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Syndecan-1 limits the progression of liver injury and promotes liver repair in acetaminophen-induced liver injury in mice.

ABSTRACT: Accidental or intentional misuse of acetaminophen (APAP) is the leading cause of acute liver failure in the Western world. Although mechanisms that trigger APAP-induced liver injury (AILI) are well known, those that halt the progression of APAP liver disease and facilitate liver recovery are less understood. Heparan sulfate proteoglycans (HSPGs) bind to and regulate various tissue injury factors through their heparan sulfate (HS) chains, but the importance of HSPGs in liver injury in vivo remains unknown. Here, we examined the role of syndecan-1, the major cell-surface HSPG of hepatocytes, in AILI. Ablation of syndecan-1 in mice led to unopposed progression of liver injury upon APAP overdose. However, direct APAP hepatoxicity and liver injury at early times post-APAP overdose were unaffected by syndecan-1, suggesting that syndecan-1 influences later mechanisms that lead to liver repair. The exuberant liver injury phenotypes in syndecan-1 null (Sdc1^-/- ) mice were traced to a deficiency in protein kinase B (Akt) activation in hepatocytes, which led to a delayed increase in glycogen synthase kinase-3β (GSK-3β)-mediated hepatocyte apoptosis. Inhibition of Akt worsened, whereas inhibition of GSK-3β and caspases protected mice from AILI. Moreover, administration of purified syndecan-1, HS, or engineered heparan compounds containing 2-O-sulfate groups rescued Sdc1^-/- mice from AILI by potentiating Akt signaling and inhibiting GSK-3β-mediated apoptosis in hepatocytes. In addition, HS showed a significantly prolonged therapeutic efficacy as compared to N-acetylcysteine.

Conclusion

These results demonstrate that 2-O-sulfated domains in syndecan-1 HS halt disease progression and promote liver repair by enhancing hepatocyte survival in AILI. We propose that syndecan-1 is a critical endogenous factor that controls the balance between prosurvival signaling and apoptosis in hepatocytes in APAP liver disease. (Hepatology 2017;66:1601-1615).

SUBMITTER: Nam EJ

PROVIDER: S-EPMC6516470 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Syndecan-1 limits the progression of liver injury and promotes liver repair in acetaminophen-induced liver injury in mice.

Nam Eon Jeong EJ Hayashida Kazutaka K Aquino Rafael S RS Couchman John R JR Kozar Rosemary A RA Liu Jian J Park Pyong Woo PW

Hepatology (Baltimore, Md.) 20170926 5

Accidental or intentional misuse of acetaminophen (APAP) is the leading cause of acute liver failure in the Western world. Although mechanisms that trigger APAP-induced liver injury (AILI) are well known, those that halt the progression of APAP liver disease and facilitate liver recovery are less understood. Heparan sulfate proteoglycans (HSPGs) bind to and regulate various tissue injury factors through their heparan sulfate (HS) chains, but the importance of HSPGs in liver injury in vivo remain ...[more]

PMID: 28543100

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Project description:Background & aimsAcetaminophen (APAP)-induced acute liver injury (AILI) is a leading cause of acute liver failure (ALF). N-acetylcysteine (NAC) is only effective within 24 h after APAP intoxication, raising an urgent need for alternative approaches to treat this disease. This study aimed to test whether cathelicidin (Camp), which is a protective factor in chronic liver diseases, protects mice against APAP-induced liver injury and ALF.MethodsA clinically relevant AILI model and an APAP-induced ALF model were generated in mice. Genetic and pharmacological approaches were used to interfere with the levels of cathelicidin in vivo.ResultsAn increase in hepatic pro-CRAMP/CRAMP (the precursor and mature forms of mouse cathelicidin) was observed in APAP-intoxicated mice. Upregulated cathelicidin was derived from liver-infiltrating neutrophils. Compared with wild-type littermates, Camp knockout had no effect on hepatic injury but dampened hepatic repair in AILI and reduced survival in APAP-induced ALF. CRAMP administration reversed impaired liver recovery observed in APAP-challenged Camp knockout mice. Delayed CRAMP, CRAMP(1-39) (the extended form of CRAMP), or LL-37 (the mature form of human cathelicidin) treatment exhibited a therapeutic benefit for AILI. Co-treatment of cathelicidin and NAC in AILI displayed a stronger hepatoprotective effect than NAC alone. A similar additive effect of CRAMP(1-39)/LL-37 and NAC was observed in APAP-induced ALF. The pro-reparative role of cathelicidin in the APAP-damaged liver was attributed to an accelerated resolution of inflammation at the onset of liver repair, possibly through enhanced neutrophil phagocytosis of necrotic cell debris in an autocrine manner.ConclusionsCathelicidin reduces APAP-induced liver injury and ALF in mice by promoting liver recovery via facilitating inflammation resolution, suggesting a therapeutic potential for late-presenting patients with AILI with or without ALF.Impact and implicationsAcetaminophen-induced acute liver injury is a leading cause of acute liver failure. The efficacy of N-acetylcysteine, the only clinically approved drug against acetaminophen-induced acute liver injury, is significantly reduced for late-presenting patients. We found that cathelicidin exhibits a great therapeutic potential in mice with acetaminophen-induced liver injury or acute liver failure, which makes up for the limitation of N-acetylcysteine therapy by specifically promoting liver repair after acetaminophen intoxication. The pro-reparative role of cathelicidin, as a key effector molecule of neutrophils, in the APAP-injured liver is attributed to an accelerated resolution of inflammation at the onset of liver repair, possibly through enhanced phagocytic function of neutrophils in an autocrine manner.

Project description:Background & aimAcetaminophen (APAP)-induced acute liver failure is associated with substantial alterations in the hemostatic system. In mice, platelets accumulate in the liver after APAP overdose and appear to promote liver injury. Interestingly, patients with acute liver injury have highly elevated levels of the platelet-adhesive protein von Willebrand factor (VWF), but a mechanistic connection between VWF and progression of liver injury has not been established. We tested the hypothesis that VWF contributes directly to experimental APAP-induced acute liver injury.MethodsWild-type mice and VWF-deficient (Vwf-/-) mice were given a hepatotoxic dose of APAP (300 mg/kg, i.p.) or vehicle (saline). VWF plasma levels were measured by ELISA, and liver necrosis or hepatocyte proliferation was measured by immunohistochemistry. Platelet and VWF deposition were measured by immunofluorescence.ResultsIn wild-type mice, VWF plasma levels, high molecular weight (HMW) VWF multimers, and VWF activity decreased 24 h after APAP challenge. These changes coupled to robust hepatic VWF and platelet deposition, although VWF deficiency had minimal effect on peak hepatic platelet accumulation or liver injury. VWF plasma levels were elevated 48 h after APAP challenge, but with relative reductions in HMW multimers and VWF activity. Whereas hepatic platelet aggregates persisted in livers of APAP-challenged wild-type mice, platelets were nearly absent in Vwf-/- mice 48 h after APAP challenge. The absence of platelet aggregates was linked to dramatically accelerated repair of the injured liver. Complementing observations in Vwf-/- mice, blocking VWF or the platelet integrin αIIbβ3 during development of injury significantly reduced hepatic platelet aggregation and accelerated liver repair in APAP-challenged wild-type mice.ConclusionThese studies are the first to suggest a mechanistic link between VWF, hepatic platelet accumulation, and liver repair. Targeting VWF might provide a novel therapeutic approach to improve repair of the APAP-injured liver.Lay summaryPatients with acute liver injury due to acetaminophen overdose have highly elevated levels of the platelet-adhesive protein von Willebrand factor. It is not known whether von Willebrand factor plays a direct role in the progression of acute liver injury. We discovered that von Willebrand factor delays repair of the acetaminophen-injured liver in mice and that targeting von Willebrand factor, even in mice with established liver injury, accelerates liver repair.

Project description:Acetaminophen (APAP) overdose leads to severe hepatotoxicity. Osthole, a natural coumarin found in traditional Chinese medicinal herbs, has therapeutic potential in the treatment of various diseases. In this study, we investigated the effects of osthole against APAP-induced hepatotoxicity in mice. Mice were administered osthole (100 mg·kg-1·d-1, ip) for 3 d, then on the fourth day APAP (300 mg/kg, ip) was co-administered with osthole. The mice were euthanized post-APAP, their serum and livers were collected for analysis. Pretreatment with osthole significantly attenuated APAP-induced hepatocyte necrosis and the increases in ALT and AST activities. Compared with the mice treated with APAP alone, osthole pretreatment significantly reduced serum MDA levels and hepatic H2O2 levels, and improved liver GSH levels and the GSSG-to-GSH ratio. Meanwhile, osthole pretreatment markedly alleviated the APAP-induced up-regulation of inflammatory cytokines in the livers, and inhibited the expression of hepatic cytochrome P450 enzymes, but it increased the expression of hepatic UDP-glucuronosyltransferases (UGTs) and sulfotransferases (SULTs). Furthermore, osthole pretreatment reversed APAP-induced reduction of hepatic cAMP levels, but pretreatment with H89, a potent selective PKA inhibitor, failed to abolish the beneficial effect of osthole, whereas pretreatment with L-buthionine sulfoximine, a GSH synthesis inhibitor, abrogated the protective effects of osthole on APAP-induced liver injury, and abolished osthole-caused alterations in APAP-metabolizing enzymes. In cultured murine primary hepatocytes and Raw264.7 cells, however, osthole (40 μmol/L) did not alleviate APAP-induced cell death, but it significantly suppressed APAP-caused elevation of inflammatory cytokines. Collectively, we have demonstrated that osthole exerts a preventive effect against APAP-induced hepatotoxicity by inhibiting the metabolic activation of APAP and enhancing its clearance through an antioxidation mechanism.

Project description:Liver injury after experimental acetaminophen treatment is mediated both by direct hepatocyte injury through a P450-generated toxic metabolite and indirectly by activated liver Kupffer cells and neutrophils. This study was designed to investigate the role of Notch signaling in the regulation of innate immune responses in acetaminophen (APAP)-induced liver injury. Using a mouse model of APAP-induced liver injury, wild-type (WT) and toll-like receptor 4 knockout (TLR4 KO) mice were injected intraperitoneally with APAP or PBS. Some animals were injected with γ-secretase inhibitor DAPT or DMSO vehicle. For the in vitro study, bone marrow-derived macrophages (BMMs) were transfected with Notch1 siRNA, TLR4 siRNA, and non-specific (NS) siRNA and stimulated with LPS. Indeed, paracetamol/acetaminophen-induced liver damage was worse after Notch blockade with DAPT in wild-type mice, which was accompanied by significantly increased ALT levels, diminished hairy and enhancer of split-1 (Hes1), and phosphorylated Stat3 and Akt but enhanced high mobility group box 1 (HMGB1), TLR4, NF-κB, and NLRP3 activation after APAP challenge. Mice receiving DAPT increased macrophage and neutrophil accumulation and hepatocellular apoptosis. However, TLR4 KO mice that received DAPT reduced APAP-induced liver damage and NF-κB, NLRP3, and cleaved caspase-1 activation. BMMs transfected with Notch1 siRNA reduced Hes1 and phosphorylated Stat3 and Akt but augmented HMGB1, TLR4, NF-κB, and NLRP3. Furthermore, TLR4 siRNA knockdown resulted in decreased NF-κB and NLRP3 and cleaved caspase-1 and IL-1β levels following LPS stimulation. These results demonstrate that Notch signaling regulates innate NLRP3 inflammasome activation through regulation of HMGB1/TLR4/NF-κB activation in APAP-induced liver injury. Our novel findings underscore the critical role of the Notch1-Hes1 signaling cascade in the regulation of innate immunity in APAP-triggered liver inflammation. This might imply a novel therapeutic potential for the drug-induced damage-associated lethal hepatitis.

Project description:Acetaminophen (APAP) overdose still poses a major clinical challenge and is a leading cause of acute liver injury (ALI). N-acetylcysteine (NAC) is the only approved antidote to treat APAP toxicity while NAC therapy can trigger side effects including severe vomiting and even shock. Thus, new insights in developing novel therapeutic drugs may pave the way for better treatment of APAP poisoning. Previous research has reported that nuciferine (Nuci) possesses anti-inflammatory and antioxidant properties. Therefore, the objective of this study was proposed to investigate the hepatoprotective effects of Nuci and explore its underlying mechanisms. Mice were intraperitoneally (i.p.) administered with APAP (300 mg/kg) and subsequently injected with Nuci (25, 50, and 100 mg/kg, i.p.) at 30 min after APAP overdose. Then, all mice were sacrificed at 12 h after APAP challenge for further analysis. Nuci-treated mice did not show any side effects and our results revealed that treating Nuci significantly attenuated APAP-induced ALI, as confirmed by histopathological examinations, biochemical analysis, and diminished hepatic oxidative stress and inflammation. The in silico prediction and mRNA-sequencing analysis were performed to explore the underlying mechanisms of Nuci. GO and KEGG enrichment of the predicted target proteins of Nuci includes reactive oxygen species, drug metabolism of cytochrome P450 (CYP450) enzymes, and autophagy. Furthermore, the mRNA-sequencing analyses indicated that Nuci can regulate glutathione metabolic processes and anti-inflammatory responses. Consistently, we found that Nuci increased the hepatic glutathione restoration but decreased APAP protein adducts in damaged livers. Western blot analysis further confirmed that Nuci effectively promoted hepatic autophagy in APAP-treated mice. However, Nuci could not affect the expression levels of the main CYP450 enzymes (CYP1A2, CYP2E1, and CYP3A11). These results demonstrated that Nuci may be a potential therapeutic drug for APAP-induced ALI via amelioration of the inflammatory response and oxidative stress, regulation of APAP metabolism, and activation of autophagy.

Project description:Background & aimsAcetaminophen (APAP)-induced liver injury is the most frequent cause of acute liver failure in the US and many other countries. Metabolism of APAP results in formation of APAP protein adducts (APAP-AD) in hepatocytes and triggers mitochondrial dysfunction and necrosis. However, the mechanisms for how APAP-AD are removed from hepatocytes remain unknown.MethodsMice or primary hepatocytes were treated with APAP. APAP-AD were determined by immunoblot, immunostaining and high pressure liquid chomatography with electrochemical detection analysis.ResultsWe found that APAP-AD were detected at 1h, peaked at approximately 2h, declined at 6h and almost full removed at 24h post treatment with APAP in mouse livers and in primary mouse hepatocytes. APAP-AD displayed a punctate pattern and were colocalized with GFP-LC3 positive autophagosomes and Lamp1 positive lysosomes in APAP-treated primary hepatocytes. Moreover, isolated autophagosomes and autolysosomes from APAP-treated mouse livers contained APAP-AD, suggesting autophagy may selectively remove APAP-AD. APAP-AD were detected in both detergent soluble and insoluble pools in APAP-treated mouse livers and hepatocytes. More importantly, pharmacological inhibition of autophagy by leupeptin or chloroquine increased whereas induction of autophagy by Torin 1 decreased serum APAP-AD levels in APAP-treated mice, which correlated with alanine aminotransferase levels and liver necrosis. Furthermore, SQSTM1/p62, an autophagy receptor protein, was recruited to APAP-AD. Adenovirus-mediated shRNA knockdown of SQSTM1/p62 led to increased APAP-AD and necrosis in primary hepatocytes.ConclusionsOur data indicate that APAP-AD are removed though selective autophagy. Pharmacological induction of autophagy may be a novel promising approach for treating APAP-induced liver injury.Lay summaryAcetaminophen overdose can form acetaminophen protein adducts and mitochondria damage in hepatocytes resulting in liver injury. Activation of autophagy-lysosomal degradation pathway can help to remove acetaminophen protein adducts. Pharmacological induction of autophagy may be a novel promising approach for treating APAP-induced liver injury.

Dataset Information

Syndecan-1 limits the progression of liver injury and promotes liver repair in acetaminophen-induced liver injury in mice.

Conclusion

Publications

Syndecan-1 limits the progression of liver injury and promotes liver repair in acetaminophen-induced liver injury in mice.

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