Hinokitiol ameliorates MASH in mice by therapeutic targeting of hepatic Nrf2 and inhibiting hepatocyte ferroptosis.

Metabolic dysfunction-associated steatohepatitis (MASH), an advanced stage of metabolic dysfunction-associated steatotic liver disease (MASLD), still lacks approved effective clinical drugs. Ferroptosis, a form of regulated cell death driven by excessive iron accumulation and uncontrollable lipid peroxidation, has been proven to be a trigger of inflammation and initiation of steatohepatitis. The pathogenic interplay is modulated by oxidative stress, while the Nrf2-mediated antioxidant response plays a regulatory role in ferroptosis. Phytochemical hinokitiol (Hino) has demonstrated positive efficacy in hepatocellular carcinoma (HCC) in the reported work, but it remains unknown whether its therapeutic effect attributes to delaying the progress of steatohepatitis to HCC. This work aimed to systemically investigate the significance of ferroptosis in the pathogenesis of MASH and to demonstrate that Hino, a bioactive monoterpene compound, attenuates the primary pathological characteristics of MASH via promotion of Nrf2/GPX4 signaling. In this work, a MASH model was established using the high-fat/high-cholesterol (HFHC) diet-fed in vivo and palmitic acid/oleic acid (PO)-stimulated hepatocytes in vitro. Biochemical indexes, pathological analysis, western blot, PCR assay, energy metabolic phenotype, molecular docking, and confirmatory assays were performed comprehensively to reveal the key link between the Nrf2/GPX4 axis and the treatment of MASH. Under MASH conditions with increased oxidative stress, we show that Nrf2 was remarkable downregulated in HFHC diet-fed mice and PO-managed hepatocytes. Mechanistically, hepatic upregulation of Nrf2 through phytochemical Hino supplementation inhibited ferroptosis, enhanced lipid metabolism, and thereby alleviated hepatic steatosis, inflammation, and fibrosis. Conversely, silencing Nrf2 in hepatocytes further promoted the accumulation of key markers of ferroptosis and aggravated MASH phenotypes. Increased ferroptosis promoted steatosis which further drove inflammation and hepatic fibrosis. Our results suggested the significance of Nrf2 in ameliorating MASH, which was regulated through Hino. Thus, targeted inhibition of ferroptosis through Hino administration is a feasible and effective approach for treating MASH.

Yin X ，Liu Z ，Li C ，Wang J ... -  《-》

ACMSD inhibition corrects fibrosis, inflammation, and DNA damage in MASLD/MASH.

Recent findings reveal the importance of tryptophan-initiated de novo nicotinamide adenine dinucleotide (NAD+) synthesis in the liver, a process previously considered secondary to biosynthesis from nicotinamide. The enzyme α-amino-β-carboxymuconate-ε-semialdehyde decarboxylase (ACMSD), primarily expressed in the liver and kidney, acts as a modulator of de novo NAD+ synthesis. Boosting NAD+ levels has previously demonstrated remarkable metabolic benefits in mouse models. In this study, we aimed to investigate the therapeutic implications of ACMSD inhibition in the treatment of metabolic dysfunction-associated steatotic liver disease/steatohepatitis (MASLD/MASH). In vitro experiments were conducted in primary rodent hepatocytes, Huh7 human liver carcinoma cells and induced pluripotent stem cell-derived human liver organoids (HLOs). C57BL/6J male mice were fed a western-style diet and housed at thermoneutrality to recapitulate key aspects of MASLD/MASH. Pharmacological ACMSD inhibition was given therapeutically, following disease onset. HLO models of steatohepatitis were used to assess the DNA damage responses to ACMSD inhibition in human contexts. Inhibiting ACMSD with a novel specific pharmacological inhibitor promotes de novo NAD+ synthesis and reduces DNA damage ex vivo, in vivo, and in HLO models. In mouse models of MASLD/MASH, de novo NAD+ biosynthesis is suppressed, and transcriptomic DNA damage signatures correlate with disease severity; in humans, Mendelian randomization-based genetic analysis suggests a notable impact of genomic stress on liver disease susceptibility. Therapeutic inhibition of ACMSD in mice increases liver NAD+ and reverses MASLD/MASH, mitigating fibrosis, inflammation, and DNA damage, as observed in HLO models of steatohepatitis. Our findings highlight the benefits of ACMSD inhibition in enhancing hepatic NAD+ levels and enabling genomic protection, underscoring its therapeutic potential in MASLD/MASH. Enhancing NAD+ levels has been shown to induce remarkable health benefits in mouse models of metabolic dysfunction-associated steatotic liver disease/steatohepatitis (MASLD/MASH), yet liver-specific NAD+ boosting strategies remain underexplored. Here, we present a novel pharmacological approach to enhance de novo synthesis of NAD+ in the liver by inhibiting α-amino-β-carboxymuconate-ε-semialdehyde decarboxylase (ACMSD), an enzyme highly expressed in the liver. Inhibiting ACMSD increases NAD+ levels, enhances mitochondrial respiration, and maintains genomic stability in hepatocytes ex vivo and in vivo. These molecular benefits prevent disease progression in both mouse and human liver organoid models of steatohepatitis. Our preclinical study identifies ACMSD as a promising target for MASLD/MASH management and lays the groundwork for developing ACMSD inhibitors as a clinical treatment.

Liu YJ ，Kimura M ，Li X ，Sulc J ，Wang Q ，Rodríguez-López S ，Scantlebery AML ，Strotjohann K ，Gallart-Ayala H ，Vijayakumar A ，Myers RP ，Ivanisevic J ，Houtkooper RH ，Subramanian GM ，Takebe T ，Auwerx J ... -  《-》

Radish red attenuates chronic kidney disease in obese mice through repressing oxidative stress and ferroptosis via Nrf2 signaling improvement.

Chronic kidney disease (CKD) presents a significant public health concern, with obesity being a prominent contributing factor to kidney disorders by inducing oxidative stress, lipotoxicity, and tubular cell injury. Natural anthocyanins extracted from red radishes (Raphanus sativus L.) exert antioxidant and anti-apoptotic functions. This study aims to employ a novel natural pigment anthocyanin, referred to as radish red (RR) isolated from red radishes, to alleviate obesity-related metabolic disturbances and kidney impairment in a CKD mouse model induced by high-fat and high-fructose diets (HFFD). The in vitro study initially demonstrated that RR treatment significantly mitigated the palmitate acid (PA)-induced injury and cytotoxicity in human tubular epithelial HK2 cells. Subsequently, RR supplementation notably improved obesity and associated metabolic dysfunctions in mice caused by HFFD. Abnormal renal function indices including serum creatinine, blood urea nitrogen (BUN), uric acid (UA), urine protein, albuminuria and urine albumin-to-creatinine ratio (UACR) were detected in HFFD-fed mice, which were effectively alleviated by RR treatment. Histologically, renal tubular cell injury, lipid deposition, tubular dilatation, and renal fibrosis induced by HFFD were markedly improved after RR administration in mice. Furthermore, RR treatment significantly alleviated oxidative stress in HFFD-fed mice, as evidenced by the decreased renal reactive oxygen species (ROS) production, 4-HNE, and NOX4 expression levels. Anti-oxidants such as superoxide dismutase-1 (SOD1), NAD (P) H: quinone oxidoreductase (NQO1), heme oxygenase-1 (HO-1) and glutamate cysteine ligase (GCLC) were highly upregulated in kidney of HFFD-fed mice with RR consumption through improving NFE2-related factor 2 (Nrf2) signaling activation. Furthermore, ferroptosis was identified in the kidneys of HFFD-fed mice, evidenced by the elevated levels of malondialdehyde (MDA), iron content, and lipid peroxidation, along with the decreased expression of glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11). These occurrences were significantly mitigated following RR treatment. Mechanistically, we further discovered that the suppressive effects of RR in restricting oxidative stress, ferroptosis, lipid accumulation, and injury of tubular epithelial cells induced by PA were significantly counteracted by Nrf2 knockdown. Collectively, our results demonstrated that dietary supplementation with RR could potentially serve as an efficacious therapeutic modality for the management of obesity-related CKD progression by enhancing Nrf2 activation to impede oxidative stress and ferroptosis.

Li Q ，Zheng Y ，Zhao J ，Wei X ，Shi Z ，Fan H ，Ge C ，Xu M ，Tan J ... -  《-》

GTPBP8 mitigates nonalcoholic steatohepatitis (NASH) by depressing hepatic oxidative stress and mitochondrial dysfunction via PGC-1α signaling.

Nonalcoholic steatohepatitis (NASH) is emerging as a major cause of liver transplantation and hepatocellular carcinoma (HCC). Regrettably, its pathological mechanisms are still not fully comprehended. GTP-binding protein 8 (GTPBP8), belonging to the GTP-binding protein superfamily, assumes a crucial role in RNA metabolism, cell proliferation, differentiation, and signal transduction. Its aberrant expression is associated with oxidative stress and mitochondrial dysfunctions. Nevertheless, its specific functions and mechanisms of action, particularly in NASH, remain elusive. In our current study, we initially discovered that human hepatocytes L02 displayed evident mitochondrial respiratory anomaly, mitochondrial damage, and dysfunction upon treatment with palmitic acids and oleic acids (PO), accompanied by significantly reduced GTPBP8 expression levels through RNA-Seq, RT-qPCR, western blotting, and immunofluorescence assays. We then demonstrated that GTPBP8 overexpression mediated by adenovirus vector (Ad-GTPBP8) markedly attenuate lipid accumulation, inflammatory response, and mitochondrial impair and dysfunction in hepatocytes stimulated by PO. Conversely, adenovirus vector-mediated GTPBP8 knockdown (Ad-shGTPBP8) significantly accelerated lipid deposition, inflammation and mitochondrial damage in PO-treated hepatocytes in vitro. Furthermore, we constructed an in vivo NASH murine model by giving a 16-week high fat high cholesterol diet (HFHC) diet to hepatocyte specific GTPBP8-knockout (GTPBP8HKO) mice. We firstly found that HFHC feeding led to metabolic disorder in mice, including high body weight, blood glucose and insulin levels, and liver dysfunctions, which were accelerated in these NASH mice with GTPBP8 deficiency in hepatocytes. Consistently, GTPBP8HKO remarkably exacerbated the progression of NASH phenotypes induced by HFHC, as proved by the anabatic lipid accumulation, inflammation, fibrosis and reactive oxygen species (ROS) production in liver tissues, which could be largely attributed to the severe mitochondrial damage and dysfunction. Mechanistically, we further identified that GTPBP8 interacted with peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) in hepatocytes. Importantly, the hepaprotective effects of GTPBP8 against mitochondrial dysfunction, oxidative stress and inflammation was largely dependent on PGC-1α expression. Collectively, GTPBP8 may exert a protective role in the progression of NASH, and targeting the GTPBP8/PGC-1α axis may represent a potential strategy for NASH treatment by improving mitochondrial functions.

Meng D ，Chang M ，Dai X ，Kuang Q ，Wang G ... -  《-》

Protocatechuic acid relieves ferroptosis in hepatic lipotoxicity and steatosis via regulating NRF2 signaling pathway.

Ferroptosis represents a newly programmed cell death, and the process is usually accompanied with iron-dependent lipid peroxidation. Importantly, ferroptosis is implicated in a myriad of diseases. Recent literature suggests a potential position of ferroptosis in the pathogenesis of metabolic dysfunction-associated fatty liver disease (MAFLD), the most widespread liver ailment worldwide. Intriguingly, several functional genes and metabolic pathways central to ferroptosis are regulated by nuclear factor erythroid-derived 2-like 2 (NRF2). In current work, we aim to identify protocatechuic acid (PCA), a primary metabolite of antioxidant polyphenols, as a potent NRF2 activator and ferroptosis inhibitor in the hepatic lipotoxicity and steatosis models. Herein, both NRF2+/+ and NRF2-/- cell lines and mice were used to analyze the importance of NRF2 in PCA function, and hepatic lipotoxicity and steatosis models were induced by palmitic acid and high-fat diet respectively. Our results indicated that ferroptosis was mitigated by PCA intervention in hepatic cells. Furthermore, PCA exhibited therapeutic efficacy against ferroptosis, as well as hepatic lipotoxicity and steatosis. The protective role of PCA was predominantly mediated through NRF2 activation, potentially elucidating a pivotal mechanism underlying PCA's therapeutic impact on MAFLD. Additionally, the augmented mitochondrial TCA cycle activity observed in hepatic lipotoxicity and steatosis models was ameliorated by PCA, in part via NRF2-dependent pathways, further bolstering PCA's anti-ferroptosis properties. Collectively, our findings underscore PCA's potential in alleviating hepatic ferroptosis, lipotoxicity and steatosis via inducing activation of NRF2 signaling pathway, offering a promising strategy for the therapy of MAFLD as well as related lipid metabolic disorders.

Feng Y ，Shi M ，Zhang Y ，Li X ，Yan L ，Xu J ，Liu C ，Li M ，Bai F ，Yuan F ，Sun Y ，Liu R ，Zhao Y ，Yang L ，Zhang Y ，Guo Y ，Zhang J ，Zhou R ，Liu P ... -  《-》