Aim To investigate the effect of tetrameth-ylpyrazine(TMP)on neuroinflammation after cerebral ischemia and hypoxia via mannose-binding lectin(MBL).Methods Patients diagnosed with ischaemic stroke at Shanxi Provincial People's Hospital were in-cluded in the study,and their clinicopathological data,as well as blood and urine samples,were collected with the consent of the patients and their families.Using these biological samples,differential proteins and tar-gets were identified by proteomic analysis and subse-quently verified with animal experiments.The mice were divided into the sham,dMCAO,and TMP(10,20,40 mg·kg-1)treatment groups.After seven days of drug administration,the modified neurological sever-ity score(mNSS)was used to assess the neurological function.TTC staining was used to detect the volume of cerebral infarction.Motor function was evaluated be-haviourally,and ELISA was used to detect MASP1,sC5b-9,TNF-α,IL-6,and IL-1β.Western blot was used to determine the expression of relevant proteins,such as MBL2,MASP2,and C3.Results Compared with the sham group,the dMCAO group exhibited in-creased neurological impairment,which was signifi-cantly ameliorated by TMP treatment.The expression levels of MBL2,C3 and MASP2 were elevated in the dMCAO group and were reduced following TMP treat-ment.Additionally,the dMCAO group showed elevat-ed expression of inflammatory factors IL-1 β,IL-6 and TNF-α,which were then suppressed by TMP treat-ment.Conclusion TMP inhibits the inflammatory re-sponse after ischemia and hypoxia by regulating MBL,thus attenuating brain injury.

To establish a method for detecting Toxoplasma gondii based on recombinant polymerase amplification(RPA)technology and apply it to clinical sample validation of pet cats.Using the 529 repeat sequence of the Toxoplasma gondii gene as the target gene sequence,primers and probes were designed,and the Rep-529 recombinant plasmid was constructed as the standard.A fluorescent RPA reaction system was established.Dilute the plasmid standard 10 times to different concentrations as the detection template for sensitivity testing;Specific testing was conducted using genomic DNA from several parasitic spe-cies,including Toxoplasma gondii,Cryptosporidium,Neosporidium,Trichinella spiralis,Giardia flagellata,Babesia bo-vis and Theileria annulata as templates;Simultaneously,fluorescence RPA and RT-PCR were used to detect 52 positive and 40 negative cats clinical samples,and the coincidence rate of the detection results of the two methods were compared and ana-lyzed.The RPA reaction system was successfully established using PTRep recombinant plasmid as the standard,ToxD-F/ToxD-R as the primer,and RepD-P as the fluorescent probe.The reaction temperature was constant at 39 ℃,the reaction time was 30 minutes,and the detection sensitivity was 1 copy/μL.There is no significant cross reaction with parasites such as Cryptosporidium,Neosporidium,Trichinella spiralis,Giardia,Babesia bovis and Theileria annulata,and the specificity is good.A total of 92 clinical fecal samples from cats were tested,and the positive coincidence rate of fluorescence RPA detection method was higher than that of conventional RT-PCR method(98.08％vs.82.69％),and the difference of the positive rate was not statistically significant(X2=1.392,P＞0.05).The fluorescence RPA detection method for Toxoplasma gondii suc-cessfully established in this study has the characteristics of being fast,sensitive,specific,accurate,and reliable.It can be used as a rapid clinical detection kit for Toxoplasma gondii in cats and other animals,providing new technical support for the subsequent epidemiological monitoring and precise clinical diagnosis of toxoplasmosis in cats,other animals,and humans in the future.

Alzheimer's disease(AD)is a multifactorial condi-tion characterized by the accumulation of toxic proteins and asso-ciated neurodegeneration.AD is distinguished by the pathologi-cal aggregation of amyloid beta(Aβ)and Tau proteins.The in-teraction between Aβ and Tau can further induce neuroinflamma-tion,mitochondrial autophagy dysfunction,and endoplasmic retic-ulum stress,exacerbating synaptic damage and neuronal death.Neuronal cells are particularly susceptible to protein misfolding due to an imbalance between protein production and degrada-tion.The ubiquitin/26S proteasome system(UPS),a major pathway for protein degradation in eukaryotic cells,plays a cruci-al role in recognizing misfolded or damaged proteins within the nervous system.In UPS,the levels of ubiquitin are tightly regu-lated by both ubiquitin ligases(E3s)and deubiquitylases(DUBs).This article reviews the involvement and mechanisms of E3s and DUBs in the pathogenesis of AD,aiming to provide novel research strategies for its treatment.

Over 40%of critically ill patients will develop coagulopathy.Once critically ill patients are complicated with coagulopathy,the incidence of bleeding and mortality can increase by more than 4 times.Early identification of coagulopathy and accurate evaluation of coagulation function are essential for correcting coagulopathy as soon as possible.Therefore,Chinese Society of Thrombosis,Hemostasis and Critical Care,Chinese Medicine Education Association,together with Chinese People's Liberation Army Professional Committee of Critical Care Medicine updated the"Chinese expert consensus on standardized assessment of severe coagulopathy(2025 Edition)"on the basis of the"Consensus of Chinese experts on standardized evaluation of coagulation dysfunction in severe patients"formulated in 2022.This consensus includes four parts:classification and typing,etiology and mechanism,assessment methods,and diagnostic criteria of severe coagulopathy,with a total of 14 recommendations,aiming to provide corresponding guidance for clinical practice.

Objective To investigate the influence of helical tomographic radiotherapy plans with different combinations of lead gate width,pitch and algorithms on threading effects.Methods A target model was established with a Cheese Phantom used as the simulated human body,then three lead gate widths(1.0,2.5,and 5.0 cm),six screw pitches(0.143,0.172,0.215,0.287,0.430,and 0.500)and two computational grids(Fine algorithm and Normal algorithm)were respectively combined for designing the helical tomography radiotherapy plans.The radiotherapy plans with a pitch of 0.143,0.172,0.215,0.287 or 0.430 were enrolled into an experimental group,and the plans with a pitch of 0.500 were divided into a control group.The dosimetric parameters including maximum dose(Dmax),minimum dose(Dmin)and mean dose(Dmean)of the target area PTV1 and PTV2 were evaluated by the dose volume histogram(DVH).The dose homogeneity index(HI)of the target area was calculated,and the single rotation time and total treatment time of each plan were recorded and counted.SPSS 27.0 software was used for statistical analysis.Results No significant threading effect appeared regardless of the pitch value when the lead gate width was 1.0 cm.The threading effects in the experimental group were weaker than those in the control group when the lead gate width was 2.5 or 5.0 cm.The threading effect gradually rose with the pitch increased when the lead gate width was 5.0 cm.The most significant difference was found between the threading effect in case of the screw pitch being 0.500 and that with the screw pitch being 0.143,with the differenes being statistically obvious(P<0.05).The lead gate width had significant effects on the Dmax,Dmin,Dmean and HI of PTV1 and PTV2.When the lead gate width was 5.0 cm,high HI value and uneven dose distribution were detected and lowered screw pitch weakened the threading effect.The single rotation time first remained constant and then increased with the screw pitch was enlarged,with the changing points occurring in case of the screw pitches of 0.287 and 0.430.With a certain lead gate width,the treatment time for plans was shortened with the decrease of the pitches in case of the pritches lower than 0.287,and tended to be constant after the screw pitches reached 0.287.The changes of the computational grid had no significant effects on the results of radiotherapy plans when the lead gate width and screw pitch were kept constant.Conclusion When designing a spiral tomotherapy plan with conventional doses,a lead gate width of 1.0 or 2.5 cm and a screw pitch of 0.287 or 0.430 should be selected in order to minimize the threading effect while ensuring the efficiency of plan implementation.[Chinese Medical Equipment Journal,2025,46(8):58-66]

Over 40%of critically ill patients will develop coagulopathy.Once critically ill patients are complicated with coagulopathy,the incidence of bleeding and mortality can increase by more than 4 times.Early identification of coagulopathy and accurate evaluation of coagulation function are essential for correcting coagulopathy as soon as possible.Therefore,Chinese Society of Thrombosis,Hemostasis and Critical Care,Chinese Medicine Education Association,together with Chinese People's Liberation Army Professional Committee of Critical Care Medicine updated the"Chinese expert consensus on standardized assessment of severe coagulopathy(2025 Edition)"on the basis of the"Consensus of Chinese experts on standardized evaluation of coagulation dysfunction in severe patients"formulated in 2022.This consensus includes four parts:classification and typing,etiology and mechanism,assessment methods,and diagnostic criteria of severe coagulopathy,with a total of 14 recommendations,aiming to provide corresponding guidance for clinical practice.

Objective:To investigate the mechanism of the synergistic anti-leukemia effect of the combination of Bcl-2 inhibitor venetoclax(VEN)and histone deacetylase(HDAC)inhibitor chidamide(CDM)in T-cell acute lymphoblastic leukemia(T-ALL).Methods:The effect of VEN combined with CDM on the proliferation of T-ALL CEM and MOLT-4 cell lines was detected by CCK-8 assay.And the effects on the cell cycle and apoptosis were detected by flow cytometry.Cell cycle protein and apoptosis-related protein expression were detected by Western blot.The key pathways of VEN combined with CDM in T-ALL were screened through network pharmacology analysis,and verifying them in T-ALL cell lines,T-ALL patient cells and public databases.Results:VEN combined with CDM displayed a synergistic effect on cell proliferation of CEM and MOLT-4 cells.In cell cycle experiment,VEN combined with CDM induced G0/G1 phase arrest in CEM and MOLT-4 cells.Western blot experiment showed that VEN combined with CDM could significantly downregulate the expression of cyclin E2 and CDK2 and upregulate the expression of p21Waf1/Cip1.In the apoptosis experiment,VEN combined with CDM could significantly induce the apoptosis of CEM and MOLT-4 cells.Western blot experiment demonstrated that VEN combined with CDM promoted endogenous apoptosis by downregulating Mcl-1 and upregulating Bax and cleaved caspase-3 protein levels.Network pharmacology analysis identified 10 hub genes.KEGG enrichment analysis revealed the cell cycle,PI3K-AKT signaling pathway,and its downstream FoxO signaling pathway were significantly enriched.GO enrichment analysis revealed the G1/S transition of mitotic cell cycle,cyclin-dependent protein kinase holoenzyme complex,and kinase activity were significantly enriched.Western blot experiment showed that VEN combined with CDM could significantly downregulate the protein level of PI3K,AKT,and p-AKT,and upregulate FoxO1 in CEM and MOLT-4 cells.In T-ALL patients,FoxO1 showed significantly lower expression compared to the normal donors,and the same result was verified in the GSE13159 and GSE26713 datasets.Conclusion:The combination of VEN and CDM exerts synergistic anti-leukemia effects by inhibiting cellular proliferation,inducing G0/G1,phase arrest and promoting apoptosis through PI3K/AKT/FoxO1 axis in T-ALL.

ObjectiveTo investigate the effects of modified Ditan tang on genes related to the transcription-translation feedback loop （TTFL） of biorhythm in the rat model of non-alcoholic fatty liver disease （NAFLD） and its mechanism for prevention and treatment of NAFLD. MethodsSixty-five healthy SPF male SD rats were randomly assigned into blank （n=20）， model （n=15）， and low-， medium-， and high-dose （2.68， 5.36， and 10.72 g·kg^-1·d^-1， respectively） modified Ditan tang （n=10） groups. Other groups except the blank group were fed a high-fat diet for 12 weeks. The modified Ditan tang groups were treated with the decoction at corresponding doses by gavage， and the blank and model groups were treated with an equal volume of normal saline from the 9th week for 4 weeks. The levels of triglyceride （TG）， total cholesterol （TC）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT） in the serum were measured by an automatic biochemical analyzer. TG and non-esterified fatty acid （NEFA） assay kits were used to measure the levels of TG and NEFA in the liver. The pathological changes in the hypothalamus and liver were observed by hematoxylin-eosin staining， and the lipid deposition in the liver was observed by oil red O staining. The levels of brain-muscle ARNT-like protein 1 （BMAL1/ARNTL） in the hypothalamus and liver were determined by immunohistochemical staining. The mRNA and protein levels of BMAL1， circadian locomotor output cycles kaput （CLOCK）， period circadian clock 2 （PER2）， and cryptochrome1 （Cry1） in the hypothalamus and liver were determined by Real-time PCR and Western blot， respectively. ResultsCompared with the blank group， the model group showed elevated levels of TG， TC， LDL-C， AST， and ALT （P<0.01） and a lowered level of HDL-C （P<0.05） in the serum， elevated levels of TG and NEFA in the liver （P<0.01）， pyknosis and deep staining of hypothalamic neuron cells， and a large number of vacuoles in the brain area. In addition， the model group showed lipid deposition in the liver， up-regulated mRNA and protein levels of CLOCK and BMAL1 （P<0.01）， and down-regulated mRNA and protein levels of Cry1 and PER2 （P<0.01） in the hypothalamus and liver. Compared with the model group， all the three modified Ditan tang groups showed lowered levels of TG， TC， LDL-C， ALT， and AST （P<0.05， P<0.01） and an elevated level of HDL-C （P<0.05） in the serum， and lowered levels of TG and NEFA （P<0.05， P<0.01） in the liver. Furthermore， the three groups showed alleviated pyknosis and deep staining of hypothalamic neuron cells， reduced lipid deposition in the liver， down-regulated mRNA and protein levels of CLOCK and BMAL1 （P<0.05， P<0.01）， and up-regulated mRNA and protein levels of Cry1 and PER2 （P<0.05， P<0.01） in the hypothalamus and liver. ConclusionModified Ditan tang can reduce lipid deposition in the liver and regulate the expression of CLOCK， BMAL1， Cry1， and PER2 in the TTFL of NAFLD rats.

This study investigated the mechanism of autophagy in the differentiation processes of MC3T3-E1 cells under osteogenic induction(physiological) and alcohol(AL) intervention(pathological), as well as the mechanism by which icariin(ICA) affected osteogenic differentiation of MC3T3-E1 cells under the pathological condition of AL intervention. Osteogenic mineralized nodule staining confirmed that the cells could differentiate into osteoblasts. After determining the appropriate concentrations of AL and ICA using the CCK-8 assay, seven groups were set up in this study: complete medium(CM) group, osteogenic induction medium(OIM) group, OIM+0.25 mol·L~(-1) AL group, OIM+0.25 mol·L~(-1) AL+1×10~(-8) mol·L~(-1) ICA group, OIM+0.25 mol·L~(-1) AL+1×10~(-7) mol·L~(-1) ICA group, OIM+0.25 mol·L~(-1) AL+1×10~(-6) mol·L~(-1) ICA group, and OIM+0.25 mol·L~(-1) AL+1×10~(-5) mol·L~(-1) ICA group, with a culture period of 7 days. Alkaline phosphatase(ALP) staining was used to detect the relative ALP area. Western blot and RT-qPCR were employed to analyze the expression of osteogenesis-and autophagy-related proteins and mRNAs. Reactive oxygen species(ROS) staining was used to detect ROS levels, and apoptosis was assessed through mitochondrial membrane potential assays. The results showed that ICA increased the relative ALP area that had been reduced by AL intervention. AL down-regulated the expression levels of Wnt family member 1(Wnt1), along with the osteogenesis-related mRNAs Wnt1, β-catenin, Runt-related transcription factor 2(Runx2), osteoprotegerin(OPG), and ALP, thereby inhibiting osteogenic differentiation. ICA up-regulated the expression levels of the osteogenesis-related proteins and mRNAs that had been inhibited by AL, promoting osteogenic differentiation. AL inhibited typical autophagy, while ICA regulated Rubicon to suppress LC3-associated phagocytosis(LAP) and promote typical autophagy. ICA also reduced the ROS levels that were elevated by AL and decreased the apoptosis of osteoblasts induced by AL intervention. In conclusion, ICA can regulate Rubicon to inhibit LAP, promote typical autophagy, eliminate ROS, reduce apoptosis, and ultimately enhance the osteogenic differentiation of MC3T3-E1 cells under the pathological condition of AL intervention by modulating the Wnt/β-catenin signaling pathway.
Autophagy/drug effects* ; Animals ; Osteogenesis/drug effects* ; Mice ; Cell Differentiation/drug effects* ; Osteoblasts/metabolism* ; Ethanol/pharmacology* ; Flavonoids/pharmacology* ; Cell Line ; Reactive Oxygen Species/metabolism* ; Drugs, Chinese Herbal/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