Journal: Cellular and Molecular Immunology

Article Title: Hepatocyte-derived LRG1 primes the liver for metastasis and impairs immunotherapy

doi: 10.1038/s41423-026-01408-9

Figure Lengend Snippet: Tumor-associated inflammation promotes the expression of LRG1 in hepatocytes by IL6/STAT3. A The relative expression of Lrg1 in AML12 cells co-cultured with various mouse tumor cell lines or treated with serum (5%) from CRC model mice. n = 3 independent experiments. Data are means ± SD. B Western blot analysis of the expression LRG1 and β-actin in AML12 cells after the indicated treatments. C Cytokine array analyses of serum from CRC orthotopic mice model (day 7, day 14, day 21 and day 28) and from CRC intrasplenic mice model (day 5, day 10, day 15 and day 21). Quantitative real-time PCR analyses of the expression Lrg1 D and ELISA analyses of media of AML12 E treated with vehicle or recombinant IL6/G-CSF/CXCL13/CCL12/TIMP1. n = 3 independent experiments. Data are means ± SD. F, G qRT-PCR and Western blot analyses of LRG1 in AML12 cells treated with cytokine combinations. n = 3 independent experiments. Data are means ± SD. H, I qRT-PCR and western blot analysis of the expression LRG1 and β-actin in AML12 cells treated with indicated serum and anti-IL6 or tocilizumab. n = 3 independent experiments. Data are means ± SD. J ELISA analyses of serum samples for IL6 from sham-group, CRC orthotopic model at day 21(PMN) and CRC intrasplenic model at day 21(Met) in BALB/c. n = 4. Data are means ± SD. K Correlation between serological IL6 and LRG1 levels of CRC patients is shown using Pearson’s correlation analysis. Dots represent individual samples. n = 161. L, M Schematic of the experimental design L . Representative images of liver metastases M in each group ( n = 6 in Lrg1 (+/+)Hep-HTVi-Ctrl group and Lrg1 (+/+)Hep-HTVi-IL6 group, n = 5 in Lrg1 (Δ/Δ)Hep-HTVi-Ctrl group and Lrg1 (Δ/Δ)Hep-HTVi-IL6 group). Scale bars, 1 cm. N Schematic of the experimental design. Representative bioluminescence images and analyses of liver metastases in CRC mouse model treated with anti-IL6 or IgG. O Schematic of the experimental design. P ELISA analyses of samples for IL6 levels from liver interstitial fluid, peripheral blood and tumor interstitial fluid of mice from different groups as indicated. n = 4. Data are means ± SD. Q ELISA analyses of samples for LRG1 levels from liver interstitial fluid and peripheral blood of mice from different groups as indicated. n = 4. Data are means ± SD. R Western blot analyses of LRG1 and β-actin in hepatocyte of mice from different groups as indicated. Statistical significance was determined using two-tailed unpaired Student’s t test A, D–F, H, and N–P

Article Snippet: ELISA kits were used to measure the levels of human LRG1 (Ray Bio), human IL6 (Boster, EK0410), mouse LRG1 (ELK Biotechnology), and mouse IL6 (Boster, EK0411) in cell culture supernatants or serum samples according to the manufacturer’s instructions.

Techniques: Expressing, Cell Culture, Western Blot, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Recombinant, Quantitative RT-PCR, Two Tailed Test