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Objectives: Depression is a widely prevalent mental disorder, and nutritional interventions play an increasingly important role in its treatment. In this paper, effects of linoleic acid (LA) on depressive behavior in mice induced by gut microbiome disorders were investigated. Methods: Fifty C57BL/6J male mice were randomly separated into five groups, control group (CK), ceftriaxone sodium group (CRO), low-dose linoleic acid group (LLA, 1 g/kg), medium-dose linoleic acid group (MLA, 2 g/kg), and high-dose linoleic acid group (HLA, 5 g/kg). In the LLA, MLA, and HLA groups, mice were treated with ceftriaxone sodium (CRO) to induce depressive behaviors, followed by LA administration. Behavioral tests were used to evaluate depressive behavior. High-throughput sequencing and Hematoxylin–eosin (H&E) staining in gut microenvironment were carried out. ELISA kits were used to measure brain inflammatory factors, and 5-hydroxy-tryptamine (5-HT). Gas chromatography and western blot were used to determine fatty acids compositions and the enzymes expression involved in lipid metabolism in brain respectively. Results: The results showed that 10 weeks CRO treatment contribute to depressive behavior, gut microbiome disturbance, and serotonin system disturbance. LLA and MLA improved the depressive-like behavior, and significantly increased the levels of 5-HT1A, 5-HTT and 5-HT in the hippocampus. LLA was found to improve the diversity of gut microbiome and alleviate colon tissue damage. Meantime, LLA increased the content of linoleic acid, improved the expression of FADS2 and COX-2, increased IL-10 levels, and decreased IL-6 levels in the brain. Discussion: LA alleviated depressive behavior in mice by improving the gut microenvironment, regulate fatty acid metabolism, and modulate inflammation. [ABSTRACT FROM AUTHOR]

Introduction: Stroke is a debilitating disease and the second leading cause of death worldwide, of which ischemic stroke is the dominant type. Carthamus tinctorius L., also known as safflower, has been used to treat cerebrovascular diseases, especially ischemic stroke in many Asian countries. However, the underlying mechanisms of safflower in preventing ischemic stroke remains elusive. This study aims to elucidate the potential of safflower as a drug candidate for the prevention of ischemic stroke and to delineate its protective effects and potential mechanisms in a rat model of cerebral ischemia-reperfusion injury (CI/RI). Methods: The aqueous extract of safflower (AESF) was verified using HPLC-UV, HPLC-MS, and TLC. The inhibitory effect of AESF on platelet aggregation was detected in vitro and in zebrafish and mice. A CI/RI model in rats was established by middle cerebral artery occlusion and reperfusion to study the protective effect of AESF on ischemic stroke. 2,3,5-triphenyltetrazolium chloride, hematoxylin and eosin, and Nissl's staining were employed to evaluate the pathological changes of brain tissue. In addition, metabolomics, ELISA, and Western blot were used to uncover the molecular alteration induced by AESF. Results: AESF significantly inhibited platelet aggregation in vitro , reduced the thrombogenesis in zebrafish, and prolonged clotting time in mice. In addition, AESF alleviated neurological dysfunction, cerebral oedema, cerebral infarct size, cerebral histopathological damage induced by ischemia-reperfusion, improved neuronal survival, increased serum levels of SOD and CAT, and decreased levels of iNOS and NO. Metabolomics revealed that AESF attenuated the metabolic disturbances in brain caused by I/R injury via regulating 38 metabolites particularly related to the arachidonic acid (AA) metabolism. Moreover, AESF elevated the serum levels of 6-keto-PGF1α, a pivotal metabolite of AA, downregulated the protein expression of p53, Bax, cleaved caspase-9, cleaved caspase-3, and cleaved caspase-8, and upregulated that of Bcl-2. Conclusion: AESF mitigated CI/RI through preventing platelet aggregation, alleviating oxidative stress, and suppressing apoptosis partially via modulating AA metabolism/p53-mediated apoptosis axis. [ABSTRACT FROM AUTHOR]

Abstract Background Drug resistance and immune escape continue to contribute to poor prognosis in AML. Increasing evidence suggests that exosomes play a crucial role in AML immune microenvironment. Methods Sanger sequencing, RNase R and fluorescence in situ hybridization were performed to confirm the existence of circ_0006896. The role of circ_0006896 in the progression of AML was assessed by in vitro and in vivo functional experiments. Flow cytometry, RT-qPCR and adoptive T cell-transfer immunotherapy were conducted to assess the function of exosomal circ_0006896 in CD8+ T cell dysfunction. RNA pull-down assay, mass spectrometry, immunofluorescence, co-immunoprecipitation and western blot were performed to identify and confirm the circ_0006896 interacting proteins. Results CircRNA expression patterns in exosomes differ significantly between AML and controls compared to lncRNAs or mRNAs. A new crucial exosomal circRNA, circ_0006896, is upregulated in both AML cells and exosomes and correlates with the prognosis and relapse of AML. In vitro and in vivo studies suggest that circ_0006896 significantly promotes AML cell proliferation, reduces chemotherapy sensitivity, and more importantly, impairs the efficacy of adoptive T cell-transfer immunotherapy. Mechanistically, circ_0006896 physically interacts with the catalytic domain of histone deacetylase HDAC1, decreasing histone H3 acetylation, and impairing the transcription of genes involved in arachidonic acid metabolism, ultimately inhibiting lipid peroxidation and ferroptosis in AML cells. Exosomal circ_0006896 disrupts CD8+ T cell function by interacting with HDAC1, impairing LEF1 transcription and subsequently decreasing the expression of cytotoxic molecules IFN-γ and Granzyme B. Conclusions We demonstrate a self-driven progression mediated by exosomal circRNAs and CD8+ T cells, highlighting the potential of targeting circRNAs in AML immunotherapy.

Background: The occult onset and rapid progression of hepatocellular carcinoma (HCC) lead to an unsatisfactory overall survival (OS) rate. Established prognostic predictive models based on tumor‐node‐metastasis staging and predictive factors do not report satisfactory predictive efficacy. Arachidonic acid plays pivotal roles in biological processes including inflammation, regeneration, immune modulation, and tumorigenesis. We, therefore, constructed a prognostic predictive model based on seven genes linked to arachidonic acid metabolism, using samples of HCC patients from databases to analyze the genomic profiles. We also assessed the predictive stability of the constructed model. Methods: Sample data of 365 patients diagnosed with HCC were extracted from The Cancer Genome Atlas (TCGA, training set) and HCCDB18, GSE14520, and GSE76427 databases (validation sets). Patient samples were clustered using ConsensusClusterPlus analysis based on the expression levels of 12 genes involved in arachidonic acid metabolism that were significantly associated with HCC prognosis. Differentially expressed genes (DEGs) within different clusters were distinguished and compared using WebGestaltR. Immunohistochemistry (IHC) analysis was performed using a human HCC tissue microarray (TMA). Tumor immune microenvironment assessment was performed using ESTIMATE, ssGSEA, and TIDE. Results: Samples of patients with HCC were classified into three clusters, with significant differences in OS. Cluster 2 showed the best prognosis, whereas cluster 1 presented the worst. The three clusters showed significant differences in immune infiltration. We then performed Cox and LASSO regression analyses, which revealed CYP2C9, G6PD, CDC20, SPP1, PON1, TRNP1, and ADH4 as prognosis‐related hub genes, making it a simplified prognostic model. TMA analysis for the seven target genes showed similar results of regression analyses. The high‐risk group showed a significantly worse prognosis and reduced immunotherapy efficacy. Our model showed stable prognostic predictive efficacy. Conclusions: This seven‐gene–based model showed stable outcomes in predicting HCC prognosis as well as responses to immunotherapy. [ABSTRACT FROM AUTHOR]

Introduction: Podophyllotoxin (PPT) is a high-content and high-activity compound extracted from the traditional Chinese medicinal plant Dysosma versipellis (DV) which exhibits various biological activities. However, its severe toxicity limits its use. In clinical settings, patients with DV poisoning often experience adverse reactions when taking large doses in a short period. The heart is an important toxic target organ, so it is necessary to conduct 24-h acute cardiac toxicity studies on PPT to understand its underlying toxicity mechanism. Methods: Based on the concept of the toxicological evidence chain (TEC), we utilized targeted metabolomic and transcriptomic analyses to reveal the mechanism of the acute cardiotoxicity of PPT. The manifestation of toxicity in Sprague-Dawley rats, including changes in weight and behavior, served as Injury Phenotype Evidence (IPE). To determine Adverse Outcomes Evidence (AOE), the hearts of the rats were evaluated through histopathological examination and by measuring myocardial enzyme and cardiac injury markers levels. Additionally, transcriptome analysis, metabolome analysis, myocardial enzymes, and cardiac injury markers were integrated to obtain Toxic Event Evidence (TEE) using correlation analysis. Results: The experiment showed significant epistaxis, hypokinesia, and hunched posture in PPT group rats within 24 h after exposure to 120 mg/kg PPT. It is found that PPT induced cardiac injury in rats within 24 h, as evidenced by increased serum myocardial enzyme levels, elevated concentrations of cardiac injury biomarkers, and altered cardiac cell morphology, all indicating some degree of cardiac toxicity. Transcriptome analysis revealed that primary altered metabolic pathway was arachidonic acid metabolism after PPT exposure. Cyp2e1, Aldob were positively correlated with differential metabolites, while DHA showed positive correlation with differential genes Fmo2 and Timd2, as well as with heart injury markers BNP and Mb. Conclusion: This study comprehensively evaluated cardiac toxicity of PPT and initially revealed the mechanism of PPT-induced acute cardiotoxicity, which involved oxidative stress, apoptosis, inflammatory response, and energy metabolism disorder. [ABSTRACT FROM AUTHOR]

Introduction: Podophyllotoxin (PPT) is a high-content and high-activity compound extracted from the traditional Chinese medicinal plant Dysosma versipellis (DV) which exhibits various biological activities. However, its severe toxicity limits its use. In clinical settings, patients with DV poisoning often experience adverse reactions when taking large doses in a short period. The heart is an important toxic target organ, so it is necessary to conduct 24-h acute cardiac toxicity studies on PPT to understand its underlying toxicity mechanism. Methods: Based on the concept of the toxicological evidence chain (TEC), we utilized targeted metabolomic and transcriptomic analyses to reveal the mechanism of the acute cardiotoxicity of PPT. The manifestation of toxicity in Sprague-Dawley rats, including changes in weight and behavior, served as Injury Phenotype Evidence (IPE). To determine Adverse Outcomes Evidence (AOE), the hearts of the rats were evaluated through histopathological examination and by measuring myocardial enzyme and cardiac injury markers levels. Additionally, transcriptome analysis, metabolome analysis, myocardial enzymes, and cardiac injury markers were integrated to obtain Toxic Event Evidence (TEE) using correlation analysis. Results: The experiment showed significant epistaxis, hypokinesia, and hunched posture in PPT group rats within 24 h after exposure to 120 mg/kg PPT. It is found that PPT induced cardiac injury in rats within 24 h, as evidenced by increased serum myocardial enzyme levels, elevated concentrations of cardiac injury biomarkers, and altered cardiac cell morphology, all indicating some degree of cardiac toxicity. Transcriptome analysis revealed that primary altered metabolic pathway was arachidonic acid metabolism after PPT exposure. Cyp2e1, Aldob were positively correlated with differential metabolites, while DHA showed positive correlation with differential genes Fmo2 and Timd2, as well as with heart injury markers BNP and Mb. Conclusion: This study comprehensively evaluated cardiac toxicity of PPT and initially revealed the mechanism of PPT-induced acute cardiotoxicity, which involved oxidative stress, apoptosis, inflammatory response, and energy metabolism disorder. [ABSTRACT FROM AUTHOR]

Background: Patients with non‐alcoholic fatty liver disease (NAFLD) often exhibit hepatic steatosis and dyslipidemia. Studies have shown that intestinal microorganisms are closely related to the occurrence of NAFLD and atherosclerosis. Our previous study has underscored the protective role of microRNA‐30a‐5p (miR‐30a‐5p) against atherosclerosis. Methods and Results: In the present study, we aimed to elucidate the effect and underlying mechanism of the intestinal microorganisms of miR‐30a‐5p knockout (KO) mice on NAFLD. Our findings demonstrated that KO exacerbated high‐fat diet (HFD)‐induced hepatic steatosis and disrupted liver function, as evidenced by elevated levels of total cholesterol, low‐density lipoprotein, alanine aminotransferase, aspartate transaminase, and total bile acids in serum. Fecal microbiota from HFD‐fed KO mice induced hepatic steatosis, dyslipidemia, and higher levels of enzymes indicative of liver damage in wild‐type mice. Remarkably, KO mice significantly intensified the above effects. 16s rDNA sequencing and metabolomics of the intestinal microbiota in the HFD‐treated KO and WT mice showed that the loss of miR‐30a‐5p resulted in intestinal microbiota imbalance and was highly related to the arachidonic acid metabolic pathway. Targeted metabolomic in the liver tissues unveiled upregulation of COX‐related (PGF2a, 8‐iso‐PGF2a and PGF2) and LOX‐related (LTB4, LTD4, 12S‐HETE and 15S‐HETE) factors in HFD‐treated KO mice. Immunohistochemistry and transcriptional analyses showed that miR‐30a‐5p affected arachidonic acid metabolism through the LOX/COX pathways. Besides, COX/LOX pathways and hepatic steatosis were reversed after reintroducing miR‐30a‐5p in HFD‐treated KO mice. Conclusions: This study reveals the pivotal mechanism by which miR‐30a‐5p and intestinal microbes regulate hepatic steatosis and abnormal lipid metabolism, offering promising avenues for NAFLD and atherosclerosis therapeutics. Highlights: MiR‐30a‐5p deletion aggravated hepatic steatosis and lipid disorder induced by an HFD in mice. Gut microbiota participated in the regulation of hepatic steatosis in the context of miR‐30a‐5p. Gut microbiota metabolism‐related arachidonic acid metabolic pathway contributed to miR‐30a‐5p‐regulated hepatic steatosis and lipid disorder. Reintroducing miR‐30a‐5p reversed hepatic steatosis and arachidonic acid metabolism disorder caused by HFD and miR‐30a‐5p deletion. [ABSTRACT FROM AUTHOR]

In vitro embryonic technology is crucial for improving farm animal reproduction but is hampered by the poor quality of oocytes and insufficient development potential. This study investigated the relationships among changes in the gut microbiota and metabolism, serum features, and the follicular fluid metabolome atlas. Correlation network maps were constructed to reveal how the metabolites affect follicular development by regulating gene expression in granulosa cells. The superovulation synchronization results showed that the number of follicle diameters from 4 to 8 mm, qualified oocyte number, cleavage, and blastocyst rates were improved in the dairy heifers (DH) compared with the non-lactating multiparous dairy cows (NDC) groups. The gut microbiota was decreased in Rikenellaceae_RC9_gut_group, Alistipes, and Bifidobacterium, but increased in Firmicutes, Cyanobacteria, Fibrobacterota, Desulfobacterota, and Verrucomicrobiota in the NDC group, which was highly associated with phospholipid-related metabolites of gut microbiota and serum. Metabolomic profiling of the gut microbiota, serum, and follicular fluid further demonstrated that the co-metabolites were phosphocholine and linoleic acid. Moreover, the expression of genes related to arachidonic acid metabolism in granulosa cells was significantly correlated with phosphocholine and linoleic acid. The results in granulosa cells showed that the levels of PLCB1 and COX2, participating in arachidonic acid metabolism, were increased in the DH group, which improved the concentrations of PGD2 and PGF2α in the follicular fluid. Finally, the expression levels of apoptosis-related proteins, cytokines, and steroidogenesis-related genes in granulosa cells and the concentrations of steroid hormones in follicular fluid were determinants of follicular development. According to our results, gut microbiota-related phosphocholine and linoleic acid participate in arachidonic acid metabolism in granulosa cells through the gut–follicle axis, which regulates follicular development. These findings hold promise for enhancing follicular development and optimizing oocyte quality in subfertile dairy cows. [ABSTRACT FROM AUTHOR]

Abstract Liver metastasis is a common cause of death in colorectal cancer (CRC) patients, but epigenetic remodeling and metabolic reprogramming for CRC liver metastasis remain unclear. The study revealed that the Lyn/RUVBL1 complex is highly expressed in CRC and is closely correlated with liver metastasis. On the one hand, ATAC‐seq and HiCut suggested that Lyn/RUVBL1 regulates the expression of TRIB3 through the POL II‐mediated chromatin conformation of TRIB3 and thus the expression of β‐catenin. This promotes the proliferation and migration of CRC through β‐catenin‐mediated upregulation of MMP9 and VEGF. On the other hand, metabolomics revealed that Lyn/RUVBL1 regulates the expression of PGE2 through the enzyme COX2, thereby promoting arachidonic acid (AA) metabolism. CUT‐Tag showed that Lyn/RUVBL1 silencing reduces the H3K27ac level in the COX2 promoter. Then, it is found that COX2 is regulated by the transcription factor FOXA1. Lyn/RUVBL1 modulates AA metabolism by regulating the chromatin accessibility of FOXA1. AA metabolism promotes the metastasis of CRC by affecting β‐catenin nuclear translocation and upregulating MMP9 and VEGF. These findings suggest that the Lyn/RUVBL1 complex mediates epigenetic remodeling to regulate the metabolic reprogramming of AA, highlighting its role in promoting the metastasis of CRC.

IntroductionStroke is a debilitating disease and the second leading cause of death worldwide, of which ischemic stroke is the dominant type. Carthamus tinctorius L., also known as safflower, has been used to treat cerebrovascular diseases, especially ischemic stroke in many Asian countries. However, the underlying mechanisms of safflower in preventing ischemic stroke remains elusive. This study aims to elucidate the potential of safflower as a drug candidate for the prevention of ischemic stroke and to delineate its protective effects and potential mechanisms in a rat model of cerebral ischemia-reperfusion injury (CI/RI).MethodsThe aqueous extract of safflower (AESF) was verified using HPLC-UV, HPLC-MS, and TLC. The inhibitory effect of AESF on platelet aggregation was detected in vitro and in zebrafish and mice. A CI/RI model in rats was established by middle cerebral artery occlusion and reperfusion to study the protective effect of AESF on ischemic stroke. 2,3,5-triphenyltetrazolium chloride, hematoxylin and eosin, and Nissl’s staining were employed to evaluate the pathological changes of brain tissue. In addition, metabolomics, ELISA, and Western blot were used to uncover the molecular alteration induced by AESF.ResultsAESF significantly inhibited platelet aggregation in vitro, reduced the thrombogenesis in zebrafish, and prolonged clotting time in mice. In addition, AESF alleviated neurological dysfunction, cerebral oedema, cerebral infarct size, cerebral histopathological damage induced by ischemia-reperfusion, improved neuronal survival, increased serum levels of SOD and CAT, and decreased levels of iNOS and NO. Metabolomics revealed that AESF attenuated the metabolic disturbances in brain caused by I/R injury via regulating 38 metabolites particularly related to the arachidonic acid (AA) metabolism. Moreover, AESF elevated the serum levels of 6-keto-PGF1α, a pivotal metabolite of AA, downregulated the protein expression of p53, Bax, cleaved caspase-9, cleaved caspase-3, and cleaved caspase-8, and upregulated that of Bcl-2.ConclusionAESF mitigated CI/RI through preventing platelet aggregation, alleviating oxidative stress, and suppressing apoptosis partially via modulating AA metabolism/p53-mediated apoptosis axis.

Abstract Background Mediators, genomic and epigenomic characteristics involving in metabolism of arachidonic acid by cyclooxygenase (COX) and lipoxygenase (ALOX) and hepatic activation of clopidogrel have been individually suggested as factors associated with resistance against aspirin and clopidogrel. The present multi-center prospective cohort study evaluated whether the mediators, genomic and epigenomic characteristics participating in arachidonic acid metabolism and clopidogrel activation could be factors that improve the prediction of the aspirin and clopidogrel resistance in addition to cardiovascular risks. Methods We enrolled 988 patients with transient ischemic attack and ischemic stroke who were evaluated for a recurrence of ischemic stroke to confirm clinical resistance, and measured aspirin (ARU) and P2Y12 reaction units (PRU) using VerifyNow to assess laboratory resistance 12 weeks after aspirin and clopidogrel administration. We investigated whether mediators, genotypes, and promoter methylation of genes involved in COX and ALOX metabolisms and clopidogrel activation could synergistically improve the prediction of ischemic stroke recurrence and the ARU and PRU levels by integrating to the established cardiovascular risk factors. Results The logistic model to predict the recurrence used thromboxane A synthase 1 (TXAS1, rs41708) A/A genotype and ALOX12 promoter methylation as independent variables, and, improved sensitivity of recurrence prediction from 3.4% before to 13.8% after adding the mediators, genomic and epigenomic variables to the cardiovascular risks. The linear model we used to predict the ARU level included leukotriene B4, COX2 (rs20417) C/G and thromboxane A2 receptor (rs1131882) A/A genotypes with the addition of COX1 and ALOX15 promoter methylations as variables. The linear PRU prediction model included G/A and prostaglandin I receptor (rs4987262) G/A genotypes, COX2 and TXAS1 promoter methylation, as well as cytochrome P450 2C19*2 (rs4244285) A/A, G/A, and *3 (rs4986893) A/A genotypes as variables. The linear models for predicting ARU (r = 0.291, R2 = 0.033, p

Background: Mediators, genomic and epigenomic characteristics involving in metabolism of arachidonic acid by cyclooxygenase (COX) and lipoxygenase (ALOX) and hepatic activation of clopidogrel have been individually suggested as factors associated with resistance against aspirin and clopidogrel. The present multi-center prospective cohort study evaluated whether the mediators, genomic and epigenomic characteristics participating in arachidonic acid metabolism and clopidogrel activation could be factors that improve the prediction of the aspirin and clopidogrel resistance in addition to cardiovascular risks. Methods: We enrolled 988 patients with transient ischemic attack and ischemic stroke who were evaluated for a recurrence of ischemic stroke to confirm clinical resistance, and measured aspirin (ARU) and P2Y12 reaction units (PRU) using VerifyNow to assess laboratory resistance 12 weeks after aspirin and clopidogrel administration. We investigated whether mediators, genotypes, and promoter methylation of genes involved in COX and ALOX metabolisms and clopidogrel activation could synergistically improve the prediction of ischemic stroke recurrence and the ARU and PRU levels by integrating to the established cardiovascular risk factors. Results: The logistic model to predict the recurrence used thromboxane A synthase 1 (TXAS1, rs41708) A/A genotype and ALOX12 promoter methylation as independent variables, and, improved sensitivity of recurrence prediction from 3.4% before to 13.8% after adding the mediators, genomic and epigenomic variables to the cardiovascular risks. The linear model we used to predict the ARU level included leukotriene B4, COX2 (rs20417) C/G and thromboxane A2 receptor (rs1131882) A/A genotypes with the addition of COX1 and ALOX15 promoter methylations as variables. The linear PRU prediction model included G/A and prostaglandin I receptor (rs4987262) G/A genotypes, COX2 and TXAS1 promoter methylation, as well as cytochrome P450 2C19*2 (rs4244285) A/A, G/A, and *3 (rs4986893) A/A genotypes as variables. The linear models for predicting ARU (r = 0.291, R2 = 0.033, p < 0.01) and PRU (r = 0.503, R2 = 0.210, p < 0.001) levels had improved prediction performance after adding the genomic and epigenomic variables to the cardiovascular risks. Conclusions: This study demonstrates that different mediators, genomic and epigenomic characteristics of arachidonic acid metabolism and clopidogrel activation synergistically improved the prediction of the aspirin and clopidogrel resistance together with the cardiovascular risk factors. Trial registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT03823274. [ABSTRACT FROM AUTHOR]

An imbalance of energy intake and expenditure is commonly considered as the fundamental cause of obesity. However, individual variations in susceptibility to obesity do indeed exist in both humans and animals, even among those with the same living environments and dietary intakes. To further explore the potential influencing factors of these individual variations, male C57BL/6J mice were used for the development of obesity-prone and obesity-resistant mice models and were fed high-fat diets for 16 weeks. Compared to the obesity-prone mice, the obesity-resistant group showed a lower body weight, liver weight, adipose accumulation and pro-inflammatory cytokine levels. 16S rRNA sequencing, which was conducted for fecal microbiota analysis, found that the fecal microbiome's structural composition and biodiversity had changed in the two groups. The genera Allobaculumbiota, SMB53, Desulfovibrio and Clostridium increased in the obesity-prone mice, and the genera Streptococcus, Odoribacter and Leuconostoc were enriched in the obesity-resistant mice. Using widely targeted metabolomics analysis, 166 differential metabolites were found, especially those products involved in arachidonic acid (AA) metabolism, which were significantly reduced in the obesity-resistant mice. Moreover, KEGG pathway analysis exhibited that AA metabolism was the most enriched pathway. Significantly altered bacteria and obesity-related parameters, as well as AA metabolites, exhibited strong correlations. Overall, the phenotypes of the obesity-prone and obesity-resistant mice were linked to gut microbiota and AA metabolism, providing new insight for developing an in-depth understanding of the driving force of obesity resistance and a scientific reference for the targeted prevention and treatment of obesity. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate the core components and mechanism of action of Rosa roxburghii fruit flavonoids in the intervention of ulcerative colitis (UC) using network pharmacology and molecular docking. We extracted flavonoids from R. roxburghii fruit by an ethanol-cellulase method and used liquid chromatography-mass spectrometry (LC-MS) to identify the major chemical components of the flavonoid extract, and then we selected 34 active flavonoid components and 146 targets in the intervention of UC on an analytical platform and constructed a “R. roxburghii flavonoids-active components-intersecting targets-UC” network. The results of gene ontology pathway analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed that the intervention of R. roxburghii flavonoids in UC mainly involved the arachidonic acid signaling pathway, the nuclear factor-kappa B signaling pathway, and cancer pathways. Molecular docking was performed on the major active ingredients and the key targets selected from the network, with docking energies all less than −6.0 kcal/mol, indicating a strong binding affinity between the core active ingredients and the key targets for UC treatment. So, the network analysis results are reliable. The results of animal experiments showed that intervention with R. roxburghii flavonoids could effectively alleviate the body mass loss and colon pathological changes of UC mice, and significantly inhibit the expression of inflammatory cytokines such as tumour necrosis factor-α, interleukin (IL)-1β and IL-6 in the UC model group (P < 0.05). The results of Western blot showed that R. roxburghii flavonoids could effectively inhibit the expression of the key target proteins associated with the metabolic pathway of arachidonic acid, such as cyclooxygenase-2 and 5-lipoxygenase. In conclusion, R. roxburghii flavonoids can intervene in UC through a multi-component, multi-target and multi-pathway mechanism. The results of this research can provide a theoretical basis for the development and utilization of functional foods from R. roxburghii.

Abstract Background Inflammatory bowel disease (IBD), comprising Crohn's disease (CD) and ulcerative colitis (UC), is a heterogeneous state of chronic intestinal inflammation. Intestinal innate immunity, including innate immune cells, defends against pathogens and excessive entry of gut microbiota, while preserving immune tolerance to resident intestinal microbiota, and may be characterized by its capacity to produce a rapid and nonspecific reaction. The association between microbiota dysbiosis and the pathogenesis of IBD is complex and dynamic. When the intestinal ecosystem is in dysbiosis, the reduced abundance and diversity of intestinal gut microbiota make the host more vulnerable to the attack of exogenous and endogenous pathogenic gut microbiota. The aim of our study was to comprehensively assess the relationship between microbial populations within UC, the signaling pathways of pathogenic gut microbe therein and the inflammatory response, as well as to understand the effects of using PE&AFWE (poppy extract [Papaver nudicaule L.] and Artemisia frigida Willd. extract) on UC modulation. Methods A UC mouse model was established by inducing SPF‐grade C57BL/6 mice using dextrose sodium sulfate (DSS). Based on metagenomic sequencing to characterize the gut microbiome, the relationship between gut microbiota dysbiosis and gut microbiota was further studied using random forest and Bayesian network analysis methods, as well as histopathological analysis. Results (1) We found that the 5 gut microbiota with the highest relative abundance of inflammatory bowel disease UC model gut microbiota were consistent with the top 5 ranked natural bacteria. There were three types of abundance changes in the model groups: increases (Chlamydiae/Proteobacteria and Deferribacteres), decreases (Firmicutes), and no significant changes (Bacteroidetes). The UC model group was significantly different from the control group, with 1308 differentially expressed species with abundance changes greater than or equal to 2‐fold. (2) The proportion of the fecal flora in the UC group decreased by 37.5% in the Firmicutes and increased by 14.29% in the proportion of Proteobacteria compared to the control group before treatment. (3) The significantly enriched and increased signaling pathways screened were the ‘arachidonic acid metabolic pathway’ and the ‘phagosomal pathway’, which both showed a decreasing trend after drug administration. (4) Based on the causal relationship between different OTUs and the UC model/PE&AFWE administration, screening for directly relevant OTU networks, the UC group was found to directly affect OTU69, followed by a cascade of effects on OTU12, OTU121, OTU93, and OTU7, which may be the pathway of action that initiated the pathological changes in normal mice. (5) We identified a causal relationship between common differentially expressed OTUs and PE&AFWE and UC in the pre‐ and post‐PE&AFWE‐treated groups. Thereby, we learned that PE&AFWE can directly affect OTU90, after which it inhibits UC, inhibiting the activity of arachidonic acid metabolic pathway by affecting OTU118, which in turn inhibits the colonization of gut microbiota by OTU93 and OTU7. (6) Histopathological observation and scoring (HS) of the colon showed that there was a significant difference between the model group and the control group (p

Background and Aim: Xuefu Zhuyu decoction (XZD), a traditional Chinese medicinal formula, was firstly recorded in the Qing dynasty of ancient China and previously demonstrated to ameliorate hepatic steatosis. In the present study, the effects of XZD on non-alcoholic fatty liver disease (NAFLD) induced by high-fat diet (HFD) were evaluated in mice and the hepatic transcriptome was detected to disclose the potential mechanisms of XZD. Experimental procedure: The effects of XZD (low- and high-dosage) on NAFLD induced by HFD for 16 weeks were evaluated. Obeticholic acid was used as control drug. Body weight, food intake and index of homeostatic model assessment for insulin resistance (HOMA-IR) were analyzed. Hepatic histology were observed in haematoxylin and eosin stained sections and quantified with NAFLD activity score (NAS). Lipid in hepatocytes was visualized by Oil red staining. Alanine aminotransferase (ALT) and hepatic triglyceride (TG) was measured. The hepatic transcriptom was detected with RNA-sequencing and validated with real-time polymerase chain reaction, western-blotting and hepatic quantitative metabolomics. Results: XZD ameliorated hepatic histology of NAFLD mice, accompanied with decreasing fasting insulin, HOMA-IR, NAS, ALT and hepatic TG. The hepatic transcriptom of NAFLD was significantly reversed by XZD treatment, especially the genes enriched in the pathways of arachidonic acid metabolism, fatty acid degradation, cytokine-cytokine receptor interaction and extracellular matrix (ECM) -receptor interaction. The hepatic quantitative metabolomics analysis confirmed fatty acid degradation as the key targeting pathway of XZD. Conclusions: XZD ameliorated NAFLD induced by HFD, which probably correlated closely to the pathways of fatty acid degradation.

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Introduction: The disruption of podocyte structure and function are the main pathological mechanism of membranous nephropathy (MN). Phospholipases A2, Group XII B (PLA2G12B) was reported involved in the regulation of MN by interfering with arachidonic acid (AA) metabolism, but there is a lack of sufficient evidence. In this study, we investigated the role and molecular mechanism of PLA2G12B in MN. Methods: C57BL/6 mice were used to establish MN model to extract primary podocytes, then divided into control, model, si-phospholipases A2 receptor (PLA2R), PLA2G12B, PLA2G12B + si-PLA2R, PLA2G12B + nuclear factor kappa-B (NF-κB) inhibitor, PLA2G12B + NF-κB inhibitor + si-PLA2R groups. Hematoxylin-eosin staining and immunofluorescence were used to detect kidney histological arrangement, serum levels of cholesterol-related indices, and AA. Genes and proteins associated with metabolism and inflammatory factors were detected by quantitative real-time PCR and Western blot. Results: The results revealed that AA metabolites were activated in the MN model mice, and the expression of PLA2G12B and NF-κB pathway levels were elevated. Besides, cellular experiments demonstrated that prostaglandin I2 (PGI2), thromboxane A2 (TXA2), leukotriene B4 (LTB4), and NF-κB pathway were significantly increased in the PLA2G12B group. Also, PLA2G12B promotes apoptosis and suppresses cell activity in podocytes, and these effects could be antagonized by NF-κB inhibitors. Furthermore, with the inference of si-PLA2R, the NF-κB inhibitors’ effects were reversed. Conclusion: Promotional effects of PLA2G12B in primary MN are associated with the regulation of AA metabolism and NF-κB pathway.

This study aimed to investigate the metabolites of conjugated linoleic acid (CLA) in vivo from the perspective of metabolomics, so as to reveal the potential mechanism of CLA in lowing blood lipids. 30 male non-adult SD rats were divided into: normal diet group, high-fat diet group, and CLA group (intragastric administration of 0.5 g/kg·BW CLA). Results indicate improvements in body weight and visceral fat deposition in obese rats treated with CLA. Metabolomic analysis reveals significant changes in the arachidonic acid (ARA) pathway and its intestinal metabolite oxylipins. Key biomarkers (PGE1, PGE2, TXB2, PGF2α, and 12S-HHTrE) catalyzed by Cox and Lox are identified, influencing blood lipid reduction. Network analysis links these biomarkers to Pparγ. PCR and western blot confirm Pparγ changes in rat liver and intestine, impacting downstream lipid metabolism. This study reveals a potential mechanism: CLA mitigates obesity through the ARA-Cox/Lox-PGE2-Pparγ pathway, offering insights for treating obesity and dyslipidemia.

Inhibition of cholesterol de novo synthesis (DNS) by statins has controversial effects on the treatment of hepatocellular carcinoma (HCC). High fatty acid conditions have been reported to limit the effect of statins on metabolism diseases. Whether high fatty acid conditions interfere with the effect of statins on HCC remains unclear. Here, we reported that inhibiting cholesterol DNS with atorvastatin promoted the oncogenic capabilities of diethylnitrosamine (DEN) in mice fed high fatty acid diets (HFD). The combined analysis of metabolomics and transcriptomics revealed that arachidonic acid (AA) metabolism was the most significant changed pathway between mice with and without atorvastatin treatment. In vitro, in the presence of AA precursor linoleic acid (LA), atorvastatin promoted the proliferation and migration ability of HCC cell lines. However, in the absence of LA, these phenomena disappeared. TCGA and tissue microarray examination revealed that prostaglandin e synthase 2 (PTGES2), a key enzyme in AA metabolism, was associated with the poor outcome of HCC patients. Overexpression of PTGES2 promoted the proliferation and migration of HCC cell lines, and knockdown of PTGES2 inhibited the proliferation and migration of cells. Additionally, atorvastatin upregulated PTGES2 expression by enhancing Sterol‐regulatory element binding protein 2 (SREBP2)‐mediated transcription. Knockdown of PTGES2 reversed the proliferation and migration ability enhanced by atorvastatin. Overall, our study reveals that a high fatty acid background is one of the possible conditions limiting the application of statins in HCC, under which statins promote the progression of HCC by enhancing SREBP2‐mediated PTGES2 transcription. [ABSTRACT FROM AUTHOR]

Arachidonic acid metabolism plays a crucial role in the development and progression of inflammatory and metabolic liver diseases. However, its role in hepatocellular carcinoma (HCC) remains unclear. In this study, we investigated the expression of key genes involved in the arachidonic acid metabolism pathway in HCC using a combination of bioinformatics, proteomics and immunohistochemistry analyses. Through a comprehensive analysis of publicly available datasets, clinical HCC tissues, and tissue microarrays, we compared the expression of hepatic arachidonic acid metabolic genes. We observed significant downregulation of cytochrome P450 (CYP450) pathway genes at both the messenger RNA and protein levels in HCC tissues compared to normal liver tissues. Furthermore, we observed a strong correlation between the deregulation of the arachidonic acid metabolism CYP450 pathway and the pathological features and prognosis of HCC. Specifically, the expression of CYP2C8/9/18/19 was significantly correlated with pathological grade (r = −.484, p <.0001), vascular invasion (r = −.402, p <.0001), aspartate transaminase (r = −.246, p =.025), gamma‐glutamyl transpeptidase (r = −.252, p =.022), alkaline phosphatase (r = −.342, p =.002), alpha‐fetoprotein (r = −.311, p =.004) and carbohydrate antigen 19‐9 (r = −.227, p =.047). Moreover, we discovered a significant association between CYP450 pathway activity and vascular invasion in HCC. Collectively, these data indicate that arachidonic acid CYP450 metabolic pathway deregulation is implicated in HCC progression and may be a potential predictive factor for early recurrence in patients with HCC. [ABSTRACT FROM AUTHOR]

Background: Xinsuning capsules is a safe and effective drug against phlegm-heat-disturbed arrhythmia (a type of rapid arrhythmia in traditional Chinese medicine. The clinical manifestations include chest tightness, irritability, vomiting, insomnia, and dreaminess) with no noticeable adverse effects. However, the molecular biological basis of Xinsuning capsules in the treatment of arrhythmia is relatively unexplored. Methods: In order to assess the antiarrhythmic effects of Xinsuning capsules, a rat arrhythmia model was constructed by injecting barium chloride into the tail vein. Metabolomic analysis was performed by UHPLC/Q-Exactive-Orbitrap mass spectrometry. The selection of differential variables in the metabolic data were necessitated fold change = 1.5 or fold change = 0.67, together with P < 0.05. Results: Barium chloride-induced arrhythmias in rats were significantly delayed in beginning and cut short in duration, according to electrocardiogram monitoring. Left ventricles of rats exhibitied significantly higher Na+-K+-ATPase activity after taking Xinsuning capsules compared to the model group. Metabolomic analysis showed that Xinsuning capsules could regulate arachidonic acid, prostaglandin F2a, 15-HETE, 15-HPETE, 12-HETE, 12(13)-DHOME, LTB4, inosine, and hypoxanthine. These metabolomics mainly involved arachidonic acid metabolism, unsaturated fat acid biosynthenosis, and purine metabolism. Conclusion: The therapeutic effects of Xinsuning capsules against arrhythmia had been investigated with metabolomics, providing the basis for further comprehensive research. [ABSTRACT FROM AUTHOR]

Ulcerative colitis (UC), often known as UC, is an inflammatory disease of the intestines that has frequent and long-lasting flare-ups. It is unknown precisely how the traditional Chinese drug Indigo Naturalis (IN) heals inflammatory bowel disease, despite its long-standing use in China and Japan. Finding new metabolite biomarkers linked to UC could improve our understanding of the disease, speed up the diagnostic process, and provide insight into how certain drugs work to treat the condition. Our work is designed to use a metabolomic method to analyze potential alterations in endogenous substances and their impact on metabolic pathways in a mouse model of UC. To determine which biomarkers and metabolisms are more frequently connected with IN's effects on UC, liquid chromatography-tandem mass spectrometry analysis of the serum metabolomics of UC mice and normal mice was performed. The outcomes demonstrated that IN boosted the health of UC mice and reduced the severity of their metabolic dysfunction. In the UC model, it was also found that IN changed the way 17 biomarkers and 3 metabolisms functioned. [ABSTRACT FROM AUTHOR]

Fluoride could cause developmental neurotoxicity and significantly affect the intelligence quotient (IQ) of children. However, the systematic mechanism of neuronal damage caused by excessive fluoride administration in offspring is largely unknown. Here, we present a comprehensive integrative transcriptome and metabolome analysis to study the mechanism of developmental neurotoxicity caused by chronic fluoride exposure. Comparing the different doses of fluoride treatments in two generations revealed the exclusive signature of metabolism pathways and gene expression profiles. In particular, neuronal development and synaptic ion transport are significantly altered at the gene expression and metabolite accumulation levels for both generations, which could act as messengers and enhancers of fluoride-induced systemic neuronal injury. Choline and arachidonic acid metabolism, which highlighted in the integrative analysis, exhibited different regulatory patterns between the two generations, particularly for synaptic vesicle formation and inflammatory factor transport. It may suggest that choline and arachidonic acid metabolism play important roles in developmental neurotoxic responses for offspring mice. Our study provides comprehensive insights into the metabolomic and transcriptomic regulation of fluoride stress responses in the mechanistic explanation of fluoride-induced developmental neurotoxicity.

Colorectal cancer (CRC) is a significant health concern and is the third most commonly diagnosed and second deadliest cancer worldwide. CRC has been steadily increasing in developing countries owing to factors such as aging and epidemics. Despite extensive research, the exact pathogenesis of CRC remains unclear, and its causes are complex and variable. Numerous in vitro, animal, and clinical trials have demonstrated the efficacy of probiotics such as Lactobacillus plantarum in reversing the adverse outcomes of CRC. These findings suggest that probiotics play vital roles in the prevention, adjuvant treatment, and prognosis of CRC. In this study, we constructed a mouse model of CRC using an intraperitoneal injection of azomethane combined with dextran sodium sulfate, while administering 5-fluorouracil as well as high- and low-doses of L. plantarum Zhang-LL live or heat-killed strains. Weight changes and disease activity indices were recorded during feeding, and the number of polyps and colon length were measured after euthanasia. HE staining was used to observe the histopathological changes in the colons of mice, and ELISA was used to detect the expression levels of IL-1β, TNF-α, and IFN-γ in serum. To investigate the specific mechanisms involved in alleviating CRC progression, gut microbial alterations were investigated using 16S rRNA amplicon sequencing and non-targeted metabolomics, and changes in genes related to CRC were assessed using eukaryotic transcriptomics. The results showed that both viable and heat-killed strains of L. plantarum Zhang-LL in high doses significantly inhibited tumorigenesis, colon shortening, adverse inflammatory reactions, intestinal tissue damage, and pro-inflammatory factor expression upregulation. Specifically, in the gut microbiota, the abundance of the dominant flora Acutalibacter muris and Lactobacillus johnsonii was regulated, PGE2 expression was significantly reduced, the arachidonic acid metabolism pathway was inhibited, and CD22-mediated B-cell receptor regulation-related gene expression was upregulated. This study showed that L. plantarum Zhang-LL live or heat-inactivated strains alleviated CRC progression by reducing the abundance of potentially pathogenic bacteria, increasing the abundance of beneficial commensal bacteria, mediating the arachidonic acid metabolism pathway, and improving host immunogenicity. [ABSTRACT FROM AUTHOR]

Background & Aims: PIEZO1 and TRPV4 are mechanically and osmotically regulated calcium‐permeable channels. The aim of this study was to determine the relevance and relationship of these channels in the contractile tone of the hepatic portal vein, which experiences mechanical and osmotic variations as it delivers blood to the liver from the intestines, gallbladder, pancreas and spleen. Methods: Wall tension was measured in freshly dissected portal veins from adult male mice, which were genetically unmodified or modified for either a non‐disruptive tag in native PIEZO1 or endothelial‐specific PIEZO1 deletion. Pharmacological agents were used to activate or inhibit PIEZO1, TRPV4 and associated pathways, including Yoda1 and Yoda2 for PIEZO1 and GSK1016790A for TRPV4 agonism, respectively. Results: PIEZO1 activation leads to nitric oxide synthase‐ and endothelium‐dependent relaxation of the portal vein. TRPV4 activation causes contraction, which is also endothelium‐dependent but independent of nitric oxide synthase. The TRPV4‐mediated contraction is suppressed by inhibitors of phospholipase A2 and cyclooxygenases and mimicked by prostaglandin E2, suggesting mediation by arachidonic acid metabolism. TRPV4 antagonism inhibits the effect of agonising TRPV4 but not PIEZO1. Increased wall stretch and hypo‐osmolality inhibit TRPV4 responses while lacking effects on or amplifying PIEZO1 responses. Conclusions: The portal vein contains independently functioning PIEZO1 channels and TRPV4 channels in the endothelium, the pharmacological activation of which leads to opposing effects of vessel relaxation (PIEZO1) and contraction (TRPV4). In mechanical and osmotic strain, the PIEZO1 mechanism dominates. Modulators of these channels could present important new opportunities for manipulating liver perfusion and regeneration in disease and surgical procedures. [ABSTRACT FROM AUTHOR]

Context The bulb of Lilium brownii F. E. Brown (Liliaceae) (LB) is a common Chinese medicine to relieve insomnia. Objective To investigate the molecular mechanism of LB relieving insomnia. Materials and methods Insomnia model was induced by intraperitoneally injection p-chlorophenylalanine (PCPA) in Wistar rats. Rats were divided into three groups: Control, PCPA (400 mg/kg, i.p. 2 days), LB (598.64 mg/kg, oral 7 days). The levels of 5-hydroxytryptamine (5-HT), norepinephrine (NE), melatonin (MT), and the expression of GABAA, 5-HT1A and MT receptors, as well as pathological changes in hypothalamus, were evaluated. 16S rDNA sequencing and UPLC-MS/MS were used to reveal the change of the intestinal flora and metabolic profile. Results The adverse changes in the abundance and diversity of intestinal flora and faecal metabolic phenotype altered by PCPA in rats were reversed after LB treatment, accompanied by the up-regulated levels of 5-HT as 8.14 ng/mL, MT as 16.16 pg/mL, 5-HT1A R and GABAA R, down-regulated level of NE as 0.47 ng/mL, and the improvement of pathological phenomena of cells in the hypothalamus. And the arachidonic acid metabolism and tryptophan metabolism pathway most significantly altered by PCPA were markedly regulated by LB. Besides, it was also found that LB reduced the levels of kynurenic acid related to psychiatric disorders and trimethylamine-N-oxide associated with cardiovascular disease. Conclusion The mechanism of LB relieving insomnia involves regulating flora and metabolites to resemble the control group. As a medicinal and edible herb, LB could be considered for development as a health-care food to relieve increasing insomniacs in the future.

Abstract Background As the second cause of cancer death in women, breast cancer has become a worldwide priority. Previous studies based on tumour cell lines demonstrated that arachidonic acid (AA) and its metabolites promote cancer development. However, recent studies based on the tumour microenvironment revealed the antitumour effect of AA metabolism. Therefore, it is essential to reevaluate and elucidate the effect of AA metabolism on breast cancer. Methods Raw data were obtained from The Cancer Genome Atlas (TCGA), Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) and Gene Expression Omnibus (GEO) databases. The AA metabolic score of each sample, enrichment of differentially expressed genes (DEGs) and immune infiltration were analysed by bioinformatics. Cox regression and least absolute shrinkage and selection operator regression were performed to establish an AA metabolism prognostic signature. An AA metabolism related nomogram for predicting the survival probability of patients was built. Result AA metabolism was related to good prognosis in the TCGA-BRCA and METABRIC cohort. DEGs enrichment suggested that the upregulated DEGs of the high AA metabolism group were significantly enriched in immune-related pathways. The high AA metabolism group was infiltrated with more CD8+ T cells and activated NK cells. An AA metabolic signature (SPINK8, KLRB1, APOD and PIGR) was constructed for breast cancer prognosis. Conclusion The study indicated that a high level of AA metabolism may be a biomarker for good prognosis in breast cancer, providing a possible explanation for the discouraging effect of cyclooxygenase inhibitors in cancer therapy. Moreover, a novel AA metabolic prognostic signature was constructed in the study, providing a novel strategy for breast cancer.

The aim of this study was to investigate the potential protective effects of walnut oligopeptides (WOPs) on indomethacin-induced gastric ulcers in rats. The rats were divided into the following groups: normal group, model group, omeprazole group (0.02 g/kg), and WOPs groups (0.22, 0.44, and 0.88 g/kg, respectively). After receiving gavage once per day for 30 consecutive days, the rats were injected intraperitoneally with indomethacin 48 mg/kg to induce gastric ulcers. Then, the serum inflammatory cytokines and gastric prostaglandin E2 (PGE2), oxidative stress-related indicators, and the RNA expression of COX-1 and COX-2 were measured. The results revealed that WOPs confer significant gastroprotection on gastric ulcers caused by indomethacin, regulating inflammatory cytokines, oxidative stress, and prostaglandins synthesis, and enhancing the expression of COX-1 and COX-2 in gastric tissue, thus exerting its protective effect on gastric mucosa. The gastroprotective mechanism may be related to the involvement of the arachidonic acid metabolism and upregulation of tryptophan, phenylalanine, tyrosine, and alpha-Linolenic acid metabolism synthesis in vivo. [ABSTRACT FROM AUTHOR]

Cellular response to inflammatory stimuli leads to the production of eicosanoids—prostanoids (PRs) and leukotrienes (LTs)—and signaling molecules—cytokines and chemokines—by macrophages. Quantitative modeling of the inflammatory response is challenging owing to a lack of knowledge of the complex regulatory processes involved. Cybernetic models address these challenges by utilizing a well-defined cybernetic goal and optimizing a coarse-grained model toward this goal. We developed a cybernetic model to study arachidonic acid (AA) metabolism, which included two branches, PRs and LTs. We utilized a priori biological knowledge to define the branch-specific cybernetic goals for PR and LT branches as the maximization of TNFα and CCL2, respectively. We estimated the model parameters by fitting data from three experimental conditions. With these parameters, we were able to capture a novel fourth independent experimental condition as part of the model validation. The cybernetic model enhanced our understanding of enzyme dynamics by predicting their profiles. The success of the model implies that the cell regulates the synthesis and activity of the associated enzymes, through cybernetic control variables, to accomplish the chosen biological goal. The results indicated that the dominant metabolites are PGD2 (a PR) and LTB4 (an LT), aligning with their corresponding known prominent biological roles during inflammation. Using heuristic arguments, we also infer that eicosanoid overproduction can lead to increased secretion of cytokines/chemokines. This novel model integrates mechanistic knowledge, known biological understanding of signaling pathways, and data-driven methods to study the dynamics of eicosanoid metabolism. [ABSTRACT FROM AUTHOR]

A critical aspect affecting the quality of life in Traumatic spinal cord injury (TSCI) patients is bladder dysfunction. Metabolities in arachidonic acid are crucial lipid signaling molecules involved innumerous physiological processes. In this study, We are the first use eicosanoid metabolomics detrusor contraction examine, to assess the effect of the arachidonic acid metabolic in bladder dysfunction following TSCI. In additon, we explore the time of inflammatory and function changes in bladder tissue. Adult male Sprague-Dawley rats were subjected to improved Weight Drop method surgeries. Detrusor contraction examination, urodynamic examination, eicosanoid metabolomics, transmission electron microscopy, Elisa and histological staining were performed to assess the change of inflammatory, metabolic and function variation over time after TSCI. Following TSCI, before the variations of bladder function, inflammatory changes including the increase of inflammatory factors, mitochondrial damage, and slight lipid peroxidation, occurred in bladder tissue. And the inflammatory changes gradually decreases over time. However, From the third day after TSCI, secondary lesions appeared in bladder tissue. Not only did inflammation-related indexes increase again, the degree of mitochondrial damage and lipid peroxidation increased, but also the contractility of detrusor began to change significantly. We also found that the content of metabolites in arachidonic acid metabolic pathway and the degree of detrusor contractility change showed a strong correlation. In addition, we found that rats had moved beyond the spinal shock stage on the seventh day after TSCI. Altogether, we are the first to demonstrate that abnormal arachidonic acid metabolism plays an important role in bladder dysfunction after TSCI. We also demonstrate that 3d is a critical juncture for changes in rat bladder tissue, which indicates it is an important juncture in the treatment of neurogenic bladder. • The following content is the focus and innovation of the manuscript. • The study focuses on the changes of detrusor itself rather than nerve after SCI. • The study discussed the correlation between metabolic changes and functional changes. • The study explored the pathological changes of detrusor at different time after SCI. [ABSTRACT FROM AUTHOR]

• Aqueous extract Sargentodoxa cuneata modulated overall serum metabolites in UC. • Multiple metabolic pathways were associated with AESc against UC. • Metabolites, like cholylleucine, birabresib, and lucidumol A, with anti-UC potential. Symptoms of ulcerative colitis (UC) are like "intestinal carbuncle" in Chinese medicine. The aqueous extract of Sargentodoxa cuneata (AESc) has good therapeutic effects on UC, but the underlying mechanism needs to be further elucidated. The mechanism of AESc against UC was studied based on metabolomics and bioinformatics in mice with UC. Dextran sodium sulfate was applied to induce a mouse model of UC. After the intervention of AESc, the general condition of the animals was recorded, and efficacy-related indicators were measured. Information on serum metabolites was determined. Multivariate analysis combined with bioinformatics methods were used to identify the differential metabolites. Furthermore, "metabolite-target-disease" network was obtained, and differential metabolites of UC were screened, and further analysis of the metabolites were performed. Molecular docking validation was also carried out. AESc improved general conditions such as blood in stool, hair of animals, and weight loss, reduced disease activity index scores and shortening of colon length in mice with UC. A total of 3445 serum metabolites were obtained, and 64 differentiated metabolites of AESc against UC were screened. Enrichment analysis showed that arachidonic acid metabolism, bile secretion, drug metabolism-other enzymes, and tyrosine metabolism were associated with AESc in the treatment of UC. In addition, based on "metabolite-target-disease" network, the serum metabolites cholylleucine, 9,10,13-TriHOME, birabresib, anthramycin methyl ether, trans -hexadec-2-enoyl carnitine, and lucidumol A were found to have the therapeutic potential for UC. Further, 14 core targets were obtained, and lipids and atherosclerosis, rheumatoid arthritis and multiple immune-inflammatory pathways were associated with AESc for the treatment of UC. AESc corrects serum metabolic disturbances in UC mice, and multiple serum metabolites have therapeutic potential for UC. AESc may treat UC by regulating biological processes such as lipid metabolism, amino acid metabolism, thereby restoring normal physiological function of the intestine. [ABSTRACT FROM AUTHOR]

As a functional fatty acid, Arachidonic acid (ARA) plays an important role in immunity due to its metabolites in mammals, which have been extensively studied in mammals. However, studies on the function of metabolites produced by dietary ARA in aquatic animals are not comprehensive and in-depth. Hence, an eight-week feeding trial was conducted to evaluated the regulatory effects of dietary ARA and its metabolites-eicosanoids on inflammation, oxidative stress and apoptosis in the liver of Acanthopagrus schlegelii with an initial weight of 0.99 ± 0.10 g. Six isonitrogenous and isolipidic (41% protein, 14% lipid) diets were formulated with gradient levels of ARA: 0% (ARA0.10), 0.5% (ARA0.59), 1.0% (ARA1.04), 1.5% (ARA1.42), 2.0% (ARA1.94), 2.5% (ARA2.42), respectively, and palmitic acid was used to regulate lipid levels. Results showed that the key enzyme contents of ARA metabolic pathway such as phospholipase A2 (PLA2), cyclooxygenase-2 (COX-2), lipoxygenase (LOX) cytochrome P450 (CYP450), prostaglandin E2 (PGE2), leukotriene C4 (LTC4) and 20-hydroxyeicosatetraenoic acid (20-HETE) in the liver were increased with dietary ARA levels increased. High levels of dietary ARA (1.94% and 2.42%) activated the gene expression levels of nuclear factor kappa B (nf-κb), which were confirmed by the results of immunofluorescence assay. Likewise, the expression levels of pro-inflammatory cytokines (tnf-α and il-1β) were up-regulated by ARA1.94 and ARA2.42 groups. Fish fed with excess ARA level has significantly low antioxidant capacity by decreasing the activity of related antioxidant parameters and down-regulating the expression level of related genes. The pro-apoptotic related genes (bax and caspase 7/9) were markedly up-regulated as dietary ARA levels increased from 1.04% to 2.42%. Collectively, excess dietary ARA levels (1.92% and 2.42%) produced excess eicosanoids, triggering inflammation via activating NF-κB expression and promoting its nuclear translocation, resulting in oxidative stress by reducing antioxidant parameters (SOD, GSH and GPX), inducing cell apoptosis through up-regulating pro-apoptotic gene expression levels.

Introduction: The disruption of podocyte structure and function are the main pathological mechanism of membranous nephropathy (MN). Phospholipases A2, Group XII B (PLA2G12B) was reported involved in the regulation of MN by interfering with arachidonic acid (AA) metabolism, but there is a lack of sufficient evidence. In this study, we investigated the role and molecular mechanism of PLA2G12B in MN. Methods: C57BL/6 mice were used to establish MN model to extract primary podocytes, then divided into control, model, si-phospholipases A2 receptor (PLA2R), PLA2G12B, PLA2G12B + si-PLA2R, PLA2G12B + nuclear factor kappa-B (NF-κB) inhibitor, PLA2G12B + NF-κB inhibitor + si-PLA2R groups. Hematoxylin-eosin staining and immunofluorescence were used to detect kidney histological arrangement, serum levels of cholesterol-related indices, and AA. Genes and proteins associated with metabolism and inflammatory factors were detected by quantitative real-time PCR and Western blot. Results: The results revealed that AA metabolites were activated in the MN model mice, and the expression of PLA2G12B and NF-κB pathway levels were elevated. Besides, cellular experiments demonstrated that prostaglandin I2 (PGI2), thromboxane A2 (TXA2), leukotriene B4 (LTB4), and NF-κB pathway were significantly increased in the PLA2G12B group. Also, PLA2G12B promotes apoptosis and suppresses cell activity in podocytes, and these effects could be antagonized by NF-κB inhibitors. Furthermore, with the inference of si-PLA2R, the NF-κB inhibitors' effects were reversed. Conclusion: Promotional effects of PLA2G12B in primary MN are associated with the regulation of AA metabolism and NF-κB pathway. [ABSTRACT FROM AUTHOR]

Pancreatic ductal adenocarcinoma (PDAC) is highly heterogeneous, making its prognosis prediction difficult. The arachidonic acid (AA) cascade is involved in carcinogenesis. Therefore, the metabolic enzymes of the AA cascade consist of lipoxygenases (LOXs), phospholipase A2s (PLA2s), and cyclooxygenases (COXs) along with their metabolic products, including leukotrienes. Nevertheless, the prognostic potential of AA metabolism-associated PDAC has not been explored. Herein, the mRNA expression patterns and the matching clinical information of individuals with PDAC were abstracted from online data resources. We employed the LASSO Cox regression model to develop a multigene clinical signature in the TCGA queue. The GEO queue and the ICGC queue were employed as the validation queue. There was differential expression of a significant number of AA metabolism-associated genes (56.8%) between PDAC and neighboring nonmalignant tissues in the TCGA queue. Univariate Cox regression demonstrated that 13 of the differentially expressed genes (DEGs) were linked to overall survival (OS) (p < 0.05). A 6-gene clinical signature was developed for stratifying the PDAC patients into two risk groups, with the high-risk group patients exhibiting remarkably lower OS than the low-risk group patients (p < 0.001 in the TCGA data set and the ICGC queue, and p = 0.001 in the GEO data set). The multivariate Cox data revealed the risk score as an independent OS predictor (HR > 1, p < 0.01). The receiver operating characteristic (ROC) curve verified the predictive potential of our signature. The expression and alteration of the six genes in PDAC were also validated using online databases. Functional analyses demonstrated that immune-linked cascades were enriched, and the immune status was remarkably different between the high- and low-risk groups. In summary, an AA metabolism-associated clinical gene signature can be applied for prognostic estimation in PDAC. [ABSTRACT FROM AUTHOR]

Background: Prostanoids are a family of lipid mediators formed from arachidonic acid by cyclooxygenase enzymes and serve as biomarkers of vascular function. Prostanoid production may be different in males and females indicating that different therapeutic approaches may be required during disease. Objectives: We examined sex-dependent differences in COX-related metabolites in genetically modified mice that produce a cyclooxygenase-2 (COX2) enzyme containing a tyrosine 385 to phenylalanine (Y385F) mutation. This mutation renders the COX2 enzyme unable to form a key intermediate radical required for complete arachidonic acid metabolism and provides a model of selective COX2 inhibition. Design and Methods: Mice heterozygous for the Y385F mutation in COX2 were mated to produce cohorts of wild-type, heterozygous, and COX2 mutant mice. We investigated whether the genotype distribution followed Mendelian genetics and studied whether sex-specific differences could be found in certain prostanoid levels measured in peritoneal macrophages and in urinary samples. Results: The inheritance of the COX2 mutation displayed a significant deviation with respect to Mendel's laws of genetics, with a lower-than-expected progeny of weaned COX2 mutant pups. In macrophages, prostaglandin E2 (PGE2) production following lipopolysaccharide (LPS) and interferon gamma (IFNγ) stimulation was COX2-dependent in both males and females, and data indicated that crosstalk between the nitric oxide (NO) and COX2 pathways may be sex specific. We observed significant differences in urinary PGE2 production by male and female COX2 mutant mice, with the loss of COX2 activity in male mice decreasing their ability to produce urinary PGE2. Finally, female mice across all 3 genotypes produced similar levels of urinary thromboxane (measured as 11-dehydro TxB2) at significantly higher levels than males, indicating a sex-related difference that is likely COX1-derived. Conclusions: Our findings clearly demonstrate that sex-related differences in COX-derived metabolites can be observed, and that other pathways (such as the NO pathway) are affected. [ABSTRACT FROM AUTHOR]

Background: Type 2 diabetes mellitus (T2DM) exhibits a bidirectional relationship with periodontitis, wherein each condition influences the progression of the other. Arachidonic acid (AA) exerts an anti-diabetic effect, while showing a protective effect by regulating the inflammatory response independently of its metabolites. However, its impact on periodontitis with T2DM remains poorly understood., Methods: The T2DM mouse model was established by combining a high-sugar and high-fat diet with streptozotocin injection, followed by silk ligation to induce periodontitis. Alterations in diabetes-associated symptoms were evaluated. Micro-computed tomography was used to measure bone-related parameters, including the distance from the cementoenamel junction to the alveolar bone crest, bone volume/total volume and bone mineral density. Targeted metabolomics analysis was conducted to evaluate the impact of exogenous AA on serum metabolite levels of AA in mice with type 2 diabetic periodontitis. 16S rRNA gene sequencing was utilized to analyze the microbial diversity. The activity of osteoclasts, levels of inflammatory factors and gene expression related to osteoclasts were investigated using TRAP staining and real-time quantitative PCR., Results: The periodontitis mouse model with T2DM was successfully established. Following two weeks of exogenous AA treatment, a reduction in fasting blood glucose levels was observed in the diabetic periodontitis mice. Exogenous AA alleviated alveolar bone loss in type 2 diabetic periodontitis mice. However, it had no substantial effect on the contents of serum AA-targeted metabolites. Exogenous AA reduced Staphylococcus in subgingival flora of type 2 diabetic periodontitis mice, but had no significant impact on microbial community structure or diversity. Furthermore, it decreased the number of osteoclasts in the alveolar bone of periodontitis with T2DM mice and increased IL-10 mRNA expression in its gingival tissue., Conclusion: Exogenous AA may alleviate alveolar bone loss in T2DM mice with periodontitis by reducing the number of osteoclasts and increasing the expression of IL-10 mRNA in periodontal tissues, rather than the change of AA-targeted metabolites in serum or the composition and diversity of microorganisms in subgingival plaque. These findings may provide a potential therapeutic approach for the prevention and treatment of periodontitis with T2DM., Competing Interests: Declarations. Ethics approval and consent to participate: All experimental procedures were approved by the Ethics Committee of Guangzhou TopBiotech Co., Ltd. Laboratory Animal Management and Use Committee (protocol code LFTOP-IACUC-2023-0019 and May 22, 2023 of approval). Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Renin plays a significant role in the regulation of blood pressure and fluid volume by modulating the renin‒angiotensin‒aldosterone (RAAS) system. Renin suppression reduces serum aldosterone levels and lowers blood pressure in addition to preserving renal function. However, exactly how renin synthesis and action are regulated and how renin suppression preserves renal function are not clear. We propose that arachidonic acid (AA) and its metabolites control renin synthesis, secretion, and action by virtue of its (AA) anti-inflammatory, cytoprotective actions and ability to regulate the secretion of renin. These findings suggest that direct renin suppression results in changes in AA metabolism. This proposal implies that AA and its metabolites may be developed as potential drugs to prevent and manage hypertension and preserve renal function., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Introduction: Sepsis-related acute liver injury involves complex immune dysfunctions. Epoxyeicosatrienoic acids (EETs), bioactive molecules derived from arachidonic acid (AA) via cytochrome P450 (CYP450) and rapidly hydrolyzed by soluble epoxide hydrolase (sEH), possess anti-inflammatory properties. Nevertheless, the impact of the sEH inhibitor TPPU on sepsis-related acute liver injury remains uncertain., Objectives: This study utilized comprehensive single-cell analysis to investigate the immunoregulatory mechanism of TPPU in alleviating sepsis-related acute liver injury., Methods: Hepatic bulk RNA sequencing and proteomics analyses were employed to investigate the mechanisms underlying sepsis-related acute liver injury induced by cecal ligation and puncture in mice. Cytometry by time-of-flight and single-cell RNA sequencing were conducted to thoroughly examine the immunoregulatory role of TPPU at single-cell resolution., Results: Downregulation of AA metabolism and the CYP450 pathway was observed during sepsis-related acute liver injury, and TPPU treatment reduced inflammatory cytokine production and mitigated sepsis-related hepatic inflammatory injury. Comprehensive single-cell analysis revealed that TPPU promotes the expansion of anti-inflammatory CD206 + CD73 + M2-like macrophages and PDL1 - CD39 - CCR2 + neutrophils, reprogramming liver neutrophils to an anti-inflammatory CAMP + NGP + CD177 + phenotype. Additionally, TPPU inhibits the CCL6-CCR1 signaling mediated by M2-like macrophages and CAMP + NGP + CD177 + neutrophils, altering intercellular communication within the septic liver immune microenvironment., Conclusion: This study demonstrated TPPU's protective efficacy against sepsis-related acute liver injury, underscoring its vital role in modulating liver macrophages and neutrophils and enhancing prospects for personalized immunomodulatory therapy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025. Published by Elsevier B.V.)

In response to organ injury in adults, macrophages often promote scarring, yet during early life, they are required for tissue regeneration. To elucidate the mechanisms underlying age-associated regeneration, we compared the macrophage injury response in newborn versus adult hearts. Single-cell analysis revealed an accumulation of tissue-resident macrophages in neonates that were selectively polarized for apoptotic cell recognition and uptake (efferocytosis). Ablation of the apoptotic cell recognition receptor Mertk in newborns prevented cardiac regeneration. These findings could be attributed to reprogramming of macrophage gene expression that was required for biosynthesis of the eicosanoid thromboxane A 2 , which unexpectedly activated parenchymal cell proliferation. Markers of thromboxane A 2 production were suppressed in adult macrophages after efferocytosis. Moreover, macrophage-neighboring neonatal cardiomyocytes expressed the thromboxane A 2 receptor, whose activation induced a metabolic shift that supported cellular proliferation. Our data reveal a fundamental age-defined macrophage response in which lipid mitogens produced during efferocytosis support receptor-mediated tissue regeneration., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: The impact of a high-salt (HS) diet on metabolic disturbances in individuals with coronary heart disease remains unclear. The arachidonic acid (AA) metabolic pathway is closely linked to the development of cardiometabolic diseases and atherosclerotic cardiovascular diseases. Furthermore, endoplasmic reticulum stress (ERS) has emerged as a major contributor to cardiometabolic diseases. AA-related inflammation and ERS are hypothesized to play a role in HS diet-induced coronary remodeling., Methods: Rats were subjected to an HS diet for 4 weeks, and the serum concentration of AA was measured via enzyme-linked immunosorbent assay. Immunofluorescence staining and vascular tension measurements were conducted on coronary arteries. In addition, AA-stimulated coronary artery smooth muscle cells (CASMCs) were treated with ERS inhibitors to explore the underlying pathway involved., Results: Increased susceptibility to myocardial infarction in the HS diet-fed rats was accompanied by increased serum AA concentrations and increased expression of the key AA metabolic enzyme cyclooxygenase-2 (COX-2). AA incubation weakened the contraction of denuded coronary arteries, reduced the expression of contraction markers, and increased the fluorescence intensity of synthetic and ERS response markers in coronary arteries. Further investigation of CASMCs revealed that AA-induced phenotypic transformation was mediated via the ERS pathway., Conclusions: ERS and AA were found to be stimulated in CASMCs following an HS diet. AA triggers an ERS response through COX-2 catalysis, and the downstream inositol requiring enzyme 1 - X-box binding protein-1 - osteopontin pathway may contribute to the AA-induced phenotypic transformation of CASMCs, resulting in dysfunctional coronary tension. This study may provide potential therapeutic targets for cardiovascular diseases associated with excessive AA-derived ERS., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

The success of arsenic trioxide (ATO) in acute promyelocytic leukemia has driven a plethora studies to investigate its efficacy in other malignancies. However, the inherent toxicity of ATO limits the expansion of its clinical applications. Such toxicity may be linked to ATO-induced metabolic derangements of endogenous substrates. Therefore, the primary objective of this study was to investigate the effect of ATO on the hepatic formation of arachidonic acid (AA) metabolites, hydroxyeicosatetraenoic acids (HETEs), as well as their most notable producing machinery, cytochrome P450 (CYP) enzymes. For this purpose, C57BL/6 mice were intraperitoneally injected with 8 mg/kg ATO for 6 and 24 h. Total RNA was extracted from harvested liver tissues for qPCR analysis of target genes. Hepatic microsomal proteins underwent incubation with AA, followed by identification/quantification of the produced HETEs. ATO downregulated Cyp2e1, while induced Cyp2j9 and most of Cyp4a and Cyp4f, and this has resulted in a significant increase in 17(S)-HETE and 18(R)-HETE, while significantly decreased 18(S)-HETE. Additionally, ATO induced Cyp4a10, Cyp4a14, Cyp4f13, Cyp4f16, and Cyp4f18, resulting in a significant elevation in 20-HETE formation. In conclusion, ATO altered hepatic AA metabolites formation through modulating the underlying network of CYP enzymes. Modifying the homeostatic production of bioactive AA metabolites, such as HETEs, may entail toxic events that can, at least partly, explain ATO-induced hepatotoxicity. Such modification can also compromise the overall body tolerability to ATO treatment in cancer patients., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Conflict of Interest: The authors declare that they have no conflict of interest., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

This study investigated the protective effects of the dietary polyphenol vanillic acid (VA) on dextran sulfate sodium-induced acute ulcerative colitis (UC) in mice, focusing on its impact on the gut microbiota and inflammatory responses. VA was supplemented following dextran sulfate sodium administration, and key indicators, including body weight, disease activity index, colon length, spleen index, and inflammatory markers, were assessed. VA supplementation significantly alleviated UC symptoms, preserved intestinal barrier integrity, and reduced pro-inflammatory cytokine levels. Additionally, VA positively altered the gut microbiota composition, promoting beneficial bacteria such as Akkermansia muciniphila while suppressing the arachidonic acid metabolism pathway. Fecal microbiota transplantation confirmed that the VA-modified gut microbiota contributed to these protective effects. VA also facilitated macrophage polarization from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype, further mitigating inflammation. These findings highlight the potential of VA as a natural dietary intervention for UC, emphasizing its role in regulating the gut microbiota and inflammatory pathways, which may have significant nutritional relevance in managing inflammatory bowel diseases., (© 2025 Wiley‐VCH GmbH.)

Gastrointestinal (GI) cancers are a major global health burden, representing 20% of all cancer diagnoses and 22.5% of global cancer-related deaths. Their aggressive nature and resistance to treatment pose a significant challenge, with late-stage survival rates below 15% at five years. Therefore, there is an urgent need to delve deeper into the mechanisms of gastrointestinal cancer progression and optimize treatment strategies. Increasing evidence highlights the active involvement of abnormal arachidonic acid (AA) metabolism in various cancers. AA is a fatty acid mainly metabolized into diverse bioactive compounds by three enzymes: cyclooxygenase, lipoxygenase, and cytochrome P450 enzymes. Abnormal AA metabolism and altered levels of its metabolites may play a pivotal role in the development of GI cancers. However, the underlying mechanisms remain unclear. This review highlights a unique perspective by focusing on the abnormal metabolism of AA and its involvement in GI cancers. We summarize the latest advancements in understanding AA metabolism in GI cancers, outlining changes in AA levels and their potential role in liver, colorectal, pancreatic, esophageal, gastric, and gallbladder cancers. Moreover, we also explore the potential of targeting abnormal AA metabolism for future therapies, considering the current need to explore AA metabolism in GI cancers and outlining promising avenues for further research. Ultimately, such investigations aim to improve treatment options for patients with GI cancers and pave the way for better cancer management in this area., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Ethical approval: Not applicable. Consent to participate: Not applicable. Consent to publication: Not applicable., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Reperfusion therapy inevitably leads to brain-blood barrier (BBB) disruption and promotes damage despite its benefits for acute ischaemic stroke (AIS). An effective brain cytoprotective treatment is still needed as an adjunct to reperfusion therapy. Here, we explore the potential benefits of therapeutic hypothermia (HT) in attenuating early BBB leakage and improving neurological outcomes. Mild HT was induced during the early and peri-recanalization stages in a mouse model of transient middle cerebral artery occlusion and reperfusion (tMCAO/R). The results showed that mild HT attenuated early BBB leakage in AIS, decreased the infarction volume, and improved functional outcomes. RNA sequencing data of the microvessels indicated that HT decreased the transcription of the actin polymerization-related pathway. We further discovered that HT attenuated the ROCK1/MLC pathway, leading to a decrease in the polymerization of G-actin to F-actin. Arachidonic acid (AA), a known structural ROCK agonist, partially counteracted the protective effects of HT in the tMCAO/R model. Our study highlights the importance of early vascular protection during reperfusion and provides a new strategy for attenuating early BBB leakage by HT treatment for ischaemic stroke., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.