Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Vagotomy promoted Ly6G + cell infiltration into eWAT. Wild‐type mice were subjected to left cervical vagotomy (VX) or sham surgery and tissues were collected at 7 days following surgery. (A) The percentage of non‐adipocyte nuclei of total nuclei per section was quantified using ImageJ ( n = 4) and right panels show representative images of paraffin sections of eWAT stained with H&E in sham and VX animals. (B) CCL2 release from eWAT was analyzed by ELISA. The bar shows the CCL2 levels from sham ( n = 4) or VX ( n = 4) mice normalized to eWAT weight: ng/mL per g ± SEM (unpaired Student's t test). (C) eWAT was collected at 1 ( n = 3), 4 ( n = 4 sham, n = 5 VX), and 7 ( n = 15) days following VX or sham surgery and the eWAT SVCs were analyzed by flow cytometry. The bar shows the % ± SEM of CD11b + Ly6G + cells from CD45 + (one‐way ANOVA, Uncorrected Fisher's LSD). (D) Graphs show representative gating for CD11b + Ly6G + cells in sham and VX eWAT at 7 days (concatenated n = 5–6). (E) Representative immunostaining of Ly6G (red) and Perilipin1 (green) in paraffin sections of eWAT. (F–H) Bone marrow neutrophils after sham ( n = 9) or VX ( n = 5) surgery were isolated using negative magnetic beads and analyzed using bulk RNAseq (DESeq2). Heatmap (F), volcano plot (G) of differentially expressed genes, and GO (Gene Ontology) (H) enrichment bar plot. ns = not significant, * p < 0.05. VX, Vagotomy; eWAT, epididymal white adipose tissue; H&E, hematoxylin–eosin; SVCs, stromal vascular cells.

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Immunostaining, Isolation, Magnetic Beads, RNA sequencing

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Neutrophil deficiency attenuated the VX‐associated reduction of eWAT weight. Ly6G cre Mcl1 fl/fl mice were subjected to VX or sham surgery and tissues were collected at 7 days following the surgery. (A) Flow cytometry analysis of CD11b + Ly6G + cells in eWAT ( n = 7–9), blood ( n = 3–6), and bone marrow ( n = 3–6) following VX or sham surgery in wild‐type (WT) and Ly6G cre Mcl1 fl/fl (KO) mice. Bars show the proportion of cells from CD45 + (one‐way ANOVA, uncorrected Fisher's LSD). (B) The graph shows the change in body weight of WT ( n = 3) and Ly6G cre Mcl1 fl/fl (KO) ( n = 3–6) mice shown as grams±SEM from body weight day 0 (two‐way ANOVA, Tukey's multiple comparisons test—Significant differences between experimental groups at each time point are indicated with a and b, and the detailed description can be found in Table ). (C) eWAT weight was recorded at 7 days following VX or sham surgery ( n = 8–10 sham, n = 9 VX). The bars depict the relative eWAT weight to sham eWAT weight in % ± SEM (one‐way ANOVA, Šídák's multiple comparisons test). (D) Flow cytometry analysis of CD11b + Ly6G − F4/80 + cells in eWAT ( n = 5–8), blood ( n = 3–6), and bone marrow ( n = 3) following VX or sham surgery in WT and Ly6G cre Mcl1 fl/fl (KO) mice. Bars show the proportion of cells from CD45 (one‐way ANOVA, Šídák's multiple comparisons test). eWAT, epididymal white adipose tissue; ns, not significant, * p < 0.05, *** p < 0.001, VX, Vagotomy.

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Flow Cytometry

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Temporary extracellular Ly6G depletion effect on VX‐associated reduction of eWAT weight. (A) Flow cytometry analysis of the frequency of extracellular Ly6G and CD11b double positive cells following one intraperitoneal injection of either anti‐Ly6G or vehicle (PBS). Bars show the % ± SEM of CD11b + Ly6G + in vehicle ( n = 4–5) and at 1 ( n = 2), 2 ( n = 4), 5 ( n = 4), and 9 ( n = 3) days following injection in eWAT, blood, and bone marrow (one‐way ANOVA, Šídák's multiple comparisons test). (B) Schematic diagram depicting the experimental setup: Wild‐type mice were intraperitoneally injected once with anti‐Ly6G antibody or IgG2a antibody 2 days before sham or VX surgery, and tissue was collected 7 days following surgery. (C) Flow cytometry analysis of CD11b + Ly6G + cells in eWAT ( n = 3) following VX or sham. Bars show the proportion of cells from CD45 + (one‐way ANOVA, uncorrected Fisher's LSD). (D) Correlation between extracellular and intracellular expression of Ly6G in flow cytometry analysis. Circles represent each sample stained for both extracellular and intracellular Ly6G in separate fluorescent channels (Pearson r correlation). (E) The mice were weighed daily. The graph shows the difference in body weight (g) of the mice from day 0 (before surgery) of each experimental group ( n = 3) in g ± SEM (two‐way ANOVA, Tukey's multiple comparisons test—Significant differences between experimental groups at each time point are indicated with a, b, and c, and the detailed description can be found in Table ). (F) eWAT weight ( n = 3) was recorded at 7 days following VX or sham surgery. The bars show the relative eWAT weight to sham eWAT weight in % ± SEM (one‐way ANOVA, uncorrected Fisher's LSD). (G) Mice were kept in separate cages according to experimental groups: Sham+IgG2a, VX + IgG2a, sham+anti‐Ly6G, VX + anti‐Ly6G ( n = 3). The food for each cage was weighed at the same time point daily. The curve shows the grams of food consumed per day per cage in g. ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. VX, Vagotomy; eWAT, epididymal white adipose tissue.

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Flow Cytometry, Injection, Expressing, Staining

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Extracellular Ly6G depletion attenuated VX‐mediated reduction of body weight. (A) Schematic of the experimental setup: Wild‐type mice were intraperitoneally injected with anti‐Ly6G antibody or IgG2a antibody 2 days before, as well as 3 and 5 days following sham or VX surgery, and tissue was collected 7 days following surgery. (B–C) Flow cytometry analysis of CD11b + Ly6G + cells in eWAT ( n = 6) and blood ( n = 6) following VX or sham surgery, with IgG2a or anti‐Ly6G treatment. Bars show the proportion of cells from CD45 + (One‐way Anova, Uncorrected Fisher's LSD). (D–E) Flow cytometry analysis of intracellular Ly6G + cells in eWAT ( n = 3) and blood ( n = 3) following VX or sham surgery, with IgG2a or anti‐Ly6G treatment. Bars show the proportion of cells from CD45 + (One‐way Anova, Uncorrected Fisher's LSD). (F–G) Flow cytometry analysis of CD11b + (IN)Ly6G − F4/80 + cells in eWAT ( n = 3) and blood ( n = 3) following VX or sham surgery, with IgG2a or anti‐Ly6G treatment. Bars show the proportion of cells from CD45 + (One‐way Anova, Uncorrected Fisher's LSD). (H) eWAT weight ( n = 6) was recorded at 7 days following VX or sham surgery. The bars depict the relative eWAT weight to sham eWAT weight in % ± SEM (one‐way ANOVA, uncorrected Fisher's LSD). (I) The mice were weighed daily. The graph shows the difference in body weight of the mice from day 0 (before initiation of surgery) of each experimental group ( n = 6) in % ± SEM (two‐way ANOVA, Tukey's multiple comparisons test—Significant differences between experimental groups at each time point are indicated with a, b, and c, and the detailed description can be found in Table ). ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. VX, Vagotomy; eWAT, epididymal white adipose tissue; BM, bone marrow; SVCs, stromal vascular cells; intracellular (IN).

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Injection, Flow Cytometry

Journal: The Korean Journal of Pain

Article Title: IL-27-Ucp2-FoxO3 axis mediating the polarization of alternatively activated macrophages and ameliorating inflammatory pain

doi: 10.3344/kjp.25307

Figure Lengend Snippet: The identification of the source of IL-27. (A–D) The intracellular co-label staining by flow cytometry was applied to determine the source of IL-27. The four APCs were labeled by dendritic cell (CD11b, CD11c, B), neutrophil (CD11b, Ly6G, C), monocyte (CD11b, Ly6C, A), and macrophage (CD11b, F4/80, D), respectively. IL-27p28 antibody labeled the IL-27. The results indicated that neutrophil/monocyte-derived IL-27 was highly expressed in serum and spleen. IL: interleukin, APCs: antigen-presenting cells, SSC-A: side scatter area.

Article Snippet: APC Anti-Mouse Ly6C , Elabscience , E-AB-F1121E.

Techniques: Staining, Flow Cytometry, Labeling, Derivative Assay