Cocaine-derived hippuric acid activates mtDNA-STING signaling in alcoholic liver disease: Implications for alcohol and cocaine co-abuse.

The simultaneous abuse of alcohol-cocaine is known to cause stronger and more unpredictable cellular damage in the liver, heart, and brain. However, the mechanistic crosstalk between cocaine and alcohol in liver injury remains unclear. The findings revealed cocaine-induced liver injury and inflammation in both marmosets and mice. Of note, co-administration of cocaine and ethanol in mice causes more severe liver damage than individual treatment. The metabolomic analysis confirmed that hippuric acid (HA) is the most abundant metabolite in marmoset serum after cocaine consumption and that is formed in primary marmoset hepatocytes. HA, a metabolite of cocaine, increases mitochondrial DNA leakage and subsequently increases the production of proinflammatory factors via STING signaling in Kupffer cells (KCs). In addition, conditioned media of cocaine-treated KC induced hepatocellular necrosis via alcohol-induced TNFR1. Finally, disruption of STING signaling in vivo ameliorated co-administration of alcohol- and cocaine-induced liver damage and inflammation. These findings postulate intervention of HA-STING-TNFR1 axis as a novel strategy for treatment of alcohol- and cocaine-induced excessive liver damage.

Kupffer cells determine intrahepatic traffic of PEGylated liposomal doxorubicin.

Intrahepatic accumulation dominates organ distribution for most nanomedicines. However, obscure intrahepatic fate largely hampers regulation on their in vivo performance. Herein, PEGylated liposomal doxorubicin is exploited to clarify the intrahepatic fate of both liposomes and the payload in male mice. Kupffer cells initiate and dominate intrahepatic capture of liposomal doxorubicin, following to deliver released doxorubicin to hepatocytes with zonated distribution along the lobule porto-central axis. Increasing Kupffer cells capture promotes doxorubicin accumulation in hepatocytes, revealing the Kupffer cells capture-payload release-hepatocytes accumulation scheme. In contrast, free doxorubicin is overlooked by Kupffer cells, instead quickly distributing into hepatocytes by directly crossing fenestrated liver sinusoid endothelium. Compared to free doxorubicin, liposomal doxorubicin exhibits sustained metabolism/excretion due to the extra capture-release process. This work unveils the pivotal role of Kupffer cells in intrahepatic traffic of PEGylated liposomal therapeutics, and quantitively describes the intrahepatic transport/distribution/elimination process, providing crucial information for guiding further development of nanomedicines.

Natural lignin nanoparticles target tumor by saturating the phagocytic capacity of Kupffer cells in the liver.

Ligand-receptor recognition serves as the fundamental driving force for active targeting, yet it is still constrained by off-target effects. Herein, we demonstrate that circumventing or blocking the mononuclear phagocyte system (MPS) are both viable strategies to address off-target effects. Naturally derived lignin nanoparticles (LNPs) show great potential to block MPS due to its good stability, low toxicity, and degradability. We further demonstrate the impact of LNPs dosage on in vivo tumor targeting and antitumor efficacy. Our results show that a high dose of LNPs (300 mg/kg) leads to significant accumulation at the tumor site for a duration of 14 days after intravenous administration. In contrast, the low-dose counterparts (e.g., 50, 150 mg/kg) result in almost all LNPs accumulating in the liver. This discovery indicates that the liver is the primary site of LNP capture, leaving only the surplus LNPs the chance to reach the tumor. In addition, although cell membrane-engineered LNPs can rapidly penetrate tumors, they are still prone to capture by the liver during subsequent circulation in the bloodstream. Excitingly, comparable therapeutic efficacy is obtained for the above two strategies. Our findings may offer valuable insights into the targeted delivery of drugs for disease treatment.

Artesunate alleviates sepsis-induced liver injury by regulating macrophage polarization via the lncRNA MALAT1/PTBP1/IFIH1 axis.

BACKGROUND: The present study aimed to explore the regulatory effects of artesunate on macrophage polarization in sepsis. METHODS: Cell models and mice models were established using lipopolysaccharide (LPS), followed by treatment with various concentrations of artesunate. The phenotype of the macrophages was determined by flow cytometry. RNA immunoprecipitation was used to confirm the binding between MALAT1 and polypyrimidine tract-binding protein 1 (PTBP1), as well as between PTBP1 and interferon-induced helicase C domain-containing protein 1 (IFIH1). RESULTS: Treatment with artesunate inhibited M1 macrophage polarization in Kupffer cells subjected to LPS stimulation by downregulating MALAT1. Furthermore, MALAT1 abolished the inhibitory effect of artesunate on M1 macrophage polarization by recruiting PTBP1 to promote IFIH. In vivo experiments confirmed that artesunate alleviated septic liver injury by affecting macrophage polarization via MALAT1. CONCLUSION: The present study showed that artesunate alleviates LPS-induced sepsis in Kupffer cells by regulating macrophage polarization via the lncRNA MALAT1/PTBP1/IFIH1 axis.

Carbon Dot-Loaded Apoptotic Vesicles Improve the Liver Kupffer Cell-Mediated Antibacterial Effect to Synergistically Alleviate Sepsis.

Sepsis is a lethal systemic inflammatory disease against infection that lacks effective therapeutic approaches. Liver resident macrophage Kupffer cell (KC)-initiated bacterial clearance is crucial for the host to defend against infection. However, it remains unclear whether this process also governs the antibacterial therapy of sepsis that would be used to improve therapeutic outcomes. Here, we found that copper-doped carbon dots (Cu-CDs) exhibited superior antibacterial capabilities in vitro but displayed limited therapeutic effects in septic mice due to their limited ability to target the liver and restore KC antimicrobial capacity. Thus, we developed a composite nanodrug of copper-doped carbon dot-loaded apoVs (CC-apoVs) that combined the antibacterial ability of Cu-CDs and liver KC targeting features of apoV. Moreover, intravenous injection of CC-apoVs markedly alleviated the systemic infection and decreased the mortality of septic mice compared to Cu-CD and apoV infusion alone. Mechanistically, CC-apoV injection rescued impaired liver KCs during sepsis and enhanced their ability to capture and kill bloodborne bacteria. In addition, apoV-promoted macrophage killing of bacteria could be blocked by the inhibition of small GTPase Rab5. This study reveals a liver KC-targeted therapeutic strategy for sepsis and provides a nanodrug CC-apoV to improve the host antibacterial defense and amplify the therapeutic effect of the nanodrug.

N-3 Polyunsaturated Fatty Acids Protect against Alcoholic Liver Steatosis by Activating FFA4 in Kupffer Cells.

Supplementation with fish oil rich in omega-3 polyunsaturated fatty acids (n-3 PUFAs) effectively reduces acute and chronic alcohol-induced hepatic steatosis. We aimed to find molecular mechanisms underlying the effects of n-3 PUFAs in alcohol-induced hepatic steatosis. Because free fatty acid receptor 4 (FFA4, also known as GPR120) has been found as a receptor for n-3 PUFAs in an ethanol-induced liver steatosis model, we investigated whether n-3 PUFAs protect against liver steatosis via FFA4 using AH7614, an FFA4 antagonist, and Ffa4 knockout (KO) mice. N-3 PUFAs and compound A (CpdA), a selective FFA4 agonist, reduced the ethanol-induced increase in lipid accumulation in hepatocytes, triglyceride content, and serum ALT levels, which were not observed in Ffa4 KO mice. N-3 PUFAs and CpdA also reduced the ethanol-induced increase in lipogenic sterol regulatory element-binding protein-1c expression in an FFA4-dependent manner. In Kupffer cells, treatment with n-3 PUFA and CpdA reversed the ethanol-induced increase in tumor necrosis factor-α, cyclooxygenase-2, and NLR family pyrin domain-containing 3 expression levels in an FFA4-dependent manner. In summary, n-3 PUFAs protect against ethanol-induced hepatic steatosis via the anti-inflammatory actions of FFA4 on Kupffer cells. Our findings suggest FFA4 as a therapeutic target for alcoholic hepatic steatosis.

Hybrid Bioactive Hydrogel Promotes Liver Regeneration through the Activation of Kupffer Cells and ECM Remodeling After Partial Hepatectomy.

Partial hepatectomy is an essential surgical technique used to treat advanced liver diseases such as liver tumors, as well as for performing liver transplants from living donors. However, postoperative complications such as bleeding, abdominal adhesions, wound infections, and inadequate liver regeneration pose significant challenges and increase morbidity and mortality rates. A self-repairing mixed hydrogel (O5H2/Cu2+/SCCK), containing stem cell derived cytokine (SCCK) derived from human umbilical cord mesenchymal stem cells (HUMSCs) treated with the traditional Chinese remedy Tanshinone IIA (TSA), is developed. This SCCK, in conjunction with O5H2, demonstrates remarkable effects on Kupffer cell activation and extracellular matrix (ECM) remodeling. This leads to the secretion of critical growth factors promoting enhanced proliferation of hepatocytes and endothelial cells, thereby facilitating liver regeneration and repair after partial hepatectomy. Furthermore, the hydrogel, featuring macrophage-regulating properties, effectively mitigates inflammation and oxidative stress damage in the incision area, creating an optimal environment for postoperative liver regeneration. The injectability and strong adhesion of the hydrogel enables rapid hemostasis at the incision site, while its physical barrier function prevents postoperative abdominal adhesions. Furthermore, the hydrogel's incorporation of Cu2+ provides comprehensive antibacterial effects, protecting against a wide range of bacteria types and reducing the chances of infections after surgery.

MDIVI-1 ALLEVIATES SEPSIS-INDUCED LIVER INJURY BY INHIBITING STING SIGNALING ACTIVATION.

ABSTRACT: Proinflammatory hyperactivation of Kupffer cells (KCs) is foremost involved in the pathogenesis of sepsis-induced liver injury. Our previous study found that stimulator of interferon genes (STING) signaling was activated in KCs in response of lipopolysaccharide (LPS) and knocking down dynamin-related protein 1 (DRP1) in KCs effectively inhibited the activation of STING signaling and the subsequent production of proinflammatory cytokines. In this study, we demonstrated that in vivo treatment with mitochondrial division inhibitor 1 (Mdivi-1), a selective inhibitor of DRP1, alleviated cecal ligation and puncture (CLP)-induced liver injury with the improvement of liver pathology and function. Moreover, we found that STING in liver was mainly concentrated in KCs and STING signaling was significantly activated in KCs after CLP. The STING deficiency effectively ameliorated liver injury and decreased the mortality of septic mice, which were reversely worsened by the enhanced activation of STING with DMXAA. The further study showed that Mdivi-1 markedly attenuated STING signaling activation in KCs and inhibited systemic inflammatory response. Importantly, DMXAA application in CLP mice blunted Mdivi-1's liver protection effect. Taken together, our study confirmed Mdivi-1 effectively alleviated CLP-induced liver injury partially through inhibiting STING signaling activation in KCs, which provides new insights and a novel potential pharmacological therapeutic target for treating septic liver injury.

6-Formylindolo[3,2-b]carbazole, a potent ligand for the aryl hydrocarbon receptor, attenuates concanavalin-induced hepatitis by limiting T-cell activation and infiltration of proinflammatory CD11b+ Kupffer cells.

FICZ (6-formylindolo[3,2-b]carbazole) is a potent aryl hydrocarbon receptor agonist that has a poorly understood function in the regulation of inflammation. In this study, we investigated the effect of aryl hydrocarbon receptor activation by FICZ in a murine model of autoimmune hepatitis induced by concanavalin A. High-throughput sequencing techniques such as single-cell RNA sequencing and assay for transposase accessible chromatin sequencing were used to explore the mechanisms through which FICZ induces its effects. FICZ treatment attenuated concanavalin A-induced hepatitis, evidenced by decreased T-cell infiltration, decreased circulating alanine transaminase levels, and suppression of proinflammatory cytokines. Concanavalin A revealed an increase in natural killer T cells, T cells, and mature B cells upon concanavalin A injection while FICZ treatment reversed the presence of these subsets. Surprisingly, concanavalin A depleted a subset of CD55+ B cells, while FICZ partially protected this subset. The immune cells showed significant dysregulation in the gene expression profiles, including diverse expression of migratory markers such as CCL4, CCL5, and CXCL2 and critical regulatory markers such as Junb. Assay for transposase accessible chromatin sequencing showed more accessible chromatin in the CD3e promoter in the concanavalin A-only group as compared to the naive and concanavalin A-exposed, FICZ-treated group. While there was overall more accessible chromatin of the Adgre1 (F4/80) promoter in the FICZ-treated group, we observed less open chromatin in the Itgam (CD11b) promoter in Kupffer cells, supporting the ability of FICZ to reduce the infiltration of proinflammatory cytokine producing CD11b+ Kupffer cells. Taken together, these data demonstrate that aryl hydrocarbon receptor activation by FICZ suppresses liver injury through the limitation of CD3+ T-cell activation and CD11b+ Kupffer cell infiltration.

Mechanism of bufalin inhibition of colon cancer liver metastasis by regulating M2-type polarization of Kupffer cells induced by highly metastatic colon cancer cells.

Patients with metastatic colorectal cancer often have poor outcomes, primarily due to hepatic metastasis. Colorectal cancer (CRC) cells have the ability to secrete cytokines and other molecules that can remodel the tumor microenvironment, facilitating the spread of cancer to the liver. Kupffer cells (KCs), which are macrophages in the liver, can be polarized to M2 type, thereby promoting the expression of adhesion molecules that aid in tumor metastasis. Our research has shown that huachanshu (with bufalin as the main active monomer) can effectively inhibit CRC metastasis. However, the underlying mechanism still needs to be thoroughly investigated. We have observed that highly metastatic CRC cells have a greater ability to induce M2-type polarization of Kupffer cells, leading to enhanced metastasis. Interestingly, we have found that inhibiting the expression of IL-6, which is highly expressed in the serum, can reverse this phenomenon. Notably, bufalin has been shown to attenuate the M2-type polarization of Kupffer cells induced by highly metastatic Colorectal cancer (mCRC) cells and down-regulate IL-6 expression, ultimately inhibiting tumor metastasis. In this project, our aim is to study how high mCRC cells induce M2-type polarization and how bufalin, via the SRC-3/IL-6 pathway, can inhibit CRC metastasis. This research will provide a theoretical foundation for understanding the anti-CRC effect of bufalin.

GO-PEG Represses the Progression of Liver Inflammation via Regulating the M1/M2 Polarization of Kupffer Cells.

As a highly promising nanomaterial, exploring the impact of the liver, a vital organ, stands out as a crucial focus in the examination of its biological effects. Kupffer cells (KCs) are one of the first immune cells to contact with exotic-substances in liver. Therefore, this study investigates the immunomodulatory effects and mechanisms of polyethylene glycol-modified graphene oxide (GO-PEG) on KCs. Initial RNA-seq and KEGG pathway analyses reveal the inhibition of the TOLL-like receptor, TNF-α and NOD-like receptor pathways in continually stimulated KCs exposed to GO-PEG. Subsequent biological experiments validate that a 48-hour exposure to GO-PEG alleviates LPS-induced KCs immune activation, characterized by a shift in polarization from M1 to M2. The underlying mechanism involves the absorption of double-stranded RNA/single-stranded RNA, inhibiting the activation of TLR3 and TLR7 in KCs. Employing a Kupffer/AML12 cell co-culture model and animal studies, it is observed that GO-PEG indirectly inhibit oxidative stress, mitochondrial dysfunction, and apoptosis in AML12 cells, partially mitigating systemic inflammation and preserving liver tissue/function. This effect is attributed to the paracrine interaction between KCs and hepatocytes. These findings suggest a meaningful and effective strategy for treating liver inflammation, particularly when combined with anti-inflammatory drugs.

Lobeglitazone inhibits LPS-induced NLRP3 inflammasome activation and inflammation in the liver.

Liver inflammation is a common feature of chronic liver disease and is often associated with increased exposure of the liver to lipopolysaccharide (LPS). Kupffer cells (KCs) are macrophages in the liver and produce various cytokines. Activation of KCs through the NLRP3 inflammasome pathway leads to release of proinflammatory cytokines and induces hepatocyte injury and hepatic stellate cell (HSC) activation. Lobeglitazone is a peroxisome proliferator-activated receptor gamma ligand and a type of thiazolidinedione that elicits anti-inflammatory effects. However, there is no clear evidence that it has direct anti-inflammatory effects in the liver. This study showed that lobeglitazone reduces LPS-induced NLPR3 inflammasome activation and production of proinflammatory cytokines in primary KCs and hepatocytes. Cytokines secreted by activated KCs increased hepatocyte inflammation and HSC activation, and lobeglitazone inhibited these responses. In addition, lobeglitazone suppressed liver fibrosis by inhibiting LPS-induced transforming growth factor (TGF)-ß secretion and TGF-ß-induced CTGF expression. The inhibitory effect of lobeglitazone on inflammasome activation was associated with suppression of liver fibrosis. These results suggest that lobeglitazone may be a treatment option for inflammation and fibrosis in the liver.

Adenosine receptor A2B mediates alcoholic hepatitis by regulating cAMP levels and the NF-KB pathway.

Alcoholic hepatitis is a serious form of liver damage. Inflammation is a key factor in alcoholic hepatitis and plays a key role in the progression of alcoholic liver disease. Adenosine receptor A2B (A2BAR) is a member of the adenosine receptor family and generally considered to be a negative regulator of the inflammatory response. We found that A2BAR was the most highly expressed adenosine receptor in ETOH-fed mouse liver tissue and was also highly expressed in primary Kupffer cells and ETOH-induced RAW264.7 cells. In addition, injection of BAY 60-6583 stimulated A2BAR, induced upregulation of the expression levels of cAMP, and reduced ETOH-induced steatosis and inflammation in mice. At the same time, knockdown of A2BAR in vitro increased the inflammatory response in RAW264.7 cells triggered by ETOH. After knockdown of A2BAR in vitro, the release of the inflammatory cytokines IL-6, IL-1ß and TNF-α was increased. After overexpression of A2BAR in vitro, the cAMP level was significantly increased, PKA expression was increased, the expression of phosphorylated proteins in the NF-kB signal transduction pathway was significantly affected, and the expression of the key phosphorylated protein p-P65 was decreased. However, after the simultaneous overexpression of A2BAR and inhibition of PKA, the expression of the key phosphorylated protein p-P65 was still significantly decreased. In addition, after the expression of A2BAR increased or decreased in RAW264.7 cells, AML-12 cells were cultured in the supernatant of RAW264.7 cells stimulated by ETOH, and the apoptosis rate was significantly changed by flow cytometry. These results suggest that A2BAR can reduce alcoholic steatohepatitis by upregulating cAMP levels and negatively regulating the NF-kB pathway. Overall, these findings suggest the significance of A2BAR-mediated inflammation in alcoholic liver disease.

Diallyl disulfide suppresses the lipopolysaccharide-driven inflammatory response of macrophages by activating the Nrf2 pathway.

Lipopolysaccharide (LPS)-driven activation of Kupffer cells plays critical roles in the development of alcoholic liver disease (ALD). Accumulating evidence has revealed that nuclear factor erythroid 2-related factor 2 (Nrf2) can modulate the polarization of macrophages. The current study aimed to investigate the roles of diallyl disulfide (DADS) in LPS-driven inflammation in vitro and in vivo. We found that DADS significantly increased the nuclear translocation of Nrf2 and the transcription of Nrf2 targets, including HO1, NQO1, and γ-GCSc, and suppressed degradation of Nrf2 protein. Besides, DADS significantly inhibited LPS-induced activation of NF-κB and MAPK, secretion of NO and TNF-α, and production of reactive oxygen species (ROS) in LPS-exposed RAW264.7 cells. In vivo study demonstrated that DADS significantly ameliorated liver damage in mice challenged with LPS, as shown by the inhibition of increases in serum aminotransferase activities, neutrophil infiltration, and NF-κB and NLRP3 inflammasome activation. Finally, knockout of Nrf2 abrogated the suppression of DADS on macrophage polarization and on liver injury induced by LPS. These findings reveal that DADS suppresses LPS-driven inflammatory response in the liver by activating Nrf2, which suggests that the protective effects of DADS against ALD may be attributed to the modulation of Kupffer cell polarization in the liver.

Elaidic acid induced NLRP3 inflammasome activation via ERS-MAPK signaling pathways in Kupffer cells.

Trans fatty acids (TFA) in food can cause liver inflammation. Activation of NOD-like receptor protein-3 (NLRP3) inflammasome is a key factor in the regulation of inflammation. Accumulating evidence suggests that ERS-induced NLRP3 inflammasome activation underlies the pathological basis of various inflammatory diseases, but the precise mechanism has not been fully elucidated. Therefore, this paper focused on TFA, represented by elaidic acid (EA), to investigate the mechanism of liver inflammation. Levels of mRNA and protein were detected by RT-qPCR and Western blotting, the release of proinflammatory cytokines was measured by ELISA, and intracellular Ca2+ levels were determined by flow cytometer using Fluo 4-AM fluorescent probes. Our research indicated that EA induced the endoplasmic reticulum stress (ERS) response in Kupffer cells (KCs), accompanied by the activation of the mitogen-activated protein kinase (MAPK) signaling pathway, which resulted in NLRP3 inflammasome formation, and eventually increased the release of inflammatory factors. NLRP3 inflammasome activation was inhibited when KCs were pretreated with ERS inhibitors (4-PBA) and MAPK selective inhibitors. Furthermore, when ERS was blocked, the MAPK pathway was inhibited.

A 3D primary human cell-based in vitro model of non-alcoholic steatohepatitis for efficacy testing of clinical drug candidates.

Non-alcoholic steatohepatitis (NASH) is a progressive and severe liver disease, characterized by lipid accumulation, inflammation, and downstream fibrosis. Despite its increasing prevalence, there is no approved treatment yet available for patients. This has been at least partially due to the lack of predictive preclinical models for studying this complex disease. Here, we present a 3D in vitro microtissue model that uses spheroidal, scaffold free co-culture of primary human hepatocytes, Kupffer cells, liver endothelial cells and hepatic stellate cells. Upon exposure to defined and clinically relevant lipotoxic and inflammatory stimuli, these microtissues develop key pathophysiological features of NASH within 10 days, including an increase of intracellular triglyceride content and lipids, and release of pro-inflammatory cytokines. Furthermore, fibrosis was evident through release of procollagen type I, and increased deposition of extracellular collagen fibers. Whole transcriptome analysis revealed changes in the regulation of pathways associated with NASH, such as lipid metabolism, inflammation and collagen processing. Importantly, treatment with anti-NASH drug candidates (Selonsertib and Firsocostat) decreased the measured specific disease parameter, in accordance with clinical observations. These drug treatments also significantly changed the gene expression patterns of the microtissues, thus providing mechanisms of action and revealing therapeutic potential. In summary, this human NASH model represents a promising drug discovery tool for understanding the underlying complex mechanisms in NASH, evaluating efficacy of anti-NASH drug candidates and identifying new approaches for therapeutic interventions.

The protective effect of cannabinoid type II receptor agonist AM1241 on ConA-induced liver injury in mice via mitogen-activated protein kinase signalling pathway.

INTRODUCTION: The endocannabinoid system plays an important role in regulating the immune responses in inflammation. At present, there are no good clinical drugs for many immune liver diseases. METHODS: We explored the protective effect of the cannabinoid type II (CB2) receptor agonist AM1241 on the liver of mice with acute liver injury caused by concanavalin from the perspective of inflammation and immunity. Pathological evaluation in hepatic tissue was examined by haematoxylin and eosin (HE) staining and the levels of biochemical parameters in the serum were measured by automatic biochemical analysis. The content of inflammatory factors was measured by enzyme-linked immunosorbent assay and real-time quantitative reverse transcription polymerase chain reaction (real-time PCR). The liver apoptosis-related proteins were observed by immunohistochemistry. The expression of liver injury-related proteins was analysed by Western blot. Immune cells were isolated from the liver of mice and studied in vitro. RESULTS: Reduced levels of alanine transaminase and aspartate transaminase were observed in ConA-induced liver injury mice treated with AM1241, together with attenuated liver damage evidenced by H&E staining. Moreover, AM1241 inhibited the protein and gene expression levels of TNF-α, IL-6 and IFN-γ in the livers of mice. The phosphorylation levels of p38, JNK, ERK1/2, P65 and cAMP response element-binding protein (CREB) in the mouse were significantly reduced in AM1241 pretreatment, while the level of p-JNK increased. In addition, the P/T-P65 and P/T-CREB of the AM1241 pretreatment group were significantly reduced. The results of immunohistochemistry measurement are consistent with those of Western blotting. The CB2-mediated effect is through macrophage-like Kupffer cells. CONCLUSION: Our study suggests that the ConA-induced liver injury model in mice is protected by CB2 agonist AM1241 by modulation of CB2 receptor-rich immune cells, for example, Kupffer cells. Reduced inflammatory responses regulate apoptosis/cell death in the liver particularly hepatocytes and other parenchymal cells.

Auranofin prevents liver fibrosis by system Xc-mediated inhibition of NLRP3 inflammasome.

Demand for a cure of liver fibrosis is rising with its increasing morbidity and mortality. Therefore, it is an urgent issue to investigate its therapeutic candidates. Liver fibrosis progresses following 'multi-hit' processes involving hepatic stellate cells, macrophages, and hepatocytes. The NOD-like receptor protein 3 (NLRP3) inflammasome is emerging as a therapeutic target in liver fibrosis. Previous studies showed that the anti-rheumatic agent auranofin inhibits the NLRP3 inflammasome; thus, this study evaluates the antifibrotic effect of auranofin in vivo and explores the underlying molecular mechanism. The antifibrotic effect of auranofin is assessed in thioacetamide- and carbon tetrachloride-induced liver fibrosis models. Moreover, hepatic stellate cell (HSC), bone marrow-derived macrophage (BMDM), kupffer cell, and hepatocyte are used to examine the underlying mechanism of auranofin. Auranofin potently inhibits activation of the NLRP3 inflammasome in BMDM and kupffer cell. It also reduces the migration of HSC. The underlying molecular mechanism was inhibition of cystine-glutamate antiporter, system Xc. Auranofin inhibits system Xc activity and instantly induced oxidative burst, which mediated inhibition of the NLRP3 inflammasome in macrophages and HSCs. Therefore, to the best of our knowledge, we propose the use of auranofin as an anti-liver fibrotic agent.

Pyrroloquinoline quinone (PQQ) alleviated sepsis-induced acute liver injury, inflammation, oxidative stress and cell apoptosis by downregulating CUL3 expression.

PQQ has anti-inflammatory and anti-oxidant effects. PQQ can relieve high glucose-induced renal cell damage by suppressing Keap1 expression. Keap1 can interact with CUL3. Upregulation of CUL3 facilitates the apoptosis of LPS-induced podocytes. Based on knowledge above, this current work was designed to explore the role of PQQ in sepsis and determine the molecular function of CUL3 in the pathogenesis of sepsis. Rats received CLP surgery to establish sepsis models in vivo. Kupffer cells were pretreated with PQQ (10, 50 and 100 nmol/L) for 2 h and then treated with 100 ng/mL LPS for 24 h, simulating sepsis-induced acute liver injury in vitro. H&E staining was performed to evaluate liver injury of SD rats. Levels of inflammatory factors and oxidative stress markers were detected to assess inflammatory response and oxidative stress. Moreover, TUNEL staining, flow cytometric analysis and western blot were applied to determine cell apoptosis. It was confirmed that PQQ treatment relieved acute liver injury, inflammatory and oxidative stress damage and apoptosis of liver tissue cells in sepsis rats. In addition, PQQ therapy could alleviate inflammation, oxidative stress and apoptosis in LPS-induced Kupffer cells. Notably, LPS stimulation enhanced CUL3 expression and PQQ repressed CUL3 expression in Kupffer cells suffered from LPS. Overall, CUL3 overexpression weakened the remission effects of PQQ on LPS-induced inflammatory and oxidative damage and apoptosis of Kupffer cells. Mechanistically, PQQ treatment may mitigate sepsis-induced acute liver injury through downregulating CUL3 expression.

Acidic Microenvironment Aggravates the Severity of Hepatic Ischemia/Reperfusion Injury by Modulating M1-Polarization Through Regulating PPAR-γ Signal.

Hepatic injury induced by ischemia and reperfusion (HIRI) is a major clinical problem after liver resection or transplantation. The polarization of macrophages plays an important role in regulating the severity of hepatic ischemia/reperfusion injury. Recent evidence had indicated that the ischemia induces an acidic microenvironment by causing increased anaerobic glycolysis and accumulation of lactic acid. We hypothesize that the acidic microenvironment might cause the imbalance of intrahepatic immunity which aggravated HIRI. The hepatic ischemia/reperfusion injury model was established to investigate the effect of the acidic microenvironment to liver injury. Liposomes were used to deplete macrophages in vivo. Macrophages were cultured under low pH conditions to analyze the polarization of macrophages in vitro. Activation of the PPAR-γ signal was determined by Western blot. PPAR-γ agonist GW1929 was administrated to functionally test the role of PPAR-γ in regulating macrophage-mediated effects in the acidic microenvironment during HIRI. We demonstrate that acidic microenvironment aggravated HIRI while NaHCO3 reduced liver injury through neutralizing the acid, besides, liposome abolished the protective ability of NaHCO3 through depleting the macrophages. In vivo and vitro experiment showed that acidic microenvironment markedly promoted M1 polarization but inhibited M2 polarization of macrophage. Furthermore, the mechanistic study proved that the PPAR-γ signal was suppressed during the polarization of macrophages under pH = 6.5 culture media. The addition of PPAR-γ agonist GW1929 inhibited M1 polarization under acidic environment and reduced HIRI. Our results indicate that acidic microenvironment is a key regulator in HIRI which promoted M1 polarization of macrophages through regulating PPAR-γ. Conversely, PPAR-γ activation reduced liver injury, which provides a novel therapeutic concept to prevent HIRI.