Journal: EMBO Molecular Medicine

Article Title: Metabolically reprogrammed eosinophils impair T cell immunity and cause chronic skin infection

doi: 10.1038/s44321-026-00392-x

Figure Lengend Snippet: ( A ) Left, representative flow cytometric analysis of foot skin cells, isolated from C57BL/6 mice and gated for CD11b + SiglecF + eosinophils on the respective days after L. mexicana infection. Right, representative clinical course of C57BL/6 mice infected with L. mexicana and influx of SSC high CD11b + SiglecF + eosinophils as measured by flow cytometry ( n = 4 mice per group, 4 independent experiments). ( B ) Quantification of Il5 mRNA expression by qRT-PCR in L. mexicana -induced skin lesions of C57BL/6 mice (day 0 and day 20: n = 3 mice each; day 6 and day 14: n = 11 mice each; 1–3 independent experiments). ( C ) Bead-based multiplex ELISA of serum cytokines from C57BL/6 mice 14 days p.i. with L. mexicana ( n = 14 mice per group, 3–4 independent experiments). ( D ) Serum IL-5 levels measured by ELISA in C57BL/6 mice infected with L. mexicana (day 0: n = 3; day 6: n = 13; day 14: n = 12; day 20: n = 10; day 56: n = 5, 2–4 independent experiments). ( E ) Right, representative flow cytometric analysis of bone marrow and blood cells isolated from naive or L. mexicana -infected C57BL/6 mice. Left, quantification of the respective population and organ (left panel day 0 n = 5, day 6 n = 9, day 14 n = 11, and day 20 n = 6; middle panel day 0 n = 4, day 6 n = 8, day 14 n = 6, and day 20 n = 6; right panel day 0 n = 5, day 6 n = 10, day 14 n = 12, and day 20 n = 5, 1–3 independent experiments). ( F ) C57BL/6 mice were treated with 500 µg of either isotype control or anti-IL-5 antibody on days 5 and 12 p.i. Right, representative flow cytometric analysis of isolated cells from bone marrow, blood and skin lesions of isotype-treated or anti-IL-5-treated mice. Left, quantification of the respective population and organ (left panel: day 0 n = 5, isotype n = 8, anti-IL-5 n = 8; middle panel: day 0 n = 5, isotype n = 8, anti-IL-5 n = 8; right panel: day 0 n = 3, isotype n = 4, anti-IL-5 n = 4, 2 independent experiments; skin data points were derived from samples pooled from two mice). ( A , E ) Minor differences in gating between the time points reflect that samples were acquired on different experimental days based on the fact that mice were infected with the same parasite batch. Data are mean ± SEM ( A ) or mean ± s.d. ( B – F ). Information regarding the exact p values and the statistical tests performed is provided in the Appendix Table . .

Article Snippet: Il5 , Mm00439646_m1.

Techniques: Isolation, Infection, Flow Cytometry, Expressing, Quantitative RT-PCR, Multiplex Assay, Enzyme-linked Immunosorbent Assay, Control, Derivative Assay

Journal: JCI Insight

Article Title: Vancomycin eliminates gut deoxycholic acid, restoring ER proteostasis in ILC2s and relieving colitis

doi: 10.1172/jci.insight.197470

Figure Lengend Snippet: ( A – C ) Large intestinal ILC2s were sorted from WT mice and cultured in the presence of IL-2, IL-7, IL-25, and IL-33 for 5 days. ILC2s were treated with DCA (100 μM) at the indicated concentrations for 24 hours and then subjected to RNA-Seq. ( A ) Experimental strategy, ( B ) volcano plot, and ( C ) heatmap of ILC2-characteristic genes. ( D – F ) Sorted large intestinal ILC2s from Il5 RFP-Cre mice were treated with DCA (100 μM) for 24 hours. ( D ) Experimental strategy. ( E ) Representative flow cytometry plots. ( F ) Percentages of RFP + ILC2s (CD45.2 + Lin – GATA3 + ) in large intestine. n = 6 wells per group. The experiment was repeated 3 times. ( G – I ) Rabbit reticulocyte lysate system. ( G ) Experimental strategy. Protein levels of Areg, IL-5, and IL-13 were analyzed by ( H ) Western blotting and ( I ) cytometric bead array in samples treated or untreated with DCA. n = 6 wells per group. The experiment was repeated twice. Data are shown as mean ± SD. Statistical analysis was performed using unpaired 2-tailed t test. *** P < 0.001, **** P < 0.0001.

Article Snippet: Il5 RFP-Cre mice (catalog R5/+) and CD45.1/CD45.1 mice (catalog 002014) were purchased from The Jackson Laboratory.

Techniques: Cell Culture, RNA Sequencing, Flow Cytometry, Western Blot

Journal: Science Advances

Article Title: Type 2 lymphocytes restrict type 3 lymphocytes during liver fibrosis and colocalize in fibroblast niches

doi: 10.1126/sciadv.aea6805

Figure Lengend Snippet: ( A ) Schematic showing CCl 4 administration in IL-33 mcherry/+ reporter mice, 0.5 μl CCl 4 /g body weight (BW) ip, three times per week for 4 weeks (4w), relevant to (B) to (N). Pooled data from three independent experiments; n ≥ 4 mice per group, unless otherwise noted. ( B and C ) Flow cytometry showing percentages (B) and total numbers (C) of ILC2s in livers from vehicle (corn oil)– or CCl 4 -treated mice. ( D to F ) Flow cytometry plots (D) and quantification [(E) and (F)] of hepatic IL-5 + lymphocytes gated on CD45 + CD11b − CD19 − NK1.1 − Thy1.2 + from Il5-tdtomato-Cre mice, highlighting ILC2s (IL-5 + CD3 − CD4 − ) and T H 2 cells (IL-5 + CD3 + CD4 + ). Total n ≥ 3 mice per group. ns, not significant. ( G to N ) Percent (G) and total γδ T cells (H), percent IL-17 + γδ T cells (I) and IL-17 + CD4 + T cells (J), percent (K) and total T H 2 cells (L), percent IL-13 + CD4 + T cells (M), and IFN-γ + CD4 + T cells (N) in livers from vehicle- or CCl 4 -treated IL-33 mcherry/+ mice. ( O ) Schematic of BDL in IL-5 + lymphocyte lineage tracker mice (IL-5tdtomato-Cre; Rosa26RFP). ( P to S ) Percent (P) and total liver IL-5 + ILC2s (Q), percent (R) and total liver γδ T cells (S) 14 days post-BDL or post–sham surgery, pooled from two experiments; n ≥ 2 mice per group. ( T to W ) Percent IL-17 + γδ T cells (T), IL-17 + CD4 + T cells (U), IL-13 + CD4 + T cells (V), and IFN-γ + CD4 + T cells (W) in livers from C57BL/6 mice 14 days post-BDL or postsham, pooled from three independent experiments; n ≥ 2 mice per group. Bar graphs indicate the means (±SE). Student’s t test [(B), (C), (G) to (N), and (P) to (W)] or one-way ANOVA with Tukey post test [(E) and (F)]; * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. See also fig. S2.

Article Snippet: Red5 (Il5-tdtomato-cre) cytokine reporter mice were used for tracking IL-5–producing T2Ls (the Jackson Laboratory, 030926) ( ).

Techniques: Flow Cytometry

Journal: Science Advances

Article Title: Type 2 lymphocytes restrict type 3 lymphocytes during liver fibrosis and colocalize in fibroblast niches

doi: 10.1126/sciadv.aea6805

Figure Lengend Snippet: ( A ) Schematic of IL-5–deleter mice ( Il5 Cre-RFP/Cre-RFP ; R26R DTA/DTA ). IL-5 deleters were homozygous for R26R DTA , and controls lacked DTA. ( B ) Flow quantification of liver IL-5 + lymphocytes from control or CCl 4 -treated Il5-tdtomato-Cre mice and IL-5–deleter mice. Pooled from three independent experiments; n ≥ 4 mice per group. ( C ) Flow plots showing IL-5 + lymphocytes (Lin − Thy1.2 + Red5 + ), IL-5 + ILC2s (Lin − Thy1.2 + CD3e − CD4 − ), and IL-5 + T H 2 cells (Lin − Thy1.2 + CD3e + CD4 + ) in livers from IL-5tdtomato-Cre or IL-5–deleter mice after 4-week CCl 4 treatment. ( D ) Liver hydroxyproline in controls or 4-week CCl 4 –treated Il5-tdtomato-Cre and IL-5–deleter mice. Pooled from three experiments. n ≥ 8 per group. ( E ) Liver expression of Col1a1 , Acta2 , and Des , normalized to GAPDH in control or 4-week CCl 4 –treated Il5-tdtomato-Cre and IL-5–deleter mice ( n ≥ 5 mice per group, two experiments). ( F ) ALT levels in same groups ( n ≥ 8 mice per group). ( G ) Confocal liver sections in control or 4-week CCl 4 –treated Il5-tdtomato-Cre mice and IL-5–deleter mice ( n = 3 or 4 per group). ( H ) Quantification of Col1 + area per tissue volume after 4-week CCl 4 ( n = 2 to 4 mice per group). ( I and J ) Sirius red staining (I) and quantification (J) in IL-5–deleter and control mice ± CCl 4 ( n ≥ 5 mice per group). ( K ) Schematic of CCl 4 and DT regimen in IL-5 DTR ( Il5 Cre-RFP/Cre-RFP ; R26R DTR/DTR ) and control mice. ( L ) Hydroxyproline in IL-5tdtomato-Cre and IL-5 DTR mice after 4-week CCl 4 treatment ( n ≥ 5 mice per group). ( M and N ) Bilirubin (M) and hydroxyproline (N) in IL-5–deleter and control mice 14 days post-BDL ( n ≥ 4 mice per group, three experiments). ( O and P ) Sirius red [(O) and (P)] and trichrome (P) staining 14 days post-BDL ( n ≥ 4 mice per group). ( Q ) Confocal liver sections 14 days post-BDL. All scale bars, 200 μm. Bar graphs indicate the means (±SE). Student’s t test [(B), (D) to (F), (H), (J), and (L) to (O)]. * P ≤ 0.05, ** P ≤ 0.01, and **** P ≤ 0.0001. See also fig. S4.

Article Snippet: Red5 (Il5-tdtomato-cre) cytokine reporter mice were used for tracking IL-5–producing T2Ls (the Jackson Laboratory, 030926) ( ).

Techniques: Control, Expressing, Staining

Journal: Science Advances

Article Title: Type 2 lymphocytes restrict type 3 lymphocytes during liver fibrosis and colocalize in fibroblast niches

doi: 10.1126/sciadv.aea6805

Figure Lengend Snippet: ( A ) CCl 4 schedule in Il5-tdtomato-Cre and IL-5–deleter ( Il5 Cre-RFP/Cre-RFP ;R26R DTA/DTA ) mice: 0.5 μl CCl 4 /g BW ip, three times per week for 2 or 4 weeks. ( B and C ) Flow quantification of percentages (B) and total numbers (C) of γδ T cells in livers from Il5-tdtomato-Cre and IL-5–deleter mice treated with vehicle or CCl 4 . Pooled data from three experiments; n ≥ 4 per group. ( D to F ) Flow quantification of percentages (D) and total numbers (E) of γδ T cells and percent IL-17 + DN T cells (F) in livers from Il5-tdtomato-Cre and IL-5 DTR ( Il5 Cre-RFP/Cre-RFP ;R26R DTR/DTR ) mice ± CCl 4 . Mice received 500 ng of DT every other day. Pooled from two experiments; n ≥ 5 per group. ( G to L ) Representative flow plots and quantitation of IL-17 [(G) and (H)] and RORγt {[(G) and (I)]; gated on CD45 + CD19 − CD11b − Thy1.2 + NK1.1 − CD4 − CD8 − ]} in hepatic γδ T cells and IFN-γ [(G) and (J)], IL-13 [(G) and (K)], and IL-17 {[(G) and (L)]; gated on CD45 + CD19 − CD11b − Thy1.2 + NK1.1 − CD4 + ]} in hepatic CD4 + T cells. n ≥ 5 per group. ( M to Q ) Flow quantification of percent B cells (M), macrophages (N), CD8 + T cells (O), CD4 + T cells (P), and neutrophils (Q) in livers ± CCl 4 . Pooled from three experiments; n ≥ 4 mice per group. ( R ) Total numbers of indicated cells at the steady state. n ≥ 5 per group. ( S ) Representative flow plots of CD9 + TREM + SAMs in livers from IL-5tdtomato-Cre mice ± CCl 4 (4 weeks). ( T to W ) Percentages [(T) and (V)] and total numbers [(U) and (W)] of CD9 + TREM + SAMs and Ly6C + CX3CRI + monocytes in livers ± CCl 4 . Pooled from two experiments; n ≥ 2 per group. ( X ) BDL schedule. ( Y and Z ) Percentages (Y) and total numbers (Z) of γδ T cells 14 days post-BDL. Pooled from three experiments; n ≥ 3 per group. Bar graphs indicate the means (±SE). Student’s t test [(B) to (F), (H) to (R), (T) to (W), (Y), and (Z)]. * P ≤ 0.05, ** P ≤ 0.01, and *** P ≤ 0.001.

Article Snippet: Red5 (Il5-tdtomato-cre) cytokine reporter mice were used for tracking IL-5–producing T2Ls (the Jackson Laboratory, 030926) ( ).

Techniques: Quantitation Assay

Journal: Science Advances

Article Title: Type 2 lymphocytes restrict type 3 lymphocytes during liver fibrosis and colocalize in fibroblast niches

doi: 10.1126/sciadv.aea6805

Figure Lengend Snippet: ( A ) Schematic of IL-17; IL-5 double-deleter mice (Il17aCre; Il5 Cre-RFP/Cre-RFP ;R26R DTA/DTA ) and IL-5 deleter controls ( Il5 Cre-RFP/Cre-RFP ;R26R DTA/DTA ). ( B ) CCl 4 treatment schedule: 0.5 μl CCl 4 /g BW ip, three times per week for 4 weeks. ( C and D ) Sirius red staining of 5-μm liver sections (C) and quantification (D) after 4-week CCl 4 treatment. Pooled from two experiments; n ≥ 4 per group. Scale bar, 200µm. ( E ) Hepatic hydroxyproline quantification; pooled from two experiments; n ≥ 5 per group. ( F ) Liver IL-6 expression normalized to 36B4 in 4-week CCl 4 –treated IL-5–deleter mice and double-deleter mice. n ≥ 8 per group, two experiments. ( G ) Flow plots showing deletion of RORγt + cells (upper) and γδ T cells (lower) in double-deleter versus IL-5–deleter mice. ( H to J ) Flow quantification of neutrophils (H), RORγt cells (I), and γδ T cells (J) in livers of Il5-tdtomato-Cre, IL-5–deleter, and double-deleter mice after 4-week CCl 4 treatment. Pooled from two experiments; n ≥ 3 per group. ( K ) GeoMx workflow. Drawn with BioRender. J. Sbierski-Kind (2025); https://biorender.com/f42j576 . ( L ) Representative staining for Col1 (fibrotic tract), α-SMA (vessels), CD45 (leukocytes) in IL-5–deleter mice, with indicated ROIs for GeoMx. ( M ) Cell abundances for parenchymal, adventitial, and fibrotic tract areas estimated using SpatialDecon. ( N to S ) AF (525 genes) [(N) to (P)] and MF [521 genes ] scores [(Q) to (S)] in vehicle-treated Il5-tdtomato-Cre mice (gray), CCl 4 -treated Il5-tdtomato-Cre mice (white), IL-5–deleter (red), and double-deleter (purple) mice. ( T to V ) Volcano plots of top 10 differentially expressed genes for AF and MF scores : CCl 4 -treated Il5-tdtomato-Cre versus vehicle (T), IL-5–deleter versus Il5-tdtomato-Cre mice (U), and double-deleter versus IL-5–deleter mice (V). ( W ) tSNE of fibrotic tract, adventitial, and parenchymal areas under indicated conditions. ( X ) Graphic scheme. Created in BioRender [J. Sbierski-Kind (2025); https://biorender.com/mx6h6dl ]. Bar graphs indicate the means (±SE). Student’s t test [(D) and (F)] and one-way ANOVA with Tukey post test [(E) and (H) to (J)]. * P ≤ 0.05, ** P ≤ 0.01, and *** P ≤ 0.001. See also fig. S7.

Article Snippet: Red5 (Il5-tdtomato-cre) cytokine reporter mice were used for tracking IL-5–producing T2Ls (the Jackson Laboratory, 030926) ( ).

Techniques: Staining, Expressing