BACKGROUND:The Guilu Erxian Glue consists of Testudinis Plastrum,Cornu Cervi,Lycii Fructus,and Ginseng Radix.In earlier clinical observations,it is discovered that using Guilu Erxian Glue to treat patients with liver-kidney deficiency type knee osteoarthritis effectively alleviated knee pain,increased the range of motion,and improved walking ability.However,the exact mechanism by which oral administration of Guilu Erxian Glue can produce local therapeutic effects on the knee joint is still unclear.OBJECTIVE:To investigate the effects of Guilu Erxian Glue on gut microbiota in rats with knee osteoarthritis and to evaluate its mechanism using 16S rDNA sequencing and machine learning analysis.METHODS:Totally 18 female SD rats were randomly divided into three groups:blank group,model group,and Guilu Erxian Glue group,with 6 rats in each group.A knee osteoarthritis model was prepared using the destabilization of the medial meniscus surgical method.After successful modeling,the Guilu Erxian Glue group was given a decoction of Guilu Erxian Glue by gavage,while the blank and model groups were given an equal amount of distilled water.After 28 days of continuous intervention,high performance liquid chromatography was used to detect the active ingredients of Guilu Erxian Glue.MRI imaging was used to observe the condition of rat knee articular cartilage.Fecal samples were collected;DNA was extracted using a kit,amplified and purified by PCR,and an Illumina sequencing library was constructed.The Illumina MiSeq platform was used for high-throughput sequencing to generate raw sequence data.After obtaining the raw data,QIIME2 software was used to process the data.Linear Discriminant Analysis Effect Size analysis and random forest algorithm were used to screen for differential species in microbial data.KEGG and MetaCyc functional pathway analyses were used to explore the association between key microbial communities and experimental groups.Linear discriminant analysis effect values and random forest algorithm were used to screen for differential species.Association networks were used to analyze the interactions between microbial communities,and machine learning methods were used to analyze the composition and changes of gut microbiota.RESULTS AND CONCLUSION:(1)LC-MS component identification was conducted on the traditional Chinese medicine formula of Guilu Erxian Glue,and a total of 7 effective ingredients were identified.(2)MRI imaging showed that synovitis scope of high-density shadows in rats of the Guilu Erxian Glue group was reduced,and the degeneration of medial femoral condyle cartilage was less than that in the model group.(3)16S rDNA sequencing showed that the model group rats exhibited significant microbial imbalance,with a significant decrease in the abundance of Firmicutes and Bacteroidetes at the phylum level,while the proportion of Proteobacteria increased significantly(P＜0.05).The gut microbiota structure of rats in the Guilu Erxian Glue group was significantly improved,and the proportion of Firmicutes and Bacteroidetes increased,restoring a more diverse microbiota composition,approaching that of the blank group(P＜0.05).(4)KEGG and MetaCyc functional pathway analysis showed that the Guilu Erxian Glue group significantly activated multiple metabolic pathways,including amino acid metabolism,lipid metabolism,and biotin synthesis pathways(P＜0.05).(5)The results indicate that Guilu Erxian Glue contains seven active ingredients,and the changes in gut microbiota of knee osteoarthritis rats were analyzed using 16S rDNA sequencing.Guilu Erxian Glue can significantly improve the imbalance of gut microbiota,restore the abundance of beneficial bacteria,and have a significant impact on the composition of gut microbiota,providing scientific basis for the efficacy and mechanism of Guilu Erxian Glue.

BACKGROUND:The Guilu Erxian Glue consists of Testudinis Plastrum,Cornu Cervi,Lycii Fructus,and Ginseng Radix.In earlier clinical observations,it is discovered that using Guilu Erxian Glue to treat patients with liver-kidney deficiency type knee osteoarthritis effectively alleviated knee pain,increased the range of motion,and improved walking ability.However,the exact mechanism by which oral administration of Guilu Erxian Glue can produce local therapeutic effects on the knee joint is still unclear.OBJECTIVE:To investigate the effects of Guilu Erxian Glue on gut microbiota in rats with knee osteoarthritis and to evaluate its mechanism using 16S rDNA sequencing and machine learning analysis.METHODS:Totally 18 female SD rats were randomly divided into three groups:blank group,model group,and Guilu Erxian Glue group,with 6 rats in each group.A knee osteoarthritis model was prepared using the destabilization of the medial meniscus surgical method.After successful modeling,the Guilu Erxian Glue group was given a decoction of Guilu Erxian Glue by gavage,while the blank and model groups were given an equal amount of distilled water.After 28 days of continuous intervention,high performance liquid chromatography was used to detect the active ingredients of Guilu Erxian Glue.MRI imaging was used to observe the condition of rat knee articular cartilage.Fecal samples were collected;DNA was extracted using a kit,amplified and purified by PCR,and an Illumina sequencing library was constructed.The Illumina MiSeq platform was used for high-throughput sequencing to generate raw sequence data.After obtaining the raw data,QIIME2 software was used to process the data.Linear Discriminant Analysis Effect Size analysis and random forest algorithm were used to screen for differential species in microbial data.KEGG and MetaCyc functional pathway analyses were used to explore the association between key microbial communities and experimental groups.Linear discriminant analysis effect values and random forest algorithm were used to screen for differential species.Association networks were used to analyze the interactions between microbial communities,and machine learning methods were used to analyze the composition and changes of gut microbiota.RESULTS AND CONCLUSION:(1)LC-MS component identification was conducted on the traditional Chinese medicine formula of Guilu Erxian Glue,and a total of 7 effective ingredients were identified.(2)MRI imaging showed that synovitis scope of high-density shadows in rats of the Guilu Erxian Glue group was reduced,and the degeneration of medial femoral condyle cartilage was less than that in the model group.(3)16S rDNA sequencing showed that the model group rats exhibited significant microbial imbalance,with a significant decrease in the abundance of Firmicutes and Bacteroidetes at the phylum level,while the proportion of Proteobacteria increased significantly(P＜0.05).The gut microbiota structure of rats in the Guilu Erxian Glue group was significantly improved,and the proportion of Firmicutes and Bacteroidetes increased,restoring a more diverse microbiota composition,approaching that of the blank group(P＜0.05).(4)KEGG and MetaCyc functional pathway analysis showed that the Guilu Erxian Glue group significantly activated multiple metabolic pathways,including amino acid metabolism,lipid metabolism,and biotin synthesis pathways(P＜0.05).(5)The results indicate that Guilu Erxian Glue contains seven active ingredients,and the changes in gut microbiota of knee osteoarthritis rats were analyzed using 16S rDNA sequencing.Guilu Erxian Glue can significantly improve the imbalance of gut microbiota,restore the abundance of beneficial bacteria,and have a significant impact on the composition of gut microbiota,providing scientific basis for the efficacy and mechanism of Guilu Erxian Glue.

Due to the immaturity of visualization and quantitative analysis methods,the utilization rate of level 3 characteristics is seriously insufficient.In this work,based on the wet-membrane method and scanning electrochemical microscopy(SECM),and the introduction of conductive black ink to enhance the visualization effect,a systematic level 3 feature quantitative method was developed.Firstly,the feasibility and effect of the multi-level characteristics extraction strategy of latent fingerprints was investigated.Then,the influences of various deposition conditions on the level 3 features were explored.The results showed that the higher the deposition pressure,the wider the ridges,and the smaller the pore size.Moreover,excessive oil content could cause the pore size to be smaller and even been covered.Subsequently,the quantitative method was established from various pore characteristics such as pore number,pore activity,pore size,pore-to-pore distance and pore-to-pore angle.The stability of the level 3 features(pore number,pore-to-pore distance and pore-to-pore angle)was confirmed via repeated experiments on the same fingerprint region.After stability test,the recognition ability of three indicators was investigated for different fingerprints,verifying the uniqueness of pore-to-pore distance and pore-to-pore angle.Finally,a multiple recognition strategy was proposed that combined frequency distribution fitting curves for pore-to-pore distance and angle with other level 3 details,and was successfully applied to incomplete fingerprint recognition.This ink-enhanced optical and electrochemical extraction method and quantitative analysis provided a new path for fingerprint recognition.

Objective To evaluate the forensic application value of used dental floss as a source of bio-logical evidence for individual identification by analyzing the effects of dental floss sample collection methods,DNA extraction methods,preservation conditions,and sampling sites on the success rate of STR typing.Methods Dental floss samples were collected using three techniques:direct cutting,cotton swab wiping,and flocked swab wiping,respectively.DNA was extracted respectively by the Chelex,spin column-based and magnetic bead-based methods.DNA quantification and STR typing were per-formed using the Qubit kit and FGI HumDNA Typing kit(Platinum),respectively.Storage environ-ments(temperature and humidity,ultraviolet radiation)and sampling locations(the floss part,the handle part)on DNA quantity and STR typing were evaluated.Results Through conducting a statisti-cal analysis of three key indicators of average DNA mass concentration,STR locus detection rate,and typing accuracy rate,the direct cutting method demonstrated the highest efficacy,followed by cotton swab wiping mothed,and the flocked swab wiping method had the lowest efficacy.Direct cutting yielded an average DNA mass concentration greater than(4.94±1.87)ng/μL,with STR locus detection and accuracy rates of 100%.Bead-based DNA extraction method produced superior DNA concentration and quality compared to spin column-based and Chelex methods,regardless of whether the sampling technique used.Preservation conditions had a significant impact on the DNA analysis of samples.Par-ticularly,the STR typing accuracy of samples preserved at 55℃/50%RH for 35 days dropped to(81.82±12.31)%,and that of samples exposed to ultraviolet radiation for 12 h dropped to(55.46±34.31)%.DNA concentration from the handle part of dental floss was extremely low,with an STR typing accuracy of only(30.91±27.35)%.Conclusion Using cotton swabs to wipe or directly cutting the thread of dental floss samples,and combining this approach with the magnetic bead method for DNA extraction,can best guarantee the concentration and quality of DNA.In addition,samples should be stored in low-temperature,low-humidity environment,protected from light and ultraviolet radiation.

In recent years,significant breakthroughs have been achieved in the immunotherapy for Alzheimer's disease.In line with global advancements,two anti-amyloid-β monoclonal antibodies have been approved and successfully launched in China for clinical use.Lecanemab and Donanemab were officially used in June 2024 and April 2025 in China,respectively.In order to standardize the rational and safe application of anti-amyloid-β monoclonal antibodies for Alzheimer's disease in China,this article integrates recom-mendations from the clinical trials and real-world experience from the author's team and domestic peers to further update the recom-mendations for the clinical use of anti-amyloid-β monoclonal antibody based on the 2024 version.It includes indications for therapy,pre-treatment evaluation and preparation,administration protocols and safety measures during treatment,and post-treatment monitor-ing strategies.

OBJECTIVE: Gastric cancer (GC) is one of the most common malignancies seen in clinic and requires novel treatment options. Morin is a natural flavonoid extracted from the flower stalk of a highly valuable medicinal plant Prunella vulgaris L., which exhibits an anti-cancer effect in multiple types of tumors. However, the therapeutic effect and underlying mechanism of morin in treating GC remains elusive. The study aims to explore the therapeutic effect and underlying molecular mechanisms of morin in GC. METHODS: For in vitro experiments, the proliferation inhibition of morin was measured by cell counting kit-8 assay and colony formation assay in human GC cell line MKN45, human gastric adenocarcinoma cell line AGS, and human gastric epithelial cell line GES-1; for apoptosis analysis, microscopic photography, Western blotting, ubiquitination analysis, quantitative polymerase chain reaction analysis, flow cytometry, and RNA interference technology were employed. For in vivo studies, immunohistochemistry, biomedical analysis, and Western blotting were used to assess the efficacy and safety of morin in a xenograft mouse model of GC. RESULTS: Morin significantly inhibited the proliferation of GC cells MKN45 and AGS in a dose- and time-dependent manner, but did not inhibit human gastric epithelial cells GES-1. Only the caspase inhibitor Z-VAD-FMK was able to significantly reverse the inhibition of proliferation by morin in both GC cells, suggesting that apoptosis was the main type of cell death during the treatment. Morin induced intrinsic apoptosis in a dose-dependent manner in GC cells, which mainly relied on B cell leukemia/lymphoma 2 (BCL-2) associated agonist of cell death (BAD) but not phorbol-12-myristate-13-acetate-induced protein 1. The upregulation of BAD by morin was due to blocking the ubiquitination degradation of BAD, rather than the transcription regulation and the phosphorylation of BAD. Furthermore, the combination of morin and BCL-2 inhibitor navitoclax (also known as ABT-737) produced a synergistic inhibitory effect in GC cells through amplifying apoptotic signals. In addition, morin treatment significantly suppressed the growth of GC in vivo by upregulating BAD and the subsequent activation of its downstream apoptosis pathway. CONCLUSION Morin suppressed GC by inducing apoptosis, which was mainly due to blocking the ubiquitination-based degradation of the pro-apoptotic protein BAD. The combination of morin and the BCL-2 inhibitor ABT-737 synergistically amplified apoptotic signals in GC cells, which may overcome the drug resistance of the BCL-2 inhibitor. These findings indicated that morin was a potent and promising agent for GC treatment. Please cite this article as: Wang Y, Sun XY, Ma FQ, Ren MM, Zhao RH, Qin MM, Zhu XH, Xu Y, Cao ND, Chen YY, Dong TG, Pan YF, Zhao AG. Morin inhibits ubiquitination degradation of BCL-2 associated agonist of cell death and synergizes with BCL-2 inhibitor in gastric cancer cells. J Integr Med. 2025; 23(3): 320-332.
Humans ; Flavonoids/therapeutic use* ; Stomach Neoplasms/pathology* ; Animals ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Cell Line, Tumor ; Apoptosis/drug effects* ; Cell Proliferation/drug effects* ; Ubiquitination/drug effects* ; Mice ; Drug Synergism ; Mice, Inbred BALB C ; Mice, Nude ; Xenograft Model Antitumor Assays ; Flavones

OBJECTIVE: To assess the efficacy of Qingda Granule (QDG) in ameliorating hypertension-induced cardiac damage and investigate the underlying mechanisms involved. METHODS: Twenty spontaneously hypertensive rats (SHRs) were used to develope a hypertension-induced cardiac damage model. Another 10 Wistar Kyoto (WKY) rats were used as normotension group. Rats were administrated intragastrically QDG [0.9 g/(kg•d)] or an equivalent volume of pure water for 8 weeks. Blood pressure, histopathological changes, cardiac function, levels of oxidative stress and inflammatory response markers were measured. Furthermore, to gain insights into the potential mechanisms underlying the protective effects of QDG against hypertension-induced cardiac injury, a network pharmacology study was conducted. Predicted results were validated by Western blot, radioimmunoassay immunohistochemistry and quantitative polymerase chain reaction, respectively. RESULTS: The administration of QDG resulted in a significant decrease in blood pressure levels in SHRs (P<0.01). Histological examinations, including hematoxylin-eosin staining and Masson trichrome staining revealed that QDG effectively attenuated hypertension-induced cardiac damage. Furthermore, echocardiography demonstrated that QDG improved hypertension-associated cardiac dysfunction. Enzyme-linked immunosorbent assay and colorimetric method indicated that QDG significantly reduced oxidative stress and inflammatory response levels in both myocardial tissue and serum (P<0.01). CONCLUSIONS Both network pharmacology and experimental investigations confirmed that QDG exerted its beneficial effects in decreasing hypertension-induced cardiac damage by regulating the angiotensin converting enzyme (ACE)/angiotensin II (Ang II)/Ang II receptor type 1 axis and ACE/Ang II/Ang II receptor type 2 axis.
Animals ; Drugs, Chinese Herbal/therapeutic use* ; Hypertension/pathology* ; Renin-Angiotensin System/drug effects* ; Rats, Inbred SHR ; Oxidative Stress/drug effects* ; Male ; Rats, Inbred WKY ; Blood Pressure/drug effects* ; Myocardium/pathology* ; Rats ; Inflammation/pathology*

BACKGROUND:Bone morphogenetic protein 2 is vital in embryonic development,bone formation,and regeneration,but its high-dose application is linked to cancer.Bone morphogenetic protein 2 osteogenic peptide L20 reduces adverse effects like cancer and boosts bone tissue regeneration.OBJECTIVE:To graft bone morphogenetic protein 2 active peptide segments onto mesopores and surfaces through a peptide mimicry strategy inspired by oysters,and explore its impact on osteogenic properties of tissue-engineered bone.METHODS:(1)Mesoporous bioactive glass was synthesized using a template method.Bone morphogenetic protein 2 osteogenic peptide L20 was loaded onto mesoporous bioactive glass using a one-step synthesis method to characterize the morphology and in vitro sustained release properties of mesoporous active glass nanoparticles loaded with bone morphogenetic protein 2 osteogenic active peptide L20.(2)Bone marrow mesenchymal stem cells were isolated and extracted from SD rats.After two generations,they were co-cultured with PBS(blank group),mesoporous bioactive glass nanoparticles(control group),and mesoporous bioactive glass nanoparticles loaded with bone morphogenetic protein 2 osteogenic active peptide L20(experimental group).Cell live/dead fluorescence staining and CCK-8 assay were used to detect cytotoxicity and cell proliferation.Scanning electron microscopy was used to observe cell adhesion.After osteogenic induction and differentiation,alkaline phosphatase staining,Alizarin red S staining,and osteogenesis-related gene expression were detected.(3)Fifteen SD rats were selected to establish bilateral femoral condyle defect models and divided into three groups using a random number table method:the blank group(n=5)was not implanted with any material;the control group(n=5)was implanted with mesoporous bioactive glass nanoparticles,and the experimental group(n=5)was implanted with mesoporous bioactive glass nanoparticles loaded with bone morphogenetic protein 2 osteogenic active peptide L20.Eight weeks after surgery,femoral Micro-CT scanning and tissue morphology observation were performed.RESULTS AND CONCLUSION:(1)Scanning electron microscopy showed that the mesoporous bioactive glass nanoparticles loaded with bone morphogenetic protein 2 osteogenic active peptide L20 were spherical and monodisperse particles.Transmission electron microscopy showed their porous structure with an average particle size of(268.10±0.58)nm,which could release L20 in vitro.(2)Mesoporous bioglass nanoparticles loaded with bone morphogenetic protein 2 osteogenic active peptide L20 were non-cytotoxic and could promote the proliferation and adhesion of bone marrow mesenchymal stem cells.Compared with the blank group and the control group,the alkaline phosphatase activity and extracellular matrix mineralization capacity of the experimental group were increased(P<0.05),and the mRNA expression levels of alkaline phosphatase,Runx2,and osteocalcin were increased(P<0.05).(3)The results of femoral Micro-CT scanning showed that compared with the blank group and the control group,the new bone mass and bone density of the experimental group were increased(P<0.05).The results of hematoxylin-eosin and Masson staining showed that compared with the blank group and the control group,the new bone formation and collagen fibers of the experimental group were increased.(4)These findings indicate that mesoporous bioactive glass loaded with bone morphogenetic protein 2 active peptide L20 exhibits excellent biocompatibility and in vitro and in vivo osteogenic properties,promoting regeneration and repair of SD rat femoral condyle defects.

Objective To characterize the features of patent foramen ovale(PFO)-related cryptogenic stroke using diffusion-weighted imaging(DWI)and to investigate the correlation between infarct size and cardiac CT characteristics of PFO.Methods A retrospective,consecutive cohort study was conducted on acute ischemic stroke patients admitted to Neurology Department of Xuanwu Hospital,Capital Medical University from January 2022 to September 2024.Patients were categorized into PFO group,arterio-arterial embolism(AAE)group,and atrial fibrillation(AF)group based on etiological diagnosis.Baseline clinical data,including age,height,body mass index,admission National Institutes of Health stroke scale(NIHSS)score,history of old cerebral infarction,hypertension,diabetes mellitus,coronary heart disease,dyslipidemia,and smoking history were collected and compared.All patients underwent head MR within 24 h of admission.DWI was used to analyze and compare infarct characteristics across the three groups,including lesion number(single or multiple),location(cortical+subcortical,deep white matter,cortical+subcortical+deep white matter,cerebellum+thalamus+brainstem),size(≥15 mm or＜15 mm,based on maximum transverse diameter;for multiple lesions,if any lesion had a maximum diameter≥15 mm,it was categorized as≥15 mm),infarcted vascular territory(anterior,posterior,or both circulations),and specific arterial supply(anterior cerebral artery,middle cerebral artery,posterior cerebral artery,basilar artery,posterior inferior cerebellar artery,superior cerebellar artery,anterior choroidal artery,or multiple arteries).Patients in the PFO group additionally underwent cardiac CT to measure PFO-related parameters:tunnel length,width,height,septum secundum thickness,and fossa ovalis length.Spearman correlation analysis was performed to evaluate the relationship between infarct size and PFO cardiac CT features.Results A total of 232 acute ischemic stroke patients were included(mean age[57±17]years,ranged 19-86 years;141 males,91 females),comprising 116 in the PFO group,36 in the AAE group,and 80 in the AF group.(1)The proportion of males in the PFO group was higher than that in the AF group,it was lower than that in the AAE group.The age,body mass index and proportions of patients with hypertension,diabetes,hyperlipidemia,coronary heart disease were all lower than those in the other two groups(both P＜0.016 7),while other baseline characteristics showed no significant differences(all P＞0.05).(2)The PFO group exhibited a higher proportion of multiple infarcts compared to the AAE group(83.62％[97/116]vs.61.11％[22/36],P＜0.016 7),but a lower proportion than the AF group(83.62％[97/116]vs.98.75％[79/80],P＜0.016 7).The PFO group also showed a significantly higher proportion of cortical+subcortical infarcts(47.41％[55/116]vs.11.11％[4/36]and 6.25％[5/80],respectively,both P＜0.016 7)and infarcts with a maximum diameter＜15 mm compared to both AAE and AF groups(66.38％[77/116]vs.36.11％[13/36]and 11.25％[9/80],respectively,both P＜0.016 7).Furthermore,the PFO group had a lower proportion of anterior circulation infarcts(27.59％[32/116]vs.69.44％[25/36]in AAE group and 67.50％[54/80]in AF group,both P＜0.016 7),but a higher proportion of posterior circulation infarcts(62.07％[72/116]vs.16.67％[6/36]in AAE group and 8.75％[7/80]in AF group,both P＜0.016 7).Specifically,middle cerebral artery infarcts were less common in the PFO group(18.97％[22/116]vs.66.67％[24/36]in AAE group and 52.50％[42/80]in AF group,both P＜0.016 7),while posterior cerebral artery infarcts were more common(48.28％[56/116]vs.8.33％[3/36]in AAE group and 8.75％[7/80]in AF group,both P＜0.016 7).(3)Spearman correlation analysis revealed that infarct size was negatively correlated with PFO tunnel length(rs=-0.429,P=0.029),fossa ovalis length(rs=-0.408,P=0.038),and septum secundum thickness(rs=-0.525,P=0.006),but not correlated with PFO width or height(both P＞0.05).Conclusions PFO-related cryptogenic stroke is predominantly characterized by multiple small infarcts,primarily located in the cortical+subcortical regions and posterior circulation.Infarct size was found to be negatively correlated with PFO tunnel length,fossa ovalis length,and septum secundum thickness.Comprehensive assessment integrating DWI and cardiac CT features may facilitate the identification of PFO-related stroke.These findings warrant further validation through larger,prospective studies.