Search

Background: Acute liver failure (ALF) is marked by a substantial generation of reactive oxygen species (ROS), which can induce both cellular senescence and a pronounced inflammatory response. Senescent cells secrete factors collectively termed the senescence-associated secretory phenotype (SASP), which exacerbate inflammation, while inflammation can reciprocally promote cellular senescence. Quercetin (Que), recognized for its ROS-scavenging capabilities, holds the potential for anti-inflammatory and anti-senescent effects. However, its extremely low aqueous solubility constrains its clinical efficacy in treating inflammation. Methods: We employed a simple and stable coordination method to synthesize ultra-small quercetin-Fe nanoparticles (QFN) by complexing quercetin with iron ions. The ROS-scavenging, anti-inflammatory, and anti-senescent effects of QFN were evaluated in vitro . A lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced ALF mice model was used to investigate the therapeutic effects of QFN in vivo , and transcriptomic analysis was conducted to elucidate the mechanisms underlying QFN-mediated hepatoprotection. Results: Our findings demonstrate that QFN possesses remarkable water solubility and highly efficient ROS-scavenging properties. In vitro , QFN effectively inhibits macrophage-mediated inflammation and mitigates hepatocyte senescence. In vivo , QFN significantly attenuates LPS/D-GalN-induced ALF by protecting against macrophage inflammation and cellular senescence, thereby disrupting the self-perpetuating cycle of inflammation and aging. Moreover, its potent ROS scavenging capacity not only suppresses cellular apoptosis but also facilitates liver regeneration. Transcriptomic analyses further reveal that QFN exerts its protective effects through the modulation of key pathways involved in cellular senescence and inflammation. Conclusions: In summary, our study characterizes QFN as a potent ROS-scavenging modulator that exhibits both anti-inflammatory and anti-senescent properties, effectively disrupting the detrimental feedback loop between inflammation and cellular senescence. QFN holds considerable potential as a therapeutic agent for the treatment of ALF and other pathologies associated with inflammation and aging., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Acute liver failure is a serious, life-threatening disease. Although the gut microbiota has been considered to play a role in liver failure, the extent to which it is involved in the pathogenesis of this disease has not been fully elucidated to date. Therefore, we here analyzed the importance of the presence of intestinal microbiota in the pathogenesis of acute liver injury, using D-galactosamine (D-GalN)/lipopolysaccharide (LPS)-treated mice, which is a widely used experimental model of acute liver injury. First, administration of the antibiotic polymyxin B markedly alleviated liver injury. Liver injury was also reduced in germ-free mice, leading to the conclusion that the presence of intestinal microbiota aggravates D-GalN/LPS-induced liver injury. The amount of bacteria and LPS transferred from the gut to the blood was not increased by D-GalN/LPS, suggesting that the worsening of liver injury was not simply owing to the entry of bacteria into the circulation. In conclusion, acute liver injury in polymyxin B-pretreated or germ-free mice was ameliorated by modulation of the gut microbiota. Modification of the gut microbiota using polymyxin B may hence have the potential to alleviate acute liver injury in human patients., (© 2024 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Acute liver failure (ALF) is a life-threatening clinical syndrome characterized by high-grade inflammation and multi-organ failure. Our previous study shows that targeting the M2 isoform of pyruvate kinase (PKM2) to inhibit macrophage inflammation may be a promising strategy for ALF treatment. however, the mechanism by which PKM2 regulates the inflammatory response is unclear. Here we demonstrate that PKM2 contributes to ALF by modulating NLRP3-mediated pyroptosis activation in liver macrophages. The specific knockout of PKM2 in myeloid cells reduces mortality and alleviates hepatic injury in D-galactosamine/LPS-induced ALF mice. Single-cell transcriptome analysis suggests that NLRP3 inflammasome activation of macrophages involves in ALF, knockout of PKM2 in macrophages reduces the expression of NLRP3, and activation of pyroptosis. Pharmacological inhibition of the PKM2 nuclear translocation, but not glycolytic activity, protects mice from ALF. Pharmacological and genetic inhibition of PKM2 attenuates NLRP3-mediated pyroptosis activation and consequently reduces the release of IL-1β and IL-18 by macrophages. Mechanistically, PKM2 translocates into the nucleus and combines with STAT3, enhancing its phosphorylation and recruitment to the NLRP3 promoter region, thereby increasing NLRP3 expression. This work defines PKM2 acts as an important nonmetabolic regulator of NLRP3 that modulates pyroptosis activation in macrophages and guides future therapeutic strategies development for ALF., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Background: Reactive oxygen species (ROS) from mitochondrial dysfunction are critical in triggering apoptosis and inflammation in acute liver failure (ALF). Quercetin (QUE), an antioxidant, is renowned for its therapeutic effects onliverdiseases. There are no studies on whether QUE regulates mitophagy level in hepatocytes to inhibit ALF., Objective: This study investigates QUE's protective effects on ALF and elucidates the mechanisms involved., Methods: The ALF and hepatocyte inflammatory injury model was established using LPS and D-Galn. To predict potential targets and mechanisms of QUE in ALF treatment, transcriptomics, network pharmacology, molecular docking techniques, and ChIP were employed. The expression level related to mitophagy, apoptosis, and signaling pathways were detected by CCK8, IHC, IF staining, TUNEL, RT-qPCR, TEM, Western blotting, ELISA, and flow cytometry., Results: Network pharmacology and transcriptomics revealed common targets between QUE and ALF. Enrichment analysis showed that the anti-ALF targets of QUE were significantly associated with mitochondria and NF-κB-related pathways. Subsequent experiments showed that QUE pretreatment significantly alleviated the loss of hepatocyte viability, enhanced mitochondrial membrane potential, activated mitophagy, and promoted the clearance of damaged mitochondria, thereby reducing ROS accumulation, significantly reducing cell apoptosis and inflammatory responses, reducing ALT and AST levels, and improving liver tissue pathology. Mechanistically, molecular docking, DARTS, and CETSA analyses confirmed that QUE directly binds to the PPARγ molecule, which reduced binding to IκB and significantly inhibit the NF-κB pathway to exert its protective effects., Conclusion: In short, our results provide the first evidence that QUE improves acute liver failure by promoting mitophagy through regulating the PPARγ/PGC-1α/NF-κB axis and inhibiting apoptosis and inflammatory responses mediated by mitochondrial dysfunction, which provides evidence for the potential of QUE in the treatment of ALF., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: As a heterogeneous and life-threatening disease, the pathogenesis of acute liver failure (ALF) is complex. Our previous study has shown that IDH1/MDH1 deacetylation promotes ALF by regulating NETosis (a novel mode of cell death). In this article, we explore the manners of IDH1/MDH1 deacetylation regulates NETosis., Methods: In vitro experiments, the formation of NETs was detected by immunofluorescence staining and Western blotting. LC3 fluorescence staining was used to detect autophagosome formation. To observe mitochondrial morphology, cells were stained by Mito-Tracker Red. Western blotting was used to detect the levels of autophagy protein and mitochondrial dynamin. In vivo experiments, the ALF model in mouse was established with LPS/D-gal, and the formation of NETs was detected by immunofluorescence staining and Western blotting. The autophagy levels were detected by Western blotting in liver samples., Results: In dHL-60 cells, Western blotting results showed that the expression of OPA1 was higher in the IDH1/MDH1 deacetylated group compared with the IDH1/MDH1 WT group. And histone deacetylase inhibitor 6 (HDAC6i, ACY1215) decreased the expression level of OPA1 in IDH1/MDH1 deacetylated group. IDH1/MDH1 deacetylation increased the expression levels of both LC3B-II and Beclin 1, while decreasing the expression level of P62. It was reversed by ACY1215. Combined with our previous experiments, IDH1/MDH1 deacetylation upregulated autophagy concomitant with the increased expression of the markers of NETs formation. In a mouse model of ALF, ACY1215 further decreased the expression levels of LC3B-II and Beclin 1, while increasing the expression level of P62 in IDH1/MDH1 deacetylated mice., Conclusions: IDH1/MDH1 deacetylation promoted NETosis by regulating autophagy and OPA1 in vitro. The regulation of neutrophil autophagy on NETosis during IDH1/MDH1 deacetylation might be masked in mice. ACY1215 might attenuate NETosis by regulating neutrophil autophagy, which alleviated ALF aggravated by IDH1/MDH1 deacetylation., 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 © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Acute liver failure (ALF) is a fulminant clinical syndrome that usually leads to multiple organ failure and high mortality. Macrophages play a crucial role in the initiation, development, and recovery of ALF. Targeting macrophages through immunotherapy holds significant promise as a therapeutic strategy. These cells exhibit remarkable plasticity, enabling them to differentiate into various subtypes based on changes in their surrounding microenvironment. M1-type macrophages are associated with a pro-inflammatory phenotype and primarily rely predominantly on glycolysis. In contrast, M2-type macrophages, which are characterized by anti-inflammatory phenotype, predominantly obtain their energy from oxidative phosphorylation (OXPHOS) and fatty acid oxidation (FAO). Shifting macrophage metabolism from glycolysis to OXPHOS inhibits M1 macrophage activation and promotes M2 macrophage activation, thereby exerting anti-inflammatory and reparative effects. This study elucidates the relationship between macrophage activation and glucose metabolism reprograming from an immunometabolism perspective. A comprehensive literature review revealed that several signaling pathways may regulate macrophage polarization through energy metabolism, including phosphatidyl-inositol 3-kinase/protein kinase B (PI3K/AKT), mammalian target of rapamycin (mTOR)/hypoxia-inducible factor 1 α (HIF-1 α ), nuclear factor- κ B (NF- κ B), and AMP-activated protein kinase (AMPK), which exhibit crosstalk with one another. Additionally, we systematically reviewed several traditional Chinese medicine (TCM) monomers that can modulate glucose metabolism reprograming and influence the polarization states of M1 and M2 macrophages. This review aimed to provide valuable insights that could contribute to the development of new therapies or drugs for ALF., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2024 Junli Zhang et al.)

Acute liver failure (ALF) is a serious global disease characterized by rapid onset and high mortality. Currently, the clinical treatment of ALF faces considerable hurdles due to limited medication options and the scarcity of liver transplants. Despite biomacromolecules such as hepatocyte growth factor (HGF) and glutathione (GSH) having been applied for ALF symptom relief in the clinic, they still face substantial challenges including poor stability, difficulty in acting on intracellular targets, and inadequate therapeutic outcome. In this work, by taking advantage of the innate targeting and regenerative capabilities of mesenchymal stem cells (MSCs), we harnessed MSC-derived exosomes as natural bioactive carriers for the simultaneous delivery of HGF and GSH, forming a fully bioactive nanodrug termed HG@Exo. Impressively, the HG@Exo demonstrated potent therapeutic effects against both carbon tetrachloride (CCl 4 )- and extreme hepatectomy-induced ALF through multiple mechanisms, including regulation of oxidative stress, reduction of inflammation, and promotion of hepatocyte regeneration, which were facilitated by its inflammation-targeting to damaged liver tissues. Furthermore, an FDA-approved near-infrared fluorescent dye, indocyanine green (ICG), has been incorporated into the exosomes (HGI@Exo) to endow them with real-time in vivo tracking capability, which showed favorable liver accumulation of the HGI@Exo in both CCl 4 - and surgery-induced ALF animal models, providing crucial insights into their biodistribution and therapeutic efficacy. Overall, the presented fully bioactive nanodrugs with targeting and theranostic abilities hold significant promise for potentiating the therapeutic efficacy of biomacromolecules for the improved treatment of ALF and other inflammatory diseases.

Acute liver failure (ALF) is a hepatology emergency with rapid hepatic destruction, multiple organ failures, and high mortality. Despite decades of research, established ALF has minimal therapeutic options. Here, we report that the small bioactive compound SCM-198 increases the survival of male ALF mice to 100%, even administered 24 hours after ALF establishment. We identify adiponectin receptor 2 (AdipoR2) as a selective target of SCM-198, with the AdipoR2 R335 residue being critical for the binding and signaling of SCM-198-AdipoR2 and AdipoR2 Y274 residue serving as a molecular switch for Ca 2+ influx. SCM-198-AdipoR2 binding causes Ca 2+ influx and elevates the phosphorylation levels of CaMKII and NOS3 in the AdipoR2-CaM-CaMKII-NOS3 complex identified in this study, rapidly inducing nitric oxide production for liver protection in murine ALF. SCM-198 also protects human ESC-derived liver organoids from APAP/TAA injuries. Thus, selectively targeting the AdipoR2-CaM-CaMKII-NOS3 axis by SCM-198 is a rapid-acting therapeutic strategy for advanced ALF., Competing Interests: Competing interests: The first author Rui Wang is the applicant and one of the inventors (Rui Wang, Yuguo Wang, Jienina Yu) of a patent (No. ZL202010091483.6) related to findings shown in Fig. 1. The patent has been authorized and covers the utilization of SCM-198 in preventing acute liver injury and promoting hepatic tissue repair and regeneration. This patent concerns only the first author, and the rest of the authors have no competing interests to declare., (© 2024. The Author(s).)

Acute liver failure results from severe hepatic injury and can lead to neurological dysfunction known as hepatic encephalopathy (HE). Thrombospondin-1 can contribute to HE by increasing cerebral edema and microglia activation in the azoxymethane (AOM) mouse model. CD47 is a receptor for TSP1 and can directly modulate inflammation in numerous disease states. However, the role of CD47 in the progression of HE is currently unknown. Therefore, the aim of this study was to assess the role of CD47 in liver and brain pathology in the AOM mouse model of HE. C57Bl/6 or CD47 knockout (CD47 -/- ) were administered AOM to induce acute liver failure and HE. Liver damage was evaluated by measuring serum transaminases and histological assessment. Neurological function was determined by evaluating the time taken to reach coma (loss of all reflexes), cerebral edema, and microglia activation. CD47 signaling, and downstream signaling pathways, were assessed by real-time PCR, western blotting, immunofluorescence, and immunohistochemistry. AOM-treated mice had increased expression of CD47 in the liver, cortex, hippocampus, and cerebellum when compared to vehicle-treated mice. CD47 -/- AOM-treated mice had reduced liver injury and apoptosis when compared to wildtype AOM-treated mice. A slower degree of neurological decline and less cerebral edema were observed in CD47 -/- mice compared to wildtype AOM-treated mice. This was associated with decreased microglia proliferation and increased SOD1 expression in CD47 -/- mice compared to wildtype AOM-treated mice. These findings support that CD47 signaling exacerbates AOM-induced acute liver failure and HE by inducing hepatic cell death, cerebral edema, and microglia activation., Competing Interests: Declarations. Ethics approval: This study was approved by the University of Texas IACUC committee. Consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background and Aim: Acute liver failure (ALF) is a fatal clinical syndrome of severe hepatic dysfunction. Chemokines promote liver diseases by recruiting and activating immune cells. We aimed to investigate the role of C-C chemokine ligand 25 (CCL25) in ALF., Methods: An ALF mouse model induced by D-galactosamine/lipopolysaccharide was evaluated through liver hematoxylin and eosin staining and serum transaminase and cytokine measurement. CCL25 expression in serum was analyzed by ELISA and in liver by immunohistochemical staining and western blot. C-C chemokine receptor 9 (CCR9)-expressing cells in the liver were identified by immunofluorescence staining. The effects of anti-CCL25 on ALF were evaluated in vivo. Cytokine expression and migration of CCL25-stimulated RAW264.7 macrophages were studied. We also investigated the role of anti-CCL25 and BMS-345541, an NF-κB signaling inhibitor, in vitro. NF-κB activation was assessed via western blot, and p65 nuclear translocation was detected using cellular immunofluorescence., Results: ALF mice showed severe histological damage and high serum levels of aminotransferase and inflammatory cytokines. Elevated CCL25 and NF-κB activation was observed in vivo. CCR9 was expressed on macrophages in ALF mouse liver. ALF was suppressed after anti-CCL25 treatment, with significant NF-κB inhibition. In vitro, CCL25 induced strong migration and cytokine release in RAW264.7 macrophages, which were eliminated by anti-CCL25 and BMS-345541. Furthermore, the NF-κB activation and p65 nuclear translocation induced by CCL25 were also inhibited by anti-CCL25 and BMS-345541., Conclusion: CCL25 contributes to ALF development by inducing macrophage-mediated inflammation via activation of the NF-κB signaling., (© 2024 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

Hepatocyte-derived liver progenitor-like cells (HepLPCs) exhibit a remarkable capacity to support liver function by detoxifying ammonia, promoting native liver regeneration, and suppressing inflammation, which leads to improvements in the recovery and survival of animals with acute liver failure (ALF). However, the mechanism through which HepLPCs promote liver regeneration is unclear. Here, we isolated HepLPC-derived extracellular vesicles (HepLPC-EVs) from conditioned media and performed microRNA sequencing analysis. Our results showed HepLPC-EVs promoted liver regeneration in mice with carbon tetrachloride or acetaminophen induced ALF. Cell cycle progression and proliferation of primary human hepatocytes were promoted after coculture with HepLPC-EVs. Exosomal miRNA sequencing confirmed that HepLPC-EVs were enriched with miR-183-5p, which played an essential role in ameliorating ALF. Mechanistically, HepLPC-derived exosomal miR-183-5p negatively regulated the expression of the target gene FoxO1, activated the Akt/GSK3β/β-catenin signaling pathway, and thereby promoted liver regeneration and restoration of normal liver function. These results indicate that during ALF, HepLPC-Exos mediate liver regeneration mainly through a paracrine exosome-dependent mechanism and these effects accelerate liver regeneration and lead to the restoration of normal liver function., Competing Interests: Declarations. This work was supported in part by Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China. H-D.Z. is a full-time employee of Shanghai Celliver Biotechnology Co. Ltd. H-X.Y. is the founder of Shanghai Celliver Biotechnology Co. Ltd., Shanghai, China and have an equity interest in Celliver Biotechnology Inc. All other authors declare that they have no competing interests. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Inflammation is critical in the development of acute liver failure (ALF). Peroxisome proliferator-activated receptor delta (PPARδ) regulates anti-inflammatory responses and is protective in several diseases such as obesity and cancer. However, the beneficial effects and underlying mechanisms of PPARδ agonist GW501516 in ALF remain unclear. This study investigated the molecular mechanisms underlying the anti-inflammatory effects of GW501516 in macrophages and assessed its protective potential against lipopolysaccharide (LPS)/galactosamine (GalN)-induced ALF. In vivo administration of GW501516 significantly reduced LPS/GalN-induced hepatotoxicity, as evidenced by lower mortality, decreased liver damage, and attenuated secretion of IL-1β, IL-6, and TNF-α. GW501516 treatment also decreased LPS-induced nitric oxide synthase 2 (NOS2) expression and nitric oxide (NO) production in RAW264.7 cells, an effect reversed by PPARδ siRNA. Additionally, GW501516 inhibited LPS-induced phosphorylation of p38 and c-Jun N-terminal kinase (JNK), suggesting that inactivation of these MAPKs contributes to its effects. The secretion of IL-6, TNF-α, and NF-κB DNA-binding activity were also suppressed by GW501516, while the nuclear translocation of the NF-κB p65 subunit was unaffected. In conclusion, our findings suggest that GW501516 exerts protective effects in ALF by inhibiting the production of inflammatory mediators. Therefore, GW501516 may act as a potential agent for developing anti-inflammatory therapies for ALF.

Acute liver failure (ALF) is a life-threatening disease, characterized by upregulated extracellular matrix deposition and inflammatory signalling, with no effective treatment options and targets. The present study was designed to investigate the preventive and therapeutic effects of berberine (BBR) and its underlying mechanism in thioacetamide (TAA)-induced ALF. Male SD rats were administered with TAA 300 mg/kg, i.p., thrice to induce ALF and pre- or post-treated with BBR. To decipher the effects of BBR LFT markers, histopathological analysis of key fibrotic and inflammatory proteins was performed. In addition, the levels of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α were assessed by ELISA. Our work showed TAA-induced ALF animals were associated with increased ALT, AST, bilirubin (LFT markers) and histopathological alterations with profuse infiltration of inflammatory cells in the liver tissue. Treatment with BBR has significantly inhibited LFT markers and histological alterations triggered by TAA. In addition, TAA animals demonstrated increased collagen accumulation and upregulated expression of TGF-β1, vimentin, and α-SMA compared to control. The excessive accumulation of collagen, TGF-β1, vimentin, and α-SMA were significantly modulated with BBR treatment. Further, the fluorescence intensity of ROS an activator of NLRP3 including the NLRP3 inflammasome, and its downstream signalling ASC, cleaved IL-1β, and other pro-inflammatory cytokines like TNF-α and IL-6 stimulated by TAA were attenuated by BBR treatment. The current work indicated that BBR significantly ameliorated TAA-induced ALF by inhibiting the extracellular matrix accumulation associated with the NLRP3/IL-1β signalling pathway and could be a viable therapeutic option to treat ALF and other fibroinflammatory diseases., 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 © 2024 Elsevier Inc. All rights reserved.)

Background and Aims: Acute liver failure (ALF) is a rare but life-threatening condition, and DILI, particularly acetaminophen toxicity, is the leading cause of ALF. Innate immune mechanisms further perpetuate liver injury, while the role of the adaptive immune system in DILI-related ALF is unclear., Approach and Results: We analyzed liver tissue from 2 independent patient cohorts with ALF and identified hepatic T cell infiltration as a prominent feature in human ALF. CD8 + T cells were characterized by zonation toward necrotic regions and an activated gene expression signature. In murine acetaminophen-induced liver injury, intravital microscopy revealed zonation of CD8 + but not CD4 + T cells at necrotic areas. Gene expression analysis exposed upregulated C-C chemokine receptor 7 (CCR7) and its ligand CCL21 in the liver as well as a broadly activated phenotype of hepatic CD8 + T cells. In 2 mouse models of ALF, Ccr7-/- mice had significantly aggravated early-phase liver damage. Functionally, CCR7 was not involved in the recruitment of CD8 + T cells, but regulated their activation profile potentially through egress to lymphatics. Ccr7-/- CD8 + T cells were characterized by elevated expression of activation, effector, and exhaustion profiles. Adoptive transfer revealed preferential homing of CCR7-deficient CD8 + T cells to the liver, and depletion of CD8 + T cells attenuated liver damage in mice., Conclusions: Our study demonstrates the involvement of the adaptive immune system in ALF in humans and mice. We identify the CCR7-CCL21 axis as an important regulatory pathway, providing downstream protection against T cell-mediated liver injury., (Copyright © 2024 American Association for the Study of Liver Diseases.)

Regulating the gut microbiota alleviates hepatic encephalopathy (HE). Whether it is imperative to withhold treatment for microbial imbalance after liver functional recovery remains unclear. The aim of this work was to elucidate the alterations in cognitive behavior, liver function, synaptic transmission, and brain metabolites in acute liver failure (ALF) mice before and after hepatic function recovery. Towards this end, thioacetamide was injected intraperitoneally to establish an ALF mouse model, which induced HE. Hierarchical clustering analysis indicated that while the liver functions normalized, cognitive dysfunction and intestinal dysbacteriosis occurred in the ALF mice 14 days after thioacetamide injection. In addition, fecal microbiota transplantation from the ALF mice with liver function recovery induced liver injury and cognitive impairment. Alterations in synaptic transmission were found in the ALF mice with liver function improvement, and the correlations between the gut bacteria and synaptic transmission in the cortex were significant. Finally, apparent alterations in the brain metabolic profiles of the ALF mice were detected after liver function improvement by performing 1 H nuclear magnetic resonance spectroscopy, suggesting a risk of HE. These results showed that intestinal dysbacteriosis in ALF mice with liver function recovery is sufficient to induce liver injury and cognitive impairment. This indicates that continuous care may be necessary for monitoring microbial imbalance even in patients with ALF-induced HE whose liver function has recovered significantly., Competing Interests: Disclosure Statement None declared., (Copyright © 2024 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Acute liver failure (ALF) is an acute liver disease with a high mortality rate in clinical practice, characterized histologically by extensive hepatocellular necrosis and massive neutrophil infiltration. However, the role of these abnormally infiltrating neutrophils during ALF development is unclear. Here, in an ALF mouse model, metabolites were identified that promote the formation of neutrophil extracellular traps (NETs) in the liver, subsequently influencing macrophage differentiation and disease progression. ALF occurs with abnormalities in hepatic and intestinal metabolites. Abnormal metabolites (LTD4 and glutathione) can directly, or indirectly via reactive oxygen species, promote NET formation of infiltrating neutrophils, which subsequently regulate macrophages in a pro-inflammatory M1-like state, inducing an amplification of the destructive effects of inflammation. Together, this study provides new insights into the role of NETs in the pathogenesis of ALF., (© 2024 Federation of European Biochemical Societies.)

Abstract: Background: Acute liver failure (ALF) is a severe clinical syndrome characterized by massive hepatocyte death in a short time due to viruses, drugs, alcohol, or other factors. Oxidative stress is an important pathogenic mechanism of ALF. LPS-induced internalization of toll-like receptor 4 (TLR4) and the subsequent activation of the toll/IL-1R domain-containing adaptor-inducing IFN-beta (TRIF) signaling pathway widely mediate inflammatory responses in a series of diseases. However, whether the TLR4-TRIF signaling pathway contributes to ALF by mediating oxidative stress processes remains unclear. Methods: An ALF mouse model was induced by lipopolysaccharide (LPS)/D-galactosamine (D-GalN). TLR4-TRIF systemic knockout mice and TLR4 conditional knockout mice were used to determine the role of the TLR4-TRIF signaling pathway in ALF. The effects of TLR4 or TRIF deficiency on oxidative stress were investigated. In addition, we examined the protective role of the clodronate liposomes (macrophage scavengers) and the antioxidant N-acetylcysteine (NAC) in ALF. Results: TLR4 or TRIF deficiency significantly alleviated LPS/D-GalN-induced lethality, hepatic dysfunction, and hepatic pathologic injury, which was dependent on myeloid-derived TLR4. Hence, macrophage clearance exhibits a similar protective effect. Mechanically, TLR4 or TRIF deficiency was observed to inhibit oxidative stress by increasing glutathione, while decreasing malondialdehyde, 8-hydroxy-2-deoxyguanosine, and γ-H2AX. Therefore, the pharmacologic antioxidant NAC exhibited significant hepato-protective effects. Conclusions: Targeting myeloid-derived TLR4-TRIF signaling pathway or antioxidant therapy may be a potential therapeutic direction to treat ALF., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 by the Shock Society.)

Liver injury and acute liver failure caused by an acetaminophen (APAP) overdose is a significant clinical problem in western countries. With the introduction of the mouse model of APAP hepatotoxicity in the 1970 s, fundamental mechanisms of cell death were discovered. This included the recognition that part of the APAP dose is metabolized by cytochrome P450 generating a reactive metabolite that is detoxified by glutathione. After the partial depletion of glutathione, the reactive metabolite will covalently bind to sulfhydryl groups of proteins, which is the initiating event of the toxicity. This insight led to the introduction of N-acetyl-L-cysteine, a glutathione precursor, as antidote against APAP overdose in the clinic. Despite substantial progress in our understanding of the pathomechanisms over the last decades viable new antidotes only emerged recently. This review will discuss the background, mechanisms of action, and the clinical prospects of the existing FDA-approved antidote N-acetylcysteine, of several new drug candidates under clinical development [4-methylpyrazole (fomepizole), calmangafodipir] and examples of additional therapeutic targets (Nrf2 activators) and regeneration promoting agents (thrombopoietin mimetics, adenosine A2B receptor agonists, Wharton's Jelly mesenchymal stem cells). Although there are clear limitations of certain therapeutic approaches, there is reason to be optimistic. The substantial progress in the understanding of the pathophysiology of APAP hepatotoxicity led to the consideration of several drugs for development as clinical antidotes against APAP overdose in recent years. Based on the currently available information, it is likely that this will result in additional drugs that could be used as adjunct treatment for N-acetylcysteine., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Hartmut Jaeschke, Steven Curry and Barry Rumack report a relationship with McNeil Consumer Health Inc that includes consulting or advisory, funding grants and travel reimbursement. Anup Ramachandran and Jephte Akakpo 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 © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Acute liver failure (ALF) is characterized by the rapidly progressive deterioration of hepatic function, which, without effective medical intervention, results in high mortality and morbidity. Here, using proteomic and transcriptomic analyses in murine ALF models, we found that the expression of multiple splicing factors was downregulated in ALF. Notably, we found that KH-type splicing regulatory protein (KHSRP) has a protective effect in ALF. Knockdown of KHSRP resulted in dramatic splicing defects, such as intron retention, and led to the exacerbation of liver injury in ALF. Moreover, we demonstrated that KHSRP directly interacts with splicing factor 3b subunit 1 (SF3B1) and enhances the binding of SF3B1 to the intronic branch sites, thereby promoting pre-mRNA splicing. Using splicing inhibitors, we found that Khsrp protects against ALF by regulating pre-mRNA splicing in vivo. Overall, our findings demonstrate that KHSRP is an important splicing activator and promotes the expression of genes associated with ALF progression by interacting with SF3B1; thus, KHSRP could be a possible target for therapeutic intervention in ALF., (© 2024. The Author(s).)

This study explores the role of SIRT2 in regulating autophagy and its interaction with AMPK in the context of acute liver failure (ALF). This study investigated the effects of SIRT2 and AMPK on autophagy in ALF mice and TAA-induced AML12 cells. The results revealed that the liver tissue in ALF model group had a lot of inflammatory cell infiltration and hepatocytes necrosis, which were reduced by SIRT2 inhibitor AGK2. In comparison to normal group, the level of SIRT2, P62, MDA, TOS in TAA group were significantly increased, which were decreased in AGK2 treatment. Compared with normal group, the expression of P-PRKAA1, Becilin1 and LC3B-II was decreased in TAA group. However, AGK2 enhanced the expression of P-PRKAA1, Becilin1 and LC3B-II in model group. Overexpression of SIRT2 in AML12 cell resulted in decreased P-PRKAA1, Becilin1 and LC3B-II level, enhanced the level of SIRT2, P62, MDA, TOS. Overexpression of PRKAA1 in AML12 cell resulted in decreased SIRT2, TOS and MDA level and triggered more autophagy. In conclusion, the data suggested the link between AMPK and SIRT2, and reveals the important role of AMPK and SIRT2 in autophagy on acute liver failure., (© 2024. The Author(s).)

Acute liver failure (ALF) is a critical condition that can lead to substantial liver dysfunction. It is characterized by complex clinical manifestations and rapid progression, presenting significant challenges in diagnosis and treatment. We investigated the protective effect of mefunidone (MFD), a novel antifibrosis pyridone agent, on ALF in mice, and explored its potential mechanism of action. MFD pretreatment can alleviate lipopolysaccharide (LPS) and d-galactosamine (D-GalN)-induced ALF, reduce hepatocyte apoptosis, and reduce inflammation and oxidative stress. Additionally, MFD alleviated LPS/D-GalN-stimulated reactive oxygen species (ROS) production and cell death in AML12 cells. RNA sequencing enrichment analysis showed that MFD significantly affected the Mitogen-Activated Protein Kinase (MAPK) pathway. In vivo and in vitro experiments showed that MFD inhibited MKK4 and JNK phosphorylation. JNK activation caused by MKK4 and JNK activators could eliminate the therapeutic effect of MFD on AML12. In addition, MFD pretreatment alleviated ConA-induced ALF, reduced inflammation and oxidative stress in mice, and reduced mouse mortality. These results suggest that MFD can potentially protect against ALF, partially by inhibiting the MKK4-JNK pathway, and is a promising new therapeutic drug for ALF., 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 © 2024 Elsevier Inc. All rights reserved.)

Acetaminophen overdose is a leading cause of acute liver failure (ALF), and effective treatment depends on early prediction of disease progression. ALF diagnosis currently requires blood collection 24-72 h after APAP ingestion, necessitating repeated tests and hospitalization. Here, we assessed earlier ALF diagnosis using positron emission tomography (PET) imaging of translocator proteins (TSPOs), which are involved in molecular transport, oxidative stress, apoptosis, and energy metabolism, with the radiotracer [ 18 F]GE180. We intraperitoneally administered propacetamol hydrochloride to male C57BL/6 mice to induce ALF. We performed in vivo PET/CT imaging 3 h later using the TSPO-specific radiotracer [ 18 F]GE180 and quantitatively analyzed the PET images by determining the averaged standardized uptake value (SUV av ) in the liver parenchyma. We assessed liver TSPO expression levels via real-time polymerase chain reaction, Western blotting, and immunohistochemistry. [ 18 F]GE180 PET imaging 3 h after propacetamol administration (1500 mg/kg) significantly increased liver SUV av compared to controls ( p = 0.001). Analyses showed a 10-fold and 4-fold increase in TSPO gene and protein expression, respectively, in the liver, 3 h after propacetamol induction compared to controls. [ 18 F]GE180 PET visualized and quantified propacetamol-induced ALF through TSPO overexpression. These findings highlight TSPO PET's potential as a non-invasive imaging biomarker for early-stage ALF.

The bioartificial liver (BAL) system can potentially rescue acute liver failure (ALF) patients by providing partial liver function until a suitable donor liver can be found or the native liver has self-regenerated. In this study, we established a suitable cryopreservation process for the development of an off-the-shelf BAL system. The viability of hepatocyte spheroids cryopreserved in liquid nitrogen was comparable to that of fresh primary hepatocyte spheroids. When hepatocyte spheroids were subjected to cryopreservation in a deep freezer, no statistically significant differences were observed in ammonia removal rate or urea secretion rate based on the cryopreservation period. However, the functional activity of the liver post-cryopreservation in a deep freezer was significantly lower than that observed following liquid nitrogen cryopreservation. Moreover, cryopreserving spheroid hydrogel beads in a deep freezer resulted in a significant decrease (approximately 30%) in both ammonia removal and urea secretion rates compared to the group cryopreserved in liquid nitrogen. The viabilities of spheroid hydrogel beads filled into the bioreactor of a BAL system were similar across all four groups. However, upon operating the BAL system for 24 h, the liver function activity was significantly higher in the group comprising hydrogel beads generated after thawing hepatocyte spheroids cryopreserved in liquid nitrogen. Consequently, the manufacturing of beads after the cryopreservation of hepatocyte spheroids is deemed the most suitable method, considering efficiency, economic feasibility, and liver function activity, for producing a BAL system., (© 2024. The Author(s).)

Acute liver failure (ALF) is a potentially fatal disorder characterized by extensive hepatocyte necrosis and rapid decline in liver function. Numerous factors, including oxidative stress, cell death, and inflammatory responses, are associated with its pathogenesis. Endotoxin tolerance (ET) refers to the phenomenon in which the body or cells exhibit low or no response to high-dose lipopolysaccharide (LPS) stimulation after pre-stimulation with low-dose LPS. However, the specific mechanism through which ET regulates LPS/D-galactosamine (D-GalN)-induced ALF remains unclear. An ALF mouse model was established by intraperitoneal injection of D-GalN (400 mg/kg) and LPS (10 mg/kg). A low dose of LPS (0.1 mg/kg/d) was continuously administered to mice for 5 d before modeling to assess the protective effect of ET. The data from this study showed that ET alleviated the inflammatory response in mice with LPS/D-GalN-induced ALF. ET inhibited LPS-induced oxidative damage and pyroptosis in macrophages in vitro. RNA sequencing analysis showed that the NF-κB/NLRP3 pathway was linked to the anti-inflammatory and antioxidative effects of ET. Furthermore, using western blot, RT-qPCR, and immunofluorescence, we verified that ET inhibited the NF-κB/NLRP3 pathway and triggered the Nrf2/HO-1 signaling pathway to attenuate oxidative stress and cell pyroptosis. Sirt1 knockdown reversed this protective effect. In summary, our research elucidates that ET prevents ALF advancement by upregulating Sirt1 levels, triggering the Nrf2/HO-1 signaling axis, and suppressing the NF-κB/NLRP3 signaling cascade to inhibit oxidative stress and cell pyroptosis. Our results provide a mechanistic explanation for the protective effect of ET against ALF., 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 © 2024 Elsevier B.V. All rights reserved.)

Acute liver failure (ALF) is a complex syndrome characterized by overactivation of innate immunity, and the recruitment and differentiation of immune cells at inflammatory sites. The present study aimed to explore the role of microRNA (miRNA/miR)‑21 and its potential mechanisms underlying inflammatory responses in ALF. Baseline serum miR‑21 was analyzed in patients with ALF and healthy controls. In addition, miR‑21 antagomir was injected via the tail vein into C57BL/6 mice, and lipopolysaccharide/D‑galactosamine (LPS/GalN) was injected into mice after 48 h. The expression levels of miR‑21, Krüppel‑like‑factor‑6 (KLF6), autophagy‑related proteins and interleukin (IL)‑23, and hepatic pathology were then assessed in the liver tissue. Furthermore, THP‑1‑derived macrophages were transfected with a miRNA negative control, miR‑21 inhibitor, miR‑21 mimics or KLF6 overexpression plasmid, followed by treatment with or without rapamycin, and the expression levels of miR‑21, KLF6, autophagy‑related proteins and IL‑23 were evaluated. The results revealed that baseline serum miR‑21 levels were significantly upregulated in patients with ALF. In addition, LPS/GalN‑induced ALF was attenuated in the antagomir‑21 mouse group. KLF6 was identified as a target of miR‑21‑5p with one putative seed match site identified by TargetScan. A subsequent luciferase activity assay demonstrated a direct interaction between miR‑21‑5p and the 3'‑UTR of KLF6 mRNA. Further experiments suggested that miR‑21 promoted the expression of IL‑23 via inhibiting KLF6, which regulated autophagy. In conclusion, in the present study, baseline serum miR‑21 levels were highly upregulated in patients with ALF, antagomir‑21 attenuated LPS/GalN‑induced ALF in a mouse model, and miR‑21 could promote the expression of IL‑23 via inhibiting KLF6.

Acute liver failure (ALF) is a deadly illness due to insufficient detoxification in liver induced by drugs, toxins, and other etiologies, and the effective treatment for ALF is very limited. Among the drug-induced ALF, acetaminophen (APAP) overdose is the most common cause. However, the molecular mechanisms underlying APAP hepatoxicity remain incompletely understood. Sirtuin 6 (Sirt6) is a stress responsive protein deacetylase and plays an important role in regulation of DNA repair, genomic stability, oxidative stress, and inflammation. Here, we report that genetic and pharmacological activation of Sirt6 protects against ALF in mice. We first observed that Sirt6 expression was significantly reduced in the liver tissues of human patients with ALF and mice treated with an overdose of APAP. Then we developed an inducible Sirt6 transgenic mice for Cre-mediated overexpression of the human Sirt6 gene in systemic (Sirt6-Tg) and hepatic-specific (Sirt6-HepTg) manners. Both Sirt6-Tg mice and Sirt6-HepTg mice exhibited the significant protection against APAP hepatoxicity. In contrast, hepatic-specific Sirt6 knockout mice exaggerated APAP-induced liver damages. Mechanistically, Sirt6 attenuated APAP-induced hepatocyte necrosis and apoptosis through downregulation of oxidative stress, inflammation, the stress-activated kinase JNK activation, and apoptotic caspase activation. Moreover, Sirt6 negatively modulated the level and activity of poly (ADP-ribose) polymerase 1 (PARP1) in APAP-treated mouse liver tissues. Importantly, the specific Sirt6 activator MDL-800 exhibited better therapeutic potential for APAP hepatoxicity than the current drug acetylcysteine. Furthermore, in the model of bile duct ligation induced ALF, hepatic Sirt6-KO exacerbated, but Sirt6-HepTg mitigated liver damage. Collectively, our results demonstrate that Sirt6 protects against ALF and suggest that targeting Sirt6 activation could be a new therapeutic strategy to alleviate ALF., (© 2024. The Author(s).)

Aim: In 2019, to examine the functions of METTL3 in liver and underlying mechanisms, we generated mice with hepatocyte-specific METTL3 homozygous knockout (METTL3Δhep) by simultaneously crossing METTL3fl/fl mice with Alb-iCre mice (GPT) or Alb-Cre mice (JAX), respectively. In this study, we explored the potential reasons why hepatocyte-specific METTL3 homozygous disruption by Alb-iCre mice (GPT), but not by Alb-Cre mice (JAX), resulted in acute liver failure (ALF) and then postnatal lethality., Main Methods: Mice with hepatocyte-specific METTL3 knockout were generated by simultaneously crossing METTL3fl/fl mice with Alb-iCre mice (GPT; Strain No. T003814) purchased from the GemPharmatech Co., Ltd., (Nanjing, China) or with Alb-Cre mice (JAX; Strain No. 003574) obtained from The Jackson Laboratory, followed by combined-phenotype analysis. The publicly available RNA-sequencing data deposited in the NCBI Gene Expression Omnibus (GEO) database under the accession No.: GSE198512 (postnatal lethality), GSE197800 (postnatal survival) and GSE176113 (postnatal survival) were mined to explore the potential reasons why hepatocyte-specific METTL3 homozygous deletion by Alb-iCre mice (GPT), but not by Alb-Cre mice (JAX), leads to ALF and then postnatal lethality., Key Findings: Firstly, we observed that hepatocyte-specific METTL3 homozygous deficiency by Alb-iCre mice (GPT) or by Alb-Cre mice (JAX) caused liver injury, abnormal lipid accumulation and apoptosis. Secondly, we are surprised to find that hepatocyte-specific METTL3 homozygous deletion by Alb-iCre mice (GPT), but not by Alb-Cre mice (JAX), led to ALF and then postnatal lethality. Our findings clearly demonstrated that METTL3Δhep mice (GPT), which are about to die, exhibited the severe destruction of liver histological structure, suggesting that METTL3Δhep mice (GPT) nearly lose normal liver function, which subsequently contributes to ALF, followed by postnatal lethality. Finally, we unexpectedly found that as the compensatory growth responses of hepatocytes to liver injury induced by METTL3Δhep (GPT), the proliferation of METTL3Δhep hepatocytes (GPT), unlike METTL3Δhep hepatocytes (JAX), was not evidenced by the significant increase of Ki67-positive hepatocytes, not accompanied by upregulation of cell-cycle-related genes. Moreover, GO analysis revealed that upregulated genes in METTL3Δhep livers (GPT), unlike METTL3Δhep livers (JAX), are not functionally enriched in terms associated with cell cycle, cell division, mitosis, microtubule cytoskeleton organization, spindle organization, chromatin segregation and organization, and nuclear division, consistent with the loss of compensatory proliferation of METTL3Δhep hepatocytes (GPT) observed in vivo . Thus, obviously, the loss of the compensatory growth capacity of METTL3Δhep hepatocytes (GPT) in response to liver injury might contribute to, at least partially, ALF and subsequently postnatal lethality of METTL3Δhep mice (GPT)., Significance: These findings from this study and other labs provide strong evidence that these phenotypes (i.e., ALF and postnatal lethality) of METTL3Δhep mice (GPT) might be not the real functions of METTL3, and closely related with Alb-iCre mice (GPT), suggesting that we should remind researchers to use Alb-iCre mice (GPT) with caution to knockout gene in hepatocytes in vivo .

There is an increasing evidence indicating the involvement of the sympathetic nervous system (SNS) in liver disease development. To achieve an extensive comprehension of the obscure process by which the SNS alleviates inflammatory damage in non-parenchymal liver cells (NPCs) during acute liver failure (ALF), we employ isoproterenol (ISO), a beta-adrenoceptor agonist, to mimic SNS signaling. ISO was administered to C57BL/6J mice to establish an acute liver failure (ALF) model using LPS/D-GalN, which was defined as ISO + ALF. Non-parenchymal cells (NPCs) were isolated from liver tissues and digested for tandem mass tag (TMT) labeled proteomics to identify differentially expressed proteins (DEPs). The administration of ISO resulted in a decreased serum levels of pro-inflammatory cytokines, e.g., TNF-α, IL-1β, and IL-6 in ALF mice, which alleviated liver damage. By using TMT analysis, it was possible to identify 1587 differentially expressed proteins (DEPs) in isolated NPCs. Notably, over 60% of the DEPs in the ISO + ALF vs. ALF comparison were shared in the Con vs. ALF comparison. According to enrichment analysis, the DEPs influenced by ISO in ALF mice were linked to biological functions of heme and fatty acid metabolism, interferon gamma response, TNFA signaling pathway, and mitochondrial oxidation function. Protein-protein interaction network analysis indicated Mapk14 and Caspase3 may serve as potentially valuable indicators of ISO intervention. In addition, the markers on activated macrophages, such as Mapk14, Casp1, Casp8, and Mrc1, were identified downregulated after ISO initiation. ISO treatment increased the abundance of anti-inflammatory markers in mouse macrophages, as evidenced by the immunohistochemistry (IHC) slides showing an increase in Arg + staining and a reduction in iNOS + staining. Furthermore, pretreatment with ISO also resulted in a reduction of LPS-stimulated inflammation signaling markers, Mapk14 and NF-κB, in human THP-1 cells. Prior treatment with ISO may have the potential to modify the biological functions of NPCs and could serve as an innovative pharmacotherapy for delaying the pathogenesis and progression of ALF., (© 2023. The Author(s).)

Acute liver failure caused by alcoholic hepatitis (AH) is only effectively treated with liver transplantation. Livers of patients with AH show a unique molecular signature characterized by defective hepatocellular redox metabolism, concurrent to hepatic infiltration of neutrophils that express myeloperoxidase (MPO) and form neutrophil extracellular traps (NETs). Exacerbated NET formation and MPO activity contribute to liver damage in mice with AH and predicts poor prognosis in AH patients. The identification of pathways that maladaptively exacerbate neutrophilic activity in liver could inform of novel therapeutic approaches to treat AH. Whether the redox defects of hepatocytes in AH directly exacerbate neutrophilic inflammation and NET formation is unclear. Here we identify that the protein content of the mitochondrial biliverdin exporter ABCB10, which increases hepatocyte-autonomous synthesis of the ROS-scavenger bilirubin, is decreased in livers from humans and mice with AH. Increasing ABCB10 expression selectively in hepatocytes of mice with AH is sufficient to decrease MPO gene expression and histone H3 citrullination, a specific marker of NET formation. These anti-inflammatory effects can be explained by ABCB10 function reducing ROS-mediated actions in liver. Accordingly, ABCB10 gain-of-function selectively increased the mitochondrial GSH/GSSG ratio and decreased hepatic 4-HNE protein adducts, without elevating mitochondrial fat expenditure capacity, nor mitigating steatosis and hepatocyte death. Thus, our study supports that ABCB10 function regulating ROS-mediated actions within surviving hepatocytes mitigates the maladaptive activation of infiltrated neutrophils in AH. Consequently, ABCB10 gain-of-function in human hepatocytes could potentially decrease acute liver failure by decreasing the inflammatory flare caused by excessive neutrophil activity., Competing Interests: Declaration of competing interest M.L. is a co-founder of Enspire Bio and was a consultant of Enspire Bio and Capacity Bio. O.S.S is a co-founder and SAB member of Enspire Bio, Senergy-Bio and Capacity-Bio, and when this study was conducted he was serving as a consultant to LUCA-Science, IMEL, Epirium, Johnson & Johnson, Pfizer, and Stealth Biotherapeutics., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Transplantation of mesenchymal stem cells (MSCs) is a newly developed strategy for treating acute liver failure (ALF). Nonetheless, the low survival rate of MSCs after transplantation and their poor homing to damaged tissues limit the clinical application of MSCs. The research assessed whether hypoxic preconditioning (HPC) can improve the biological activity of human amniotic mesenchymal stem cells (hA-MSCs), promote their homing ability to the liver of mice with ALF, and influence liver tissue repair., Methods: Flow cytometry, CCK8, Transwell, and Western blotting assays were conducted to assess the effects of hypoxic preconditioning on the phenotype, proliferation, and migration of hA-MSCs and the changes in the c-Met and CXCR4 gene expression levels were studied. To evaluate the effects of the transplantation of hypoxic preconditioning of hA-MSCs on the homing and repair of D-galactosamine (D-GalN)/LPS-induced ALF, the mechanism was elucidated by adding c-Met, CXCR4-specific blockers (SU11274 and AMD3100)., Results: After hypoxia pretreatment (1% oxygen volume fraction), hA-MSCs maintained the morphological characteristics of adherence and vortex colony growth and showed high CD44, CD90, and CD105 and low CD31, CD34, and CD45 expression levels. Hypoxic preconditioning of hA-MSCs significantly increased their proliferation and migration and highly expressed the c-Met and CXCR4 genes. In vivo and in vitro, this migration-promoting effect was suppressed by the c-Met specific blocker SU11274. In the acute liver failure mouse model, the HGF expression level was considerably elevated in the liver than that in the serum, lungs and kidneys. The transplantation of hypoxic preconditioned hA-MSCs introduced a remarkable improvement in the liver function and survival rate of mice with ALF and enhanced the anti-apoptosis ability of liver cells. The anti-apoptotic enhancing effect of hypoxic preconditioning was suppressed by the c-Met specific blocker SU11274. Hypoxic hA-MSCs administration was observed to have considerably increased the fluorescent cells in the liver than that recorded after administering normal oxygen-hA-MSCs. The number of hepatic fluorescent cells decreased remarkably after adding the c-Met inhibitor SU11274, compared to that recorded after hypoxic pretreatment, whereas the effect of c-Met inhibitor SU11274 on normal oxygen-hA-MSCs was not significant., Conclusions: Hypoxic preconditioning depicted no impact on the morphology and phenotype features of the human amniotic mesenchymal stem cells, but it can promote their proliferation, migration, anti-apoptotic effect, and homing rate and improve the repair of acute liver failure, which might be mediated by the HGF/c-Met signaling axis., Competing Interests: Declaration of Competing Interest Not applicable., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Fulminant hepatic failure (FHF) is the terminal phase of acute liver injury, which is characterized by massive hepatocyte necrosis and rapid hepatic dysfunction in patients without preexisting liver disease. There are currently no therapeutic options for such a life-threatening hepatic failure except liver transplantation; therefore, the terminal phase of the underlying acute liver injury should be avoided. Tomatidine (TOM), asteroidal alkaloid, may have different biological activities, including antioxidant and anti-inflammatory effects. Herein, the lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced FHF mouse model was established to explore the protective potential of TOM and the underlying mechanisms of action. TOM pretreatment significantly inhibited hepatocyte necrosis and decreased serum aminotransferase activities in LPS/D-GalN-stimulated mice. TOM further increased the level of different antioxidant enzymes while reducing lipid peroxidation biomarkers in the liver. These beneficial effects of TOM were shown to be associated with targeting of NF-κB signaling pathways, where TOM repressed NF-κB activation and decreased LPS/D-GalN-induced TNF-α, IL-6, IL-1β, and iNOS production. Moreover, TOM prevented LPS/D-GalN-induced upregulation of Keap1 expression and downregulation of Nrf2 and HO-1 expression, leading to increased Nrf2-binding activity and HO-1 levels. Besides, TOM pretreatment repressed LPS/D-GalN-induced upregulation of proliferating cell nuclear antigen (PCNA) expression, which spared the hepatocytes from damage and subsequent repair following the LPS/D-GalN challenge. Collectively, our findings revealed that TOM has a protective effect on LPS/D-GalN-induced FHF in mice, showing powerful antioxidant and anti-inflammatory effects, primarily mediated via modulating Keap1/Nrf2/HO-1 and NF-κB/TNF-α/IL-6/IL-1β/iNOS signaling pathways., 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 © 2024 Elsevier B.V. All rights reserved.)

With the increasing influx of microplastics (MPs) into the environment, their potential toxicity represents an increasing threat to human health. However, there is a lack of relevant research surrounding the biological toxicity associated with pre-exposure to MPs under pathological conditions. To fill this gap, we established a mouse model of fulminant hepatic failure after 14 days of pre-exposure to polystyrene (PS) MPs and investigated its biological response process under combined stimulation with lipopolysaccharide (LPS)/d-galactosamine (d-GalN) and PS-MPs. The results indicated that the stress response from exposure to PS-MPs exacerbated the death induced by LPS/d-GalN and reinforced the potential of liver damage in mice. The dominant roles of inflammation promotion, reactive oxygen species (ROS), and neutrophil extracellular traps in this process were confirmed by cellular reactive oxygen species assays and experiments on oxidative stress and inflammatory responses in the liver. Transcriptomic analysis revealed that PS-MPs exacerbated the expression levels of neutrophil extracellular traps in mice treated with LPS/d-GalN, and weakened the expression of genes involved in pathways related to peroxisome, taurine, and hypotaurine metabolism, which was further validated by reverse-transcription quantitative polymerase chain reaction. This study addresses the knowledge gap regarding the adverse effects caused by a pathological state upon exposure to MPs and provides a theoretical reference for further assessment of the underlying health risks of MPs., 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 © 2023 Elsevier B.V. All rights reserved.)

Acetaminophen overuse is a common cause of acute liver failure (ALF). During ALF, toxins are metabolized by enzymes such as CYP2E1 and transformed into reactive species, leading to oxidative damage and liver failure. Here, we found that oral magnesium (Mg) alleviated acetaminophen-induced ALF through metabolic changes in gut microbiota that inhibit CYP2E1. The gut microbiota from Mg-supplemented humans prevented acetaminophen-induced ALF in mice. Mg exposure modulated Bifidobacterium metabolism and enriched indole-3-carboxylic acid (I3C) levels. Formate C-acetyltransferase (pflB) was identified as a key Bifidobacterium enzyme involved in I3C generation. Accordingly, a Bifidobacterium pflB knockout showed diminished I3C generation and reduced the beneficial effects of Mg. Conversely, treatment with I3C or an engineered bacteria overexpressing Bifidobacterium pflB protected against ALF. Mechanistically, I3C bound and inactivated CYP2E1, thus suppressing formation of harmful reactive intermediates and diminishing hepatocyte oxidative damage. These findings highlight how interactions between Mg and gut microbiota may help combat ALF., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Acute liver failure (ALF) is a life-threatening disease, but its pathogenesis is not fully understood. NETosis is a novel mode of cell death. Although the formation of neutrophil extracellular traps (NETs) has been found in various liver diseases, the specific mechanism by which NETosis regulates the development of ALF is unclear. In this article, we explore the role and mechanism of NETosis in the pathogenesis of ALF., Methods: Clinically, we evaluated NETs-related markers in the liver and peripheral neutrophils of patients with ALF. In in vitro experiments, HL-60 cells were first induced to differentiate into neutrophil-like cells (dHL-60 cells) with dimethyl sulfoxide (DMSO). NETs were formed by inducing dHL-60 cells with PMA. In in vivo experiments, the ALF model in mice was established with LPS/D-gal, and the release of NETs was detected by immunofluorescence staining and western blotting. Finally, the acetylation levels of IDH1 and MDH1 were detected in dHL-60 cells and liver samples by immunoprecipitation., Results: Clinically, increased release of NETs in liver tissue was observed in patients with ALF, and NETs formation was detected in neutrophils from patients with liver failure. In dHL-60 cells, mutations at IDH1-K93 and MDH1-K118 deacetylate IDH1 and MDH1, which promotes the formation of NETs. In a mouse model of ALF, deacetylation of IDH1 and MDH1 resulted in NETosis and promoted the progression of acute liver failure., Conclusions: Deacetylation of IDH1 and MDH1 reduces their activity and promotes the formation of NETs. This change aggravates the progression of acute liver failure., (© 2024. The Author(s).)

In this paper, we generated a short hairpin RNA growth differentiation factor-11 (sh-GDF11) and evaluated the effects of sh-GDF11 on the pathogenesis of acute liver failure (ALF) in vitro and in vivo. Through bioinformatics study, the key gene related to ALF was assayed. Lipopolysaccharide (LPS) and D-galactoamine (D-GalN) were applied to establish the mouse model of LPS/D-GalN-induced liver injury, and TNF-α and D-Gal were used to construct an in vitro cell model, followed by treatment of sh-GDF11 for analysis of liver cell proliferation. Bioinformatics analysis showed that the protective effect of sh-GDF11 on ALF may be mediated by phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway. The results of in vitro study found that sh-GDF11 could promote cell proliferation and inhibit death by blocking the PI3K/Akt/mTOR signaling pathway. In vivo animal experiments further confirmed that sh-GDF11 could suppress hepatocyte apoptosis by inhibiting the PI3K/Akt/mTOR signaling pathway. sh-GDF11 relieved LPS/D-GalN-induced ALF by blocking the PI3K/Akt/mTOR signaling pathway, emphasizing its critical role in LPS/D-GalN-induced ALF treatment., (© 2023 Wiley Periodicals LLC.)

Background & Aims: The function of cholinergic anti-inflammatory pathway (CAP) in acute liver failure (ALF) with inflammatory storm remains indefinite. The liver-gut axis has been proved to be crucial for liver homeostasis. Investigation about CAP regulation on liver-gut axis would enrich our understanding over cholinergic anti-inflammatory mechanism., Methods: Co-injection of lipopolysaccharide and D-galactosamine was used to establish the model of ALF. PNU-282987 was used to activate the CAP. Histological staining, real-time polymerase chain reaction, Western blotting, RNA sequencing, and flow cytometry were conducted. Liver biopsy specimens and patients' serum from patients with liver failure were also analyzed., Results: We confirmed that activating the CAP alleviated hepatocyte destruction, accompanied by a significant decrease in hepatocyte apoptosis, pro-inflammatory cytokines, and NLRP3 inflammasome activation. Moreover, hepatic MAdCAM1 and serum MAdCAM1 levels were induced in ALF, and MAdCAM1 levels were positively correlated with the extent of liver damage and the expression of pro-inflammatory markers. Furthermore, activating the CAP mainly downregulated ectopic expression of MAdCAM1 on endothelial cells, and inhibition of NF-κB p65 nuclear translocation was partly attributed to the decreased MAdCAM1. Notably, in ALF, the aberrant hepatic expression of MAdCAM1 subsequently recruited gut-derived α4β7+ CD4+T cells to the liver, which exhibited an augmented IFN-γ-secreting and IL-17-producing phenotype. Finally, we revealed that the levels of serum and hepatic MAdCAM1 were elevated in patients with liver failure and closely correlated with clinical course. Increasing hepatic infiltration of β7+ cells were also confirmed in patients., Conclusions: Activating the CAP attenuated liver injury by inhibiting MAdCAM1/α4β7 -mediated gut-derived proinflammatory lymphocytes infiltration, which provides a potential therapeutic target for ALF., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The E3 ubiquitin ligase RING finger protein 115 (RNF115), also known as breast cancer-associated gene 2 (BCA2), has been linked with the growth of some cancers and immune regulation, which is negatively correlated with prognosis. Here, it is demonstrated that the RNF115 deletion can protect mice from acute liver injury (ALI) induced by the treatment of lipopolysaccharide (LPS)/D-galactosamine (D-GalN), as evidenced by decreased levels of alanine aminotransaminase, aspartate transaminase, inflammatory cytokines (e.g., tumor necrosis factor α and interleukin-6), chemokines (e.g., MCP1/CCL2) and inflammatory cell (e.g., monocytes and neutrophils) infiltration. Moreover, it was found that the autophagy activity in Rnf115 -/- livers was increased, which resulted in the removal of damaged mitochondria and hepatocyte apoptosis. However, the administration of adeno-associated virus Rnf115 or autophagy inhibitor 3-MA impaired autophagy and aggravated liver injury in Rnf115 -/- mice with ALI. Further experiments proved that RNF115 interacts with LC3B, downregulates LC3B protein levels and cell autophagy. Additionally, Rnf115 deletion inhibited M1 type macrophage activation via NF-κB and Jnk signaling pathways. Elimination of macrophages narrowed the difference in liver damage between Rnf115 +/+ and Rnf115 -/- mice, indicating that macrophages were linked in the ALI induced by LPS/D-GalN. Collectively, for the first time, we have proved that Rnf115 inactivation ameliorated LPS/D-GalN-induced ALI in mice by promoting autophagy and attenuating inflammatory responses. This study provides new evidence for the involvement of autophagy mechanisms in the protection against acute liver injury., (© 2023. The Author(s).)

Background: Hepatic encephalopathy (HE) symptoms associated with liver insufficiency are linked to the neurotoxic effects of ammonia and other toxic metabolites reaching the brain via the blood-brain barrier (BBB), further aggravated by the inflammatory response. Cumulative evidence documents that the non-coding single-stranded RNAs, micro RNAs (miRs) control the BBB functioning. However, miRs' involvement in BBB breakdown in HE is still underexplored. Here, we hypothesized that in rats with acute liver failure (ALF) or rats subjected to hyperammonemia, altered circulating miRs affect BBB composing proteins., Methods: Transmission electron microscopy was employed to delineate structural alterations of the BBB in rats with ALF (thioacetamide (TAA) intraperitoneal (ip.) administration) or hyperammonemia (ammonium acetate (OA) ip. administration). The BBB permeability was determined with Evans blue dye and sodium fluorescein assay. Plasma MiRs were profiled by Next Generation Sequencing (NGS), followed by in silico analysis. Selected miRs, verified by qRT-PCR, were examined in cultured rat brain endothelial cells. Targeted protein alterations were elucidated with immunofluorescence, western blotting, and, after selected miR mimics transfection, through an in vitro resistance measurement., Results: Changes in BBB structure and increased permeability were observed in the prefrontal cortex of TAA rats but not in the brains of OA rats. The NGS results revealed divergently changed miRNA-ome in the plasma of both rat models. The in silico analysis led to the selection of miR-122-5p and miR-183-5p with their target genes occludin and integrin β1, respectively, as potential contributors to BBB alterations. Both proteins were reduced in isolated brain vessels and cortical homogenates in TAA rats. We documented in cultured primary brain endothelial cells that ammonia alone and, in combination with TNFα increases the relative expression of NGS-selected miRs with a less pronounced effect of TNFα when added alone. The in vitro study also confirmed miR-122-5p-dependent decrease in occludin and miR-183-5p-related reduction in integrin β1 expression., Conclusion: This work identified, to our knowledge for the first time, potential functional links between alterations in miRs residing in brain endothelium and BBB dysfunction in ALF., (© 2023. The Author(s).)

Aims: Purinergic signaling-mediated mitochondria dysfunction and innate immune-mediated inflammation act as triggers during acetaminophen (APAP)-induced liver injury (AILI). However, the underlying mechanisms by which purinoceptor regulates mitochondria function and inflammation response in the progression of AILI remains unclear., Methods: First, the hepatic level of purinergic receptor P2X 1 (P2RX1) was identified in the DILI patients and APAP-induced WT mice. P2rx1 knockout (KO) mice (P2rx1 -/- ) with 300 mg/kg APAP challenge were used for the analysis of the potential role of P2RX1 in the progression of AILI. Administration of DMX, the activator of stimulator of interferon genes (STING), was performed to investigate the effects of the STING-related pathway on APAP-treated P2rx1 -/- mice., Results: The elevated hepatic P2RX1 levels were found in DILI patients and the AILI mice. P2rx1 depletion offered protection against the initial stages of AILI, mainly by inhibiting cell death and promoting inflammation resolution, which was associated with alleviating mitochondria dysfunction. Mechanistically, P2rx1 depletion could inhibit STING-TANK-binding kinase 1 (TBK1)-P65 signaling pathways in vivo. We then showed that DMX-mediated STING activation could greatly aggravate the liver injury of P2rx1 -/- mice treated with APAP., Conclusion: Our data confirmed that P2RX1 was inducted during AILI, identified P2RX1 as a novel regulator in mitochondria dysfunction and STING pathways, and suggested a promising therapeutic approach for AILI involving the blockade of P2RX1. 1. It first demonstrated the protective effects of P2rx1 deficiency on acetaminophen-induced liver injury (AILI). 2. P2rx1 knockout alleviates mitochondria function and promotes inflammation resolution after APAP treatment. 3. It first reported the regulation of P2RX1 on the STING signaling pathway in the progress of AILI. 4. P2RX1 blockade is a promising therapeutic strategy for AILI., (© 2023. The Author(s).)

As a clinical emergency with a high mortality rate, the treatment of acute liver failure has been paid attention to by society. At present, liver transplantation is the most effective treatment for acute liver failure, but there is still an insufficient supply of liver sources and a poor prognosis. In view of the current therapeutic development of this disease, more researchers have turned their attention to the research of drugs related to the NF-κB pathway. The NF-κB canonical pathway has been proven to play a role in a variety of diseases, regulating inflammation, apoptosis, and other physiological processes. More and more evidence shows that the NF-κB canonical pathway regulates the pathogenesis of acute liver failure. In this review, we will summarize the regulation process of the NF-κB canonical pathway on acute liver failure, and develop a new way to treat acute liver failure by targeting the components of the pathway.

Cell-based liver therapies based on retrieving and steadying failed metabolic function(s) for acute and chronic diseases could be a valuable substitute for liver transplants, even though they are limited by the low engraftment capability and reduced functional quality of primary human hepatocytes (PHH). In this paper we propose the use of gelatin-hyaluronic acid (Gel-HA) scaffolds seeded with PHH for the treatment of liver failure. We first optimized the composition using Gel-HA hydrogels, looking for the mechanical properties closer to the human liver and determining HepG2 cells functionality. Gel-HA scaffolds with interconnected porosity (pore size 102 μm) were prepared and used for PHH culture and evaluation of key hepatic functions. PHH cultured in Gel-HA scaffolds exhibited increased albumin and urea secretion and metabolic capacity (CYP and UGT activity levels) compared to standard monolayer cultures. The transplant of the scaffold containing PHH led to an improvement in liver function (transaminase levels, necrosis) and ameliorated damage in a mouse model of acetaminophen (APAP)-induced liver failure. The study provided a mechanistic understanding of APAP-induced liver injury and the impact of transplantation by analyzing cytokine production and oxidative stress induction to find suitable biomarkers of cell therapy effectiveness., 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. The funders had no role in either the design of the study, the collection, analyses, or interpretation of data in writing the manuscript or in the decision to publish the results., (Crown Copyright © 2023. Published by Elsevier B.V. All rights reserved.)

MicroRNAs (miRNAs) control liver diseases, but the role of microRNA-181a-5p in acute liver failure (ALF) is unclear. In this study, the ALF model was generated by injection of D-galactosamine (D-GalN) and lipopolysaccharide (LPS). The levels of miRNAs were assessed by microarray and qRT-PCR. The expression of caspase 3 was detected as the marker of cell apoptosis in ALF by immunohistochemistry and western blot. The targeting of microRNA-181a-5p on the high mobility group box 1 (HMGB1) was verified by dual luciferase assay. The impact of microRNA-181a-5p and HMGB1 was explored by flow cytometry. Results showed that microRNA-181a-5p was significantly down-regulated by D-GalN/LPS in vivo and in vitro, while the level of HMGB1 was up-regulated after the challenge. Furthermore, microRNA-181a-5p overexpression attenuated cell apoptosis in D-GalN/TNF-treated BNLCL2 cells. MicroRNA-181a-5p could directly target HMGB1 mRNA and repress its expressions, in further HMGB1 is involved in microRNA-181a-5p effect on cell apoptosis of ALF. In conclusion, these findings demonstrate that microRNA-181a-5p regulates hepatocyte apoptosis via HMGB1 in the development of ALF, which may provide potential therapeutic targets for ALF. However, the precise underlying mechanism that connects microRNA-181a-5p and HMGB1 remains to be explored.

Purpose: Acute liver failure (ALF) is a clinically fatal disease that leads to the rapid loss of normal liver function. Acetaminophen (APAP) is a leading cause of drug-induced ALF. Ferroptosis, defined as iron-dependent cell death associated with lipid peroxide accumulation, has been shown to be strongly associated with APAP-induced liver injury. Growth arrest-specific 1 (GAS1) is a growth arrest-specific gene, which is closely related to the inhibition of cell growth and promotion of apoptosis. However, the functional role and underlying mechanism of GAS1 in APAP-induced ferroptosis remain unknown. Methods: We established liver-specific overexpression of GAS1 (GAS1 AAV8-OE) mice and the control (GAS1 AAV8-vector ) mice by tail vein injection of male mice with adeno-associated virus. APAP at 500 mg/kg was intraperitoneally injected into these two groups of mice to induce acute liver failure. The shRNA packaged by the lentivirus inhibits GAS1 gene expression in human hepatoma cell line HepaRG (HepaRG-shNC and HepaRG-shGAS1-2) and primary hepatocytes of mice with liver-specific overexpression of GAS1 were isolated and induced by APAP in vitro to further investigate the regulatory role of GAS1 in APAP-induced acute liver failure. Results: APAP-induced upregulation of ferroptosis, levels of lipid peroxides and reactive oxygen species, and depletion of glutathione were effectively alleviated by the ferroptosis inhibitor, ferrostatin-1, and downregulation of GAS1 expression. GAS1 overexpression promoted ferroptosis-induced lipid peroxide accumulation via p53, inhibiting its downstream target, solute carrier family 7 member 11. Conclusion: Collectively, our findings suggest that GAS1 overexpression plays a key role in aggravating APAP-induced acute liver injury by promoting ferroptosis-induced accumulation of lipid peroxides., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Acute liver failure (ALF) is a severe liver disease caused by many reasons. One of them is the overdosed acetaminophen (APAP), which is metabolized into N-acetyl-p-benzoquinone imine (NAPQI), an excessive toxic metabolite, by CYP2E1, resulting in excessive reactive oxygen species (ROS), exhausted glutathione (GSH), and thereafter hepatocyte necrosis. N-acetylcysteine is the Food and Drug Administration-approved drug for detoxification of APAP, but it has limited clinical application due to the short therapeutic time window and concentration-related adverse effects. In this study, a carrier-free and bilirubin dotted nanoparticle (B/BG@N) is developed, which is formed using bilirubin and 18β-Glycyrrhetinic acid, and bovine serum albumin (BSA) is then adsorbed to mimic the in vivo behavior of the conjugated bilirubin for hitchhiking. The results demonstrate that B/BG@N can effectively reduce the production of NAPQI as well as exhibit antioxidant effects against intracellular oxidative stress via regulating the nuclear factor erythroid 2-related factor 2/heme oxygenase-1 signal axis and reducing the production of inflammatory factors. In vivo study shows that B/BG@N can effectively improve the clinical symptom of the mice model. This study suggests that B/BG@N own increases circulation half-life, improves accumulation in the liver, and dual detoxification, providing a promising strategy for clinical ALF treatment., (© 2023 Wiley-VCH GmbH.)