Journal: Cell death & disease

Article Title: Cellular crosstalk mediated by Meteorin-like regulating hepatic stellate cell activation during hepatic fibrosis.

doi: 10.1038/s41419-025-07734-6

Figure Lengend Snippet: Fig. 4 Metrnl regulates HSCs activation in vitro. A The level of Metrnl in the cultured medium, secreted by LX-2 cells overexpressing Metrnl (OE-Met) and knockdown Metrnl (sh-Met) induced by adenovirus, was measured using ELISA (n = 3 per group). Representative Western blot images and quantification analysis of FN, Col4, and α-SMA expression in LX-2 cells transfected with adenovirus-mediated Metrnl knockdown (B), rMetrnl treatment (200 ng/mL) (C), and adenovirus-mediated Metrnl overexpression (D) combined with TGF-β (2 ng/mL) treatment for 48 h (n = 3 blots). E Volcano plot analysis was performed on the RNA sequencing results of the indicated samples. A stringent set of genes (p < 0.05) was identified in Metrnl overexpressing LX-2 cells compared to vector cells treated with TGF-β for 48 h (n = 3 per group). F Heat map of representative, differentially expressed genes associated with matrix remodeling, inflammation, and fibrosis is shown. The analysis includes genes involved in fibrosis-related signaling pathways (vector and OE-Met, both with TGF-β, n = 3 per group). G Changes in relative mRNA levels of genes related to matrix remodeling, inflammation, and fibrosis were observed in Metrnl overexpressing LX-2 cells. These mRNA levels were normalized to that of GAPDH (n = 3 per group). H Relative mRNA levels of genes related to fibrosis in primary HSCs isolated from C57BL/ 6 mice and treated with TGF-β (2 ng/mL) for 48 h. These mRNA levels were normalized to that of GAPDH (n = 3 per group). All data are mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001.

Article Snippet: The PDGFB and FN content of the medium supernatant from cells was measured using a commercially available Human PDGFB ELISA Kit (Proteintech; KE100161) and Human Fibronectin/FN1 ELISA Kit (BOSTER; EK0349) individually, following the manufacturer’s instructions.

Techniques: Activation Assay, In Vitro, Cell Culture, Knockdown, Enzyme-linked Immunosorbent Assay, Western Blot, Expressing, Transfection, Over Expression, RNA Sequencing, Plasmid Preparation, Protein-Protein interactions, Isolation

Journal: Cell death & disease

Article Title: Cellular crosstalk mediated by Meteorin-like regulating hepatic stellate cell activation during hepatic fibrosis.

doi: 10.1038/s41419-025-07734-6

Figure Lengend Snippet: Fig. 7 Metrnl regulates PDGFB release via EGR1 from hepatocytes to activate PDGFRβ signaling pathway. ELISA-quantified PDGFB levels in culture medium (CM) (A) and mRNA expression level (B) of primary hepatocytes transfected with adenovirus-mediated Metrnl overexpression (OE-Met) and primary hepatocytes from Alb-Metrnl−/−mice (n = 3 per group). The mRNA expression (C) and Western blot images (D) and its quantification analysis (down panel) of α-SMA and Col4 in LX-2 cells after incubating CM from LO2 cells transfected with OE-Metrnl virus, and treated with or without PDGFB neutralization antibody (PDGFB-Nabs) or control IgG, which was used to block PDGFB in the CM (n = 3 per group). E A Venn diagram created by the jvenn tool to display potential transcription factors of PDGFB predicted by TF- Target Finder. F A Venn diagram showing the intersection of seven potential transcription factors (CTCF, EGR1, et.al) with RNA-Seq data from Metrnl-overexpressing LX-2 cells. G The mRNA expression of EGR1 in primary hepatocytes from C57BL/6 mice transfected with adenovirus- mediated OE-Metrnl constructs (n = 5 per group). H The mRNA expression of EGR1 in the liver from CCl4-induced mice injected with AAV- vector or AAV-Metrnl virus (n = 6 per group). Representative Western blot images (I) and quantification analysis of EGR1, also the relative mRNA expression (J) in the liver from WT and Metrnl−/−mice induced by CCl4 for 8 weeks (n = 4 per group). ELISA-quantified PDGFB levels in CM (K) and the mRNA expression levels (L) of primary hepatocytes transfected with OE-Metrnl or OE-EGR1 overexpression virus, or both viruses combined (n = 3–4 per group). M RT-qPCR analysis of PDGFB mRNA levels with EGR1 antibody or IgG antibody respectively by ChIP assay in Metrnl-knockdown or control LO2 cells (n = 3 per group). All data are mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001.

Article Snippet: The PDGFB and FN content of the medium supernatant from cells was measured using a commercially available Human PDGFB ELISA Kit (Proteintech; KE100161) and Human Fibronectin/FN1 ELISA Kit (BOSTER; EK0349) individually, following the manufacturer’s instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Transfection, Over Expression, Western Blot, Virus, Neutralization, Control, Blocking Assay, RNA Sequencing, Construct, Injection, Plasmid Preparation, Quantitative RT-PCR, Knockdown

Journal: Cell death & disease

Article Title: Cellular crosstalk mediated by Meteorin-like regulating hepatic stellate cell activation during hepatic fibrosis.

doi: 10.1038/s41419-025-07734-6

Figure Lengend Snippet: Fig. 9 Schematic summary of the possible effect of Metrnl on HSC activation during liver fibrosis. Hepatic Metrnl expression was significantly reduced in the fibrotic liver. The deficiency of Metrnl had a dual effect, resulting in the acceleration of HSC activation. Specifically, the absence of Metrnl in hepatocytes promotes the release of fibrogenic cytokine PDGFB through upregulating transcription factor EGR1, which activates PDGFRβ signaling in HSCs through paracrine regulation. Additionally, the absence of Metrnl in hepatocytes and HSCs results in the downregulation of E3 ubiquitin ligase HECW2, inhibiting K48-linked ubiquitination of FN on proteasomal degradation, thus promoting FN secretion from HSCs. These effects contribute to the deposition of extracellular matrix (ECM) and HSCs activation, ultimately exacerbating liver fibrosis. These findings highlight Metrnl as a novel regulator of liver fibrosis that mediates communication between hepatocytes and HSCs. The mechanistic illustration was created with Adobe Illustrator.

Article Snippet: The PDGFB and FN content of the medium supernatant from cells was measured using a commercially available Human PDGFB ELISA Kit (Proteintech; KE100161) and Human Fibronectin/FN1 ELISA Kit (BOSTER; EK0349) individually, following the manufacturer’s instructions.

Techniques: Activation Assay, Expressing, Ubiquitin Proteomics