Objective:To evaluate the potential causal relationship between tinnitus and the risk of Alzheimer's disease(AD)onset using the two-sample Mendelian randomization(MR)method and to clarify its mechanism of action,so as to provide new ideas for early warning of AD.Methods:Genome-wide association study(GWAS)database was used to search the keywords"tinnitus"and"Alzheimer"to obtain the related datasets of exposure factor tinnitus and outcome AD;the tinnitus datasets included ukb-d-4803_11,ukb-d-4803_12,ukb-d-4803_13,ukb-b-14254 and ukb-a-384;the AD datasets included ieu-b-5067,ieu-b-2,ieu-a-297,ebi-a-GCST90027158 and ebi-a-GCST002245.The single nucleotide polymorphisms(SNPs)closely and independently associated with tinnitus were screened as instrumental variables(IVs),and the SNPs associated with AD were used as outcomes.Inverse variance weighted(IVW)method was used to conduct MR analysis to evaluate its odds ratio(OR)value,95%confidence interval(CI)and P value;P<0.05 indicated significant causal relationship.Sensitivity detection used Cochran's Q test to detect the heterogeneity of IVs to evaluate its Q value,df value and P value;when IVW method P>0.05,it indicated no significant heterogeneity;MR-Egger intercept was used to detect horizontal pleiotropy;when the intercept was 0 or close to 0 and P>0.05,it indicated no significant horizontal pleiotropy;meanwhile,leave-one-out method was used for sensitivity analysis.Finally,visualization results were performed using forest plot,scatter plot,funnel plot and leave-one-out plot.Results:A total of 286 SNPs were screened as IVs.All instrumental variables satisfied F>10,suggesting no weak instrumental variable;after screening by PhenoScanner web tool,all SNPs were unrelated to confounding factors.When the tinnitus and AD datasets were ukb-d-4803 and ebi-a-GCST90027158 respectively,there was a significant positive correlation between tinnitus and the risk of AD onset(IVW:OR=1.842,95%CI:1.065-3.188,P=0.029);Cochran's Q test suggested no significant heterogeneity of IVs(Q=9.788,df=10.000,P=0.459);MR-Egger intercept indicated no horizontal pleiotropy(Egger intercept=-0.006,P=0.147);leave-one-out method showed stable results,and no SNP with significant influence on the results was detected.Conclusion:There is a positive causal relationship between tinnitus and the risk of AD onset.Neuroinflammation accompanied by persistent microglial activation to varying degrees may be the common pathogenesis of tinnitus and AD;in addition,depression may also act as an upstream factor to hyperactivate the hypothalamic-pituitary-adrenal(HPA)axis,leading to the progression of relationship between tinnitus and AD.

Objective To explore the relationship between pathogenic microorganism types,age and season in 2 188 children with respiratory tract infections.Methods A total of 2 188 children with respiratory tract in-fections admitted to the Department of Pediatrics,962 Hospital,Joint Logistic Support Force of PLA from June 2023 to May 2024 were selected as the study subjects.Targeted next generation sequencing(tNGS)tech-nology was used to detect 107 common pathogenic microorganism in children with respiratory tract infections,including Haemophilus influenzae,rhinovirus,Moraxella catarrhalis,Mycoplasma pneumoniae,Staphylococcus aureus,Streptococcus pneumoniae,human parainfluenza virus,human respiratory syncytial virus,etc.The re-spiratory tract infection situation and epidemiological characteristics of children in Harbin were analyzed.Re-sults Among 2 188 pediatric patients,98.5%(2 156/2 188)tested positive for pathogenic microorganism,with Haemophilus influenzae accounting for the highest proportion of 33.5%(732/2 188),followed by rhino-virus of 25.0%(547/2 188)and Moraxella catarrhalis of 24.8%(543/2 188).The positive rates of Hae-mophilus influenzae and human adenovirus in male children were higher than those in female children(P<0.05),while there were no statistically significant differences in the positive positive rates of other pathogenic microorganism between male and female children(P>0.05).Except for human adenovirus and influenza A virus,which showed no statistically significant differences in positive rates among different age groups(P>0.05),there were statistically significant differences in the positive rates of other pathogenic microorganism a-mong different age groups(P<0.05).The positive rates of pathogenic microorganism in preschool children were relatively high.There were no statistically significant differences in the positive rates of Streptococcus and Staphylococcus aureus in different seasons(P>0.05),while there were statistically significant differences in the positive rates of other pathogenic microorganism in different seasons(P<0.05).The positive rates of Haemophilus influenzae,Streptococcus pneumoniae,human metapneumovirus,human parainfluenza virus and SARS-Cov-2 were the highest in summer(P<0.05).Conclusion 2 188 children with respiratory tract infec-tions were mainly caused by pathogenic microorganism such as Haemophilus influenzae,rhinovirus,and Moraxella catarrhalis,etc.Preschool children is a susceptible group,and the prevalence of pathogenic microor-ganism varies seasonally.In clinical practice,relevant prevention and control measures should be developed based on this characteristic to reduce the incidence of diseases.

Objective To establish a 13C isotope-labeled hyperinsulinemic-euglycemic clamp model for the assessment of insulin sensitivity in mice.Methods The mouse model of insulin resistance was established by high-fat diet feeding.The phosphorylation level of downstream insulin signaling protein,Protein Kinase,also known as Akt was assessed.Glucose metabolism was evaluated using glucose tolerance test,insulin tolerance test,and pyruvate tol-erance tests and the area under the curve of the respective testes over 2 hours was calculated to quantify overall blood glucose level.Mice underwent jugular vein catheterization surgery,and a 13C isotope-labeled hyperinsulinemic-eugly-cemic clamp experiment was conducted to monitor blood glucose levels and to calculate the glucose infusion rate(GIR).Tail vein serum was collected for liquid chromatography-tandem mass spectrometry(LC-MS/MS)analysis to determine glucose disposal rate(GDR)and hepatic gluconeogenesis rate(HGP).Following the inhibition of glycog-enolysis in mice,Akt phosphorylation level was measured to evaluate insulin signaling.The clamp test was repeated to calculate GIR,and tail vein blood serum was analyzed by LC-MS/MS to determine GDR and HGP.Results After one week of high-fat diet feeding,mice exhibited significantly elevated blood glucose level(P＜0.001)accompanied by reduced p-Akt level in liver and muscle.Glucose tolerance tests,insulin tolerance tests and pyruvate tolerance test demonstrated a significant increase in blood glucose level(P＜0.05)and a higher area under curves(AUC)(P＜0.001)in high-fat-fed mice.During the 13C-labeled hyperinsulinemic-euglycemic clamp experiment,after the blood glucose levels were stable,the GIR of high-fat-fed mice was significantly reduced(P＜0.001),GDR was de-creased(P＜0.0001)and hepatic gluconeogenesis rate was increased(P＜0.01).After pharmacological inhibition of glycogenolysis,mice showed elevated blood glucose level(P＜0.001)and further reductions in p-Akt level in liver and muscle.The 13C-labeled clamp experiment revealed that in the treated group,the GIR decreased(P＜0.01)while GDR was reduced(P＜0.000 1)and HGP increased(P＜0.01).Conclusions An improved hyperinsulinemic-eugly-cemic clamp model was developed to assess insulin sensitivity in mice.

Objective:To observe the effects of acupuncture and moxibustion on interleukin(IL)-9/IL-9 receptor(IL-9R)in the colon tissue of rats with ulcerative colitis(UC)and investigate the protective mechanism of acupuncture and moxibustion on the intestinal mucosal barrier in UC rats. Methods:Male Sprague-Dawley rats were randomly divided into a normal control(NC)group and a modeling group.UC models were prepared by giving 4%dextran sulfate sodium(DSS)water for 7 d.After the successful construction of the UC rat model,the modeling group was randomly divided into a UC group,a herb-insulated moxibustion(HM)group,and an electroacupuncture(EA)group.HM and EA interventions at bilateral Tianshu(ST25)were performed once a day for 7 d.Hematoxylin-eosin(HE)staining was used to observe the histopathological changes in the colon.The serum concentrations of IL-9,IL-6,IL-1β,and hemoglobin-H(HbH)were determined by enzyme-linked immunosorbent assay.The protein expression levels of IL-9,IL-9R,claudin-2,zonula occludens-1(ZO-1),and occludin in the colon tissue were measured by Western blotting or immuno-histochemistry.Immunofluorescence was used to detect the co-expression of PU.1 and CD4 with the IL-9 protein. Results:Compared with the NC group,the colon tissue of UC rats was severely damaged and ulcerated with congestion and edema,and the colonic histopathological score increased significantly(P<0.01).The serum HbH concentration decreased significantly(P<0.01),while the serum concentrations of IL-9,IL-6,and IL-1β increased(P<0.01).The protein expression of colonic ZO-1 and occludin decreased significantly(P<0.01),while the protein expression of colonic IL-9 and IL-9R increased(P<0.05).The positive co-expression levels of IL-9/PU.1 and IL-9/CD4 increased in the colon tissue(P<0.05).Compared with the UC group,the colonic mucosal structures were gradually repaired in both HM group and EA group,and healed ulcers could be observed,the colonic histopathological score decreased significantly(P<0.05).The serum concentration of HbH increased(P<0.01),while the serum concentrations of IL-9,IL-6,and IL-1β decreased(P<0.05).The protein expression levels of ZO-1 and occludin increased(P<0.05),while the protein expression levels of IL-9 and IL-9R decreased(P<0.01).The positive co-expression levels of IL-9/PU.1 and IL-9/CD4 decreased in the colon tissue(P<0.05). Conclusion:Both HM and EA can inhibit the protein expression levels of IL-9 and IL-9R in the UC colon by regulating the transcription factor PU.1,promote the repair of intestinal mucosal barrier,and down-regulate protein contents of proinflammatory factors IL-9,IL-6,and IL-1β in the serum,which may be one of the key mechanisms of acupuncture and moxibustion in reducing the inflammation of UC colonic mucosa and protecting the intestinal mucosal barrier.

Cerebral organoids are three-dimensional nerve cultures induced by embryonic stem cells(ESCs)or induced pluripotent stem cells(iPSCs)that mimic the structure and function of human brain.With the continuous optimization of cerebral organoid culture technology and the combination with emerging technologies such as organ transplantation,gene editing and organoids-on-chip,complex brain tissue structures such as functional vascular structures and neural circuits have been produced,which provides new methods and ideas for studying human brain development and diseases.This article reviews the latest advances in brain organoid technology,describes its application in neurological diseases and advances in stroke modeling and transplantation treatment.

Post-stroke cognitive impairment(PSCI)is mainly manifested as learning and memory disorders.Highly enriched RNA m6A methylation modification in mammalian brain is involved in glial cell-mediated neuroinflammation.Given that neuroinflammation is the main mechanism for neural damage and spatial and memory impairment of PSCI,it is speculated that RNA m6A methylation modification can regulate the inflammatory response of glial cells after stroke to improve PSCI.This review summarizes and analyzes the role of RNA m6A methylation modification in the development of PSCI and analyzes its detailed mechanism of regulating glial cell-mediated inflammation,which will provide reference for researchers in this field.

BACKGROUND:Long non-coding RNAs(lncRNAs),as important regulators of the inflammatory response,are involved in the immune-inflammation-brain crosstalk mechanism after ischemic stroke and have the potential to become a therapeutic agent for neurological dysfunction after ischemic stroke. OBJECTIVE:To analyze and summarize the molecular mechanism of lncRNA acting on glial cells involved in the neuroimmuno-inflammatory cascade response after ischemic stroke and the associated signaling pathways,pointing out that lncRNAs have the potential to regulate inflammation after ischemic stroke. METHODS:PubMed was searched using the search terms of"ischemic stroke,long non-coding RNA,neuroinflammation,immune function,signal pathway,microglia,astrocytes,oligodendrocyte,mechanism,"and 63 relevant documents were finally included for review. RESULTS AND CONCLUSION:In the early stage of ischemic stroke,the death of nerve cells due to ischemia and hypoxia activates the innate immune response of the brain,promoting the secretion of inflammatory factors and inducing blood-brain barrier damage and a series of inflammatory cascades responses.As an important pathogenesis factor in ischemic stroke,the neuroimmuno-inflammatory cascade has been proved to seriously affect the prognosis of patients with ischemic stroke,and it needs to be suppressed promptly in the early stage.Neuroinflammation after ischemic stroke usually induces abnormal expression of a large number of lncRNAs that mediate a series of neuro-immune-inflammatory crosstalk mechanisms through regulating the polarization of microglia,astrocytes and oligodendrocytes to exert post-stroke neuroprotective effects.LncRNAs,as important regulatory factors of the inflammatory response,inhibit the neuroimmuno-inflammatory cascade response after ischemic stroke through regulating nuclear factor-κB,lncRNA-miRNA-mRNA axis,Rho-ROCK,MAPK,AKT,ERK and other signaling pathways to effectively improve neurological impairment after ischemic stroke.Most of experimental studies on the interaction between lncRNAs and ischemic stroke are based on a middle cerebral artery occlusion model or a cerebral ischemia-reperfusion injury model,but no clinical trials have been conducted.Therefore,it remains to be further explored about whether lncRNAs can be safely applied in clinical practice.At present,there are many therapeutic drugs for the treatment of ischemic stroke,but there are relatively few studies on the application of lncRNAs,exosomes and other transplantation technologies for the treatment of ischemic stroke using tissue engineering technology,which need to be further explored.lncRNA has become an important target for the treatment of ischemic stroke with its relative stability and high specificity.In future studies,more types of inflammatory lncRNAs that function under ischemic-hypoxia conditions should continue to be explored,in order to provide new research directions for the treatment of neuroinflammation after ischemic stroke.

Post-stroke cognitive impairment(PSCI)is a common complication after stroke,which significantly affects quality of life.However,the pathogenesis has not been fully explained.Increasing evidence has shown that the mechanism of programmed cell death(PCD)is related to PSCI,including apoptosis,necroptosis,pyroptosis,PANoptosis,parthanatos,and ferroptosis.Therefore,it is crucial to clearly understand the various mechanisms of PCD and their relationship with PSCI,and to elucidate the role of PCD in PSCI pathogenesis.The article reviews six PCD pathways related to PSCI,summarizes their mechanisms of action in PSCI,and elucidates the possible crosstalk among pathways to provide a basis for clinical targeting of regulatory factors in the PCD pathway for PSCI treatment.

Objective:To compare the antiplatelet effects of vicagrel and clopidogrel in patients with different cytochrome P450 (CYP) 2C19 genotypes.Methods:This is a post-hoc analysis of a phase Ⅱ clinical trial of vicagrel, which included patients with coronary heart disease who underwent percutaneous coronary intervention from August 2018 to June 2019 in 18 centers. Patients were categorized based on the presence of CYP 2C19 *2 or *3 loss-of-function (LOF) alleles into LOF carrier group ( n=111) and non-LOF carrier group ( n=90). Each group included patients received vicagrel 5 mg, 6 mg, 7.5 mg, or clopidogrel 75 mg for 28 days per study protocol. P2Y 12 reaction units (PRU) were measured using VerifyNow at baseline, 6 to 8 hours after loading dose, 7 to 10 days after randomization, and 28 days after randomization and the percentage inhibition of platelet aggregation (%IPA) was calculated. The primary endpoint was %IPA on day 28. Within the patients from the General Hospital of Northern Theater Command, 8 to 12 patients in each study arms were enrolled in a prespecified pharmacokinetic sub-study, measuring the time to reach maximum plasma concentration (T max), peak plasma concentration (C max), and area under the plasma concentration-time curve (AUC). Results:Among 201 patients, the age was (58.8±8.5) years, and 139 (69.2%) were male. In non-LOF carriers, there was no significant differences in PRU values and %IPA between the vicagrel 5 mg, 6 mg, 7 mg, and clopidogrel groups at all time points (all P>0.05). In LOF carriers, %IPA was significantly higher in the vicagrel-treated groups than in the clopidogrel group at 6-8 hours after loading dose (22.9 (14.2, 31.5)% vs. 19.8 (11.0, 28.6)% vs. 29.5 (20.9, 38.0)% vs. 12.9 (3.9, 21.9)%, P=0.038) and 7-10 days after randomization (22.4 (14.2, 30.5)% vs. 34.4 (26.1, 42.6)% vs. 39.8 (31.8, 47.9)% vs. 24.7 (16.3, 33.2)%, P=0.001), with a trend towards higher %IPA in the vicagrel-treated groups at day 28 (30.4 (21.3, 39.6)% vs. 36.5 (27.2, 45.7)% vs. 40.8 (31.8, 49.8)% vs. 30.7(21.2, 40.2)%, P=0.056). Pharmacokinetic results of 35 patients showed that the C max and AUC of the active metabolite M15-2 of vicagrel was similar to that of clopidogrel in non-LOF carriers, but AUC between vicagrel 5 mg, 6 mg, 7 mg and clopidogrel were significantly different in LOF carriers ((5.6±0.6) h·μg -1·L -1 vs. (6.8±2.7) h·μg -1·L -1 vs. (9.2±3.3) h·μg -1·L -1 vs. (4.2±1.9) h·μg -1·ml -1, P=0.020). Conclusion:Vicagrel and clopidogrel have similar antiplatelet effects in non-LOF carriers, but vicagrel exhibits superior antiplatelet effects in LOF carriers.

The aim of this study was to prepare a high drug-carrying capacity micellar drug delivery system(CTD@Sol)of the polymer zebra tetracycline and to preliminarily investigate the feasibility of this drug delivery system for the treatment of breast cancer.Firstly,CTD@Sol was prepared using sol as the carrier material and CTD as the model drug,and its pharmacological properties such as appearance and morphology,particle size,potential and in vitro release were evaluated.The growth inhibitory and apoptotic effects of CTD@Sol on breast cancer(4T1)cells were investigated by MTT assay and Annexin V-FITC/PI double staining assay;the uptake efficiency of 4T1 to this delivery system was investigated by flow cytometry;and the in vivo tissue distribution of the delivery system and the targeting of tumour tissues were investigated by small animal in vivo imaging technique.The results showed that CTD@Sol appeared as a light pale blue creamy white colour,with an average particle size of(159.73±1.96)nm,a PDI of 0.198±0.006,Zeta potential of-(47.60±1.77)mV,an encapsulation rate of(90.29±1.69)%and a drug loading capacity of(45.00±0.84)%;the in vitro release and haemolysis experiments showed that the drug release rate of CTD@Sol in acidic environment(pH 5.5)was significantly faster than that in neutral environment(pH 7.4),suggesting that the system is acid-sensitive and has good biosafety under endocytosed pH conditions.Cellular uptake,cytotoxicity and apoptosis experiments showed that CTD@Sol was more lethal to 4T1 cells,and the sol-gel polymer micelles as a drug delivery vehicle could significantly improve the cellular uptake efficiency of the drug;in vivo experiments showed that the delivery system had a significant targeting effect on tumour tissues.In conclusion,this study has successfully produced a CTD@Sol drug delivery system with high drug loading capacity(>45%),good pharmacological performance,strong targeting and biosafety,which has the potential to be used in the treatment of breast cancer.