Liraglutide Alleviates Hepatic Steatosis by Activating the TFEB-Regulated Autophagy-Lysosomal Pathway.
TL;DR: The findings suggest that liraglutide attenuated hepatic steatosis via restoring autophagic flux, specifically the GLP-1R-TFEB-mediated autophagy-lysosomal pathway.
Abstract: Liraglutide, a glucagon-like peptide-1 receptor agonist (GLP-1RA), has been demonstrated to alleviate non-alcoholic fatty liver disease (NAFLD). However, the underlying mechanism has not been fully elucidated. Increasing evidence suggests that autophagy is involved in the pathogenesis of hepatic steatosis. In this study, we examined whether liraglutide could alleviate hepatic steatosis through autophagy-dependent lipid degradation and investigated the underlying mechanisms. Herein, the effects of liraglutide on NAFLD were evaluated in a high-fat diet (HFD)-induced mouse model of NAFLD as well as in mouse primary and HepG2 hepatocytes exposed to palmitic acid (PA). The expression of the GLP-1 receptor (GLP-1R) was measured in vivo and in vitro. Oil red O staining was performed to detect lipid accumulation in hepatocytes. Electron microscopy was used to observe the morphology of autophagic vesicles and autolysosomes. Autophagic flux activity was measured by infecting HepG2 cells with mRFP-GFP-LC3 adenovirus. The roles of GLP-1R and transcription factor EB (TFEB) in autophagy-lysosomal activation were explored using small interfering RNA. Liraglutide treatment alleviated hepatic steatosis in vivo and in vitro. In models of hepatic steatosis, microtubule-associated protein 1B light chain-3-II (LC3-II) and SQSTM1/P62 levels were elevated in parallel to blockade of autophagic flux. Liraglutide treatment restored autophagic activity by improving lysosomal function. Furthermore, treatment with autophagy inhibitor chloroquine weakened liraglutide-induced autophagy activation and lipid degradation. TFEB has been identified as a key regulator of lysosome biogenesis and autophagy. The protein levels of nuclear TFEB and its downstream targets CTSB and LAMP1 were decreased in hepatocytes treated with PA, and these decreases were reversed by liraglutide treatment. Knockdown of TFEB expression compromised the effects of liraglutide on lysosome biogenesis and hepatic lipid accumulation. Mechanistically, GLP-1R expression was decreased in HFD mouse livers as well as PA-stimulated hepatocytes, and liraglutide treatment reversed the downregulation of GLP-1R expression in vivo and in vitro. Moreover, GLP-1R inhibition could mimic the effect of the TFEB downregulation-mediated decrease in lysosome biogenesis. Thus, our findings suggest that liraglutide attenuated hepatic steatosis via restoring autophagic flux, specifically the GLP-1R-TFEB-mediated autophagy-lysosomal pathway.
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TL;DR: The role of autophagy in the pathogenesis of metabolic diseases associated with or occurring in the context of ageing, including insulin resistance, T2DM and sarcopenic obesity, was discussed in this article.
Abstract: Autophagy is an evolutionarily conserved, lysosome-dependent catabolic process whereby cytoplasmic components, including damaged organelles, protein aggregates and lipid droplets, are degraded and their components recycled. Autophagy has an essential role in maintaining cellular homeostasis in response to intracellular stress; however, the efficiency of autophagy declines with age and overnutrition can interfere with the autophagic process. Therefore, conditions such as sarcopenic obesity, insulin resistance and type 2 diabetes mellitus (T2DM) that are characterized by metabolic derangement and intracellular stresses (including oxidative stress, inflammation and endoplasmic reticulum stress) also involve the accumulation of damaged cellular components. These conditions are prevalent in ageing populations. For example, sarcopenia is an age-related loss of skeletal muscle mass and strength that is involved in the pathogenesis of both insulin resistance and T2DM, particularly in elderly people. Impairment of autophagy results in further aggravation of diabetes-related metabolic derangements in insulin target tissues, including the liver, skeletal muscle and adipose tissue, as well as in pancreatic β-cells. This Review summarizes the role of autophagy in the pathogenesis of metabolic diseases associated with or occurring in the context of ageing, including insulin resistance, T2DM and sarcopenic obesity, and describes its potential as a therapeutic target. The cellular consequences of dysfunctional autophagy contribute to numerous diseases. In this Review, Kitada and Koya consider the relationship between impaired autophagy and age-related metabolic derangements, including insulin resistance, type 2 diabetes mellitus and sarcopenic obesity, and discuss candidate autophagy-based therapies.
109 citations
TL;DR: In this paper, the authors reviewed the therapeutic effects and possible mechanisms of action of GLP-1RAs in the nervous, cardiovascular, and endocrine systems and their correlation with metabolism, tumours and other diseases.
Abstract: Glucagon like peptide-1 (GLP-1) is an incretin secretory molecule. GLP-1 receptor agonists (GLP-1RAs) are widely used in the treatment of type 2 diabetes (T2DM) due to their attributes such as body weight loss, protection of islet β cells, promotion of islet β cell proliferation and minimal side effects. Studies have found that GLP-1R is widely distributed on pancreatic and other tissues and has multiple biological effects, such as reducing neuroinflammation, promoting nerve growth, improving heart function, suppressing appetite, delaying gastric emptying, regulating blood lipid metabolism and reducing fat deposition. Moreover, GLP-1RAs have neuroprotective, anti-infectious, cardiovascular protective, and metabolic regulatory effects, exhibiting good application prospects. Growing attention has been paid to the relationship between GLP-1RAs and tumorigenesis, development and prognosis in patient with T2DM. Here, we reviewed the therapeutic effects and possible mechanisms of action of GLP-1RAs in the nervous, cardiovascular, and endocrine systems and their correlation with metabolism, tumours and other diseases.
49 citations
TL;DR: In this article, the authors identify the underlying pathways contributing to enhanced autophagic flux and identify the RAPGEF4/EPAC2-calcium-PPP3/calcineurin-TFEB axis as a key mediator of autophagous flux, lysosomal function and cell survival in pancreatic β-cells.
Abstract: Macroautophagy/autophagy is critical for the regulation of pancreatic β-cell mass and its deregulation has been implicated in the pathogenesis of type 2 diabetes (T2D). We have previously shown that treatment of pancreatic β-cells with the GLP1R (glucagon like peptide 1 receptor) agonist exendin-4 stimulates autophagic flux in a setting of chronic nutrient excess. The aim of this study was to identify the underlying pathways contributing to enhanced autophagic flux.Pancreatic β-cells (INS-1E),mouse and human islets were treated with glucolipotoxic stress (0.5 mM palmitate and 25 mM glucose) in the presence of exendin-4. Consistent with our previous work, exendin-4 stimulated autophagic flux. Using chemical inhibitors and siRNA knockdown, we identified RAPGEF4/EPAC2 (Rap guanine nucleotide exchange factor 4) and downstream calcium signaling to be essential for regulation of autophagic flux by exendin-4. This pathway was independent of AMPK and MTOR signaling. Further analysis identified PPP3/calcineurin and its downstream regulator TFEB (transcription factor EB) as key proteins mediating exendin-4 induced autophagy. Importantly, inhibition of this pathway prevented exendin-4-mediated cell survival and overexpression of TFEB mimicked the cell protective effects of exendin-4 in INS-1E and human islets. Moreover, treatment of db/db mice with exendin-4 for 21 days increased the expression of lysosomal markers within the pancreatic islets. Collectively our data identify the RAPGEF4/EPAC2-calcium-PPP3/calcineurin-TFEB axis as a key mediator of autophagic flux, lysosomal function and cell survival in pancreatic β-cells. Pharmacological modulation of this axis may offer a novel therapeutic target for the treatment of T2D.Abbreviations: AKT1/protein kinase B: AKT serine/threonine kinase 1; AMPK: 5' AMP-activated protein kinase; CAMKK: calcium/calmodulin-dependent protein kinase kinase; cAMP: cyclic adenosine monophosphate; CASP3: caspase 3; CREB: cAMP response element-binding protein; CTSD: cathepsin D; Ex4: exendin-4(1-39); GLP-1: glucagon like peptide 1; GLP1R: glucagon like peptide 1 receptor; GLT: glucolipotoxicity; INS: insulin; MTOR: mechanistic target of rapamycin kinase; NFAT: nuclear factor of activated T-cells; PPP3/calcineurin: protein phosphatase 3; PRKA/PKA: protein kinase cAMP activated; RAPGEF3/EPAC1: Rap guanine nucleotide exchange factor 3; RAPGEF4/EPAC2: Rap guanine nucleotide exchange factor 4; SQSTM1/p62: sequestosome 1; T2D: type 2 diabetes; TFEB: transcription factor EB.
17 citations
TL;DR: In this paper, the authors investigated the transcriptional regulation of choline kinase alpha (Chka) in hepatocytes in the context of liver regeneration and found that Chka expression was significantly upregulated in the regenerating livers in the PHx model and the acetaminophen (APAP) injection model.
Abstract: Liver regeneration is a key compensatory process in response to liver injury serving to contain damages and to rescue liver functions. Hepatocytes, having temporarily exited the cell cycle after embryogenesis, resume proliferation to regenerate the injured liver parenchyma. In the present study we investigated the transcriptional regulation of choline kinase alpha (Chka) in hepatocytes in the context of liver regeneration. We report that Chka expression was significantly up-regulated in the regenerating livers in the partial hepatectomy (PHx) model and the acetaminophen (APAP) injection model. In addition, treatment with hepatocyte growth factor (HGF), a strong pro-proliferative cue, stimulated Chka expression in primary hepatocytes. Chka depletion attenuated HGF-induced proliferation of hepatocytes as evidenced by quantitative PCR and Western blotting measurements of pro-proliferative genes as well as EdU incorporation into replicating DNA. Of interest, deletion of Brahma-related gene 1 (Brg1), a chromatin remodeling protein, attenuated Chka induction in the regenerating livers in mice and in cultured hepatocytes. Further analysis revealed that Brg1 interacted with hypoxia-inducible factor 1 alpha (HIF-1α) to directly bind to the Chka promoter and activate Chka transcription. Finally, examination of human acute liver failure (ALF) specimens identified a positive correlation between Chka expression and Brg1 expression. In conclusion, our data suggest that Brg1-dependent trans-activation of Chka expression may contribute to liver regeneration.
14 citations
TL;DR: In this paper, the authors demonstrate that the activity of Transcription Factor EB (TFEB) is reduced in the liver of mice fed a high-fat diet and showed that TFEB acts as a mediator, linking metformin with autophagy to reverse NAFLD.
Abstract: Nonalcoholic fatty liver disease (NAFLD) results from an abnormal accumulation of lipids within hepatocytes, and is commonly associated with obesity, insulin resistance, and hyperlipidemia. Metformin is commonly used to treat type 2 diabetes mellitus and, in recent years, it was found to play a potential role in the amelioration of NAFLD. However, the mechanisms underlying the protective effect of metformin against NAFLD remain largely unknown. Transcription factor EB (TFEB) is a master transcriptional regulator of lysosomal biogenesis and autophagy and, when activated, is effective against disorders of lipid metabolism. However, the role of TFEB in hepatic steatosis is not well understood. In this report, we demonstrate that the activity of TFEB is reduced in the liver of mice fed a high-fat diet. Metformin treatment significantly reverses the activity of TFEB, and the protective effect of metformin against hepatic steatosis and insulin resistance is dependent on TFEB. We show that metformin-induced autophagy is regulated by TFEB, and our findings reveal that TFEB acts as a mediator, linking metformin with autophagy to reverse NAFLD, and highlight that TFEB may be a promising molecular target for the treatment of NAFLD.
13 citations
References
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TL;DR: This chapter discusses preparation of isolated rat liver cells by incubation of rat liver minces with pronase, which results in most of the liver parenchyma is digested, while nonparenchymal cells remain intact and can be recovered from the incubate.
Abstract: Publisher Summary This chapter discusses preparation of isolated rat liver cells The early mechanical and chemical methods for liver-cell preparation were relatively successful in converting liver tissue to a suspension of isolated cells The successful preparation of intact liver cells by perfusion with collagenase is technically quite difficult The major method for preparation of nonparenchymal liver cells is based on the selective sensitivity of parenchymal cells toward proteases By incubation of rat liver minces with pronase, most of the liver parenchyma is digested, while nonparenchymal cells remain intact and can be recovered from the incubate Similar results have been reported with trypsin digestion of collagenase-dispersed liver minces, but pronase appears to be more effective The most common procedure is to perfuse the liver briefly with pronase before it is minced and incubated with the enzyme Such direct pronase methods have been used by several investigators with yields of nonparenchymal liver cells reported to be in the range 2–15 × 10 6 cells/gm liver
5,285 citations
"Liraglutide Alleviates Hepatic Stea..." refers methods in this paper
...Murine primary hepatocytes were isolated from male C57BL/6J mice (8 weeks old) using the collagenase IV perfusion technique as previously described (Seglen, 1976; Klaunig et al., 1981)....
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TL;DR: A previously unknown function for autophagy in regulating intracellular lipid stores (macrolipophagy) is identified that could have important implications for human diseases with lipid over-accumulation such as those that comprise the metabolic syndrome.
Abstract: The intracellular storage and utilization of lipids are critical to maintain cellular energy homeostasis. During nutrient deprivation, cellular lipids stored as triglycerides in lipid droplets are hydrolysed into fatty acids for energy. A second cellular response to starvation is the induction of autophagy, which delivers intracellular proteins and organelles sequestered in double-membrane vesicles (autophagosomes) to lysosomes for degradation and use as an energy source. Lipolysis and autophagy share similarities in regulation and function but are not known to be interrelated. Here we show a previously unknown function for autophagy in regulating intracellular lipid stores (macrolipophagy). Lipid droplets and autophagic components associated during nutrient deprivation, and inhibition of autophagy in cultured hepatocytes and mouse liver increased triglyceride storage in lipid droplets. This study identifies a critical function for autophagy in lipid metabolism that could have important implications for human diseases with lipid over-accumulation such as those that comprise the metabolic syndrome.
3,091 citations
"Liraglutide Alleviates Hepatic Stea..." refers background in this paper
...Increasing evidence has demonstrated that autophagy is necessary for regulating lipid metabolism because lipid droplets are degraded through lipophagy (Singh et al., 2009; Cui et al., 2020)....
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TL;DR: Aliment Pharmacol Ther 2011; 34: 274–285
Abstract: SUMMARY Background Non-alcoholic fatty liver disease (NAFLD) is a common cause of chronic liver disease, and its worldwide prevalence continues to increase with the growing obesity epidemic. This study assesses the epidemiology of NAFLD in adults based on clinical literature published over the past 30 years. Aim To review epidemiology and natural history of non-alcoholic fatty liver disease and non-alcoholic steatohepatitis in adults based on clinical literature published over the past 30 years. Methods An in-depth search of PubMed (1980–2010) was based on five search terms: ‘nonalcoholic fatty liver disease’ OR ‘non-alcoholic steatohepatitis’ OR ‘fatty liver’ OR ‘steatosis’ AND ‘incidence’ [MeSH Terms] OR ‘prevalence’ [MeSH Terms] OR ‘natural history’. Studies of paediatric cohorts were excluded. Articles were categorised by topic and summarised, noting generalisations concerning their content.
2,679 citations
TL;DR: Recent advances in identifying and understanding the core molecular machinery and signaling pathways that are involved in mammalian autophagy are highlighted.
1,782 citations
TL;DR: Liraglutide was safe, well tolerated, and led to histological resolution of non-alcoholic steatohepatitis, warranting extensive, longer-term studies.
1,285 citations