Journal: Life Metabolism

Article Title: Propionic acid secreted by Akkermansia muciniphila alleviates hepatic fibrosis by antioxidant regulation across the gut–liver axis

doi: 10.1093/lifemeta/loaf036

Figure Lengend Snippet: Propionic acid inhibits the activation of HSCs and oxidative stress. (a) Immunofluorescence analysis (Scale bar = 50 μm). (b) ROS visualized by microscopy using the 2’,7’-dichlorodihydrofluorescein diacetate (DCFH-DA) probe at a final concentration of 10 μmol/L (Scale bar = 50 μm). (c) Fibrosis-related markers such as TIMP-1, α-SMA, and Col1α1 and TGF-β1 protein expression assayed by western blotting. (d) Oxidative stress-related Keap1/Nrf2 pathway protein levels assayed by western blotting. (e–h) MDA level (e), CAT activity (f), SOD activity (g), and GSH-Px activity (h). (i) In vitro experiments conducted to assess the effects of TGF-β, GPR41 inhibitor (SHB), GPR43 inhibitor (GLPG0974), and propionic acid pretreatment on oxidative stress-related Keap1/Nrf2 signaling pathway in LX2 cells through immunoblot analysis. (j) Western blot analysis of oxidative stress-related Keap1/Nrf2 signaling pathway performed in LX2 cells transfected with sh GPR41 , followed by stimulation with TGF-β and pretreatment with propionic acid. Data are presented as the mean ± SEM. Statistical significance was determined using ANOVA for multiple-group comparisons. ** P < 0.01, *** P < 0.05, compared with the control group. # P < 0.05, ## P < 0.01, compared with the model group.

Article Snippet: The study utilized various reagents and kits, including sodium propionate (MedChemExpress, HY-B1773A), 4’,6-diamidino-2-phenylindole (DAPI; Thermo, D3571), MDA, CAT, SOD, and GSH-Px assay kits (Thermo, EEA015; Jianglai Biotechnology, JL18163, JL12237, JL-T0879, respectively), propionic acid ELISA kit (Zhuocai Biotechnology, ZC-56375-J), and antibodies such as Col1α1 (Abcam, RRID: AB_138492), α-SMA (Abcam, RRID: AB_5694), TGF-β1 (Proteintech, 21898-1-AP), SMAD2, p-SMAD2, SMAD3, p-SMAD3 (Cell Signaling Technology, 5339, 18338, 9523, 9520, respectively), CAT (Atlas Antibodies, HPA051282), SOD (Fitzgerald, 70R-13887), Nrf-2 (Cell Signaling Technology, 80593-1-RR), Keap1 (Proteintech, 80744-1-RR), GPR41 (Proteintech, 66811-1-Ig), GPR43 (Proteintech, 19952-1-AP), and Trizol reagent (Beyotime, R0016).

Techniques: Activation Assay, Immunofluorescence, Microscopy, Concentration Assay, Expressing, Western Blot, Activity Assay, In Vitro, Transfection, Control

Journal: Nature Communications

Article Title: Genetic diversity of Collaborative Cross mice implicates FFAR3 as a target for ILC2 anti-inflammatory reprogramming

doi: 10.1038/s41467-025-67813-2

Figure Lengend Snippet: A ILC2s from lungs of C57BL/6J and CC030 after 4 days of culture with IL-33, TSLP, and IL-2 (n = 3 per strain) ** p = 0.0016. B CFSE dilution between ILC2s after 48 h of culture with IL-33, TSLP, and IL-2. C Proliferation index and D division index after 48 h culture as in ( B ) (n = 4 per strain) **** p < 0.0001. E IL-13 in supernatant of ILC2s cultured per (A) (n = 3 per strain) ** p = 0.0018. F IL-13 expression by intracellular cytokine staining. MFI = mean fluorescence intensity (n = 3 per strain) * p = 0.011. G Representative flow plots of active caspase+ ILC2s, pre-gated on live cells (numbers above gates = %). H Quantification of active caspase+ ILC2s as in (G) (n = 4 per strain) *** p = 0.0004. I Representative flow plot of early/late apoptosis by Annexin V and propidium iodide (PI) staining on ILC2s 6 days post culture. J Early and late apoptotic K ILC2s (n = 5 per strain) *** p = 0.0004, ** p = 0.0019. L Apoptotic pathways enriched in naïve C57BL/6J vs CC030 ILC2s. M ILC2s as % of live lung cells and N as % of innate lymphocytes (Lineage⁻, CD90⁺) (n = 3 per strain) ** p = 0.0027, **** p < 0.0001. O IL-5 and P IL-13 in lung homogenate after 4 days of Alt Ex challenge (n = 4 C57BL/6J, 5 CC030) *** p = 0.0006, **** p < 0.0001. Q Total eosinophils in bronchoalveolar lavage (BAL) fluid after 4 days of Alt Ex challenge (n = 5 per strain, also in Supplementary Fig. C) ** p = 0.0027. R Diagram implicating candidate genes in Ch.7 ILC2 locus. S Ffar3 expression by normalized counts from bulk RNA-seq (DESeq2 DGE analysis, Wald test (two-tailed) with Benjamini–Hochberg correction; **** p = 1.76 × 10 −12 ). T qPCR for Ffar3 from RNA of flow-sorted naïve CC030 ILC2s from various tissues (n = 5 mice; one sample each from bone marrow, adipose, and colon had insufficient RNA). A , C – F , H , J , K , M – Q , S , T : bar height = mean, error bars = SD. A , C – F , M – P , S – T : each representative of two independent experiments, H , J , K , Q , S , T : each representative of one independent experiment. Significance for ( A ), ( C – F ), ( H ), ( J , K ), ( L – Q ), assessed by two-tailed unpaired Student’s t -test; for ( T ) by one-way ANOVA with Tukey multiple comparisons (two-tailed).

Article Snippet: Quantitative real-time PCR was carried out with TaqMan Gene Expression assays: Gapdh (Mm99999915) and Ffar3 (Mm02621638_s1) on a QuantStudio 5 Real-Time PCR System (Applied Biosystems) instrument for 45 cycles.

Techniques: Cell Culture, Expressing, Staining, Fluorescence, RNA Sequencing, Two Tailed Test

Journal: Nature Communications

Article Title: Genetic diversity of Collaborative Cross mice implicates FFAR3 as a target for ILC2 anti-inflammatory reprogramming

doi: 10.1038/s41467-025-67813-2

Figure Lengend Snippet: A Schematic for vehicle- vs. AR420626 (10 µM)–treated ILC2s by bulk RNA-sequencing. B Volcano plot of differentially expressed genes from ( A ). Egfr encircled. Labels = 10 most positively and negatively differentially expressed genes by log2 fold change (p-value threshold = 5 × 10⁻⁴; fold change threshold = 4; n = 5 paired biological replicates, representative of one experiment for ( A – C ). C Normalized EGFR counts from RNA-seq data in vehicle vs agonist-treated cells. Significance determined for ( B , C ) by DESeq2 DGE analysis (Wald test (two-tailed) with Benjamini–Hochberg correction); ***p < 2×10 −52 . (Bar height = mean, error bars = SD). D Representative flow plots of EGFR expression on ILC2s with and without FFAR3 agonist. Secondary-only control: no α-EGFR primary antibody. E EGFR expression by median fluorescence intensity (MFI) (n = 5 paired biological replicates; representative of one experiment) **** p < 0.0001. F Total ILC2s after 6-day culture with DMSO AR420626 (10 µM) and gefitinib (5 µM), groups = A–C left to right. (A-B *** p = 0.0005) (B-C ** p = 0.0018). G IL-10 levels in the supernatant; two vehicle ( AR420626 -/gefitinib-) samples were below detection. (A-B **** p < 0.0001) (A–C ** p = 0.0070) (B-C ** p = 0.0097). ( F , G ): n = 5 paired biological replicates; representative of two independent experiments). H Representative intracellular cytokine staining of IL-5 (gefitinib = 10 µM). I Quantification of IL-5 MFI (n = 4 for Vehicle and AR420626 (1 sample each used as FMOs); n = 5 for AR420626 + gefitinib). IL-5 quantification for vehicle and AR420626 is shown in Fig. (A-B *** p = 0.0007) (A–C *** p = 0.0002) (B-C **** p < 0.0001). J Amphiregulin in the supernatant of ILC2s with and without 10 µM AR420626 measured by ELISA (n = 5 paired biological replicates; representative of one experiment) ** p = 0.0012. K IL-10 in supernatant of ILC2s cultured with IL-33, TSLP, IL-2 + AR420626 + α-Amphiregulin antibody or isotype control (n = 5 paired biological replicates; representative of one experiment) * p = 0.033. C , E – G , I , J bar height = mean, error bars = SD). Significance for ( E ), ( J ), and ( K ) was assessed by two-tailed paired Student’s t-test; for ( F ), ( G ), and ( I ) by two-way ANOVA with Tukey’s multiple comparisons test. ns not significant.

Article Snippet: Quantitative real-time PCR was carried out with TaqMan Gene Expression assays: Gapdh (Mm99999915) and Ffar3 (Mm02621638_s1) on a QuantStudio 5 Real-Time PCR System (Applied Biosystems) instrument for 45 cycles.

Techniques: RNA Sequencing, Two Tailed Test, Expressing, Control, Fluorescence, Staining, Enzyme-linked Immunosorbent Assay, Cell Culture