Liver fibrosis, a common outcome of chronic liver injury, is characterized by excessive inflammation and oxidative stress. Rutin, a bioactive flavonoid with known antioxidant and anti-inflammatory properties, has not been thoroughly investigated for its potential anti-fibrotic mechanisms. This study aimed to elucidate the role of Rutin in liver fibrosis and its underlying molecular pathways. A carbon tetrachloride (CCl₄)-induced murine liver fibrosis model was employed. Liver injury, fibrotic deposition, inflammatory response, and oxidative stress were evaluated through histopathological examination, Western blotting, quantitative real-time PCR, and RNA sequence. The involvement of immune-responsive gene 1 (IRG1) was investigated using IRG1-knockout mice, while molecular docking and cellular thermal shift assay (CETSA) were performed to assess Rutin-IRG1 binding. The results showed that Rutin treatment significantly attenuated CCl₄-induced hepatic injury and collagen accumulation, accompanied by reduced markers of fibrosis. Mechanistically, Rutin activated the IRG1-itaconate axis, leading to a notable decrease in reactive oxygen species and pro-inflammatory cytokine release by Nrf2 activation and NLRP3 inflammasome containment. Molecular analyses confirmed direct binding of Rutin to IRG1, stabilizing its structure and enhancing its functional activity. The protective effects of Rutin were abolished in IRG1-deficient mice, underscoring the essential role of IRG1 in its anti-fibrotic action. In conclusion, Rutin ameliorates liver fibrosis by mitigating oxidative stress and suppressing NLRP3 inflammasome activation through targeting the IRG1/itaconate pathway, revealing a novel immunometabolic mechanism for its hepatoprotective effect.

Liver fibrosis is a common consequence of chronic liver injury, driven by persistent inflammation, oxidative stress, and excessive extracellular matrix deposition. The natural flavonoid rutin possesses antioxidant and anti-inflammatory properties and has shown hepatoprotective potential; however, its anti-fibrotic mechanism remains incompletely understood. This study aimed to investigate the protective effects of rutin and its underlying molecular mechanism in a carbon tetrachloride (CCl₄)-induced mouse model of liver fibrosis, with a particular focus on the role of immune-responsive gene 1 (IRG1). Liver fibrosis was induced in mice by CCl₄ administration. Hepatic injury, collagen deposition, inflammatory activation, and oxidative stress were evaluated using histological, biochemical, molecular, and transcriptomic approaches. The involvement of IRG1 was assessed by employing IRG1-deficient mice. Direct binding between rutin and IRG1 was examined through molecular docking, molecular dynamics simulations, and the cellular thermal shift assay (CETSA). Rutin treatment markedly alleviated CCl₄-induced hepatic injury, fibrotic deposition, inflammatory cytokine production, and reactive oxygen species accumulation. Mechanistically, rutin directly bound to and stabilized IRG1, which was associated with enhanced IRG1 enzymatic activity and increased endogenous itaconate production. This led to activation of Nrf2-mediated antioxidant signaling and suppression of NLRP3 inflammasome activation. Importantly, the protective effects of rutin were largely abolished in IRG1-deficient mice, confirming that IRG1 is essential for rutin-mediated hepatoprotection. These findings identify rutin as a potential anti-fibrotic agent that targets the IRG1/itaconate axis to coordinate antioxidant and anti-inflammatory responses during liver fibrosis.

Background Bloom's taxonomy is widely used to classify examination items by cognitive level, yet its predictive validity for item discrimination remains understudied in pharmacology education. This study investigated whether cognitive level coding predicts item discrimination in an anesthetic pharmacology examination. Methods This retrospective document analysis examined 58 items from a final anesthetic pharmacology examination completed by 194 undergraduate students. Two independent raters coded items using a modified three-level Bloom's taxonomy (recall, interpretation, problem-solving). Item difficulty (P) and discrimination (D) indices were extracted from the official test analysis report. Predictive validity was assessed using Spearman correlation, linear regression, and one‑way ANOVA. Items with D < 0.10 underwent distractor analysis and cognitive attribution. Results The examination demonstrated excellent reliability (Cronbach's α = 0.91), moderate difficulty (P = 0.68), and good overall discrimination (D = 0.45). Cognitive level was positively correlated with item discrimination (rₛ = 0.42, 95% CI [0.18, 0.61], P < 0.01), explaining 16.8% of the variance (R² = 0.168). Problem‑solving items exhibited higher discrimination (0.49 ± 0.18) than recall items (0.34 ± 0.22) (η² = 0.14, P = 0.03). Five items (8.6%) showed low discrimination (D < 0.10), predominantly at the recall level, and were compromised by ceiling effects, dysfunctional distractors, or strong‑distractor‑induced empirical error. Conclusions Cognitive level coding significantly predicts item discrimination in anesthetic pharmacology assessments, with problem‑solving items demonstrating superior discriminatory power. However, predictive validity depends on item‑writing quality. Integrating cognitive blueprints into examination design and refining distractors using empirical data are recommended to enhance assessment quality in pharmacology education. Clinical trial number Not applicable.

A growing body of evidence suggests that enhanced glycolysis profoundly reprograms inflammatory response in acute lung injury (ALI)/ acute respiratory distress syndrome (ARDS), but the underlying mechanisms largely remain unclear. Lactate is the end-product of glycolysis, which has been recently found to function as a bioactive metabolite via G-protein coupled receptor 81 (GPR81). In the present study, the potential roles of lactate-GPR81 axis were investigated in mice with lipopolysaccharide (LPS)-induced ALI. The results indicated that LPS challenge increased the level of lactate in bronchoalveolar lavage fluid (BALF). Pharmacological suppression of glycolysis or inhibition of lactate dehydrogenase decreased lactate level and alleviated lung injury, but supplementation with lactate or a GPR81 agonist exacerbated lung injury. Global deletion of GPR81 or endothelial-specific deletion of GPR81, but not myeloid-specific deletion of GPR81, resulted in alleviated lung injury. In endothelial cells, the differentially expressed genes (DEGs) in RNA-seq after lactate supplementation were enriched in cAMP signaling pathway. Consistently, supplementation with lactate resulted in decline of cAMP, reduction of VE-cadherin and enhanced phosphorylation of myosin light chain 2 (MLC2) in endothelial cells, whereas knockdown of GPR81 reversed these effects. In addition, the modulation of lactate/GPR81 on cAMP, VE-cadherin and MLC2 was validated in GPR81 knockout mice. Importantly, the elevation of lactate is positively correlated with the degree of protein leakage, the level of proinflammatory cytokines in BALF and APACHE II score from patients with ARDS. Taken together, the present study suggests that endothelial GPR81 molecularly bridges glycolysis and inflammation, which drives microvascular hyperpermeability and the development of ALI.

The metabolic basis of tumor-associated macrophages (TAMs)-driven immune checkpoint blockade (ICB) resistance remains poorly understood. Here, in patients with immunotherapy-resistant cancer, we identify significant enrichment of TAMs marked by elevated aldehyde dehydrogenase 2 (ALDH2) expression. Myeloid-restricted ALDH2 ablation converts TAMs from a pro-tumorigenic phenotype to immunostimulatory regulators, concomitantly amplifying CD8+ T cell infiltration and cytotoxicity to improve ICB responsiveness. Mechanistically, ALDH2 deficiency induces the intracellular accumulation of reactive aldehydes, specifically 4-hydroxynonenal, which activates the PI3K-AKT signaling axis. This pathway phosphorylates and suppresses EZH2 methyltransferase activity, leading to the erosion of H3K27me3-mediated epigenetic silencing at CXCL9 promoter region. Subsequent CXCL9 derepression in TAMs facilitates persistent CD8+ T cell infiltration and enhances their cytotoxic effector functions. Clinical validation confirms that pronounced ALDH2 elevation in TAMs correlates with accelerated immunotherapy failure. Therapeutically, as a clinically approved ALDH2 inhibitor, disulfiram exerts its anti-tumor effect by selective reprogramming TAMs metabolism. Overall, our findings delineate a druggable ALDH2-metabolism-epigenetics axis in antitumor immunity, nominating ALDH2 inhibition for combination immunotherapy.

Liver fibrosis, a common pathological endpoint of chronic liver diseases, is critically shaped by Notch signaling in establishing a pro-fibrotic microenvironment. However, the mechanisms by which Notch signaling orchestrates the transition from pro-inflammatory to pro-fibrotic microenvironments, particularly through crosstalk between inflammatory macrophages and fibrogenic hepatic stellate cells (HSCs), remain poorly defined. Here, using mouse models of methionine-choline-deficiency (MCD) diet-induced metabolic dysfunction-associated steatohepatitis (MASH) and CCl4-induced liver fibrosis, we demonstrate that macrophage-derived Delta-like ligand 4 (DLL4) exacerbates liver fibrosis by activating HSCs. Macrophage-specific knockout Dll4 markedly attenuated the hepatitis and liver fibrosis, whereas Dll4 overexpression aggravated these pathologies. Mechanistically, macrophage-derived DLL4 activates the Notch2 receptor on HSCs, driving their activation and proliferation, as evidenced by upregulated α-SMA and COL1A1 and increased EdU+ cells. Critically, Notch2 knockdown in HSCs reversed DLL4-induced HSCs activation and proliferation. Notably, macrophage DLL4 expression is regulated by the TLR4-NF-κB pathway, as LPS stimulation increases DLL4 expression, while NF-κB inhibition inhibits its expression. Collectively, our findings identify the DLL4-Notch2 axis as a key mediator of the interaction between macrophages and HSCs during fibrosis, highlighting its therapeutic potential to block the progression of chronic liver diseases.

T cell dysfunction enables tumor immune evasion, understanding its mechanism is crucial for improving immunotherapy. Here we show, by RNA-sequencing analysis of human colon adenocarcinoma and triple-negative breast cancer tissues, that expression of Adipocyte Enhancer-Binding Protein 1 (AEBP1) positively correlates with T cell dysfunction and indicative of unfavorable patient outcomes. Subsequent single-cell RNA sequencing identifies cancer-associated fibroblasts (CAF) as the primary AEBP1 source. Fibroblast-specific AEBP1 deletion in mice enhances T cell cytotoxicity and suppresses tumor growth. Mechanistically, autocrine AEBP1 binds CKAP4 on CAFs, activating AKT/PD-L1 signaling to drive T cell dysfunction. By molecular-docking-based virtual screening we identify Chem-0199, a drug that disrupts the interaction between AEBP1 and CKAP4, thereby enhancing antitumor immunity. Both genetic and pharmacological AEBP1 inhibition synergize with immune checkpoint blockade in syngeneic models. Our study establishes AEBP1 as a key regulator of CAF-mediated T cell dysfunction and a therapeutic target.

BACKGROUND:Liver fibrosis is a progressive disorder resulting from chronic liver injury, and its molecular mechanism remains incompletely elucidated. METHODS:The role of PROM2 was assessed by integrating transcriptomic datasets with in vivo liver fibrosis models. Functional analyses were performed using PROM2 knockdown or overexpression strategies, along with NLRP3 knockout models, to investigate its mechanistic involvement in epithelial-mesenchymal transition (EMT) and inflammasome activation. RESULTS:Transcriptomic analysis revealed that PROM2 expression was significantly upregulated in fibrotic livers and positively correlated with fibrosis- and EMT-related gene signatures. PROM2 knockdown significantly attenuated hepatic inflammation, inhibited EMT, and reduced fibrotic remodeling. In contrast, PROM2 overexpression aggravated these pathological features. Notably, in NLRP3-deficient mice, PROM2 overexpression failed to induce inflammation, EMT, or fibrosis, indicating that the pro-fibrotic effects of PROM2 are mediated through NLRP3 signaling. CONCLUSION:PROM2 contributes to liver fibrosis by promoting EMT and activating NLRP3-dependent inflammatory pathways, suggesting it as a potential therapeutic target for liver fibrosis.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease, but effective therapeutic drugs are still lacking. Dihydrotanshinone I (DHTS), a natural product isolated from Salvia miltiorrhiza , has been shown to have ameliorative effects on NAFLD. The aim of this study was to investigate the hepatoprotective effect of DHTS on NAFLD and its mechanism. A model of NAFLD and DHTS treatment was established using a Western diet to observe the effect of DHTS on NAFLD, which were detected by immunohistochemical, immunofluorescence, and other experiments. The mechanism was further explored by constructing immune responsive gene 1 (IRG1) knockout mice, RNA sequence, and molecular docking. The results revealed that DHTS significantly improved diet-induced metabolic disorders in mice, notably alleviating liver inflammation, oxidative stress, and fibrosis. Further analysis revealed that the intervention of DHTS was associated with the activation of IRG1. Subsequent experiments confirmed that IRG1 gene deletion reversed the above protective effects of DHTS in NAFLD. Mechanistically, DHTS enhanced the antioxidant nuclear factor-erythroid 2-related factor 2 (Nrf2) pathway through IRG1/itaconate and blocked the oxidative stress response in the liver. In addition, DHTS also inhibited the activation of NACHT-, leucine-rich repeat (LRR)-, and pyrin domain (PYD)-containing protein 3 (NLRP3) inflammasome via IRG1/itaconate, blocking the inflammatory amplification effect in the liver. The study suggests that DHTS may be a potential drug for the treatment of NAFLD, which exerts protective regulatory effects mainly through the IRG1/itaconate molecular pathway.

T cell exclusion is crucial in enabling tumor immune evasion and immunotherapy resistance. However, the key genes driving this process remain unclear. We uncovered a notable increase of insulin-like growth factor 2 (IGF2) in immune- excluded tumors, predominantly secreted by cancer-associated fibroblasts (CAFs). Using mice with systemic or fibroblast- specific deletion of IGF2, we demonstrated that IGF2 deficiency enhanced the infiltration and cytotoxic activity of CD8+ T cells, leading to a reduction in tumor burden. Integration of spatial and single-cell transcriptomics revealed that IGF2 promoted interaction between CAFs and T cells via CXCL12 and programmed death ligand 1 (PD-L1). Mechanistically, autocrine IGF2 activated PI3K/AKT signaling by binding to the IGF1 receptor (IGF1R) on CAFs, which was required for the immunosuppressive functions of CAFs. Furthermore, genetic ablation of IGF2 or targeted inhibition of the IGF2/IGF1R axis with the inhibitor linsitinib markedly boosted the response to immune checkpoint blockade. Clinically, elevated levels of IGF2 in tumors or plasma correlated with an adverse prognosis and reduced efficacy of anti-programmed death 1 treatment. Together, these results highlight the pivotal role of IGF2 in promoting CAF-mediated immunoevasion, indicating its potential as a biomarker and therapeutic target in immunotherapy.

Excessive inflammatory response and increased oxidative stress play an essential role in the pathophysiology of ischemia/reperfusion (I/R)-induced acute kidney injury (IRI-AKI). Emerging evidence suggests that lipoxin A4 (LXA4), as an endogenous negative regulator in inflammation, can ameliorate several I/R injuries. However, the mechanisms and effects of LXA4 on IRI-AKI remain unknown. In this study, A bilateral renal I/R mouse model was used to evaluate the role of LXA4 in wild-type, IRG1 knockout, and IRAK-M knockout mice. Our results showed that LXA4, as well as 5-LOX and ALXR, were quickly induced, and subsequently decreased by renal I/R. LXA4 pretreatment improved renal I/R-induced renal function impairment and renal damage and inhibited inflammatory responses and oxidative stresses in mice kidneys. Notably, LXA4 inhibited I/R-induced the activation of TLR4 signal pathway including decreased phosphorylation of TAK1, p36, and p65, but did not affect TLR4 and p-IRAK-1. The analysis of transcriptomic sequencing data and immunoblotting suggested that innate immune signal molecules interleukin-1 receptor-associated kinase-M (IRAK-M) and immunoresponsive gene 1 (IRG1) might be the key targets of LXA4. Further, the knockout of IRG1 or IRAK-M abolished the beneficial effects of LXA4 on IRI-AKI. In addition, IRG1 deficiency reversed the up-regulation of IRAK-M by LXA4, while IRAK-M knockout had no impact on the IRG1 expression, indicating that IRAK-M is a downstream molecule of IRG1. Mechanistically, we found that LXA4-promoted IRG1-itaconate not only enhanced Nrf2 activation and increased HO-1 and NQO1, but also upregulated IRAK-M, which interacted with TRAF6 by competing with IRAK-1, resulting in deactivation of TLR4 downstream signal in IRI-AKI. These data suggested that LXA4 protected against IRI-AKI via promoting IRG1/Itaconate-Nrf2 and IRAK-M-TRAF6 signaling pathways, providing the rationale for a novel strategy for preventing and treating IRI-AKI.

AbstractNon-alcoholic steatohepatitis (NASH) is a predominant metabolic liver disease, typically characterized by hepatic steatosis, oxidative stress, and inflammation. The traditional Chinese medicine Cornus officinalis possesses anti-inflammatory and hepatoprotective pharmacological properties and has shown ameliorative effects on NASH. however, its mechanism of action remains unclear. This study aims to elucidate the mechanisms by which C. officinalis ameliorates NASH. The active components of C. officinalis were analyzed using the Traditional Chinese Medicine Systems Pharmacology Database (TCMSP), and the corresponding targets were predicted. Subsequently, the DisGeNET, GeneCards, and GEO databases were employed to identify NASH-related targets. Venn diagrams were used to intersect the C. officinalis targets with the NASH targets. Protein–protein interaction (PPI) networks were constructed using the STRING database, and PPI network analysis was performed using Cytoscape. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were conducted using the Database for Annotation, Visualization, and Integrated Discovery (DAVID), followed by molecular docking validation. Cornus officinalis was found to contain 20 major active ingredients corresponding to 672 potential targets, 61 of which overlapped with NASH targets. PPI network, GO, and KEGG pathway analyses identified four targets with the highest correlation, and molecular docking results indicated that the active ingredients of C. officinalis exhibited strong binding affinities to NASH targets. The treatment of NASH with C. officinalis is characterized by multiple active ingredients and multiple targets, underscoring the major advantage of traditional Chinese medicine in treating NASH.

Background & aimsUnrestricted endoplasmic reticulum (ER) stress and the continuous activation of ER associated protein degradation (ERAD) pathway might lead to the aggravation of non-alcoholic steatohepatitis (NASH). Derlin-1 has been considered to be an integral part of the ERAD pathway, which is involved in the regulation of the transport and excretion of protein degradation products within ER. However, the regulatory role and mechanism of Derlin-1 in NASH remains unclear.MethodsThe expression of Derlin-1 was firstly detected in the liver of normal and NASH animal model and patient. Then, western diet (WD)-induced NASH mice were administrated with the lentivirus-mediated Derlin-1 knockdown or overexpression. Finally, RIPK3 knockout mice were used to explore the mechanism. The liver injury, hepatic steatosis, inflammation, and fibrosis as well as ER stress signal pathway were evaluated.ResultsThe levels of Derlin-1 were significantly elevated in the liver of WD-fed mice and NASH patients when compared to the control group. Furthermore, Derlin-1 knockdown attenuated WD-induced liver injury, lipid accumulation, inflammatory response, and fibrosis. Conversely, overexpression of Derlin-1 presented the completely opposite results. Mechanistically, Derlin-1 enhanced ER stress pathways and led to necroptosis, and RIPK3 knockout dramatically reduced Derlin-1 expression and reversed the progression of NASH aggravated by Derlin-1.ConclusionsNotably, Derlin-1 is a critical modulator in NASH. It may accelerate the progression of NASH by regulating the activation of the ERAD pathway and further aggravating the ER stress, which might be involved in RIPK3-mediated necroptosis. Therefore, targeting Derlin-1 as a novel intervention point holds the potential to delay or even reverse NASH.

BackgroundNon-alcoholic steatohepatitis (NASH) is a metabolic dysregulation-related disorder that is generally characterized by lipid metabolism dysfunction and an excessive inflammatory response. Currently, there are no authorized pharmacological interventions specifically designed to manage NASH. It has been reported that Ginkgolide C exhibits anti-inflammatory effects and modulates lipid metabolism. However, the impact and function of Ginkgolide C in diet-induced NASH are unclear.MethodsIn this study, mice were induced by a Western Diet (WD) with different doses of Ginkgolide C with or without Compound C (adenosine 5 '-monophosphate (AMP)-activated protein kinase (AMPK) inhibitor). The effects of Ginkgolide C were evaluated by assessing liver damage, steatosis, fibrosis, and AMPK expression.ResultsThe results showed that Ginkgolide C significantly alleviated liver damage, steatosis, and fibrosis in the WD-induced mice. In addition, Ginkgolide C markedly improved insulin resistance and attenuated hepatic inflammation. Importantly, Ginkgolide C exerted protective effects by activating the AMPK signaling pathway, which was reversed by AMPK inhibition.ConclusionGinkgolide C alleviated NASH induced by WD in mice, potentially via activating the AMPK signaling pathway.

Piperine, the major active substance in black pepper, has been shown to have anti-inflammatory and antioxidant effects in several ischemic diseases. However, the role of piperine in hepatic ischemia/reperfusion injury (HIRI) and its underlying mechanisms remain unclear. In this study, the mice were administered piperine (30 mg/kg) intragastric administration before surgery. After 24 h of hepatic ischemia-reperfusion, liver histopathological evaluation, serum transaminase measurements, and TUNEL analysis were performed. The infiltration of inflammatory cells and production of inflammatory mediators in the liver tissue were determined by immunofluorescence and immunohistochemical staining. The protein levels of toll-like receptor 4 (TLR4) and related proteins such as nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), interleukin-1 receptor-associated kinase 1 (IRAK1), p65, and p38 were detected by western blotting. The results showed that plasma aminotransferase (ALT), aspartate aminotransferase (AST), hepatocyte apoptosis, oxidative stress, and inflammatory cell infiltration significantly increased in HIRI mice. Piperine pretreatment notably repaired liver function, improved the histopathology and apoptosis of liver cells, alleviated oxidative stress injury, and reduced inflammatory cell infiltration. Further analysis showed that piperine attenuated tumor necrosis factor-a (TNF-α) and interleukin 6 (IL-6) production and reduced TLR4 activation and phosphorylation of IRAK1, p38, and NF-κB in HIRI. Piperine has a protective effect against HIRI through the TLR4/IRAK1/NF-κB signaling pathway and may be a safer option for future clinical treatment and prevention of ischemia-related diseases.

Non-alcoholic steatohepatitis (NASH) is a prevalent metabolic disease, characterized by the hepatic steatosis, inflammation, and fibrosis, which is lack of effective treatment currently. Protectin D1 (PTD1), a lipid mediator from omega-3 fatty acid docosahexaenoic acid (DHA), has displayed wide pharmacological actions including anti-inflammation in a variety of diseases, but the role of PTD1 on NASH remains unclear. In this study, using the methionine and choline deficient (MCD) fed NASH model, we explored the effect and underlying mechanism of PTD1 on NASH in mice. Our results showed PTD1 improved MCD-induced steatosis, hepatocellular injury, inflammation and fibrosis. Furthermore, PTD1 inhibited MCD-induced activation of TLR4 downstream molecules (TAK1, p38 and p65) without affecting the levels of TLR4 and phosphorylated IRAK-1. Notably, the levels of IRAK-M protein and the binding between IRAK-M and TRAF6 in the liver were also increased by PTD1 in NASH mice. Moreover, IRAK-M knockout remarkedly reverted the beneficial effects of PTD1 on the NASH in mice. Thus, these results demonstrated that PTD1 could protect mice from NASH by inhibiting the activation of TLR4 downstream signaling pathway, which might be related to the upregulation of IRAK-M, indicating that PTD1 may provide a new treatment for NASH.

Background and Objectives. The prognostic role of adjacent nontumor tissue in patients with breast cancer (BC) is still unclear. The activity changes in immunologic and hallmark gene sets in normal tissues adjacent to BC may play a crucial role in predicting the prognosis of BC patients. The aim of this study was to identify BC subtypes and ribosome-associated prognostic genes based on activity changes of immunologic and hallmark gene sets in tumor and adjacent nontumor tissues to improve patient prognosis. Materials and Methods. Gene set variation analysis (GSVA) was applied to assess immunoreactivity changes in the overall sample and three immune-related BC subtypes were identified by non-negative matrix factorization (NMF). KEGG (Kyoto Encyclopedia of Genes and Genomes) and GO (Gene Ontology) analyses were after determining the prognostic gene set using the least absolute shrinkage and selection operator (LASSO) method. Ribosome-related genes were identified by PPI (protein-protein interaction) analysis, and finally a prognostic risk model was constructed based on the expression of five ribosomal genes (RPS18, RPL11, PRLP1, RPL27A, and RPL38). Results. A comprehensive analysis of immune and marker genomic activity changes in normal breast tissue and BC tissue identified three immune-related BC subtypes. BC subtype 1 has the best prognosis, and subtype 3 has the worst overall survival rate. We identified a prognostic gene set in nontumor tissue by the least absolute shrinkage and selection operator (LASSO) method. We found that the results of both KEGG and GO analyses were indistinguishable from those of ribosome-associated genes. Finally, we determined that genes associated with ribosomes exhibit potential as a reliable predictor of overall survival in breast cancer patients. Conclusions. Our research provides an important guidance for the treatment of BC. After a mastectomy, the changes in gene set activity of both BC tissues and the nontumor tissues adjacent to it should be thoroughly evaluated, with special attention to changes in ribosome-related genes in the nontumor tissues.

Background and Objectives: Non-alcoholic steatohepatitis (NASH) is a significant risk factor for hepatocellular carcinoma (HCC) development. Timely treatment during the NASH stage is essential to minimize the possibility of disease progression to HCC. Cuproptosis is a newly identified form of cellular death that could impact the progression of various diseases and cancers. Materials and Methods: Transcriptome and single-cell sequencing datasets were utilized to investigate the role of cuproptosis-related genes (CRGs) in NASH progression to HCC. FDX1, LIPT1, and PDHP were identified as CRGs in NASH patients, and FDX1, DBT, GCSH, SLC31A1, and DLAT were identified as CRGs in patients with NASH progressing to HCC. FDX1 was found to play a significant role in both NASH patients and patients with NASH progressing to HCC. This study constructed cuproptosis-related clusters (CRCs) using the Nonnegative Matrix Factorization algorithm, and they were linked to fatty acid metabolism and the PPAR signaling pathway in both NASH CRCs and HCC CRCs. The Weighted Correlation Network Analysis algorithm identified CRP, CRC, TAT, CXCL10, and ACTA1 as highly relevant genes in NASH CRCs and HCC CRCs. The expression of FDX1 was validated in both mouse models and human NASH samples. Results: The investigation highlights FDX1 as a pivotal CRG in both NASH and NASH progression to HCC. The comprehensive characterization of CRGs sheds light on their potential biofunctional importance in the context of NASH and HCC. Our experimental results show that FDX1 expression was significantly increased in NASH patients. Conclusions: The present study identified key CRGs, revealing their potential impact on NASH and HCC. Meanwhile, targeting FDX1 may prevent the progression of NASH to HCC.

目的 探讨过氧化物酶体增殖物激活受体共激活因子-1α(peroxisome proliferator activating receptor coactivator-1α,PGC-1α)激动剂2-(4-叔丁基苯基)苯并咪唑(2-(4-tert-butylphenyl)benzimidazole,ZLN005)对蛋氨酸胆碱缺乏饮食(choline methionine deficient diet,MCD)诱导的小鼠非酒精性脂肪性肝炎(non-alcoholic steatohepatitis,NASH)的作用及机制.方法 用MCD饲料喂养小鼠构建NASH模型,给小鼠每日口服15 mg/mL的ZLN005,8周后检测小鼠血清丙氨酸氨基转移酶(alanine aminotransferase,ALT)、天门冬氨酸氨基转移酶(aspartate aminotransferase,AST)水平,对小鼠肝组织进行HE和油红O染色,丙二醛(malondialdehyde,MDA)和超氧化物歧化酶(superoxide dismutase,SOD)活性分析,RT-PCR检测肝组织白介素-1β(leucoides-1β)mRNA和肿瘤坏死因子(tumor necrosis factor alpha,TNF-α)mRNA 的表达,Western blot检测小鼠肝组织内PGC-1α、血红素氧合酶1(heme oxygenase,HO-1)的蛋白表达.结果 ZLN005处理,显著降低了血清ALT、AST水平,减轻了MCD诱导的NASH小鼠肝损伤、脂肪变性;减少了小鼠肝组织炎性细胞浸润;同时,ZLN005降低了IL-1β和TNF-α的mRNA水平;降低了肝脏MDA的含量,提高了SOD活性;上调了PGC-1α/HO-1的表达.结论 ZLN005可缓解MCD诱导的NASH,其机制可能与激活PGC-1α/HO-1通路,发挥抗氧化应激和抗炎作用有关.

Objectives Non-alcoholic steatohepatitis (NASH) is a chronic liver disease histologically characterized by liver steatosis, hepatocellular injury, inflammation and fibrosis, resulting in cirrhosis and hepatocellular carcinoma, but effective measures and obvious pathogenesis for NASH remain elusive. Chrysin (CH) has been reported to have anti-inflammatory effects but shows lower bioavailability.Methods In this study, a chrysin nanoliposome (CH-NL) was first prepared and characterized. Then, we used the methionine-choline-deficient (MCD) diet to induce a mouse model of NASH. Finally, the effects of CH and CH-NL on NASH were evaluated in the liver of NASH mice.Key findings The results showed that CH or CH-NL significantly reduced the accumulation of lipids in hepatocytes, alleviated liver injury, decreased the generation of radical oxygen species, and attenuated the accumulation of collagen fibre in the liver of NASH mice. In addition, CH and its nano-liposomes markedly inhibited the production of inflammatory cytokines and inflammatory cell infiltration in the liver of NASH mice. Further studies found that CH-NL and CH-NL downregulated the MCD diet-induced activation of Toll-like receptor 4 (TLR4) signalling pathway in the liver of mice.Conclusions CH and its nanoliposome alleviated MCD diet-induced NASH in mice, which might be through inhibiting TLR4 signalling pathway.