The Gly/N-degron pathway is a branch of the proteasomal degradation pathway that specifically targets proteins initiated with an N-terminal glycine. The E3 ligase substrate adaptors ZYG11B and ZER1 have been identified as being responsible for recognizing the target proteins of the Gly/N-degron pathway. Previously, it has been shown that the Gly/N-degron pathway activates the human NLRP1 inflammasome by degrading the autoinhibitory N-terminal fragment of NLRP1 after cleavage by the enteroviral 3C protease. However, the recognition of the NLRP1 Gly/N-degron is not yet fully understood. Here, we determined the X-ray crystal structure of ZER1 bound to the NLRP1 Gly/N-degron at a resolution of 2.2 Å. The structural information revealed that ZER1 uses its ARM repeats to form a conserved cavity that engages the N-terminal glycine (G1) through hydrogen bonds to Asp556, Asn597 and Glu600. Structural comparisons show a shared recognition mode for Gly/N-degrons despite subtle differences in side-chain interactions. However, ZER1 exhibits weaker affinity for the NLRP1 Gly/N-degron than ZYG11B, likely due to distinct local environments surrounding position 3. This study elucidates the molecular basis of NLRP1 recognition by ZER1 and provides insights into targeting this pathway in inflammatory diseases.

Background:Sepsis-associated liver injury (SALI) is a significant risk factor for mortality in patients with sepsis. Magnesium, as an essential electrolyte, has a correlation with adverse outcomes in critical illness when deficient, yet the therapeutic efficacy of magnesium sulfate in SALI remains undetermined. This study was designed to evaluate the association between magnesium sulfate therapy and prognosis in SALI patients. Method:This retrospective cohort study utilized data from the Medical Information Mart for Intensive Care IV (MIMIC-IV) database, with the primary endpoint being 30-day all-cause mortality.Propensity score matching (PSM) achieved covariate balance, Kaplan-Meier survival curves and Cox regression were employed to analyze the magnesium sulfate-mortality relationship in SALI patients. The study results were externally validated using the eICU 2.0 database. Result:The present study was conducted on 648 SALI patients. After PSM, the 30-day all-cause mortality rate was significantly reduced in the magnesium sulfate group versus the non-magnesium sulfate group (30.4% vs. 44.6%, P = 0.002). Kaplan-Meier survival analysis demonstrated superior 30-day cumulative survival rates in the magnesium sulfate group after PSM (P < 0.001). Both multivariable Cox regression (HR, 0.62, 95% CI, 0.47-0.82, P < 0.001) and inverse probability weighting (IPW) analysis (HR, 0.69, 95% CI, 0.53-0.89, P = 0.005) indicated that magnesium sulfate treatment was an independent protective factor for 30-day all-cause mortality risk. Conclusion:The use of magnesium sulfate is associated with a reduction in all-cause mortality among SALI patients. Future research should consider individual patient variations to explore its true effectiveness.

The crosstalk between lactylation and autophagy within the hepatocellular carcinoma (HCC) microenvironment is a burgeoning field with profound implications. By integrating multi-omics data from public cohorts, we delineated two molecular subtypes of HCC with divergent clinical outcomes and established a lactylation-autophagy-related prognostic signature. This signature highlighted CLEC3B as a pivotal gene. Subsequent single-cell RNA sequencing and experimental validation unequivocally pinpointed liver sinusoidal endothelial cells (LSECs) as the principal cellular source of CLEC3B, which was significantly downregulated in HCC tissues. Functionally, conditioned media derived from CLEC3B-overexpressing LSECs potently inhibited HCC cell proliferation. Mechanistic investigations revealed that this tumor-suppressive effect was orchestrated through the concurrent suppression of autophagy and diminution of lactylation levels. Our findings position LSEC-secreted CLEC3B as a novel metabolic mediator in HCC, bridging two key pathways in tumor suppression, and endorse its clinical value both as a prognostic indicator and a promising therapeutic target.

Neutrophil extracellular traps (NETs) drive severe acute pancreatitis (SAP) progression by promoting pancreatic injury, duct obstruction, and systemic inflammation. Reactive oxygen species (ROS) are critical for NETs formation, while NETs degradation remains therapeutically challenging. This study investigates whether combined N-acetylcysteine (NAC) and deoxyribonuclease I (DNase I) therapy mitigates SAP and associated lung injury by suppressing NETs formation and degradation, respectively, and explores the underlying molecular mechanisms. NETs were elevated in SAP pancreatic tissue. In vitro, NAC reduced NETs formation by inhibiting oxidative stress, while DNase I degraded preformed NETs. Combined therapy surpassed monotherapy efficacy, synergistically attenuating NETs burden. In vivo, Early dual intervention degraded NETs, reduced neutrophil infiltration and apoptosis, and lowered inflammatory cytokines, thereby alleviating pancreatitis and lung injury. Mechanistically, dual therapy suppressed NF-κB activation in pancreatic tissue, decreasing CXCL3 release and subsequent CXCR2-positive neutrophil recruitment, ultimately ameliorating SAP. NAC and DNase I synergistically target NETs generation and clearance, offering a promising redox-based therapeutic strategy for SAP. This schematic diagram illustrates the in vivo mechanisms of combined NAC andDNase I administration in ameliorating SAP. Schematic Diagram of the Mechanism In Vivo, Created by BioRender.

Purpose To investigate the association and causality between physical activity and hepatitis C virus (HCV) infection. Methods In this cross-sectional study, we conducted a bidirectional two-sample Mendelian Randomization (MR) analysis using data from the GWAS database. The analytic population was divided into three categories based on hepatitis C virus infection outcomes, and statistical differences were analyzed with covariates. Additionally, Restricted Cubic Spline (RCS) analysis was applied to examine the nonlinear relationship between physical exercise and hepatitis C virus infection. Instrumental variables were selected with a threshold of P < 5*10⁻⁸, and the causal relationships were assessed using IVW, MR-Egger, Weighted Median, Weighted Mode, and Simple Mode methods. Cochran's Q test was used to assess and exclude data heterogeneity. Results In the population data analysis, the moderate-to-vigorous work (MVW) variable showed a significant association with HCV infection (p = 0.006). MVW demonstrated a positive association with HCV infection (OR = 2.46, 95% CI = 1.02–5.92). RCS analysis indicated that the risk of HCV infection significantly increased when MVW exceeded 810 minutes per week. MR analysis indicated that MVW had a unidirectional effect on HCV infection (IVW model: OR = 1.366, 95% CI: 1.121–1.663, p = 0.002), and heterogeneity was excluded from the analysis. Conclusions In the context of HCV infection, the primary factor influencing risk is not the duration of physical activity, but the type. High-intensity physical activity significantly elevates the risk of HCV infection. It may be prudent to manage work hours responsibly to minimize physical strain.

AIM:There is no consensus regarding the minimally invasive treatment method for concomitant cholecystolithiasis and choledocholithiasis. Therefore, this study aimed to develop a universal classification system for minimally invasive surgeries, thereby supporting development of consensus in guidelines for diagnosing choledocholithiasis. METHODS:This retrospective study included 1044 consecutive patients with concomitant cholecystolithiasis and choledocholithiasis who underwent different minimally invasive surgical treatments at the Zhongnan Hospital of Wuhan University, China, between January 2014 and April 2021. To identify the key factors influencing the choice of different minimally invasive surgical procedures, clinical data for all hospitalized patients were analyzed. The patients were followed up through outpatient visits or telephonic calls at 1 week, 6 weeks, 3 months, 6 months, and 1 year or immediately if symptoms developed following discharge from the hospital. This information was integrated in the form of a new disease classification model, and the optimal treatment approaches were screened. RESULTS:A significant correlation was observed between the choice of minimally invasive surgical procedures and the concomitant common bile duct (CBD) (p < 0.001), stone size (p < 0.001), or stone number (p < 0.001). A new clinical classification model was developed for patients with concomitant gallbladder (GB) and CBD stones based on the CBD diameter, stone sizes, and stone numbers, and the patients were sorted into Type I, II, III, and Ⅳ, respectively. Three invasive surgical methods were performed in patients with type I patients, revealing the laparoscopic cholecystectomy + Laparoscopic Transcystic Common Bile Duct Exploration (LC + LTCBDE) method as a preferred option for these patients. Furthermore, five surgical methods were performed on patients with type II CBD stones, demonstrating LC + LTCBDE as the viable option for these patients. Additionally, among the four minimally invasive surgical methods applied in patients with type III, the LC + laparoscopic choledochotomy for common bile duct exploration (LCCBDE) + Duodenoscope or LC + LCCBDE + primary closure demonstrated favorable results in this group of patients. Among the three methods applied in type IV patients, LC + laparoscopic choledocholithotomy and T-tube drainage (LCTD) were found to be more favorable. CONCLUSIONS:In summary, this novel and simple clinical classification system, which is based on CBD diameter, stone sizes, and stone numbers, can assist clinicians in selecting a minimally invasive treatment approach for managing concomitant GB and CBD stones.

Hepatocellular carcinoma (HCC) remains a lethal malignancy with a persistently poor prognosis. While our previous studies established the anti-tumor function of Deoxyribonuclease I Like Protein 3 (DNASE1L3) in HCC, the underlying mechanisms involving the immune microenvironment are less understood. Here, we demonstrate that loss of DNASE1L3 accelerated HCC progression by impairing M1-type macrophage polarization in Dnase1l3 knockout (KO) mice, co-culture models, RNA-seq, and comprehensive molecular/cellular analyses. Mechanistically, DNASE1L3 deletion suppresses tumor-associated macrophages (TAMs) polarization toward the M1 phenotype and inhibits pyroptosis by attenuating the NLRP3 inflammasome/gasdermin D (GSDMD) pathway in vitro and in vivo, thereby reducing pyroptosis in HCC cells. This regulation involves impaired nuclear translocation of NF-κB p65. Crucially, NLRP3 agonism partially reversed DNase1L3-deletion-induced suppression of the NLRP3-GSDMD axis and restored M1 polarization. Our findings reveal DNase1L3 as a pivotal regulator of TAM phenotype via the NF-κB/NLRP3-GSDMD axis and highlight its potential for immunotherapy targeting macrophage reprogramming in HCC.

Chemodynamic therapy (CDT) plays a crucial role in transforming the tumor microenvironment by inducing immunogenic cell death (ICD) to eliminate cancer cells. Nonetheless, the effectiveness of CDT in eliciting antitumor immunity is somewhat constrained. The persistent presence of immunosuppressive cells, such as myeloid-derived suppressor cells (MDSCs), in the tumor offsets some of the ICD effects triggered by CDT. Moreover, extracellular adenosine triphosphate (eATP), a crucial damage-related molecular pattern that initiates ICD, is quickly degraded into adenosine, an immunosuppressive metabolite, by CD39 molecules prevalent in the tumor environment, thereby evading immune destruction. In this report, we introduce a nanomaterial, CP@HMM (copper-doped carbon dots and POM1 encapsulated by a hybrid membrane composed of Hepa1-6 and a MDSCs membrane), which targets liver cancer cells and MDSCs and inhibits the ATP-adenosine metabolic pathway. The hybrid membrane, derived from hepatocellular carcinoma (HCC) cells and MDSCs, facilitates the targeted delivery of copper-doped carbon dots (Cu-CDs) to these cells. The potent Fenton-like reactions and the cytotoxicity of copper ions allow CP@HMM to not only kill tumor cells but also eradicate intratumoral MDSCs. Additionally, the CD39 inhibitor POM1 within the system prevents the degradation of eATP induced by the Cu-CDs treatment. This leads to increased eATP levels and drives antitumor immunity activation, including macrophage pyroptosis and dendritic cell maturation, which suppresses primary tumor progression and distant metastases while fostering immune memory to prevent tumor recurrence. Our findings suggest that CP@HMM is an effective drug-delivery system and offers a potential therapeutic alternative for patients with HCC, promising advancements in combined tumor immunotherapy strategies.

Deoxyribonuclease 1-like 3 (DNASE1L3) is a secreted endonuclease essential for degrading extracellular DNA and maintaining immune tolerance, but its role in hepatic immune-metabolic regulation remains unclear. Using multi-omics analyses combined with immunophenotyping, we demonstrate that Dnase1l3-deficient (knockout, KO) mice exhibit disrupted myeloid differentiation, Kupffer cell M1 polarization (M1), and progressive hepatic steatosis. Integrated transcriptomic, proteomic, and metabolomic profiling revealed activation of damage-associated molecular pattern (DAMP) sensing, endoplasmic reticulum stress, redox imbalance, ferroptosis susceptibility, and lipid metabolic reprogramming. Whole-genome resequencing further identified chromosome 4 mutation hotspots linked to iron metabolism, oxidative stress, and inflammation. These findings establish DNASE1L3 as a key regulator of hepatic immune-metabolic homeostasis and suggest that its deficiency drives a pathological cascade involving pattern recognition receptor activation, endoplasmic reticulum stress, and ferroptosis, ultimately leading to non-alcoholic fatty liver disease (NAFLD). This study provides a mechanistic framework for nucleic acid-driven immunometabolic dysregulation in chronic liver disease.

The tumor microenvironment (TME) plays a pivotal role in tumor progression, immune evasion, and therapeutic responses. Among its key components, endothelial cells (ECs) are crucial regulators of angiogenesis, immune cell trafficking, and metabolic adaptations. This study integrates single-cell and transcriptomic analyses to identify tumor-specific endothelial cell signatures in hepatocellular carcinoma (HCC) and stratify tumors into three distinct molecular subtypes. These subtypes exhibit unique immune landscapes and biological characteristics, including pathway activation and differential responses to immunotherapy and targeted treatments. Using machine learning, we developed a robust prognostic scoring model to predict patient outcomes and therapy responsiveness, which was validated across independent cohorts. Our findings highlight the critical role of endothelial cells in modulating the TME and underscore the potential of targeting EC-specific molecular features to enhance the efficacy of immunotherapy and optimize personalized cancer treatment.

BACKGROUND:Hepatocellular carcinoma (HCC) remains a significant cause of cancer-related mortality, highlighting the need for novel therapeutic strategies. Identifying key proteins and potential therapeutic agents is critical for improving treatment outcomes. METHODS:We employed Mendelian randomization to identify proteins associated with HCC risk and utilized drug enrichment and molecular docking analyses to discover potential therapeutic agents. The efficacy of identified drugs was evaluated in vitro using immune-tumor co-culture systems and in vivo in a murine HCC model. Single-cell expression profiling and clinical sample analyses were conducted to explore expression patterns. RESULTS:Our analyses identified 16 proteins linked to HCC pathogenesis. Among the therapeutic agents tested, Belinostat significantly enhanced T cell-mediated cytotoxicity against HCC cells and effectively reduced tumor growth in vivo. Single-cell analysis revealed significant modulation of immune cells within the tumor microenvironment, suggesting potential mechanisms for the observed therapeutic effects. CONCLUSION:This study highlights the potential of Belinostat as a promising therapeutic agent for HCC. By modulating immune responses and tumor growth, Belinostat offers a novel approach to HCC treatment, warranting further clinical investigation to validate its efficacy and therapeutic potential.

Abstract Background Although gut microbiota and serum metabolite composition have been observed to be altered in patients with non-alcoholic fatty liver disease (NAFLD), previous observational studies have demonstrated inconsistent results. As this may be influenced by factors such as confounders and reverse causality, we used Mendelian randomization to clarify the causal effect of gut microbiota and blood metabolites on NAFLD. Methods In this research, we performed a two-step Mendelian randomization analysis by utilizing genome-wide association study (GWAS) data obtained from MiBioGen and UK Biobank. To mitigate potential errors, we employed False Discovery Rate (FDR) correction and linkage unbalanced regression (LDSC) analysis. Sensitivity analyses including cML-MA and bidirectional Mendelian randomization were performed to ensure the robustness of the results. Results In this study, a total of nine gut microbiota and seven metabolites were found to be significantly associated with NAFLD. MR analysis of the above findings revealed a causal relationship between Ruminococcus2 and cysteine-glutathione disulfide (OR = 1.17, 95%CI = 1.006–1.369, P = 0.041), as well as 3-indoleglyoxylic acid (OR = 1.18, 95%CI = 1.011–1.370, P = 0.036). For each incremental standard deviation in Ruminococcus2 abundance, there was a corresponding 26% reduction in NAFLD risk (OR = 0.74, 95%CI = 0.61–0.89, P = 0.0012), accompanied by a 17% increase in cysteine-glutathione disulfide levels (OR = 1.17, 95%CI = 1.01–1.37, P = 0.041) and an 18% increase in 3-indoleglyoxylic acid levels (OR = 1.18, 95%CI = 0.81-1.00, P = 0.036). The proportion mediated by cysteine-glutathione disulfide is 11.2%, while the proportion mediated by 3-indoleglyoxylic acid is 7.5%. Conclusion Our study suggests that increased abundance of specific gut microbiota may reduce the risk of developing NAFLD, and this relationship could potentially be mediated through blood metabolites.

SAP05, a secreted effector by the obligate parasitic bacteria phytoplasma, bridges host SPL and GATA transcription factors (TFs) to the 26 S proteasome subunit RPN10 for ubiquitination-independent degradation. Here, we report the crystal structures of SAP05 in complex with SPL5, GATA18 and RPN10, which provide detailed insights into the protein-protein interactions involving SAP05. SAP05 employs two opposing lobes with an acidic path and a hydrophobic path to contact TFs and RPN10, respectively. Our crystal structures, in conjunction with mutagenesis and degradation assays, reveal that SAP05 targets plant GATAs but not animal GATAs dependent on their direct salt-bridged electrostatic interactions. Additionally, SAP05 hijacks plant RPN10 but not animal RPN10 due to structural steric hindrance and the key hydrophobic interactions. This study provides valuable molecular-level information into the modulation of host proteins to prevent insect-borne diseases.

The ribosome-associated quality-control (RQC) pathway degrades aberrant nascent polypeptides arising from ribosome stalling during translation. In mammals, the E3 ligase Pirh2 mediates the degradation of aberrant nascent polypeptides by targeting the C-terminal polyalanine degrons (polyAla/C-degrons). Here, we present the crystal structure of Pirh2 bound to the polyAla/C-degron, which shows that the N-terminal domain and the RING domain of Pirh2 form a narrow groove encapsulating the alanine residues of the polyAla/C-degron. Affinity measurements in vitro and global protein stability assays in cells further demonstrate that Pirh2 recognizes a C-terminal A/S-X-A-A motif for substrate degradation. Taken together, our study provides the molecular basis underlying polyAla/C-degron recognition by Pirh2 and expands the substrate recognition spectrum of Pirh2.

目的:探讨术前炎症指标包括外周血中性粒细胞与淋巴细胞比值(NLR)、血小板与淋巴细胞比值(PLR)、单核细胞与淋巴细胞比值(MLR)对肝细胞癌(HCC)患者术后复发的预测价值.方法:收集234例肝细胞癌患者的术前炎症指标资料及术后3年随访资料.根据受试者工作特征(ROC)曲线确定预测复发的截距值并分组.采用单因素和多因素COX模型分析肝细胞癌患者复发的危险因素.使用倾向性评分匹配(PSM)均衡分组后比较组间差异,并进行组间生存分析.结果:ROC曲线分析得到NLR预测复发的最佳截距值2.88,PLR的最佳截距值138.09,MLR的最佳截距值是0.48.据此,分别将纳入患者分为高/低NLR组、高/低PLR组和高/低MLR组.单因素分析显示甲胎蛋白(AFP)、NLR、PLR和MLR是肝细胞癌患者复发的危险因素.多因素分析显示AFP、NLR和PLR是肝细胞癌患者术后复发的独立危险因素.倾向性评分匹配后亚组间差异均衡(P＞0.05).匹配后组间生存分析显示,高NLR组3年复发率22.8％,低NLR组3年复发率10.4％(P＜0.05);高PLR组3年复发率32.3％,低PLR组3年复发率17.2％(P＜0.05).结论:术前NLR值、PLR值是影响肝细胞癌患者术后复发的独立危险因素.NLR＞2.88、PLR＞138.09的患者术后更容易复发.

We investigated the change of Genome-wide methylation patterns, DNA methylation patterns, miR-1297 expression after SNHG6 knockdown or overexpression in hepatoma cells. q-PCR was performed to simultaneously detect MAT1A mRNA in the nucleus and cytoplasm after FUS overexpression in Huh7 and Hep3B cells. RIP with q-PCR analysis was utilized to verify the direct interaction between the FUS protein and MAT1A mRNA. RNA FISH was used to study the subcellular localization and visual molecular abundance of SNHG6, miR-1297, FUS mRNA and MAT2A mRNA in Huh7 and Hep3B cells.We also identified that miR-1297 inhibits FUS expression by binding to its 3'UTR.

Supplementary Tables 1-3 from Hypoxia Induces Genomic DNA Demethylation through the Activation of HIF-1α and Transcriptional Upregulation of MAT2A in Hepatoma Cells