Mouse Model Of Liver Injury Research Articles

Monocyte-derived macrophages contribute to the deterioration of immunological liver injury in mice

Background & aimsAutoimmune hepatitis (AIH) is characterized by hepatocyte destruction, leading to lymphocyte and macrophage accumulation in the liver. However, the specific mechanisms of how macrophages participate in the occurrence and development of AIH are still unclear. In this study, we investigated the effect of monocyte-derived macrophages on Con A-induced immunological liver injury in mice and we hypothesized that inhibition of CCR2 with the dual CCR2/5 inhibitor, cenicriviroc (CVC), would attenuate Con A-induced hepatitis in mice by reducing the recruitment of monocytes into the liver. MethodsMurine experimental AIH was established by concanavalin A (Con A) injection intravenously. Macrophages were depleted by injection of clodronate liposomes in Con A-treated mice. Moreover, inhibition of the CCR2/5 signaling pathway in Con A mice is achieved by CVC. Liver injury and infiltration of monocyte-derived macrophages were assessed by serum transaminase levels, histopathology, immunohistochemistry, flow cytometry, RT–qPCR, ELISA, TUNEL assay and dihydroethidium staining. ResultsThe number of macrophages in the mouse livers increased in the Con A-induced hepatitis mouse model, and flow cytometry showed a significant increase in the proportion of F4/80loCD11bhi monocyte-derived macrophages, while there was no significant change in the proportion of F4/80hiCD11blo Kupffer cells. After the depletion of liver macrophages by clodronate liposomes, the levels of serum ALT and AST, and the degree of liver tissue damage were alleviated in Con A-treated mice. Furthermore, Con A leaded an increase in the expression of a group of CC chemokines in mouse livers, and the elevation of CCL2 was prevented with the depletion of macrophages. Additionally, CVC reduced macrophage infiltration in the liver and ameliorated Con A-induced liver injury. Meanwhile, CVC reduced the apoptosis and oxidative damage of hepatocytes caused by Con A. ConclusionsOur research demonstrates that there is an increase in monocyte-derived macrophages in the livers due to the monocyte infiltration resulted from the activation of the CCL2-CCR2 axis in Con A-induced liver injury mouse model. Pharmacological inhibition of CCR2 monocyte recruitment by CVC efficiently ameliorates the hepatic inflammation, indicating the therapeutic potential of CVC in patients with AIH.

Hepatocyte survival and proliferation by fibroblast growth factor 7 attenuates liver inflammation, and fibrogenesis during acute liver injury via paracrine mechanisms

Hepatocyte damage during liver injury instigates activation of macrophages and hepatic stellate cells (HSCs) resulting in liver inflammation and fibrosis respectively. Improving hepatocyte survival and proliferation thereby ameliorating inflammation and fibrosis represents a promising approach for the treatment of liver injury. In the liver, fibroblast growth factors (FGFs) play a crucial role in promoting hepatocyte proliferation and tissue regeneration. Among 22 FGFs, FGF7 induces hepatocyte survival and liver regeneration as shown previously in mouse models of cholestatic liver injury and partial hepatectomy. We hypothesized that FGF7 promotes hepatocyte survival and proliferation by interacting with FGFR2b, expressed on hepatocytes, and ameliorates liver injury (inflammation and early fibrogenesis) via paracrine mechanisms. To prove this hypothesis and to study the effect of FGF7 on hepatocytes and liver injury, we administered FGF7 exogenously to mice with acute carbon tetrachloride (CCl4)-induced liver injury. We thereafter studied the underlying mechanisms and the effect of exogenous FGF7 on hepatocyte survival and proliferation, and the consequent paracrine effects on macrophage-induced inflammation, and HSCs activation in vitro and in vivo. We observed that the expression of FGF7 as well as FGFR2 is upregulated during acute liver injury. Co-immunostaining of FGF7 and collagen-I confirmed that FGF7 is expressed by HSCs and is possibly captured by the secreted ECM. Immunohistochemical analysis of liver sections showed increased hepatocyte proliferation upon exogenous FGF7 treatment as determined by Ki67 expression. Mechanistically, exogenous FGF7 improved hepatocyte survival (and increased drug detoxification) via AKT and ERK pathways while maintaining hepatocyte quiescence restricting hepatocarcinogenesis via P27 pathways. Flow cytometry analysis revealed that improved hepatocyte survival and proliferation leads to a decrease in infiltrated monocytes-derived macrophages, as a result of reduced CCL2 (and CXCL8) expression by hepatocytes. Moreover, conditioned medium studies showed reduced collagen-I secretion by HSCs (indicative of HSCs activation) upon treatment with FGF7-treated hepatocytes conditioned medium. Altogether, we show that exogenous administration of FGF7 induces hepatocyte survival and proliferation and leads to amelioration of inflammatory response and fibrosis in acute liver injury via paracrine mechanisms. Our study further demonstrates that FGF7, FGF7 derivatives, or nano-engineered FGF7 may benefit patients with hepatic dysfunction.

Hepatoprotective effects of the mixed decoction of Inula cappa and Serissa japonica in acute liver injury mouse model

Hepatoprotective effects of the mixed decoction of Inula cappa and Serissa japonica in acute liver injury mouse model

Characterisation of forkhead box protein A3 as a key transcription factor for hepatocyte regeneration

Liver regeneration is vital for the recovery of liver function after injury, yet the underlying mechanism remains to be elucidated. Forkhead box protein A3 (FOXA3), a member of the forkhead box family, plays important roles in endoplasmic reticulum stress sensing, and lipid and glucose homoeostasis, yet its functions in liver regeneration are unknown. Here, we explored whether Foxa3 regulates liver regeneration via acute and chronic liver injury mice models. We further characterised the molecular mechanism by chromatin immunoprecipitation sequencing and rescue experiments invivo and invitro. Then, we assessed the impact of Foxa3 pharmacological activation on progression and termination of liver regeneration. Finally, we confirmed the Foxa3-Cebpb axis in human liver samples. Foxa3 is dominantly expressed in hepatocytes and cholangiocytes and is induced upon partial hepatectomy (PH) or carbon tetrachloride (CCl4) administration. Foxa3 deficiency in mice decreased cyclin gene levels and delayed liver regeneration after PH, or acute or chronic i.p. CCl4 injection. Conversely, hepatocyte-specific Foxa3 overexpression accelerated hepatocytes proliferation and attenuated liver damage in an CCl4-induced acute model. Mechanistically, Foxa3 directly regulates Cebpb transcription, which is involved in hepatocyte division and apoptosis both invivo and invitro. Of note, Cebpb overexpression in livers of Foxa3-deficient mice rescued their defects in cell proliferation and regeneration upon CCl4 treatment. In addition, pharmacological induction of Foxa3 via cardamonin speeded up hepatocyte proliferation after PH, without interfering with liver regeneration termination. Finally, Cebpb and Ki67 levels had a positive correlation with Foxa3 expression in human chronic disease livers. These data characterise Foxa3 as a vital regulator of liver regeneration, which may represent an essential factor to maintain liver mass after liver injury by governing Cebpb transcription. Liver regeneration is vital for the recovery of liver function after chemical insults or hepatectomy, yet the underlying mechanism remains to be elucidated. Herein, via invitro and invivo models and analysis, we demonstrated that Forkhead box protein A3 (FOXA3), a Forkhead box family member, maintained normal liver regeneration progression by governing Cebpb transcription and proposed cardamonin as a lead compound to induce Foxa3 and accelerate liver repair, which signified that FOXA3 may be a potential therapeutic target for further preclinical study on treating liver injury.

An orally deliverable ornithine-based self-assembling polymer nanomedicine ameliorates hyperammonemia in acetaminophen-induced acute liver injury.

l-Ornithine (Orn) is a core amino acid responsible for ammonia detoxification in the body via the hepatic urea cycle. Clinical studies in Orn therapy have focused on interventions for hyperammonemia-associated diseases, such as hepatic encephalopathy (HE), a life-threatening neurological symptom affecting more than 80% of patients with liver cirrhosis. However, its low molecular weight (LMW) causes Orn to diffuse nonspecifically and be rapidly eliminated from the body after oral administration, resulting in unfavorable therapeutic efficacy. Hence, Orn is constantly supplied by intravenous infusion in many clinical settings; however, this treatment inevitably decreases patient compliance and limits its application in long-term management. To improve the performance of Orn, we designed self-assembling polyOrn-based nanoparticles for oral administration through ring-opening polymerization of Orn-N-carboxy anhydride initiated with amino-ended poly(ethylene glycol), followed by acylation of free amino groups in the main chain of the polyOrn segment. The obtained amphiphilic block copolymers, poly(ethylene glycol)-block-polyOrn(acyl) (PEG-block-POrn(acyl)), enabled the formation of stable nanoparticles (NanoOrn(acyl)) in aqueous media. We employed the isobutyryl (iBu) group for acyl derivatization in this study (NanoOrn(iBu)). In the healthy mice, daily oral administration of NanoOrn(iBu) for one week did not induce any abnormalities. In the mice exhibiting acetaminophen (APAP)-induced acute liver injury, oral pretreatment with NanoOrn(iBu) effectively reduced systemic ammonia and transaminases levels compared to the LMW Orn and untreated groups. The results suggest that the application of NanoOrn(iBu) is of significant clinical value with the feasibility of oral delivery and improvement in APAP-induced hepatic pathogenesis. STATEMENT OF SIGNIFICANCE: Liver injury is often accompanied by hyperammonemia, a life-threatening condition characterized by elevated blood ammonia levels. Current clinical treatments for reducing ammonia typically entail the invasive approach of intravenous infusion, involving the administration of l-ornithine (Orn) or a combination of Orn and L-aspartate. This method is employed due to the poor pharmacokinetics associated with these compounds. In our pursuit of enhancing therapy, we have developed an orally administrable nanomedicine based on Orn-based self-assembling nanoparticle (NanoOrn(iBu)), which provides sustained Orn supply to the injured liver. Oral administration of NanoOrn(iBu) to healthy mice did not cause any toxic effects. In a mouse model of acetaminophen-induced acute liver injury, oral administration of NanoOrn(iBu) surpassed Orn in reducing systemic ammonia levels and liver damage, thereby establishing NanoOrn(iBu) as a safe and effective therapeutic option.

Chuanxiong Rhizoma extracts prevent cholestatic liver injury by targeting H3K9ac-mediated and cholangiocyte-derived secretory protein PAI-1 and FN.

Chuanxiong Rhizoma (CX, the dried rhizome of Ligusticum wallichii Franch.), a well-known traditional Chinese medicine, is clinically used for treating cardiovascular, cerebrovascular and hepatobiliary diseases. Cholestatic liver damage is one of the chronic liver diseases with limited effective therapeutic strategies. Currently, little is known about the mechanism links between CX-induced anti-cholestatic action and intercellular communication between cholangiocytes and hepatic stellate cells (HSCs). The study aimed to evaluate the hepatoprotective activity of different CX extracts including the aqueous, alkaloid, phenolic acid and phthalide extracts of CX (CXAE, CXAL, CXPA and CXPHL) and investigate the intercellular communication-related mechanisms by which the most effective extracts work on cholestatic liver injury. The active compounds of different CX extracts were identified by UPLC-MS/MS. A cholestatic liver injury mouse model induced by bile duct ligation (BDL), and transforming growth factor-β (TGF-β)-treated human intrahepatic biliary epithelial cholangiocytes (HIBECs) and HSC cell line (LX-2 cells) were used for in vivo and in vitro studies. Histological and other biological techniques were also applied. The results indicated that CXAE, CXAL and CXPHL significantly reduced ductular reaction (DR) and improved liver fibrosis in the BDL mice. Meanwhile, both CXAE and CXPHL suppressed DR in injured HIBECs and reduced collagen contraction force and the expression of fibrosis biomarkers in LX-2 cells treated with TGF-β. CXPHL suppressed the transcription and transfer of plasminogen activator inhibitor-1 (PAI-1) and fibronectin (FN) from the 'DR-like' cholangiocytes to activated HSCs. Mechanistically, the inhibition of PAI-1 and FN by CXPHL was attributed to the untight combination of the acetyltransferase KAT2A and SMAD3, followdd by the suppression of histone 3 lysine 9 acetylation (H3K9ac)-mediated transcription in cholangiocytes. In conclusion, CXPHL exerts stronger anti-cholestatic activity in vivo and in vitro than other CX extracts, and its protective effect on the intracellular communication between cholangiocytes and HSCs is achieved by reducing KAT2A/H3K9ac-mediated transcription and release of PAI-1 and FN.

Protective effects of Emblica officinalis polysaccharide against lead induced liver injury in mice model

Heavy metal contamination especially lead (Pb) causes a serious threat to global public health. In the current study, we explored the protective and regulating effects of Emblica officinalis polysaccharide (EOP) in the liver against Pb-induced toxicity. According to our findings, EOP decreased the Pb-induced pathological lesions of liver and overall weight index in mice (p < 0.05). Following treatment with EOP, the levels of biological biomarkers for liver hepatic function (i.e., ALT and AST) were significantly decreased (p < 0.01) in a dose-dependent manner, consisted with histopathological changes. The key proteins involved in hepatic oxidative stress and apoptosis, including Nrf2, HO-1, Bcl-2, and Bax were quantified, which indicated EOP as an effective approach in protecting against the liver injury. Moreover, EOP treatment ameliorated the negative changes of liver metabolic profile (i.e., metabolites concentrations and metabolic patterns). In conclusion, EOP could protect the liver against oxidative stress and apoptosis induced by Pb poisoning, associated with the efficacy of ameliorating the negative changes in liver metabolic profile. Hence, the current findings recommend EOP as an efficient way for alleviating liver injury in lead poisoning.

A reactive oxygen/nitrogen species-eliminating natural product-based nanomedicine for prevention and treatment of inflammatory liver injury

Reactive oxygen and nitrogen species (RONS) are key signaling molecules that play an important role in the progression of inflammatory diseases. Although various antioxidants have RONS scavenging properties, they are difficult to be used to cope with acute inflammation which requires inflammatory site-targeting, effective anti-inflammatory activity, and low toxicity. In this work, we developed a safe and effective nano-antioxidant ZMF formed by simply self-assembly of natural plant protein zein and natural drug mangiferin for the synergistic treatment of inflammation-related acute liver injury. ZMF could eliminate RONS, inhibit the activation of macrophages, and modulate the level of inflammatory cytokines through the synergistic effects of mangiferin and zein. In addition, in the mouse models of inflammation-related acute liver injury, ZMF showed superior efficacy in reducing inflammatory reactions. Importantly, the preliminary in vitro and in vivo experiments proved the good safety of ZMF. This study proved the remarkable anti-inflammatory activities of nanomedicine self-assembled by RONS-eliminating natural products, which could provide a reference for the green development of efficient nano-antioxidant from natural products for the treatment of inflammation-related diseases.

"Yajieshaba" prevents acute alcoholic liver injury and repairs the intestinal mucosal barrier

Ethnopharmacological relevanceAn essential factor related to the acute alcoholic liver injury is damage to the intestinal mucosal barrier. Yajieshaba (YJSB) is a commonly used formulation of Dai people in China and protects the liver. Aim of the studyThis study investigated whether YJSB can prevent acute alcoholic liver injury by regulating the intestinal mucosal barrier. Materials and methodsThe mice received 0.39 g/kg, 1.17 g/kg, and 3.51 g/kg dose YJSB for 7 days, a mouse model of acute alcoholic liver injury was established by a single instillation of 56% alcohol. Plasma biochemical markers were analyzed, liver injury was identified by histopathology, lipopolysaccharide (LPS), nuclear factor-k-gene binding (NF-κB), hepatic inflammatory factors, oxidative stress factors and reactive oxygen species (ROS) content was analyzed. The morphological changes of intestinal histology were observed by H&E staining, and the ultrastructure of ileal cells was observed by transmission electron microscopy. Immunofluorescence and Western blot was used to determine the expression levels of transporters and enzymes involved in Claudin 1, Occludin and zona occludens 1 (ZO-1) homeostasis in the liver and intestine. ResultsThe findings showed that YJSB reduced the levels of aspartate aminotr ansferase (AST), alanine aminotransferase (ALT) and total bile acid (TBA), both of which are indicators of liver function and had a protective effect against liver injury. In the liver homogenate, YJSB reduced the level of LPS, NF-κB, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6), decreased the level of superoxide dismutase (SOD), malondialdehyde (MDA), glutathione peroxidase (GSH-PX) and catalase (CAT) and ROS. The results of hematoxylin and eosin (H&E) staining and transmission electron microscopy analysis revealed that YJSB reduced the degree of damage to intestinal tissue and intracellular organelles, implying that YJSB can reduce the "attack factor" that causes intestinal barrier damage, increase the "defense factor" that protects the intestinal barrier. The results of immunohistochemistry and Western blotting analysis showed that YJSB could increase the expression of claudin 1, occludin, and ZO-1 proteins, suggesting that the mechanism of action of YJSB against acute alcohol liver injury involves the upregulation of the expression of the intestinal barrier-related proteins and the repair of the damaged intestinal barrier. ConclusionsYJSB can block LPS, oxidative stress factors, and other harmful substances in the blood and protect the liver resisting acute alcoholic liver injury.

The Effect of Bergenin on Isonicotinic Acid Hydrazide and Rifampicin-Induced Liver Injury Revealed by RNA Sequencing.

Bergenin (BER), a natural component of polyphenols, has a variety of pharmacological activities, especially in improving drug metabolism, reducing cholestasis, anti-oxidative stress and inhibiting inflammatory responses. The aim of this study was to investigate the effects of BER on liver injury induced by isonicotinic acid hydrazide (INH) and rifampicin (RIF) in mice. The mice model of liver injury was established with INH (100 mg/kg)+RIF (100 mg/kg), and then different doses of BER were used to intervene. The pathological morphology and biochemical indicators of mice were detected. Meanwhile, RNA sequencing was performed to screen the differentially expressed genes and signaling pathways. Finally, critical differentially expressed genes were verified by qRT-PCR and Western blot. RNA sequencing results showed that 707 genes were significantly changed in the INH+RIF group compared with the Control group, and 496 genes were significantly changed after the BER intervention. These differentially expressed genes were mainly enriched in the drug metabolism, bile acid metabolism, Nrf2 pathway and TLR4 pathway. The validation results of qRT-PCR and Western blot were consistent with the RNA sequencing. Therefore, BER alleviated INH+RIF-induced liver injury in mice. The mechanism of BER improving INH+RIF-induced liver injury was related to regulating drug metabolism enzymes, bile acid metabolism, Nrf2 pathway and TLR4 pathway.

A Simple ICT-Based Fluorescent Probe for HOCl and Bioimaging Applications.

Over the past few decades, drug-induced liver damage (DILI) has become a serious public health problem due to drug abuse. Among multifarious reactive oxygen species, mounting evidence attests that ClO- has been used as a potential biomarker in DILI. In this work, a new "turn-on" fluorescent probe 1 was designed and synthesized by modifying 4'-hydroxybiphenyl-4-carbonitrile (dye 2) with N, N-dimethylthiocarbamate as a response site for detecting ClO-. Probe 1 displayed a low detection limit (72 nM), fast response time (30 s), wide pH operating range (6-8), great tissue penetration, large Stokes shift (125 nm) and 291-fold fluorescence enhancement at 475 nm in the mapping of ClO-. Probe 1 could trace amounts of exogenous and endogenous ClO- with high sensitivity in MCF-7 cells and HeLa cells. Expectantly, the fluoxetine-induced liver injury model is successfully established, and probe 1 has been used for detecting the fluctuation of ClO- levels in the mouse model of fluoxetine-induced liver injury. All in all, probe 1 with its high specificity, good biological compatibility and liver tissue penetration ability is expected to assist with the early diagnosis of DILI and the clinical screening of various new drugs. We expect that probe 1 could be efficiently used as a powerful molecular tool to predict clinical DILI and explore molecular mechanisms between molecules and disease.

Molecular magnetic resonance imaging of liver inflammation using an oxidatively activated probe

Many liver diseases are driven by inflammation, but imaging to non-invasively diagnose and quantify liver inflammation has been underdeveloped. The inflammatory liver microenvironment is aberrantly oxidising owing in part to reactive oxygen species generated by myeloid leucocytes. We hypothesised that magnetic resonance imaging using the oxidatively activated probe Fe-PyC3A will provide a non-invasive biomarker of liver inflammation. A mouse model of drug-induced liver injury was generated through intraperitoneal injection of a hepatoxic dose of acetaminophen. A mouse model of steatohepatitis was generated via a choline-deficient, l-amino acid defined high-fat diet (CDAHFD). Images were acquired dynamically before and after intravenous injection of Fe-PyC3A. The contrast agent gadoterate meglumine was used as a non-oxidatively activated negative control probe in mice fed CDAHFD. The (post-pre) Fe-PyC3A injection change in liver vs. muscle contrast-to-noise ratio (ΔCNR) recorded 2min post-injection was correlated with liver function test values, histologic scoring assigned using the NASH Clinical Research Network criteria, and intrahepatic myeloid leucocyte composition determined by flow cytometry. For mice receiving i.p. injections of acetaminophen, intrahepatic neutrophil composition correlated poorly with liver test values but positively and significantly with ΔCNR (r= 0.64, p <0.0001). For mice fed CDAHFD, ΔCNR generated by Fe-PyC3A in the left lobe was significantly greater in mice meeting histologic criteria strongly associated with a diagnosis NASH compared to mice where histology was consistent with likely non-NASH (p= 0.0001), whereas no differential effect was observed using gadoterate meglumine. In mice fed CDAHFD, ΔCNR did not correlate strongly with fractional composition of any specific myeloid cell subpopulation as determined by flow cytometry. Magnetic resonance imaging using Fe-PyC3A merits further evaluation as a non-invasive biomarker for liver inflammation. Non-invasive tests to diagnose and measure liver inflammation are underdeveloped. Inflammatory cells such as neutrophils release reactive oxygen species which creates an inflammatory liver microenvironment that can drive chemical oxidation. We recently invented a new class of magnetic resonance imaging probe that is made visible to the scanner only after chemical oxidation. Here, we demonstrate how this imaging technology could be applied as a non-invasive biomarker for liver inflammation.

Alisma Shugan Decoction attenuates hepatic fibrosis and endoplasmic reticulum stress in mice with carbon tetrachloride-induced fibrosis.

Over the years, Alisma Shugan Decoction (ASD), because of its potent anti-inflammation activity, has been used in traditional Chinese medicine (TCM) for treatment of many inflammation-associated disorders including those of the heart, blood vessel and brain. Herein, we examined the probable therapeutic effect of ASD in carbon tetrachloride (CCl4)-induced liver injury and fibrosis mice models. Our results demonstrate that ASD dose-dependently reduced the fibrosis-related increased collagen deposition secondary to liver tissue exposure to CCl4. Data from our biochemical analyses showed significantly less liver damage biomarkers including ALT, AST and hydroxyproline in the ASD-treated samples, suggesting hepato-protective effect of ASD. Furthermore, we demonstrated that treatment with ASD significantly reversed CCl4-induced elevation of TNF-α, IL-6, IL-1β and MP-1. Interestingly, NF-κB signalling, a principal regulator of inflammation was markedly suppressed by ASD treatment. In addition, treatment with ASD deregulated stress signalling pathways by suppressing the expression of markers of unfolded protein response, such as ATF6, IRE and GRP78. In conclusion, the present study provides preclinical evidence for the use of ASD as an efficacious therapeutic option in cases of chemical-induced liver damage and/or fibrosis. Further large-cohort validation of these findings is warranted.

An Extracellular miRNA-Responsive Artificial Receptor via Dynamic DNA Nano-assembly for Biomarker-Driven Therapy.

MicroRNAs (miRNAs) have emerged as promising diagnostic biomarkers and therapeutic targets in various diseases. However, there is currently a lack of molecular strategies that can effectively use disease-associated extracellular miRNAs as input signals to drive therapeutic functions. Herein, we present a modular and programmable miRNA-responsive chimeric DNA receptor (miRNA-CDR) capable of biomarker-driven therapy. By grafting a miRNA-responsive DNA nanodevice on a natural membrane receptor via aptamer anchoring, miRNA-CDR can sense extracellular miRNA levels and autonomously induce dimerization-mediated receptor activation via the complementary-mediated strand displacement reaction-induced dynamic DNA assembly. The sequence programmability of miRNA-CDR allows it to sense and respond to a user-defined miRNA with tunable sensitivity. Moreover, the miRNA-CDR is versatile and customizable to reprogram desirable signaling output via adapting a designated receptor, such as MET and FGFR1. Using a mouse model of drug-induced acute liver injury (DILI), we demonstrate the functionality of a designer miRNA-CDR in rewiring the recognition of the DILI-elevated miR-122 to promote MET signaling of hepatocytes for biomarker-driven in situ repair and liver function restoration. Our synthetic miRNA-CDR platform provides a novel molecular device enabling biomarker-driven therapeutic cellular response, potentially paving the way for improving the precision of cell therapy in regenerative medicine.

Development of KEAP1-targeting PROTAC and its antioxidant properties: In vitro and in vivo

Oxidative stress due to abnormal accumulation of reactive oxygen species (ROS) is an initiator of a large number of human diseases, and thus, the elimination and prevention of excessive ROS are important aspects of preventing the development of such diseases. Nuclear factor erythroid 2-related factor 2 (NRF2) is an essential transcription factor that defends against oxidative stress, and its function is negatively controlled by Kelch-like ECH-associated protein 1 (KEAP1). Therefore, activating NRF2 by inhibiting KEAP1 is viewed as a strategy for combating oxidative stress-related diseases. Here, we generated a cereblon (CRBN)-based proteolysis-targeting chimera (PROTAC), which we named SD2267, that induces the proteasomal degradation of KEAP1 and leads to NRF2 activation. As was intended, SD2267 bound to KEAP1, recruited CRBN, and induced the degradation of KEAP1. Furthermore, the KEAP1 degradation efficacy of SD2267 was diminished by MG132 (a proteasomal degradation inhibitor) but not by chloroquine (an autophagy inhibitor), which suggested that KEAP1 degradation by SD2267 was proteasomal degradation-dependent and autophagy-independent. Following KEAP1 degradation, SD2267 induced the nuclear translocation of NRF2, which led to the expression of NRF2 target genes and attenuated ROS accumulation induced by acetaminophen (APAP) in hepatocytes. Based on in vivo pharmacokinetic study, SD2267 was injected intraperitoneally at 1 or 3 mg/kg in APAP-induced liver injury mouse model. We observed that SD2267 degraded hepatic KEAP1 and attenuated APAP-induced liver damage. Summarizing, we described the synthesis of a KEAP1-targeting PROTAC (SD2267) and its efficacy and mode of action in vitro and in vivo. The results obtained suggest that SD2267 could be used to treat hepatic diseases related to oxidative stress.

Viscosity-responsive NIR-II fluorescent probe with aggregation-induced emission features for early diagnosis of liver injury

As the primary organ for drug metabolism and detoxification, the liver is susceptible to damage and seriously impaired function. In situ diagnosing and real-time monitoring of liver damage are thus of great significance but remain limited owing to the lack of reliable in vivo visualization protocols with minimal invasion. Herein, we reported for the first time an aggregation-induced emission (AIE) probe, namely DPXBI, emitting light in the second near-infrared window (NIR-II) for early diagnosis liver injury. DPXBI featured by strong intramolecular rotations, excellent aqueous solubility and robust chemical stability, is powerfully sensitive to viscosity alteration affording rapid response and high selectivity, through NIR-Ⅱ fluorescence intensity changes. The prominent viscosity-responsive performance enables DPXBI to accurately monitor both drug-induced liver injury (DILI) and hepatic ischemia-reperfusion injury (HIRI) with excellent image contrast to the background. By using the presented strategy, the detection of liver injury in mouse model can be achieved at least several hours earlier than typical clinical assays. Moreover, DPXBI is able to dynamically track the liver improvement process in vivo in the case of DILI when the hepatotoxicity is alleviated by using hepatoprotective medication. All these results demonstrate that DPXBI is a promising probe for investigating viscosity-associated pathological and physiological processes.

WED-437 - Anti-inflammatory and anti-fibrotic effects by simultaneous activation of glucagon, GIP, and GLP-1 of efocipegtrutide (HM15211) in thioacetamide-induced mouse model of liver injury and fibrosis

WED-437 - Anti-inflammatory and anti-fibrotic effects by simultaneous activation of glucagon, GIP, and GLP-1 of efocipegtrutide (HM15211) in thioacetamide-induced mouse model of liver injury and fibrosis

SAT-355 - Tunable host senescence determines the success of hepatocyte transplantation in a mouse model of liver injury

SAT-355 - Tunable host senescence determines the success of hepatocyte transplantation in a mouse model of liver injury

WED-473 - Improvement of liver fibrosis by a novel long acting Glucagon/GLP-1/GIP triple agonist, efocipegtrutide (HM15211) in carbon tetrachloride-induced mouse model of liver injury and fibrosis

WED-473 - Improvement of liver fibrosis by a novel long acting Glucagon/GLP-1/GIP triple agonist, efocipegtrutide (HM15211) in carbon tetrachloride-induced mouse model of liver injury and fibrosis

Lipid nanoparticle delivery of siRNA targeting Cyp2e1 gene attenuates subacute alcoholic liver injury in mice.

To investigate the effect and mechanism of lipid nanoparticle (LNP) delivery of small interfering RNA (siRNA) targeting Cyp2e1 gene on subacute alcoholic liver injury in mice. siRNA targeting Cyp2e1 gene was encapsulated in LNP (si-Cyp2e1 LNP) by microfluidic technique and the resulting LNPs were characterized. The optimal dose of si-Cyp2e1 LNP administration was screened. Forty female C57BL/6N mice were randomly divided into blank control group, model control group, si-Cyp2e1 LNP group, LNP control group and metadoxine group. The subacute alcoholic liver injury mouse model was induced by ethanol feeding for 10 d plus ethanol gavage for the last 3 d. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, and the superoxide dismutase (SOD) activity as well as malondialdehyde, reactive oxygen species, glutathione, triacylglycerol, total cholesterol contents in liver tissue were measured in each group, and liver index was calculated. The expression of genes related to oxidative stress, lipid synthesis and inflammation in each group of mice were measured by realtime RT-PCR. Compared with the model control group, the levels of liver index, serum ALT, AST activities, malondialdehyde, reactive oxygen species, triacylglycerol, total cholesterol contents in liver tissue decreased, but the SOD activity as well as glutathione increased in the si-Cyp2e1 LNP group (all P<0.01). Hematoxylin-eosin staining result showed disorganized hepatocytes with sparse cytoplasm and a large number of fat vacuoles and necrosis in the model control group, while the si-Cyp2e1 LNP group had uniformly sized and arranged hepatocytes with normal liver tissue morphology and structure. Oil red O staining result showed si-Cyp2e1 LNP group had lower fat content of the liver compared to the model control group (P<0.01), and no fat droplets accumulated. Anti-F4/80 monoclonal antibody fluorescence immunohistochemistry showed that the si-Cyp2e1 LNP group had lower cumulative optical density values compared to the model control group (P<0.01) and no significant inflammatory reaction. Compared with the model control group, the expression of catalytic genes P47phox, P67phox and Gp91phox were reduced (all P<0.01), while the expression of the antioxidant enzyme genes Sod1, Gsh-rd and Gsh-px were increased (all P<0.01). The mRNA expression of the lipid metabolism genes Pgc-1α and Cpt1 were increased (all P<0.01) and the lipid synthesis-related genes Srebp1c, Acc and Fasn were decreased (all P<0.01); the expression of liver inflammation-related genes Tgf-β, Tnf-α and Il-6 were decreased (all P<0.01). The si-Cyp2e1 LNP may attenuate subacute alcoholic liver injury in mice mainly by reducing reactive oxygen levels, increasing antioxidant activity, blocking oxidative stress pathways and reducing ethanol-induced steatosis and inflammation.