Hypercholesterolemia is a significant risk factor for the development of atherosclerosis. 2',3',5'-Tri-O-acetyl-N ⁶-(3-hydroxyphenyl) adenosine (IMM-H007), a novel AMPK agonist, has shown protective effects in metabolic diseases. However, its impact on cholesterol and triglyceride metabolism in hypercholesterolemia remains unclear. In this study, we aimed to elucidate the effects and specific mechanisms by which IMM-H007 regulates cholesterol and triglyceride metabolism. To achieve this goal, we used Apoe ^-/- and Ldlr ^-/- mice to establish a hypercholesterolemia/atherosclerosis model. Additionally, hepatocyte-specific Ampka1/2 knockout mice were subjected to a 5-week high-cholesterol diet to establish hypercholesterolemia, while atherosclerosis was induced via AAV-PCSK9 injection combined with a 16-week high-cholesterol diet. Our results demonstrated that IMM-H007 improved cholesterol and triglyceride metabolism in mice with hypercholesterolemia. Mechanistically, IMM-H007 modulated the AMPKα1/2-LDLR signaling pathway, increasing cholesterol uptake in the liver. Furthermore, IMM-H007 activated the AMPKα1-FXR pathway, promoting the conversion of hepatic cholesterol to bile acids. Additionally, IMM-H007 prevented hepatic steatosis by activating the AMPKα1/2-ATGL pathway. In conclusion, our study suggests that IMM-H007 is a promising therapeutic agent for improving hypercholesterolemia and atherosclerosis through the activation of AMPKα.

OBJECTIVE: CRISPR-Cas protects bacteria from exogenous DNA invasion and is associated with bacterial biofilm formation and pathogenicity. METHODS: We analyzed the type I-F CRISPR-Cas system of Legionella pneumophila WX48, including Cas1, Cas2-Cas3, Csy1, Csy2, Csy3, and Cas6f, along with downstream CRISPR arrays. We explored the effects of the CRISPR-Cas system on the in vitro growth, biofilm-forming ability, and pathogenicity of L. pneumophila through constructing gene deletion mutants. RESULTS: The type I-F CRISPR-Cas system did not affect the in vitro growth of wild-type or mutant strains. The biofilm formation and intracellular proliferation of the mutant strains were weaker than those of the wild type owing to the regulation of type IV pili and Dot/Icm type IV secretion systems. In particular, Cas6f deletion strongly inhibited these processes. CONCLUSION The type I-F CRISPR-Cas system may reduce biofilm formation and intracellular proliferation in L. pneumophila.
Legionella pneumophila/pathogenicity* ; CRISPR-Cas Systems ; Biofilms/growth & development* ; Phenotype ; Bacterial Proteins/metabolism* ; Gene Deletion

A method for simultaneous determination of 21 kinds of Aconitum alkaloids(ATS)in biological specimens and infused liquor using ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)was developed.The biological samples were pretreated with methanol-acetonitrile(1∶2,V/V)for protein precipitation,while infused liquors were diluted 100-fold with acetonitrile,followed by centrifugation,and filtration by a 0.22-μm membrane.Chromatographic separation was carried out on an EC-C18 column using gradient elution with the mixture of 10 mmol/L ammonium acetate and 0.2%formic acid as mobile phase A and acetonitrile as mobile phase B.With this method,all the analytes were separated within 9.5 min.The samples were detected in positive ESI mode with dynamic multiple reaction monitoring(MRM)and quantified via external standard calibration.The results showed that the concentrations of the analytes in the range of 2-1000 ng/mL had excellent linearity(R2>0.9992)with the peak area.The developed method was successfully used for detection of 21 kinds of aconitum alkaloids,with limits of detection of 0.5-2 ng/mL,quantification limits of 2-6 ng/mL,intra/inter-day precision≤6.0%,spiked recoveries of 89.4%-100.9%,extraction recoveries of 74.2%-104.4%,and matrix effects ranging from-11.1%to 9.2%in blood/urine.The method was applied to detection of 12 samples from 4 fatal aconite poisoning cases,and all 21 kinds of ATS with total alkaloid concentrations of 0.04-4.18 μg/mL in blood and 154.96-422.83 μg/mL in medicinal liquors were detected.Tissue distribution revealed that the order of concentrations from highest to lowest is as follows:urine(157.22 μg/mL)>gastric contents(51.37 μg/mL)>kidney(21.6 μg/g)>whole blood(4.18 μg/mL)>liver(0.03 μg/g).This method showed many advantages such as simple pretreatment,low detection limits,accurate quantification,broad analyte coverage,and superior anti-interference capability in complex matrices,proving ideal for forensic and toxicological analysis of aconitum alkaloids.

Objective To explore the correlation between gallbladder stones and small intestinal bacterial overgrowth(SIBO).Methods A retrospective analysis was conducted on the clinical data of 393 patients who attended the Department of Gastroenterology of the Sixth Medical Center of Chinese PLA General Hospital from January 2021 to September 2023.They were divided into gallbladder stones group(n=190)and control group(n=203)based on the presence of gallbladder stones.Their general clinical data,laboratory test results,and abdominal symptoms were compared.Multivariate logistic regression was used to analyze the risk factors for gallbladder stones.Additionally,the total population was divided into SIBO-positive group(n=239)and SIBO-negative group(n=154),and their clinical characteristics were analyzed by logistic regression to explore the risk factors for SIBO.Results Univariate analysis revealed that gallbladder stones group had a higher rate of age,body mass index(BMI),fasting plasma glucose(FPG),glutaminase levels,prevalence of hypertension,diabetes,coronary heart disease,non-alcoholic fatty liver disease,gallbladder polyps,and SIBO,as well as a higher prevalence of CH4-positive and H2-positive in SIBO group than control group(P＜0.05).In terms of abdominal symptoms,the incidence of bad breath(48.4%vs.35.5%),dyspepsia(38.4%vs.28.6%),abdominal pain(30.5%vs.14.8%),bloating(42.1%vs.28.6%),diarrhea(20.5%vs.7.4%),and more exhaustion(46.8%vs.34.5%)were significantly higher in gallbladder stones group than those in control group(P＜0.05).Multivariate logistic regression analysis showed that independent positive determinants for incident gallbladder stones were age,BMI,FPG,total bilirubin(TBIL),coronary heart disease,gallbladder polyps,and SIBO.Univariate analysis revealed that age,prevalence of gallbladder stones,proportion of single stones,triglycerides(TG),total cholesterol(TC),and low-density lipoprotein cholesterol(LDL-C)were significantly higher in SIBO-positive group than those in SIBO-negative group(P＜0.05).Multivariate logistic regression analysis showed that the risk factors for SIBO were age,coronary heart disease,and gallbladder stones,while the protective factor for SIBO was high-density lipoprotein cholesterol(HDL-C).Conclusion There is a significant correlation between gallbladder stones and small SIBO;interventions on related factors of gallbladder stones and small SIBO may help reduce their incidence.

Objective:To analyze the effects of different doses of butorphanol on prolactin, serotonin and lipid hydrogen peroxide after cesarean section.Methods:A total of 124 women who underwent cesarean section in the Affiliated Hospital of Jining Medical University from January 2023 to January 2024 were prospectatively selected as study subjects, and divided into the study group 1 (41 cases), study group 2 (41 cases) and study group 3 (42 cases) according to random number table method. All three groups underwent patient-controlled analgesia after surgery, and 0.2 mg fentanyl was used as analgesic drug. However, the doses of butorphanol were 100 mg/L in the study group 1, 120 mg/L in the study group 2, and 140 mg/L in the study group 3. The level of PRL and first lactation time before and after surgery were compared among the three groups at different time points, serotonin and LHP levels were compared among the three groups at different time points after surgery, Ramsay and visual analog scale (VAS) scores were compared among the three groups at different time points after surgery, and the occurrence of adverse reactions and satisfaction with analgesic effect were compared among the three groups.Results:At 24, 48 and 72 h after operation, the level of PRL in the study group 3 was higher than that in the study group 2 and study group 1: (383.02 ± 47.57) μg/L vs. (376.39 ± 46.83), (302.88 ± 41.38) μg/L; (412.38 ± 40.22) μg/L vs. (394.82 ± 38.30), (315.09 ± 37.93) μg/L; (434.39 ± 39.39) μg/L vs. (427.48 ± 40.27), (344.39 ± 42.78) μg/L, there were statistical differences ( P<0.05). The first lactation time in the study group 2 and study group 3 was lower than that in the study group 1: (50.31 ± 6.52), (49.54 ± 6.27) h vs. (53.91 ± 8.42) h, there was statistical difference ( P<0.05). At 2 h (T 1), 12 h (T 2), 24 h (T 3) and 48 h (T 4) after surgery, the levels of serotonin in the study group 3, study group 2 and study group 1 were increased successively, and the levels of LHP were decreased successively, there were statistical differences ( P<0.05). At 30 min after surgery (T 0), T 1, T 2, T 3 and T 4, the Ramsay score in the study group 3, study group 2 and study group 1 were increased successively, and the VAS score were decreased successively, there were statistical differences ( P<0.05). There was no significant difference in the incidence of adverse reactions among the three groups ( P>0.05). The total satisfaction of study group 3 was higher than that of study group 2 and study group 1: 92.86%(39/42) vs. 73.17%(30/41), 68.29%(28/41), there was statistical difference ( Z = 2.52, P = 0.008). Conclusions:Butorphanol 140 mg/L has a more significant analgesic effect after cesarean section, and can improve the levels of serum PRL, serotonin and LHP, reduce the occurrence of adverse reactions, and improve the satisfaction of analgesia

This study was aimed at establishing a method of ultra-fast quantitative PCR for Brucella detection.We used an exogenous recombinant plasmid as the internal reference and targeted the T4SS secretion system,an important Brucella viru-lence factor,to design specific primers and probes.The sensitivity,specificity,and repeatability of this method were evaluated,and a standard curve was constructed.The coincidence rate of detection findings with this method versus quantitative PCR was determined.This method markedly decreased the detection time to only 10 minutes.The standard curve demonstrated a good linear relationship(Y=-3.410 7x+38.357,R2=0.998 5)with a low minimum detection limit of 10 copies/μL.The method exhibited good specificity and did not specifically amplify several common clinical bacteria other than Brucella.The de-tection of three concentrations of positive plasmids yielded coefficients of variation(CVs)of 0.20％to 0.91％,thus demonstra-ting the method's excellent repeatability.Furthermore,140 clinical samples were analyzed concurrently with the fluorescence PCR method,which yielded a 100％compliance rate and consistent results.Our findings indicated that the Brucella ultra-fast quantitative PCR was ultrafast;had high sensitivity,high specificity,and good specificity;and can be used for the clinical de-tection of Brucella and emergency investigation of epidemics.Therefore,this method is valuable for the early diagnosis of Bru-cella.

This study was aimed at establishing a method of ultra-fast quantitative PCR for Brucella detection.We used an exogenous recombinant plasmid as the internal reference and targeted the T4SS secretion system,an important Brucella viru-lence factor,to design specific primers and probes.The sensitivity,specificity,and repeatability of this method were evaluated,and a standard curve was constructed.The coincidence rate of detection findings with this method versus quantitative PCR was determined.This method markedly decreased the detection time to only 10 minutes.The standard curve demonstrated a good linear relationship(Y=-3.410 7x+38.357,R2=0.998 5)with a low minimum detection limit of 10 copies/μL.The method exhibited good specificity and did not specifically amplify several common clinical bacteria other than Brucella.The de-tection of three concentrations of positive plasmids yielded coefficients of variation(CVs)of 0.20％to 0.91％,thus demonstra-ting the method's excellent repeatability.Furthermore,140 clinical samples were analyzed concurrently with the fluorescence PCR method,which yielded a 100％compliance rate and consistent results.Our findings indicated that the Brucella ultra-fast quantitative PCR was ultrafast;had high sensitivity,high specificity,and good specificity;and can be used for the clinical de-tection of Brucella and emergency investigation of epidemics.Therefore,this method is valuable for the early diagnosis of Bru-cella.

Objective:To analyze the effects of different doses of butorphanol on prolactin, serotonin and lipid hydrogen peroxide after cesarean section.Methods:A total of 124 women who underwent cesarean section in the Affiliated Hospital of Jining Medical University from January 2023 to January 2024 were prospectatively selected as study subjects, and divided into the study group 1 (41 cases), study group 2 (41 cases) and study group 3 (42 cases) according to random number table method. All three groups underwent patient-controlled analgesia after surgery, and 0.2 mg fentanyl was used as analgesic drug. However, the doses of butorphanol were 100 mg/L in the study group 1, 120 mg/L in the study group 2, and 140 mg/L in the study group 3. The level of PRL and first lactation time before and after surgery were compared among the three groups at different time points, serotonin and LHP levels were compared among the three groups at different time points after surgery, Ramsay and visual analog scale (VAS) scores were compared among the three groups at different time points after surgery, and the occurrence of adverse reactions and satisfaction with analgesic effect were compared among the three groups.Results:At 24, 48 and 72 h after operation, the level of PRL in the study group 3 was higher than that in the study group 2 and study group 1: (383.02 ± 47.57) μg/L vs. (376.39 ± 46.83), (302.88 ± 41.38) μg/L; (412.38 ± 40.22) μg/L vs. (394.82 ± 38.30), (315.09 ± 37.93) μg/L; (434.39 ± 39.39) μg/L vs. (427.48 ± 40.27), (344.39 ± 42.78) μg/L, there were statistical differences ( P<0.05). The first lactation time in the study group 2 and study group 3 was lower than that in the study group 1: (50.31 ± 6.52), (49.54 ± 6.27) h vs. (53.91 ± 8.42) h, there was statistical difference ( P<0.05). At 2 h (T 1), 12 h (T 2), 24 h (T 3) and 48 h (T 4) after surgery, the levels of serotonin in the study group 3, study group 2 and study group 1 were increased successively, and the levels of LHP were decreased successively, there were statistical differences ( P<0.05). At 30 min after surgery (T 0), T 1, T 2, T 3 and T 4, the Ramsay score in the study group 3, study group 2 and study group 1 were increased successively, and the VAS score were decreased successively, there were statistical differences ( P<0.05). There was no significant difference in the incidence of adverse reactions among the three groups ( P>0.05). The total satisfaction of study group 3 was higher than that of study group 2 and study group 1: 92.86%(39/42) vs. 73.17%(30/41), 68.29%(28/41), there was statistical difference ( Z = 2.52, P = 0.008). Conclusions:Butorphanol 140 mg/L has a more significant analgesic effect after cesarean section, and can improve the levels of serum PRL, serotonin and LHP, reduce the occurrence of adverse reactions, and improve the satisfaction of analgesia

italic>Polygonum capitatum is a characteristic Miao medicine in Guizhou, commonly used in clinical practice to treat gastrointestinal and urinary tract infections. Research has found that it has good antibacterial and anti-inflammatory effects, and its main active ingredient is flavonoids. Lavonoid O-methyltransferase (FOMT) is a key enzyme for oxymethylation modification of flavonoid compounds. In order to understand the function and properties of FOMT protein in Polygonum capitatum, the transcriptome was sequenced by Illumina HiSeq 4000 high throughput sequencing technology. Then, the obtained transcripts were annotated and analyzed, and the whole genome of the FOMT gene family in Polygon capitalis was mined and identified. A total of 99 298 Unigenes were obtained, of which 71 514 were successfully annotated by the public database. In the genome of Polygonum capitatum, a total of 50 FOMT genes were identified. The phylogenetic tree showed that FOMT genes were divided into two subfamilies: caffeoyl CoA O-methyltransferase (CCoAOMT) and caffeic acid O-methyltransferase (COMT). Gene sequence analysis showed that the number of FOMT encoded amino acids ranged from 99 to 1 053 aa, the molecular weight ranged from 11 224.91 to 86 687.42 Da, and the isoelectric point ranged from 4.79 to 9.45. The 50 FOMT family members were hydrophobic proteins. Subcellular localization results showed that 54% of FOMT subfamily CCoAOMT and COMT members were located in cytoplasm and 28% were located in chloroplasts. The FOMT gene was tissue specific and highly expressed in the flowers of Polygonum capitatum, followed by the stems, and the least expressed in the roots. In this study, the FOMT gene family of Polygonum capitatum was identified and analyzed to provide theoretical basis for further study of FOMT function and biosynthesis of methylated flavonoids.

Ubiquitination, a diverse post-translational modification, is carried out by enzymes including E1-activating enzymes, E2-conjugating enzymes, E3 ligases, and deubiquitinating enzymes (DUBs). Ubiquitin itself possesses 7 lysine residues and N-terminal methionine, allowing for the formation of polyubiquitin chains with different lengths and linkages. These chains exhibit various topologies that can be recognized by proteins containing ubiquitin-binding domain, thereby transmitting distinct cellular signals. To unravel the physiological mechanisms associated with ubiquitin, numerous ubiquitin probes have been developed. This review provides an overview of recent advancements in the field of ubiquitin probes, focusing on activity-based and affinity-based probes. Activity-based probes are designed to covalently bind to DUBs, E1s, or E3s, enabling the identification and characterization of these enzymes. Affinity-based probes, on the other hand, selectively bind to ubiquitin-binding domains, facilitating the identification of proteins that interact with ubiquitin. Moreover, this review comprehensively discusses the synthetic methodologies employed for the acquisition of ubiquitin probes. These includes meticulous discussions on the synthesis of individual monomeric modules, the establishment of isopeptide linkages, as well as the incorporation of reactive functional groups. Additionally, the review explores the emerging area of cell-penetrating ubiquitin probes and highlights their latest applications in living cells. These probes incorporate cell-penetrating peptides to enable their internalization into cells, allowing for direct visualization and manipulation of ubiquitin-modified proteins within their native environment. Overall, this review offers insights into the design, synthesis, and applications of ubiquitin probes, highlighting their significance in elucidating ubiquitin-mediated cellular processes.