Mouse Liver Research Articles

Helicobacter pylori infection exacerbates metabolic dysfunction-associated steatotic liver disease through lipid metabolic pathways: a transcriptomic study

BackgroundThe relationship between Helicobacter pylori (H. pylori) infection and metabolic dysfunction-associated steatotic liver disease (MASLD) has attracted increased clinical attention. However, most of those current studies involve cross-sectional studies and meta-analyses, and experimental mechanistic exploration still needs to be improved. This study aimed to investigate the mechanisms by which H. pylori impacts MASLD.MethodsWe established two H. pylori-infected (Cag A positive and Cag A negative) mouse models with 16 weeks of chow diet (CD) or high-fat diet (HFD) feeding. Body weight, liver triglyceride, blood glucose, serum biochemical parameters, inflammatory factors, and insulin resistance were measured, and histological analysis of liver tissues was performed. Mouse livers were subjected to transcriptome RNA sequencing analysis.ResultsAlthough H. pylori infection could not significantly affect serum inflammatory factor levels and serum biochemical parameters in mice, serum insulin and homeostatic model assessment for insulin resistance levels increased in CD mode. In contrast, H. pylori Cag A + infection significantly aggravated hepatic pathological steatosis induced by HFD and elevated serum inflammatory factors and lipid metabolism parameters. Hepatic transcriptomic analysis in the CD groups revealed 767 differentially expressed genes (DEGs) in the H. pylori Cag A + infected group and 1473 DEGs in the H. pylori Cag A- infected group, and the “nonalcoholic fatty liver disease” pathway was significantly enriched in KEGG analysis. There were 578 DEGs in H. pylori Cag A + infection combined with the HFD feeding group and 820 DEGs in the H. pylori Cag A- infected group. DEGs in the HFD groups were significantly enriched in “fatty acid degradation” and “PPAR pathway.” Exploring the effect of different Cag A statuses on mouse liver revealed that fatty acid binding protein 5 was differentially expressed in Cag A- H. pylori. DEG enrichment pathways were concentrated in the “PPAR pathway” and “fatty acid degradation.”ConclusionsClinicians are expected to comprehend the impact of H. pylori on MASLD and better understand and manage MASLD. H. pylori infection may exacerbate the development of MASLD by regulating hepatic lipid metabolism, and the H. pylori virulence factor Cag A plays a vital role in this regulation.

Gene body DNA hydroxymethylation restricts the magnitude of transcriptional changes during aging

DNA hydroxymethylation (5hmC), the most abundant oxidative derivative of DNA methylation, is typically enriched at enhancers and gene bodies of transcriptionally active and tissue-specific genes. Although aberrant genomic 5hmC has been implicated in age-related diseases, its functional role in aging remains unknown. Here, using mouse liver and cerebellum as model organs, we show that 5hmC accumulates in gene bodies associated with tissue-specific function and restricts the magnitude of gene expression changes with age. Mechanistically, 5hmC decreases the binding of splicing associated factors and correlates with age-related alternative splicing events. We found that various age-related contexts, such as prolonged quiescence and senescence, drive the accumulation of 5hmC with age. We provide evidence that this age-related transcriptionally restrictive function is conserved in mouse and human tissues. Our findings reveal that 5hmC regulates tissue-specific function and may play a role in longevity.

Galectin 3-binding protein (LGALS3BP) depletion attenuates hepatic fibrosis by reducing transforming growth factor-β1 (TGF-β1) availability and inhibits hepatocarcinogenesis.

Increased Galectin 3-binding protein (LGALS3BP) serum levels have been used to assess hepatic fibrosis stages and the severity of hepatocellular carcinoma (HCC). Considering the crucial role of transforming growth factor-β1 (TGF-β1) in the emergence of these diseases, the present study tested the hypothesis that LGALS3BP regulates the TGF-β1 signaling pathway. The expression levels of LGALS3BP and TGFB1 were analyzed in patients with metabolic dysfunction-associated steatohepatitis (MASH) and HCC. Multiple omics techniques, such as RNA-sequencing, transposase-accessible chromatin-sequencing assay, and liquid chromatography-tandem mass spectrometry proteomics, were used to identify the regulatory mechanisms for the LGALS3BP-TGF-β1 axis. The effects of altered TGF-β1 signaling by LGALS3BP were investigated in conditional LGALS3BP-knockin and LGALS3BP-knockout mice. In patients with MASH and HCC, the levels of LGALS3BP and TGFB1 exhibited positive correlations. Stimulation of LGALS3BP by the inflammatory cytokine interferon α in HCC cells or ectopic overexpression of LGALS3BP in hepatocytes promoted the expression levels of TGFB1. Aggravated fibrosis was observed in the livers of hepatocyte-specific LGALS3BP-knockin mice, with increased TGFB1 levels. LGALS3BP directly bound to and assembled integrin αV, an integral mediator required for releasing active TGF-β1 from extracellular latent complex with the rearranged F-actin cytoskeleton. The released TGF-β1 activated JunB transcription factor, which in turn promoted the TGF-β1 positive feedback loop. LGALS3BP deletion in the hepatocytes downregulated TGF-β1 signaling and CCl4 induced fibrosis. Moreover, LGALS3BP depletion hindered hepatocarcinogenesis by limiting the availability of fibrogenic TGF-β1. LGALS3BP plays a crucial role in hepatic fibrosis and carcinogenesis by controlling the TGF-β1 signaling pathway, making it a promising therapeutic target in TGF-β1-related diseases.

Molecular exploration of the diurnal alteration of glycogen structural fragility and stability in time-restricted-feeding mouse liver

The structure of glycogen α particles in healthy mouse liver has two states: stability and fragility. In contrast, glycogen α particles in diabetic liver present consistent fragility, which may exacerbate hyperglycemia. Currently, the molecular mechanism behind glycogen structural alteration is still unclear. In this study, we characterized the fine molecular structure of liver glycogen α particles in healthy mice under time-restricted feeding (TRF) mode during a 24-h cycle. Then, differentially expressed genes (DEGs) in the liver during daytime and nighttime were revealed via transcriptomics, which identified that the key downregulated DEGs were mainly related to insulin secretion in daytime. Furthermore, GO annotation and KEGG pathway enrichment found that negative regulation of the glycogen catabolic process and insulin secretion process were significantly downregulated in the daytime. Therefore, transcriptomic analyses indicated that the structural stability of glycogen α particles might be correlated with the glycogen degradation process via insulin secretion downregulation. Further molecular experiments confirmed the significant upregulation of glycogen phosphorylase (PYGL), phosphorylated PYGL (p-PYGL), and glycogen debranching enzyme (AGL) at the protein level during the daytime. Overall, we concluded that the downregulation of insulin secretion in the daytime under TRF mode facilitated glycogenolysis, contributing to the structural stability of glycogen α-particles.

Jiawei Taohe Chengqi Decoction attenuates CCl4 induced hepatic fibrosis by inhibiting HSCs activation via TGF-β1/CUGBP1 and IFN-γ/Smad7 pathway

BackgroundHepatic fibrosis (HF) is an essential stage in the progression of different chronic liver conditions to cirrhosis and even hepatocellular carcinoma. The activation of hepatic stellate cells (HSCs) plays a crucial role in the progression of HF. IFN- γ/Smad7 pathway can inhibit HSCs activation, while TGF-β1/CUGBP1 pathway can inhibit IFN-γ/Smad7 pathway transduction and promote HSCs activation. Thus, inhibiting the TGF-β1/CUGBP1 pathway and activating the IFN-γ/Smad7 pathway reverses HSCs activation and inhibits HF. Jiawei Taohe Chengqi Decoction (JTCD) was derived from the Taohe Chengqi Tang in the ancient Chinese medical text titled “Treatise on Febrile Diseases”. We found several anti-HF components in JTCD including ginsenoside Rb1 and others, but the specific mechanism of anti-HF in JTCD is not clear. PurposeTo elucidate the specific mechanism by which JTCD reverses HF by inhibiting the activation of HSCs, and to establish a scientific foundation for treating HF with Traditional Chinese medicine (TCM). MethodsWe constructed a CCl4-induced mice HF model in vivo and activated human hepatic stellate cell line (LX-2) with TGF-β1 in vitro, after which they were treated with JTCD and the corresponding inhibitors. We examined the expression of pivotal molecules in the two pathways mentioned above by immunofluorescence staining, Western blotting and RT-PCR. ResultsJTCD attenuated liver injury and reduced serum ALT and AST levels in mice. In addition, JTCD attenuated CCl4-induced HF by decreasing the expression of α-SMA, COL1A1 and other markers of HSCs activation in mice liver tissue. Moreover, JTCD effectively suppressed the levels of TGF-β1, p-Smad3, p-p38MAPK, p-ATF2, and CUGBP1 in vivo and in vitro and upregulated the levels of IFN-γ, p-STAT1, and Smad7. Mechanically, after using the inhibitors of both pathways in vitro, we found that JTCD inhibited the activation of HSCs by restoring the balance of the TGF-β1/CUGBP1 and IFN-γ/Smad7 pathways. ConclusionWe demonstrated that JTCD inhibited HSCs activation and reversed HF by inhibiting the TGF-β1/CUGBP1 signalling pathway and upregulating the IFN-γ/Smad7 signalling pathway. Moreover, we have identified specific links where JTCD interferes with both pathways to inhibit HSCs activation. JTCD is an effective candidate for the clinical treatment of HF.

The role of C5aR1-mediated hepatic macrophage efferocytosis in NASH

Non-alcoholic fatty liver disease (NAFLD) has become the first major chronic liver disease in developed countries. 10–20% of NAFLD patients will progress to non-alcoholic steatohepatitis (NASH), and up to 25% of NASH patients may develop cirrhosis within 10 years. Therefore, it is critical to find key targets that may treat this disease. Here, we identified C5aR1 as a highly-expressed gene in NASH mouse model through analyzing Gene Expression Omnibus (GEO) database and confirmed its higher expression in livers of NASH patients than that of NAFL patients. Meanwhile, we verified its positive correlation with patients’ serum alanine transaminase (ALT) and aspartate transaminase (AST) levels. In vivo and in vitro experiments revealed that knocking down C5aR1 in liver significantly reduced liver weight ratio and serum ALT and AST levels and attenuated inflammatory cell infiltration and cell apoptosis in the liver of NASH mice as well as enhanced the efferocytotic ability of liver macrophages, suggesting that C5aR1 may play a crucial role in the efferocytosis of liver macrophages. Furthermore, we also found that the expression levels of nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing protein 3 (NLRP3), caspase-1, IL-1β and other inflammation-related factors in the liver were significantly reduced. Our work demonstrates a potential mechanism of how C5aR1 deficiency protects against diet-induced NASH by coordinating the regulation of inflammatory factors and affecting hepatic macrophage efferocytosis.

Effect of Placental Derived Nucleoproteins on liver regeneration in DEN-induced liver fibrosis model

BackgroundPlacental Derived Nucleoproteins (PDNs) is commonly associated with the process of angiogenesis, and doesn't affect the healthy vasculature. PDNs are clinically estimated for the treatment of cancer cases and severe hepatic injuries. Thus, the pathophysiological effects of PDNs targeting liver fibrosis is a concern. ObjectivesTo assess the molecular, histopathological, and chemical impact of PDNs on liver regeneration in Diethylnitrosamine (DEN)-induced mice liver fibrosis. MethodsNormal untreated reference group of ten mice and two groups of induced liver cirrhosis using the recommended weekly dose of Diethylnitrosamine in total of eleven doses, initially 20 mg/kg body weight, and then 30 mg/kg in the third week, followed by 50 mg/kg for the last eight weeks, one of them combined treatment aligned with injection with total dose of extracted PDNs 25 mg/kg, in comparison to PDNs only treated group. An autopsy was performed after 22 weeks of the initial dose of DEN in each group. Molecular characterization of Alpha smooth muscle actin, TGFβ and NF-κB biomarkers for liver then liver function panel were analyzed and finally hepatopathological changes were observed using H&E stain and Sirius red stain. ResultsLiver enzymes, total bilirubin and total proteins in tissue in PDNs-DEN treated models were controlled in the direction of normal group and 50 % reduction of fibrosis in comparing to DEN-treated models. The cellular arrangement of fibrosis in the DEN entire groups were differentiated with high significant impact on the survival of mice. Increased levels of the biochemical markers in liver homogenate, loss of tissue architecture, and proliferation were observed in induced groups and down regulation of alpha smooth muscle actin, TGFβ and NF-κB. ConclusionThis finding demonstrates an improvement of Liver tissue induced fibrosis using DEN combined with PDNs. This strategy is to generate an animal model with a lower occurrence of fibrosis in a short time treatment regarding liver regeneration.

Effect of electroacupuncture of "Zusanli" (ST36) combined with capeOX on apoptosis and ferroptosis in nude mice with colorectal cancer.

To investigate the impact of combined treatment of colorectal cancer (CRC) with electroacupuncture (EA) and capeOX (combined administration of fluorouracil, oxaliplatin and capecitabine) on the tumor volume, weight, spleen coefficient, apoptosis and ferroptosis of tumor tissue, and liver and kidney functions in nude mice with CRC, so as to explore its mechanisms underlying inhibiting CRC and alleviating toxic reactions of capeOX. Female Balb/c nude mice were randomly assigned to 3 groups：model, capeOX, and EA+capeOX, with 8 nude mice in each group. The CRC model was established by subcutaneous injection of colon cancer cells at the right inguinal region. Nude mice of the capeOX group received intraperitoneal injection of oxaliplatin for 1 day and gavage of capecitabine from day 2 to day 7. EA (1 mA, 2 Hz/100 Hz) was applied to bilateral "Zusanli" (ST36) for 20 min, once daily for 7 days. During the interven-tion, the tumor volume and weight were measured every day, and at the end of intervention, the weight of the tumor tissue and spleen were measured, with tumor volume difference and spleen coefficient calculated. The proportion of apoptotic cells was measured by flow cytometry, and the contents of serum malondialdehyde (MDA), alanine aninotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), and creatinine (Cr) were detected using ELISA. The expression level of glutathione peroxidase 4 (GPX4, a key regulator for ferroptosis) protein of the tumor tissue was determined using Western blot. Compared to the model group, both the capeOX group and EA+capeOX group showed a decrease in the tumor volume (on day 3 and 4 in the capeOX group, and from day 2 to 7 in the EA+capeOX group) and body weight (P<0.05, on day 3 to 7 in the EA+capeOX group and on day 2 to 7 in the capeOX group), being evidently lower in the tumor volume on day 7 in the EA+capeOX than in the capeOX group (P<0.05), and evidently higher in the body weight on day 6 and 7 in the EA+capeOX group than in the capeOX group (P<0.05). In comparison with the model group, the tumor volume difference, tumor weight and spleen coefficient in both capeOX and EA+capeOX groups were significantly decreased (P<0.05), and MDA content in EA+capeOX group was significantly decreased (P<0.05), while the contents of ALT, BUN and Cr in the capeOX group, the proportion of apoptotic cells in both capeOX and EA+capeOX groups, and the GPX4 expression level in the EA+capeOX group were all significantly increased (P<0.05). The tumor volume difference, tumor weight, and contents of MDA, ALT, AST, BUN and Cr in the EA+capeOX group were markedly lower than in the capeOX group (P<0.05), while the spleen coefficient, proportion of apoptotic cells and GPX4 expression level in the EA+capeOX group were markedly higher than those in the capeOX group (P<0.05). EA of ST36 can enhance the effect of capeOX in inhibiting colorectal cancer growth in nude mice with CRC, which may be related with its functions in promoting tumor cell apoptosis, inhibiting ferroptosis, and modulating immune tolerance. In addition, EA can lower the side effects of capeOX in hematopoietic and immune, liver, and kidney functions.

Rutin, a natural flavonoid glycoside, ameliorates zearalenone induced liver inflammation via inhibiting lipopolysaccharide gut leakage and NF-κB signaling pathway in mice

Zearalenone (ZEN) poses a potential threat on human and animal health partly through the nuclear factor (NF)-κB signaling pathway. In silico study suggested that rutin effective against TLR4 and NF-κB. A wetting test was designed to evaluate the effect and underlying mechanism of rutin in alleviating ZEN-induced inflammation in animals. Twenty-four female mice were randomly divided into 4 groups: control (basal diet), ZEN group (basal diet + ZEN), rutin group (basic diet + rutin), Z + R group (basal diet + rutin + ZEN). Results showed that rutin effectively alleviated ZEN-induced inflammation and damage of liver and jejunum in mice. Rutin addition reduced the content of lipopolysaccharide (LPS) in serum and liver mainly by improving the intestinal barrier function resulted from the production increase of short-chain fatty acids (SCFA). In sum, this study showed that rutin alleviated ZEN-induced liver inflammation and injury by modulating the gut microbiota, increasing the production of SCFA and improving intestinal barrier function, leading to the decrease of LPS in liver and the inhibition of MyD88 independent NF-κB signaling pathway in mice. Specifically, these findings may provide useful insights into the screening of functional natural compounds and its action mechanism to alleviate ZEN induced liver inflammation.

Circular RNA RRM2 alleviates metabolic dysfunction-associated steatotic liver disease by targeting miR-142-5p to increase NRG1 expression

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a prevalent chronic liver condition worldwide, demanding further investigation into its pathogenesis. Circular RNAs (circRNAs) are emerging as pivotal regulators in MASLD processes, yet their pathological implications in MASLD remain poorly understood. This study focused on elucidating the role of circular RNA ribonucleotide reductase subunit M2 (circRRM2) in MASLD progression. In this study, we used both in vitro and in vivo MASLD models, utilizing long-chain-free fatty acid (FFA)-treated hepatocytes and high-fat diet (HFD)-induced MASLD in mice, respectively. We determined the expression patterns of circRRM2, microRNA-142-5p (miR-142-5p), and NRG1 in livers of MASLD-afflicted mice and MASLD hepatocytes by RT-qPCR. Dual-luciferase reporter assays verified the binding relationships among circRRM2, miR-142-5p, and NRG1. We conducted further analyses of their roles in MASLD hepatocytes and modulated circRRM2, miR-142-5p, and NRG1 expression in vitro by transfection. Our findings were validated in vivo. The results demonstrated reduced levels of circRRM2 and NRG1, along with elevated miR-142-5p expression in MASLD livers and hepatocytes. Overexpression of circRRM2 downregulated lipogenesis-related genes and decreased triglycerides accumulation in livers of MASLD mice. MiR-142-5p, which interacts with circRRM2, effectively counteracted the effects of circRRM2 in MASLD hepatocytes. Furthermore, NRG1 was identified as a miR-142-5p target, and its overexpression mitigated the regulatory impact of miR-142-5p on MASLD hepatocytes. In conclusion, circRRM2, via its role as a miR-142-5p sponge, upregulating NRG1, possibly influenced triglycerides accumulation in both in vitro and in vivo MASLD models.

Fabrication of a Near-Infrared-Emissive Probe for Detecting Dipeptidyl Peptidase 4 in the Liver of Diabetic Mice and Clinical Serum.

Dipeptidyl peptidase 4 (DPP4) plays a key role in glucose metabolism, which has been a close target for diabetes pathology and treatment. It is significant for the evaluation of cellular DPP4 activity in various biological systems. Fluorescence imaging technology is currently a popular method for detecting enzymes in living cells due to its advantages of high selectivity, high sensitivity, high spatiotemporal resolution, and real-time visualization. Herein, a near-infrared (NIR)-emissive probe NEDP with a large Stokes shift (153 nm) was developed for the assay of DPP4 activity. Upon addition of DPP4, NEDP can emit a significant turn-on NIR fluorescence signal (673 nm) with high sensitivity and specificity. Moreover, NEDP can successfully be used for imaging of intracellular DPP4, confirming the regulation of DPP4 expression in hyperglucose and its treatment in living cells. Most importantly, NEDP can not only monitor the changes of DPP4 in vivo but also show that DPP4 in diabetes is mainly up-regulated in the liver, and the level of DPP4 is positively correlated with the pathological damage of the liver. In addition, NEDP can identify the serum of diabetic patients from healthy people through the fluorescence response to DPP4. These results demonstrated that the designed probe NEDP provides a prospective visual tool to explore the relationship between DPP4 and diabetes and would be applied for detecting serum of diabetes in the clinic.

Bioanalysis of free antisense oligonucleotide payload from antibody-oligonucleotide conjugate by hybridization LC-MS/MS.

Background: Antisense oligonucleotides (ASOs) have been conjugated to various moieties, such as peptides, antibodies or Fab regions of antibodies, to enhance their delivery to target tissues. The quantitation of free ASO (ASO payload) is critical to characterize its pharmacokinetics/pharmacodynamics (PK/PD) properties and biodistribution after delivery of the peptide/antibody/Fab ASO conjugates. Results: We developed a hybridization-based LC-MS/MS methodology for quantification of free ASO in tissues in the presence of Fab-ASO and ASO with linker (ASO-linker). Conclusion: The developed method was applied to measure accurately the free ASO concentrations in liver and gastrocnemius in mice that were dosed with Fab-ASO. This methodology has also been applied to free ASO bioanalysis for other antibody-ASO and Fab-ASO conjugates in various tissues and plasma/serum samples.

PACAP ameliorates obesity-induced insulin resistance through FAIM/Rictor/AKT axis.

Obesity and obesity-related insulin resistance have been a research hotspot. Pituitary adenylate cyclase activating polypeptide (PACAP) has emerged as playing a significant role in energy metabolism, holding promising potential for attenuating insulin resistance. However, the precise mechanism is not fully understood. Palmitic acid and a high-fat diet (HFD) were used to establish insulin resistance model in Alpha mouse liver 12 cell line and C57BL/6 mice, respectively. Subsequently, we assessed the effects of PACAP both in vivo and in vitro. Lentivirus vectors were used to explore the signaling pathway through which PACAP may ameliorate insulin resistance. PACAP was found to selectively bind to the PACAP type I receptor receptor and ameliorate insulin resistance, which was characterized by increased glycogen synthesis and the suppression of gluconeogenesis in the insulin-resistant cell model and HFD-fed mice. These effects were linked to the activation of the Fas apoptotic inhibitory molecule/rapamycin-insensitive companion of mammalian target of rapamycin/RAC-alpha serine/threonine-protein kinase (FAIM/Rictor/AKT) axis. Furthermore, PACAP ameliorated insulin resistance by increasing solute carrier family 2, facilitated glucose transporter members 2/4 and inhibiting gluconeogenesis-related proteins glucose 6-phosphatase catalytic subunit 1 and phosphoenolpyruvate carboxykinase 2 expression. Meanwhile, the phosphorylation of hepatic AKT/glycogen synthase kinase 3β was promoted both in vivo and in vitro by PACAP. Additionally, PACAP treatment decreased body weight, food intake and blood glucose levels in obese mice. Our study shows that PACAP ameliorated insulin resistance through the FAIM/Rictor/AKT axis, presenting it as a promising drug candidate for the treatment of obesity-related insulin resistance.

Exploring the Anti-inflammatory and Immunomodulatory Potential of Licochalcone B against Psoralidin-Induced Liver Injury

Ethnopharmacological relevanceHerb-induced liver injury (HILI) represents an exacerbated inflammatory response, with Psoraleae fructus (PF) and its preparations recently associated with hepatotoxicity. Licorice, historically recognized as a detoxifying herbal remedy, is considered to possess hepatoprotective properties. Our previous research identified bavachin, bakuchiol, and psoralidin (PSO) as potential toxic constituents in PF, while licochalcone B (LCB) and echinatin were identified as bioactive components in licorice. However, evidence regarding the interactions of active compounds in herbs and their underlying mechanisms remains limited. Aim of the studyThe objective of this study is to assess the potential mechanisms through which LCB modulates immunological and anti-inflammatory responses to treat PSO-induced liver injury by using human hepatocyte cells (L02) and LPS-primed mice. MethodsThe ameliorative effects of LCB and echinatin on bavachin, bakuchiol, and PSO-induced liver injury were demonstrated in L02 cells. Subsequently, the efficacy of LCB on PSO-induced idiosyncratic liver injury was further validated in C57BL/6 mice under moderate inflammatory stress induced by LPS priming. The mechanisms were preliminarily explored with an integrated strategy of molecular docking, RT-PCR verification, and untargeted metabolomics. ResultsThe study shows that LCB significantly reduced cell injury induced by the three chemicals in PF and provided substantial protection against PSO-induced hepatic damage, as indicated by the levels of ALT, AST, and LDH. LCB normalized liver function and remarkedly alleviated hepatic lesions and inflammation caused by PSO in mice under moderate inflammatory stress. The mRNA profiles of both L02 cells and mice liver tissue revealed that LCB mitigated PSO-induced hepatotoxicity by regulating the gene expression of pro-inflammatory cytokines IL1B and TNF, as well as immunoinflammatory genes PIK3CA, AKT1, NFKB1, and NLRP3. Furthermore, untargeted metabolomics of liver tissue indicated that LCB could reverse the abnormal expression of 11 discriminatory metabolites, with the interrelationship between differential metabolites and target genes primarily clustering in glycerophospholipid metabolism, arachidonic acid metabolism, and phosphatidylinositol signaling system. ConclusionLCB demonstrated a superior anti-inflammatory and immunomodulatory effect on PSO-induced hepatotoxicity by modulating the inflammatory response and metabolic signaling system. Key interactive targets included phosphatidylcholine, phosphatidic acid, and subunit isoforms of PI3K.

Liver microRNA transcriptome reveals miR-182 as link between type 2 diabetes and fatty liver disease in obesity

Background:The development of obesity-associated comorbidities such as type 2 diabetes (T2D) and hepatic steatosis has been linked to selected microRNAs in individual studies; however, an unbiased genome-wide approach to map T2D induced changes in the miRNAs landscape in human liver samples, and a subsequent robust identification and validation of target genes are still missing.Methods:Liver biopsies from age- and gender-matched obese individuals with (n=20) or without (n=20) T2D were used for microRNA microarray analysis. The candidate microRNA and target genes were validated in 85 human liver samples, and subsequently mechanistically characterized in hepatic cells as well as by dietary interventions and hepatic overexpression in mice.Results:Here, we present the human hepatic microRNA transcriptome of type 2 diabetes in liver biopsies and use a novel seed prediction tool to robustly identify microRNA target genes, which were then validated in a unique cohort of 85 human livers. Subsequent mouse studies identified a distinct signature of T2D-associated miRNAs, partly conserved in both species. Of those, human-murine miR-182–5 p was the most associated with whole-body glucose homeostasis and hepatic lipid metabolism. Its target gene LRP6 was consistently lower expressed in livers of obese T2D humans and mice as well as under conditions of miR-182–5 p overexpression. Weight loss in obese mice decreased hepatic miR-182–5 p and restored Lrp6 expression and other miR-182–5 p target genes. Hepatic overexpression of miR-182–5 p in mice rapidly decreased LRP6 protein levels and increased liver triglycerides and fasting insulin under obesogenic conditions after only seven days.Conclusions:By mapping the hepatic miRNA-transcriptome of type 2 diabetic obese subjects, validating conserved miRNAs in diet-induced mice, and establishing a novel miRNA prediction tool, we provide a robust and unique resource that will pave the way for future studies in the field. As proof of concept, we revealed that the repression of LRP6 by miR-182–5 p, which promotes lipogenesis and impairs glucose homeostasis, provides a novel mechanistic link between T2D and non-alcoholic fatty liver disease, and demonstrate in vivo that miR-182–5 p can serve as a future drug target for the treatment of obesity-driven hepatic steatosis.Funding:This work was supported by research funding from the Deutsche Forschungsgemeinschaft (KI 1887/2-1, KI 1887/2-2, KI 1887/3-1 and CRC-TR296), the European Research Council (ERC, CoG Yoyo LepReSens no. 101002247; PTP), the Helmholtz Association (Initiative and Networking Fund International Helmholtz Research School for Diabetes; MB) and the German Center for Diabetes Research (DZD Next Grant 82DZD09D1G).

Liver microRNA transcriptome reveals miR-182 as link between type 2 diabetes and fatty liver disease in obesity.

The development of obesity-associated comorbidities such as type 2 diabetes (T2D) and hepatic steatosis has been linked to selected microRNAs in individual studies; however, an unbiased genome-wide approach to map T2D induced changes in the miRNAs landscape in human liver samples, and a subsequent robust identification and validation of target genes are still missing. Liver biopsies from age- and gender-matched obese individuals with (n=20) or without (n=20) T2D were used for microRNA microarray analysis. The candidate microRNA and target genes were validated in 85 human liver samples, and subsequently mechanistically characterized in hepatic cells as well as by dietary interventions and hepatic overexpression in mice. Here, we present the human hepatic microRNA transcriptome of type 2 diabetes in liver biopsies and use a novel seed prediction tool to robustly identify microRNA target genes, which were then validated in a unique cohort of 85 human livers. Subsequent mouse studies identified a distinct signature of T2D-associated miRNAs, partly conserved in both species. Of those, human-murine miR-182-5 p was the most associated with whole-body glucose homeostasis and hepatic lipid metabolism. Its target gene LRP6 was consistently lower expressed in livers of obese T2D humans and mice as well as under conditions of miR-182-5 p overexpression. Weight loss in obese mice decreased hepatic miR-182-5 p and restored Lrp6 expression and other miR-182-5 p target genes. Hepatic overexpression of miR-182-5 p in mice rapidly decreased LRP6 protein levels and increased liver triglycerides and fasting insulin under obesogenic conditions after only seven days. By mapping the hepatic miRNA-transcriptome of type 2 diabetic obese subjects, validating conserved miRNAs in diet-induced mice, and establishing a novel miRNA prediction tool, we provide a robust and unique resource that will pave the way for future studies in the field. As proof of concept, we revealed that the repression of LRP6 by miR-182-5 p, which promotes lipogenesis and impairs glucose homeostasis, provides a novel mechanistic link between T2D and non-alcoholic fatty liver disease, and demonstrate in vivo that miR-182-5 p can serve as a future drug target for the treatment of obesity-driven hepatic steatosis. This work was supported by research funding from the Deutsche Forschungsgemeinschaft (KI 1887/2-1, KI 1887/2-2, KI 1887/3-1 and CRC-TR296), the European Research Council (ERC, CoG Yoyo LepReSens no. 101002247; PTP), the Helmholtz Association (Initiative and Networking Fund International Helmholtz Research School for Diabetes; MB) and the German Center for Diabetes Research (DZD Next Grant 82DZD09D1G).

Glutathione synthesis in the mouse liver supports lipid abundance through NRF2 repression

Cells rely on antioxidants to survive. The most abundant antioxidant is glutathione (GSH). The synthesis of GSH is non-redundantly controlled by the glutamate-cysteine ligase catalytic subunit (GCLC). GSH imbalance is implicated in many diseases, but the requirement for GSH in adult tissues is unclear. To interrogate this, we have developed a series of in vivo models to induce Gclc deletion in adult animals. We find that GSH is essential to lipid abundance in vivo. GSH levels are highest in liver tissue, which is also a hub for lipid production. While the loss of GSH does not cause liver failure, it decreases lipogenic enzyme expression, circulating triglyceride levels, and fat stores. Mechanistically, we find that GSH promotes lipid abundance by repressing NRF2, a transcription factor induced by oxidative stress. These studies identify GSH as a fulcrum in the liver’s balance of redox buffering and triglyceride production.

Apolipoprotein H deficiency exacerbates alcohol-induced liver injury via gut Dysbiosis and altered bile acid metabolism

BackgroundAPOH plays an essential role in lipid metabolism and the transport of lipids in the circulation. Previous studies have shown that APOH deficiency causes fatty liver and gut microbiota dysbiosis in mouse models. However, the role and potential mechanisms of APOH deficiency in the pathogenesis of alcoholic liver disease remain unclear. MethodsC57BL/6 WT and ApoH−/− mice were used to construct the binge-on-chronic alcohol feeding model. Mouse liver transcriptome, targeted bile acid metabolome, and 16S gut bacterial taxa were assayed and analyzed. Open-source human liver transcriptome dataset was analyzed. ResultsApoH−/− mice fed with alcohol showed severe hepatic steatosis. Liver RNAseq and RT-qPCR data indicated that APOH deficiency predominantly impacts hepatic lipid metabolism by disrupting de novo lipogenesis, cholesterol processing, and bile acid metabolism. A targeted bile acid metabolomics assay indicated significant changes in bile acid composition, including increased percentages of TCA in the liver and DCA in the gut of alcohol-fed ApoH−/− mice. The concentrations of CA, NorCA, and HCA in the liver were higher in ApoH−/− mice on an ethanol diet compared to the control mice (p < 0.05). Additionally, APOH deficiency altered the composition of gut flora, which correlated with changes in the liver bile acid composition in the ethanol-feeding mouse model. Finally, open-source transcript-level data from human ALD livers highlighted a remarkable link between APOH downregulation and steatohepatitis, as well as bile acid metabolism. ConclusionAPOH deficiency aggravates alcohol induced hepatic steatosis through the disruption of gut microbiota homeostasis and bile acid metabolism in mice.

Leaf extract of Kentut (Paederia foetida L.) as a preventive measure against interleukin-6 expression in the liver of mice in a sepsis model injected with Escherichia coli

Kentut leaves (Paederia foetida L.) are a type of medicinal plant that can be used as a preventative medicine against sepsis. This plant contains secondary metabolite compounds such as alkaloids, flavonoids, triterpenoids, saponins, and other active compounds. This research aims to determine the influence and effective dosage of Kentut leaf extract as a preventive measure against IL-6 expression in the livers of mice in a sepsis model injected with E. coli. The method used was a Completely Randomized Design (CRD). The study involved 24 white male mice divided into 6 groups. Data analysis was performed using One Way ANOVA. The average values of IL-6 expression in the mouse livers for each group are as follows: KN at 7.09%±0.06; K+ at 26.36%±0.02; K- at 72.60%±0.05; PI (100mg/kgBW) at 71.04%±0.04; PII (300mg/kgBW) at 62.22%±0.02; and PIII (500mg/kgBW) at 40.92%±0.01. The research results indicate that there is an influence of kentut leaf extract as a preventive measure against IL-6 expression in the livers of mice in the sepsis model injected with E. coli, with a significance value of 0.000 or p-value 0.005. The effective dosage of kentut leaf extract as a preventive measure against IL-6 expression is the PIII dosage of 500mg/kgBW. The anti-inflammatory mechanism in sepsis is thought to be caused by the presence of flavonoids, alkaloids, phenolic acids, and terpenoid compounds. The most likely anti-inflammatory mechanism is believed to involve flavonoids inhibiting cyclooxygenase (COX) and lipoxygenase (LOX) enzymes involved in the synthesis of inflammatory mediators such as prostaglandins and leukotrienes, which can trigger IL-6 production.

New findings about neuropathological outcomes in the PKU mouse throughout lifespan

Phenylketonuria (PKU, OMIM 261600) is a genetic disorder caused by a deficiency of the hepatic enzyme phenylalanine hydroxylase (PAH). If left untreated, PKU leads to systemic phenylalanine (Phe) accumulation, which can result in irreversible brain damage and intellectual disabilities. In the last 60 years, early and strict dietary restriction of phenylalanine (Phe) intake proved to prevent the severe clinical phenotype of untreated PKU. While the specific mechanisms through which phenylalanine causes brain damage are still poorly understood, preclinical models have been deeply explored to characterize the neurotoxic effect of Phe on neurodevelopmental processes. At the same time, that on the aging brain still needs to be explored. In the brain of untreated PAHEnu2(−/−) mouse, we previously reported a reduction of myelin basic protein (MBP) during postnatal development up to 60 PND. Later in the diseased mouse's life, a spontaneous and persistent restoration of MBP was detected. In this present longitudinal study, ranging from 14 to 540 post-natal days (PND) of untreated PAHEnu2(−/−) mice, we further investigated: a) the long-life consistency of two Phe-related brain metabolic alterations, such as large neutral amino acids (LNAA) and biogenic amine neurotransmitters' depletion; b) the outcome of locomotor functions during the same life span; c) the integrity of myelin as assessed ex vivo by central (hippocampus) and peripheral (extensor digitorum longus-sciatic nerve) action potential conduction velocities. In contrast with the results of other studies, brain Leu, Ile, and Val concentrations were not significantly altered in the brain PAHEnu2(−/−) mouse. On the other hand, 3-O-Methyldopa (3-OMD, a biomarker of L-DOPA), serotonin, and its associated metabolites were reduced throughout most of the considered time points, with consistent reductions observed prevalently from 14 to 60 PND. Normal saltatory conduction was restored after 60 PND and remained normal at the last examination at 360 PND, resulting nonetheless in a persistent locomotor impairment throughout a lifetime. These new findings contribute to laying the foundations for the preclinical characterization of aging in PKU, confirming neurotransmitter defects as consistent metabolic traits. LNAAs have a minor role, if any, in brain damage pathogenesis. Transient myelin synthesis failure may impact brain connectivity during postnatal development but not nervous signal conduction.