OBJECTIVE To form an expert consensus on the clinical application of parenteral direct thrombin inhibitors （DTIs） in patients during the perioperative period. METHODS Led by Sichuan Academy of Medical Sciences & Sichuan Provincial People’s Hospital （the Affiliated Hospital of UESTC）， a multidisciplinary working group was established. Through literature review and the Delphi method， clinical questions related to the rational perioperative use of parenteral DTIs were identified. A structured design was adopted using the “Population-Intervention-Comparison-Outcome” framework； systematic searches were conducted in CNKI， Medline， Embase and other databases. Relevant evidence from randomized controlled trials and cohort studies was included and synthesized. Evidence quality was assessed using the Grades of Recommendations Assessment，Development and Evaluation （GRADE） approach， and recommendations were formulated through multiple rounds of Delphi surveys and expert consensus meetings. RESULTS &CONCLUSIONS Seven recommendations （each with an expert consensus rate exceeding 90%） on the use of parenteral DTIs in perioperative patients were developed. These recommendations specify drug selection， dosing ranges， key monitoring points， and safety management strategies for parenteral DTIs in various scenarios， including the perioperative period of ventricular assist device implantation， the perioperative period of cardiac surgery， perioperative patients with lower-extremity atherosclerotic disease， the perioperative period of percutaneous coronary intervention in patients with acute coronary syndrome， the perioperative period of carotid artery stenting in patients with carotid stenosis， the perioperative period of patients with right heart thrombosis， and patients who develop related thrombosis and dysfunction after a central venous catheter insertion. In addition， warning and management pathways for perioperative bleeding and thrombotic events were proposed. This expert consensus， which is formulated based on the best available evidence， provides evidence-based guidance for standardized and individualized use of parenteral DTIs in perioperative period.

ObjectiveTo investigate the role and mechanism of Lck/Yes-related novel protein tyrosine kinase （Lyn） on lipopolysaccharide （LPS）-induced M1-type polarization of macrophage. MethodsThe LentiCRISPR-V2 plasmid was digested with the restriction endonuclease BSMBI-V2， and the digested DNA fragments were recovered. The digested plasmid was ligated with Lyn-sgRNA using T4 ligase to generate the Lenti-Lyn-gRNA lentivirus. THP-1 cells were infected with the Lenti-Lyn-gRNA lentivirus to obtain a stable cell line with Lyn knockout， and a monoclonal THP-1 cell line with complete Lyn knockout （Lyn^⁻/⁻） was established subsequently. Wild-type Lyn （Lyn^WT） and Lyn^⁻/⁻ THP-1 cells were induced with 100 ng/mL phorbol myristate acetate （PMA） for 48 h to differentiate into M0 macrophages， which were further polarized into M1 macrophages by stimulation with 100 ng/mL LPS for 24 h. Quantitative real-time polymerase chain reaction （qPCR） was performed to detect the expression of M0 macrophage markers， including integrin αM （CD11b）， macrophage antigen （CD68）， and monocyte differentiation antigen （CD14）. The expression of Lyn in M1 macrophages differentiated from wild-type THP-1 cells （Lyn^WT-M1） was measured by qPCR， and the ratio of phosphorylated Lyn to total Lyn （P-Lyn/Lyn） in Lyn^WT-M1 cells was determined by Western blot. In M1 macrophages differentiated from Lyn-knockout THP-1 cells （Lyn^⁻/⁻-M1）， qPCR was used to detect the mRNA expression of inducible nitric oxide synthase （iNOS）， interleukin-6 （IL-6）， and chemokine （C-X-C motif） ligand 10 （CXCL-10）. Western blot was conducted to assess the protein expression of iNOS， as well as the protein levels of molecules related to the Janus kinase 1 （JAK1）-signal transducer and activator of transcription 1 （STAT1）signaling pathway， including JAK1， phosphorylated JAK1 （P-JAK1）， STAT1， and phosphorylated STAT1 （P-STAT1）. Additionally， the expression of the M1 macrophage marker cluster of differentiation 80 （CD80） was analyzed by flow cytometry. ResultsThe Lyn^-/- monoclonal cell line was successfully constructed. The expression of CD11b was significantly elevated in Lyn^-/- M0 macrophages， and the differentiation of M1 macrophages was successful. Knockdown of Lyn inhibited mRNA expression of iNOS， IL⁃6， CXCL⁃10， protein expression of iNOS and CD80 expression in M1 macrophages （P<0.05）. Western blot assay showed that Lyn knockdown inhibited protein expression of JAK1 and P-STAT1 （P<0.01）. ConclusionAfter CRISPR/Cas9-mediated Lyn knockout， the expression levels of JAK1 and P-STAT1， the key molecules in the JAK/STAT signaling pathway of M1 macrophages， are significantly downregulated； concomitantly， the expression of M1 macrophage-specific secretory factors （iNOS， IL⁃6， CXCL⁃10 mRNA） and CD80 is also downregulated， which may be achieved via targeted regulation of the JAK1/P-STAT1-mediated JAK/STAT signaling pathway.

ObjectiveTo explore the effects of different concentrations of oleic acid on human osteosarcoma cell lines 143B and HOS， as well as the impacts of the optimal concentration of oleic acid on cellular lipid droplet synthesis and cell functions. MethodsThe 143B and HOS cells were treated with varying concentrations of oleic acid （0， 25， 50， 100， and 200 µmol/L） for 48 hours. Following treatment， oil red O staining and BODIPY staining were performed to determine the optimal concentration. Subsequently， CCK-8 assays and colony formation experiments were conducted to assess the effect of this optimal concentration of oleic acid on the cell proliferation of both cell lines. Transwell migration assays were utilized to evaluate the influence of the optimal concentration on migratory capacity and Transwell invasion assays were utilized to evaluate the invasive ability. Additionally， Western blot analysis was employed to examine the expression levels of epithelial-mesenchymal transition （EMT） markers Epithelial cadherin （E-cadherin） and Neural cadherin （N-cadherin） in response to treatment with the optimal concentration of oleic acid. ResultsTreatment with oleic acid did not induce significant cell death in either 143B or HOS cells； however， an increase in intracellular lipid droplets was observed alongside enhanced proliferation， migration， invasion capabilities as well as EMT transformation potential （P<0.05）. ConclusionOleic acid induces lipid droplet synthesis in osteosarcoma cells which subsequently promotes their proliferation， migration and invasion abilities along with EMT transformation.

[摘 要] 目的：探讨X连锁锌指蛋白（ZFX）通过Nectin-4表达影响食管鳞状细胞癌（ESCC）进展的分子机制及其对PI3K/AKT信号通路的激活作用。方法：收集2022年8月至2023年7月在南充市中心医院手术切除的30对ESCC组织及癌旁组织标本。采用人食管上皮细胞HET-1A及ESCC细胞KYSE-30、KYSE-150、KYSE-410、KYSE-510和TE-1。基于2018年至2019年收集的6例ESCC配对标本转录组测序数据筛选Nectin-4，通过TIMER2.0数据库、RT-qPCR和WB法、免疫组织化学（IHC）检测ESCC组织及细胞中Nectin-4的表达水平。采用shRNA技术在KYSE-410和KYSE-510细胞中敲低Nectin-4，通过CCK-8、克隆形成、划痕愈合及Transwell实验检测敲低Nectin-4对细胞增殖、迁移和侵袭能力的影响，采用WB法检测敲低Nectin-4对细胞PI3K/AKT通路及EMT相关蛋白表达水平的影响。通过生物信息学预测并结合双萤光素酶报告基因实验，鉴定并验证ZFX作为Nectin-4的上游转录调控因子。结果：综合分析显示，ESCC组织及细胞系中Nectin-4表达显著高于癌旁组织及HET-1A细胞（均P < 0.01）。功能研究表明，敲低Nectin-4显著抑制KYSE-410和KYSE-510细胞的增殖活性、克隆形成能力，以及迁移和侵袭能力（均P < 0.01）。机制方面，敲低Nectin-4时细胞中E-cadherin表达上调、N-cadherin下调（均P < 0.01），并抑制PI3K/AKT通路相关蛋白的磷酸化水平（P < 0.01）。结论： ZFX通过上调Nectin-4表达激活PI3K/AKT信号通路促进ESCC进展，为ESCC的治疗提供了潜在的新靶点。

Symptom and syndrome efficacy evaluation scales are indispensable tools for clinical efficacy assessment in traditional Chinese medicine （TCM）， and hold significant value at all stages of new drug research development for spleen and stomach diseases. These scales can provide scientific basis for clinical positioning， efficacy evaluation， and expansion of indications of new drugs. By analyzing the current hotspots and difficulties in research， this study aims to explore the important significance of these scales in the development of new drugs for spleen and stomach diseases， summarize the domestic research progress， and conduct comparative analyses with international studies. Future development trends are also discussed in order to promote the application of symptom and syndrome efficacy evaluation scales in the development of new drugs for spleen and stomach diseases and to advance the moder-nization process of TCM.

BACKGROUND: The clinical impact of post-percutaneous coronary intervention (PCI) quantitative flow ratio (QFR) in patients treated with PCI for chronic total occlusion (CTO) was still undetermined. METHODS: All CTO vessels treated with successful anatomical PCI in patients from PANDA III trial were retrospectively measured for post-PCI QFR. The primary outcome was 2-year vessel-oriented composite endpoints (VOCEs, composite of target vessel-related cardiac death, target vessel-related myocardial infarction, and ischemia-driven target vessel revascularization). Receiver operator characteristic curve analysis was conducted to identify optimal cutoff value of post-PCI QFR for predicting the 2-year VOCEs, and all vessels were stratified by this optimal cutoff value. Cox proportional hazards models were employed to calculate the hazard ratio (HR) with 95% CI. RESULTS: Among 428 CTO vessels treated with PCI, 353 vessels (82.5%) were analyzable for post-PCI QFR. 31 VOCEs (8.7%) occurred at 2 years. Mean value of post-PCI QFR was 0.92 ± 0.13. Receiver operator characteristic curve analysis shown the optimal cutoff value of post-PCI QFR for predicting 2-year VOCEs was 0.91. The incidence of 2-year VOCEs in the vessel with post-PCI QFR < 0.91 (n = 91) was significantly higher compared with the vessels with post-PCI QFR ≥ 0.91 (n = 262) (22.0% vs. 4.2%, HR = 4.98, 95% CI: 2.32-10.70). CONCLUSIONS Higher post-PCI QFR values were associated with improved prognosis in the PCI practice for coronary CTO. Achieving functionally optimal PCI results (post-PCI QFR value ≥ 0.91) tends to get better prognosis for patients with CTO lesions.

Objective: To explore the efficacy of vonoprazan fumarate in the treatment of laryngopharyngeal reflux disease（LPRD） evaluated by the Chinese version of the RSS-12 scale. Methods:A total of 100 LPRD patients treated in the otolaryngology-head and neck surgery outpatient clinic of our hospital were randomly divided into two groups（50 cases each）. The observation group was treated with vonoprazan fumarate（20 mg, once daily）, and the control group was treated with esomeprazole enteric-coated capsules（20 mg, twice daily） for 12 weeks. The selected observation indicators in this study included RSI, the Chinese version of RSS-12, and RFS scores before and after treatment. Results:Prior to treatment, there was no statistically significant difference in the Chinese version of RSS-12, RSI, and RFS between the two groups（P>0.05）. After 8 and 12 weeks of antacid treatment, both the Chinese version of RSS-12 and RSI significantly decreased in both group （P<0.05）. The changes in symptoms were particularly noticeable between 0-8 weeks of treatment according to the Chinese version of RSS-12 and RSI. After 12 weeks of treatment, the RFS scores significantly decreased compared to pretreatment levels （P<0.05）. After 12 weeks of antacid treatment, according to the Chinese version of RSS-12, 80% of patients in the observation group showed a good therapeutic response, compared to 64% in the control group; according to RSI, 90% of patients in the observation group showed a good therapeutic response, compared to 84% in the control group. There was no statistically significant difference in the treatment effect between the two groups after 12 weeks of treatment（P>0.05）. Conclusion:Vonoprazan fumarate can significantly improve the symptoms and signs of laryngopharyngeal reflux, and their treatment effect is not inferior to proton pump inhibitors. Compared with RSI, the Chinese version of RSS-12 can serve as a new screening tool for clinical diagnosis of LPRD in China.
Humans ; Sulfonamides/therapeutic use* ; Male ; Pyrroles/therapeutic use* ; Female ; Laryngopharyngeal Reflux/drug therapy* ; Treatment Outcome ; Middle Aged ; Esomeprazole/therapeutic use* ; Proton Pump Inhibitors/therapeutic use* ; Adult

Metastasis is the leading cause of death from cutaneous melanoma. Identifying metastasis-related targets and developing corresponding therapeutic strategies are major areas of focus. While functional genomics strategies provide powerful tools for target discovery, investigations at the protein level can directly decode the bioactive epitopes on functional proteins. Aptamers present a promising avenue as they can explore membrane proteomes and have the potential to interfere with cell function. Herein, we developed a target and epitope discovery platform, termed functional aptamer evolution-enabled target identification (FAETI), by integrating affinity aptamer acquisition with phenotype screening and target protein identification. Utilizing the aptamer XH3C, which was screened for its migration-inhibitory function, we identified the Chondroitin Sulfate Proteoglycan 4 (CSPG4), as a potential target involved in melanoma migration. Further evidence demonstrated that XH3C induces cytoskeletal rearrangement by blocking the interaction between the bioactive epitope of CSPG4 and integrin α4. Taken together, our study demonstrates the robustness of aptamer-based molecular tools for target and epitope discovery. Additionally, XH3C is an affinity and functional molecule that selectively binds to a unique epitope on CSPG4, enabling the development of innovative therapeutic strategies.

OBJECTIVES: To study the mechanism mediating the protective effect of asiaticoside (AS) against myocardial ischemia-reperfusion injury (MIRI) in rats. METHODS: Fifty SD rats were randomized into sham-operated group, MIRI model group and AS treatment group. AS treatment was administered at low, moderate and high doses by daily gavage for 2 weeks before MIRI modeling (n=10). Serum levels of lactate dehydrogenase (LDH), creatine kinase isoenzyme (CK-MB), interleukin-18 (IL-18) and IL-1β, the volume of myocardial infarction and ischemia, and myocardial pathologies of the rats were determined or observed. The protein expression levels of NLRP3, ASC, caspase-1, GSDMD, GSDMD-N, IL-1β and IL-18 in the myocardial tissues were detected using Western blotting. The changes in the expression levels of these proteins were also detected in H9C2 cells with AS pretreatment prior to hypoxia-reoxygenation (H/R) injury. RESULTS: The rats models of MIRI exhibited significant myocardial infarction and ischemia with increased serum levels of LDH and CK-MB and myocardial expressions of NLRP3, ASC, caspase-1, GSDMD, GSDMD-N, IL-1β and IL-18. AS pretreatment effectively reduced myocardial infarction volume in the rat models and significantly reduced serum LDH and CK-MB levels and the protein levels in the myocardial tissue in a dose-dependent manner. In the H9C2 cell model of H/R injury, AS pretreatment significantly suppressed the elevation of the protein expressions of NLRP3, ASC, caspase-1, GSDMD, GSDMD-N, IL-1β and IL-18. Molecular docking studies showed that AS had a strong binding affinity with NLRP3. CONCLUSIONS Asiaticoside can alleviate MIRI in rats possibly by inhibiting NLRP3 inflammasome-mediated pyroptosis.
Animals ; Myocardial Reperfusion Injury/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Pyroptosis/drug effects* ; Rats, Sprague-Dawley ; Rats ; Inflammasomes/metabolism* ; Triterpenes/pharmacology* ; Interleukin-18/metabolism* ; Male ; Interleukin-1beta/metabolism* ; Caspase 1/metabolism*