cancer treatment.

Aim To explore the effect of hydroxysafflor yellow A(HSYA)on lipocalin 2(LCN2)-induced fer-roptosis in HT22 cells and the related mechanism.Methods Thirty male Sprague-Dawley(SD)rats were used to establish the middle cerebral artery occlu-sion/reperfusion(MCAO/R)model by the suture method.The rats were randomly divided into the Sham group,the MCAO/R group,and the MCAO/R+HSYA group.The infarct area was measured by TTC staining,and the degree of neurological deficit was evaluated by the Z-Longa scoring method.The expressions of LCN2 and 24P3R in brain tissues were detected by Western blot.LCN2 protein was added to HT-22 cells,and the cells were divided into the normal group,the LCN2 group,and the LCN2+HSYA group.The optimal con-centration of LCN2-induced neuronal ferroptosis was screened by LDH assay and Western blot,and the ex-pression levels of ferritin,FPN1,GPX4,SLC7A11,COX2,and 24P3R were detected.LCN2 was knocked down by siRNA transfection,and the expressions of GPX4 and ferritin were detected.The contents of glu-tathione(GSH),malondialdehyde(MDA),GPX4,and Fe2+were determined by colorimetry,and the expres-sion of GPX4 was detected by immunofluorescence.The binding force between HSYA and LCN2 was ana-lyzed by molecular docking technology.Results Ani-mal experiments showed that HSYA could reduce the cerebral infarction area and decrease the neurological function score of MCAO/R rats.Compared with the sham group,the levels of LCN2 and 24P3R increased in the MCAO/R group,while HSYA inhibited their ex-pressions.Cell experiments showed that the optimal concentration of LCN2 to induce ferroptosis in HT22 cells was 2 μmol·L-1.After knocking down LCN2 by siRNA transfection,compared with the LCN2 group,the expression levels of GPX4 and ferritin in the siLCN2 group increased significantly.Compared with the nor-mal group,the expressions of SLC7A11,GPX4,FPN1,ferritin,and GSH in the LCN2 group decreased signifi-cantly,while the concentration of Fe2+,and the expres-sions of MDA,COX2,and 24P3R increased.HSYA could increase the expressions of SLC7A11,GPX4,FPN1,ferritin,and GSH,reduce the contents of Fe2+and MDA,and inhibit the expressions of COX2 and 24P3R.Molecular docking showed that the binding en-ergy between HSYA and LCN2 was-8.0 kJ·mol-1.Conclusion HSYA can inhibit LCN2-induced ferrop-tosis in HT22 cells through the SLC7A11/GPX4 signa-ling pathway.

Gastric cancer remains one of the most prevalent and lethal malignancies of the digestive tract worldwide,underscoring the urgent need for more effective targeted therapeutic strategies.Poly(ADP-ri-bose)polymerase(PARP)inhibitors have demonstrated remarkable efficacy in tumors with homologous recombination repair(HRR)deficiency;however,their clinical application in gastric cancer remains limited.Clinical evidence suggests that patients harboring Helicobacter pylori infection in combination with HRR gene mutations exhibit a significantly elevated risk of developing gastric cancer,thereby supporting the potential benefit of PARP inhibition in this setting.In this study,a kinase inhibitor library was screened in combination with the PARP inhibitor olaparib in gastric cancer cells.And we identify the cy-clin-dependent kinase 8/19(CDK8/19)inhibitor Senexin A as a compound that synergistically enhances the cytotoxic effect of PARP inhibition(P<0.05).Phenotypic validation using CCK-8 and colony for-mation assays demonstrated that the combination treatment significantly suppressed cellular proliferation and clonogenic potential compared to either monotherapy(P<0.0001).Mechanistically,alkaline comet assays revealed a significant increase in DNA damage in the combination treatment group relative to either single-agent group(P<0.0001),suggesting that the synergistic effect results from the exacerbation of DNA damage via impaired DNA repair mechanisms.In addition,treatment with CDK8/19 inhibitors a-lone markedly increased the formation of γH2AX and 53BP1 foci in irradiated gastric cancer cells(P<0.0001),indicating inhibition of DNA damage repair pathways.Transcriptome sequencing further re-vealed that CDK8/19 inhibition impacts critical cellular pathways,including DNA repair,cell cycle reg-ulation,and RNA splicing.Co-immunoprecipitation assays confirmed that inhibition of CDK8/19 kinase activity significantly reduces the phosphorylation level of PARP1,suggesting a potential regulatory inter-action.Immunohistochemical analysis of tumor and adjacent non-tumor tissues from gastric cancer pa-tients demonstrated that CDK8 is significantly overexpressed in tumor tissues,supporting its potential as both a prognostic biomarker and a therapeutic target.Collectively,this study elucidates a mechanistic ba-sis by which CDK8/19 inhibition enhances the sensitivity of gastric cancer cells to PARP inhibitors.These findings provide a strong rationale for the combined use of CDK8/19 and PARP inhibitors as a tar-geted therapeutic strategy and offer promising translational implications for advancing personalized medi-cine in gastric cancer treatment.