Targeted inhibition of autophagy in hepatic stellate cells by hydroxychloroquine: An effective therapeutic approach for the treatment of liver fibrosis.

BACKGROUND AND PURPOSE: Liver fibrosis is a wound-healing reaction which is the main cause of chronic liver diseases worldwide. The activated hepatic stellate cell (aHSC) is the main driving factor in the development of liver fibrosis. Inhibiting autophagy of aHSC can prevent the progression of liver fibrosis, but inhibiting autophagy of other liver cells has opposite effects. Hence, targeted inhibition of autophagy in aHSC is quite necessary for the treatment of liver fibrosis, which prompts us to explore the targeted delivery system of small molecule autophagy inhibitor hydroxychloroquine (HCQ) that can target aHSC and alleviate the liver fibrosis. METHODS: The delivery system of HCQ@retinol-liposome nanoparticles (HCQ@ROL-LNPs) targeting aHSC was constructed by the film dispersion and pH-gradient method. TGF-ß-induced HSC activation and thioacetamide (TAA)-induced liver fibrosis mice model were established, and the targeting ability and therapeutic effect of HCQ@ROL-LNPs in liver fibrosis were studied subsequently in vitro and in vivo. RESULTS: HCQ@ROL-LNPs have good homogeneity and stability. They inhibited the autophagy of aHSC selectively by HCQ and reduced the deposition of extracellular matrix (ECM) and the damage to other liver cells. Compared with the free HCQ and HCQ@LNPs, HCQ@ROL-LNPs had good targeting ability, showing enhanced therapeutic effect and low toxicity to other organs. CONCLUSION: Construction of HCQ@ROL-LNPs delivery system lays a theoretical and experimental foundation for the treatment of liver fibrosis and promotes the development of clinical therapeutic drugs for liver diseases.

Sesamol as a potential candidate for the treatment of hepatic fibrosis, based on its regulation of FXR/LXR axis-mediated inhibition of autophagy through crosstalk between hepatic cells and macrophage.

BACKGROUND: Sesamol (SEM), a natural lignan compound isolated from sesame, has strong anti-oxidant property, regulating lipid metabolism, decreasing cholesterol and hepatoprotection. However, its anti-hepatic fibrosis effect and mechanisms have not been comprehensively elucidated. HYPOTHESIS/PURPOSE: This study aims to investigate the anti-hepatic fibrosis of SEM and its underlying mechanisms. METHOD: C57BL/6 mice with hepatic fibrosis were induced by TAA, then administrated with SEM or curcumin, respectively. HSCs were stimulated by TGF-ß or conditioned medium, and then cultured with SEM, GW4064, GW3965, Rapamycin (RA) or 3-methyladenine (3-MA), respectively. Mice with hepatic fibrosis also were administrated with SEM, RA or 3-MA to estimate the effect of SEM on autophagy. RESULTS: In vitro, SEM significantly inhibited extracellular matrix deposition, P2 × 7r-NLRP3, and inflammatory cytokines. SEM increased FXR and LXRα/ß expressions and decreased MAPLC3α/ß and P62 expressions, functioning as 3-MA (autophagy inhibitor). In vivo, SEM reduced serum transaminase, histopathology changes, fibrogenesis, autophagy markers and inflammatory cytokines caused by TAA. LX-2 were activated with conditioned medium from LPS-primed THP-1, which resulted in significant enhance of autophagy markers and inflammatory cytokines and decrease of FXR and LXRα/ß expressions. SEM could reverse above these changes and function as 3-MA, GW4064, or GW3965. Deficiency of FXR or LXR attenuated the regulation of SEM on α-SMA, MAPLC3α/ß, P62 and IL-1ß in activated LX-2. In activated THP-1, deficiency of FXR could decrease the expression of LXR, and vice versa. Deficiency of FXR or LXR in activated MΦ decreased the expressions of FXR and LXR in activated LX-2. Deficiency FXR or LXR in activated MΦ also attenuated the regulation of SEM on α-SMA, MAPLC3α/ß, P62, caspase-1 and IL-1ß. In vivo, SEM significantly reversed hepatic fibrosis via FXR/LXR and autophagy. CONCLUSION: SEM could regulate hepatic fibrosis by inhibiting fibrogenesis, autophagy and inflammation. FXR/LXR axis-mediated inhibition of autophagy contributed to the regulation of SEM against hepatic fibrosis, especially based on involving in the crosstalk of HSCs-macrophage. SEM might be a prospective therapeutic candidate, and its mechanism would be a new direction or strategy for hepatic fibrosis treatment.

Two-Membrane Hybrid Nanobiomimetic Delivery System for Targeted Autophagy Inhibition of Activated Hepatic Stellate Cells To Synergistically Treat Liver Fibrosis.

Liver fibrosis is one of the most common and highly prevalent chronic liver diseases caused by multiple pathogenic factors, and there is still no effective therapeutic drugs up to now. The activated hepatic stellate cells (aHSCs) are the main executor in liver fibrosis, and the autophagy plays a key role in the proliferation and differentiation of aHSCs, which promotes the development of liver fibrosis. However, autophagy has the opposite effect on the different kinds of liver cells in the development of liver fibrosis, and the clinical treatment has been limited by the poor selectivity and inefficient drug delivery to aHSCs. Therefore, in this study, a liposome (Lip) and exosome (Exo) two-membrane hybrid nanobiomimetic delivery system HCQ@VA-Lip-Exo was designed, which was modified by vitamin A (VA) to target the aHSCs and carried the autophagy inhibitor hydroxychloroquine (HCQ). The experimental results in vitro and in vivo revealed that the constructed aHSC-targeted hybrid delivery system HCQ@VA-Lip-Exo combined with the benefits of HCQ and exosomes derived from bone marrow mesenchymal stem cells. HCQ@VA-Lip-Exo had good aHSC-targeted delivery ability, effective autophagy inhibition, and synergistical anti-liver fibrosis performance, thus reducing the production and deposition of the extracellular matrix to inhibit the liver fibrosis. This combined strategy provided a potential idea for the construction and clinical application of a two-membrane hybrid delivery system as an effective targeted therapy of liver fibrosis.

Bone targeted miRNA delivery system for miR-34a with enhanced anti-tumor efficacy to bone-associated metastatic breast cancer.

Bone metastatic cancer is the most common occurrence in breast cancer, and the treatment is also facing great challenges. MicroRNA-34a (miRNA-34a) is a promising anti-cancer miRNA for gene therapy to bone metastatic cancer patients. However, the lack of specificity to bone and low accumulation at the site of bone tumor remains the major challenge when used bone-associated tumor. To solve this problem, a bone-targeted vector for delivery of miR-34a to bone metastatic breast cancer was constructed by using the commonly used gene vector branched polyethylenimine 25 k (BPEI 25 k) as the skeleton and linking with alendronate (ALN) moieties for bone targeting group. The constructed gene delivery system PCA/miR-34a can efficiently prevent miR-34a from degradation during blood circulation and enhance the specific bone delivery and distribution. PCA/miR-34a nanoparticles can be uptake into tumor cells through clathrin and caveolae-mediated endocytosis, and directly regulate the expression of oncogenes, thus promoting tumor cell apoptosis and relieving bone tissue erosion. The results of experiments in vitro and in vivo confirmed that the constructed bone-targeted miRNA delivery system PCA/miR-34a can enhance the anti-tumor efficacy in bone metastatic cancer, and provide a potential strategy for gene therapy in bone metastatic cancer.

The regulatory role and mechanism of autophagy in energy metabolism-related hepatic fibrosis.

Hepatic fibrosis is a key pathological process of chronic liver diseases, caused by alcohol, toxic and aberrant energy metabolism. It progresses to cirrhosis or even hepatic carcinoma without effective treatment. Studies have shown that autophagy has important regulatory effects on hepatic stellate cells (HSCs) energy metabolism, and then affect the activation state of HSCs. Autophagy maintains hepatic energy homeostasis, and the dysregulation of autophagy can lead to the activation of HSCs and the occurrence and development of hepatic fibrosis. It is necessary to explore the mechanism of autophagy in energy metabolism-related hepatic fibrosis. Herein, the current study summarizes the regulating mechanisms of autophagy through different targets and signal pathways in energy metabolism-related hepatic fibrosis, and discusses the regulatory effect of autophagy by natural plant-derived, endogenous and synthetic compounds for the treatment of hepatic fibrosis. A better comprehension of autophagy in hepatic stellate cells energy metabolism-related hepatic fibrosis may provide effective intervention of hepatic fibrosis, explore the potential clinical strategies and promote the drug treatment of hepatic fibrosis.

Sustainable and Clean Utilization of Yellow Phosphorus Slag (YPS): Activation and Preparation of Granular Rice Fertilizer.

Yellow phosphorus slag (YPS) is a typical industrial solid waste, while it contains abundant silicon micronutrient required for the growth of rice. The key scientific problem to use the YPS as rice fertilizer is how to activate the slag efficiently during the phosphorite reduction smelting process. In this work, an alkaline rice fertilizer from the activated YPS was successfully prepared to use the micronutrients. Thermodynamic analyses of SiO2-CaO, SiO2-CaO-Al2O3, and SiO2-CaO-Al2O3-MgO systems were discussed to optimize the acidity for reduction smelting. Results showed that the reduction smelting followed by the water quenching process can realize the reduction of phosphorite and activation of YPS synchronously. Ternary acidity m(SiO2)/(m(CaO) + m(MgO)) of 0.92 is suitable for the reduction smelting and activation of the slag. After smelting, the molten YPS can be effectively activated by water quenching, and 78.28% P, 90.03% Ca, and 77.12% Si in the YPS are activated, which can be readily absorbed by the rice roots. Finally, high-strength granular rice fertilizers with a particle size of Φ2-4 mm were successfully prepared from the powdery nitrogen-phosphorus-potassium (NPK) and activated YPS mixture.

20S-Protopanaxatriol Ameliorates Hepatic Fibrosis, Potentially Involving FXR-Mediated Inflammatory Signaling Cascades.

Ginseng has been used as a functional food and tonic for enhancing immune power. Here, the potential protective effect of 20S-protopanaxatriol (M4), the metabolite of protopanaxatriol, against hepatic fibrosis is investigated, which could provide nutritional interventions for disease treatment. M4 could inhibit extracellular matrix (ECM) deposition and reduce the levels of proinflammatory cytokines such as caspase 1, interleukin 1 ß (IL-1ß), interleukin 1 receptor type 1 (IL1R1), and interleukin 6 (IL-6). M4 also significantly increased the expression of farnesoid X receptor (FXR), suppressed the purinergic ligand-gated ion channel 7 receptor (P2X7r) signaling pathway, and works as an FXR agonist, GW4064. In thioacetamide (TAA)-induced mice, M4 could attenuate the histopathological changes and significantly regulate the expression levels of FXR and P2X7r. M4 ameliorated TAA-induced hepatic fibrosis due to the reduction of P2X7r secretion, inhibition of hepatic stellate cell (HSCs) activation, and inflammation, which were all associated with FXR activation. Hence, M4 might be useful a nutritional preventive approach in antihepatic fibrosis and antihepatic inflammation.

Acanthoic acid modulates lipogenesis in nonalcoholic fatty liver disease via FXR/LXRs-dependent manner.

Acanthoic acid (AA) is a pimaradiene diterpene isolated from Acanthopanax koreanum Nakai (Araliaceae), with anti-inflammatory and hepatic-protective effects. The present study intended to reveal the effect and mechanism of AA on nonalcoholic fatty liver disease (NAFLD) associated with lipid accumulation by activating Farnesoid X receptor (FXR) and liver X receptors (LXRs) signaling. C57BL/6 mice were received a modified Lieber-DeCarli diet with 71% high-fat (L-D) and treated with AA (20 and 40 mg/kg) or equal volume of saline for 12 weeks. The regulation of AA on lipid accumulation was also detected in pro-steatotic stimulated AML12 cells with palmitic acid (PA). When L-D diet-fed mice were treated with AA, loss in body weight, liver index, and liver lipid droplet were observed along with reduced triglyceride (TG) and serum transaminase. Furthermore, AA decreased sterol regulatory element binding protein 1 (SREBP-1) and target genes expression, regulated PPARα and PPARγ expressions, ameliorated hepatic fibrosis markers, enhanced hepatic FXR and LXR, and regulated AMPK-LKB1 and SIRT1 signaling pathway. Moreover, AA attenuated lipid accumulation via FXR and LXR activation in steatotic AML-12 cells, which was confirmed by guggulsterones (FXR antagonist) or GW3965 (LXR agonist). Activation of FXR and LXR signaling caused by AA might increase AMPK-SIRT1 signaling and then contribute to modulating lipid accumulation and fatty acid synthesis, which suggested that activated FXR-LXR axis by AA represented an effective strategy for relieving NAFLD.

Ginsenoside 25-OCH3-PPD Promotes Activity of LXRs To Ameliorate P2X7R-Mediated NLRP3 Inflammasome in the Development of Hepatic Fibrosis.

Ginseng is widely used in energy drinks, dietary supplements, and herbal medicines, and its pharmacological actions are related with energy metabolism. As an important modulating energy metabolism pathway, liver X receptors (LXRs) can promote the resolving of hepatic fibrosis and inflammation. The present study aims to evaluate the regulation of 25-OCH3-PPD, a ginsenoside isolated from Panax ginseng, against hepatic fibrosis and inflammation in thioacetamide (TAA)-stimulated mice by activating the LXRs pathway. 25-OCH3-PPD decreases serum ALT/AST levels and improves the histological pathology of liver in TAA-induced mice; attenuates transcripts of pro-fibrogenic markers associated with hepatic stellate cell activation; attenuates the levels of pro-Inflammatory cytokines and blocks apoptosis happened in liver; inhibits NLRP3 inflammasome by affecting P2X7R activation; and regulates PI3K/Akt and LKB1/AMPK-SIRT1. 25-OCH3-PPD also facilitates LX25Rs and FXR activities decreased by TAA stimulation. 25-OCH3-PPD also decreases α-SMA via regulation of LXRs and P2X7R-NLRP3 in vitro. Our data suggest the possibility that 25-OCH3-PPD promotes activity of LXRs to ameliorate P2X7R-mediated NLRP3 inflammasome in the development of hepatic fibrosis.

Acanthoic acid suppresses lipin1/2 via TLR4 and IRAK4 signalling pathways in EtOH- and lipopolysaccharide-induced hepatic lipogenesis.

OBJECTIVES: In alcoholic liver disease, alcohol and lipopolysaccharide (LPS) are major stimulation factors of hepatic lipogenesis. Our objective was to determine the protective mechanism of acanthoic acid (AA) in EtOH- and LPS-induced hepatic lipogenesis. METHODS: HSC-T6 cells were treated with ethanol (200 mm) plus LPS (1 µg/ml) for 1 h, followed by AA (10 or 20 µm) for another 6 h. C57BL/6 mice were pretreated with of AA (20 and 40 mg/kg) or equal volume of saline and then exposed to three doses of ethanol (5 g/kg body weight) within 24 h. The mice were sacrificed at 6 h after the last ethanol dosing. KEY FINDINGS: Acanthoic acid significantly decreased the expressions of α-SMA, collagen-I, SREBP-1, and lipin1/2 induced, also decreased fat droplets caused by EtOH/LPS. AA treatment decreased the protein expressions of TLR4, CD14, IRAK4, TRAF3, p-TAK1 and NF-κB increased by EtOH/LPS on HSC cells. Results in vivo were consistent with results in vitro. CONCLUSIONS: Our data demonstrated that AA might modulate hepatic fibrosis and lipid deposition in HSC-T6 cell stimulated with ethanol combined with LPS by decreasing lipin1/2 via TLR4 and IRAK4 signalling pathways, and AA might be considered as a potential therapeutic candidate for alcoholic liver disease.