Journal: Molecular cell

Article Title: Deciphering functional tumor-immune crosstalk through highly multiplexed imaging and deep visual proteomics.

doi: 10.1016/j.molcel.2024.12.023

Figure Lengend Snippet: Figure 3. T cell subsets have distinctive spatial proteome signatures in colorectal tumor microenvironment (A and B) Ranking of protein quantified in cytotoxic T cells (CTLs) (A) and helper T cells (TH) (B), ranked by median transformed intensity values detected by mass spectrometry, with cell-type-specific markers highlighted. (C and D) Volcano plots illustrating proteomic comparisons of CTLs (C) and TH (D) between the lamina propria and tumor epithelial regions, with T cell-activity- related markers highlighted. Significantly enriched proteins are denoted as yellow or green dots (two-sided t test, false discovery rate [FDR] < 0.05, S0 = 0.1, n = 3). (E and F) Gene Ontology (GO) term analysis of significantly altered proteins in CTLs (E) and TH (F) residing in the lamina propria and tumor epithelial regions.

Article Snippet: For CD4 + ARG1 staining, the sections were incubated overnight at 4 C with primary antibodies (anti-CD4-FITC, 1:100, R&D FAB8165G; anti-ARG1-PE, 1:200,Miltenyi Biotec 130-127-839) in a humidified chamber.

Techniques: Transformation Assay, Mass Spectrometry, Activity Assay

Journal: Molecular cell

Article Title: Deciphering functional tumor-immune crosstalk through highly multiplexed imaging and deep visual proteomics.

doi: 10.1016/j.molcel.2024.12.023

Figure Lengend Snippet: Figure 5. Cytotoxic T cells (CTLs) exhibit close proximity to tumors with a suppressive impact on tumor proliferation (A) Density gradient in the background indicates distance to tumor islands, with brighter yellow representing greater distance from tumor cells. Left panel only has the gradient whereas the middle, zoomed panel additionally depicts yellow masks of the CTLs and the right panel red dots of helper T cells (TH), respectively. (B) Representative images showcasing markers for T cell phenotyping. (C) T cell gating based on the expression of CD3, CD8, CD45RA, CD45RO, and PD1 to analyze the corresponding T cell subpopulations. (D) Representative images showing CD3- and CD8-positive CTLs in tumor parenchyma (TIL-CTLs) with absent CD45RA and CD45RO expression. (E) Distance heatmap depicting proximity and interactions between tumor cells and T and B lymphocyte subtypes, with lower values indicating higher proximity as in Figure 4E. (F) Histogram of Ki67/Ki67+ tumor cell number as a function of distance from RARO CTLs. Data represent three technical replicates. Statistical significance was determined using one-way ANOVA. ***p < 0.001, **p < 0.01. (G) Representative image displaying masks of tumor cells at varying proximities to RARO TIL-CTLs.

Article Snippet: For CD4 + ARG1 staining, the sections were incubated overnight at 4 C with primary antibodies (anti-CD4-FITC, 1:100, R&D FAB8165G; anti-ARG1-PE, 1:200,Miltenyi Biotec 130-127-839) in a humidified chamber.

Techniques: Expressing

Journal: Clinical and Experimental Pharmacology & Physiology

Article Title: Mechanisms of IL‐17A Neutralisation in Alleviating Renal Fibrosis and Inflammation in Spontaneously Hypertensive Rats

doi: 10.1111/1440-1681.70116

Figure Lengend Snippet: Effects of IL‐17A neutralisation on macrophage polarisation in SHR renal tissues. (A) Representative IHC staining of M1 macrophage markers (iNOS and CD86) with quantitative analysis of their positive areas. (B) Representative IHC staining of M2 macrophage markers (Arg‐1 and CD163) with quantitative analysis of their positive areas. (C) Representative immunoblots and relative expression levels of iNOS, CD86, Arg‐1, and CD163 proteins. (D) Proportion of CD86 + and CD163 + cells among CD68 + macrophages. (E) mRNA expression levels of iNOS, CD86, Arg‐1, and CD163. Data are presented as mean ± SD ( n = 6).

Article Snippet: Sections were then incubated overnight at 4°C with primary antibodies against: E‐cadherin (Boster, China), Collagen III (Boster, China), inducible nitric oxide synthase (iNOS) (Boster, China), CD86 (Boster, China), arginase‐1 (Arg‐1) (Boster, China), CD163 (Boster, China), α‐smooth muscle actin (α‐SMA) (Cell Signalling Technology, USA).

Techniques: Immunohistochemistry, Western Blot, Expressing

Journal: Oncotarget

Article Title: Deletion or inhibition of soluble epoxide hydrolase protects against brain damage and reduces microglia-mediated neuroinflammation in traumatic brain injury

doi: 10.18632/oncotarget.21139

Figure Lengend Snippet: (A) Representative double-immunofluorescence of CD16/32 (classical activation marker; red), CD206 or arginase 1 (ARG1, alternative activation markers; red) and Iba1 (green) in the contusion margin (see the red box in the brain atlas coronal brain section in Figure ) of WT and sEH KO mice at 1 day and 4 days following TBI. Arrowheads indicate the cells of colocalization. The inset images represent higher magnification of the boxed region in the corresponding images. The scale bar is 100 μm. The bar graph shows the degree of Iba1 and CD16/32, CD206 or arginase 1 colocalization in gray pixel intensity. Deletion of sEH resulted in a significant decrease of CD16/32-Iba1double-positive signal at 1 day and 4 days and an increase of CD206-Iba1 or arginase-1 double-positive signal at 4 days post-TBI. (B) Bar graphs of IL-4 protein level as assessed by ELISA in ipsilateral hemispheres of WT and sEH KO mice at 1 day and 4 days following sham surgery or TBI. Deletion of sEH caused a significant elevation of IL-4 level at 4 day. Values are mean ± S.E.M; ### , ††† P < 0.001 vs. sham group; * P < 0.05, ** P < 0.01, *** P < 0.001 vs. WT group (n = 5 mice / group for CD16/32, CD 206 or arginase 1 and Iba1 double stainings, Student’s t -test; n = 6 mice / group for IL-4 ELISA, repeated measures two-way ANOVA).

Article Snippet: To assess proinflammatory (M1) and anti-inflammatory (M2) microglia/macrophages, sections were incubated overnight at 4°C with rabbit anti-Iba1 (1:1,000; Wako), together with rat anti-CD16/32 (a classical M1 activation marker; 1:100; BD Biosciences), mouse anti-CD206 (an alternative M2 activation marker; 1:100; Bio-Rad Laboratories, Hercules, CA, USA) or rabbit anti-arginase 1 (an alternative M2 activation marker; 1:100; BD Biosciences).

Techniques: Immunofluorescence, Activation Assay, Marker, Enzyme-linked Immunosorbent Assay

Journal: Mediators of Inflammation

Article Title: Anti-Inflammatory Effect of Curcumin on the Mouse Model of Myocardial Infarction through Regulating Macrophage Polarization

doi: 10.1155/2021/9976912

Figure Lengend Snippet: Curcumin decreases M1 macrophage marker expression (iNOS, CCL2, and CD86) and increases M2 marker mRNA expression (Arg1, CD163, and CD206) in homogenized hearts. Data were shown as the mean ± SEM. # was P < 0.05, and ns was P > 0.05 vs. the MI group. And P value was calculated by Student's t -test. Baseline was defined as the gene expression in a normal mouse myocardium.

Article Snippet: We performed intracellular staining after cells were fixed and permeabilized, i.e., incubation with following antibodies for 30 min on ice: anti-iNOS (PE, 14792, Cell Signaling Technology, USA) and anti-arginase 1 (FITC, 554001, BD, USA).

Techniques: Marker, Expressing

Journal: Mediators of Inflammation

Article Title: Anti-Inflammatory Effect of Curcumin on the Mouse Model of Myocardial Infarction through Regulating Macrophage Polarization

doi: 10.1155/2021/9976912

Figure Lengend Snippet: Effect of curcumin on M1 and M2 macrophage marker protein expression in the myocardium at 7 days post-MI. (a) Representative immunohistochemistry detection of M1 macrophage marker (iNOS) expression; (b) representative immunohistochemistry detection of M2 macrophage marker (Arg1) expression; (c) quantification of iNOS-positive cells; (d) quantification of Arg1-positive cells. Seven mice in each group. Data were shown as the mean ± SEM; ∗ was P < 0.05 vs. the sham group, and # was P < 0.05 vs. the MI group. And P value was calculated by Student's t -test.

Article Snippet: We performed intracellular staining after cells were fixed and permeabilized, i.e., incubation with following antibodies for 30 min on ice: anti-iNOS (PE, 14792, Cell Signaling Technology, USA) and anti-arginase 1 (FITC, 554001, BD, USA).

Techniques: Marker, Expressing, Immunohistochemistry

Journal: Mediators of Inflammation

Article Title: Anti-Inflammatory Effect of Curcumin on the Mouse Model of Myocardial Infarction through Regulating Macrophage Polarization

doi: 10.1155/2021/9976912

Figure Lengend Snippet: Effect of curcumin on LPS/IFN γ -induced M1 and M2 macrophage marker expression in BMMs. mRNA expression of iNOS (a), CD86 (b), Arg1 (c), and CD163 (d) in BMMs using qPCR after being stimulated with different stimuli. Representative protein bands (e) and quantification of the grayscale value of iNOS (f), CD86 (g), Arg1 (h), and CD163 (i) protein. Each experiment was repeated 3 times independently. Data were shown as the mean ± SEM. ∗ was P < 0.05 vs. the LPS+IFN γ stimulation group, and # was P < 0.05 vs. the unstimulated group. P value was calculated by post hoc comparisons.

Article Snippet: We performed intracellular staining after cells were fixed and permeabilized, i.e., incubation with following antibodies for 30 min on ice: anti-iNOS (PE, 14792, Cell Signaling Technology, USA) and anti-arginase 1 (FITC, 554001, BD, USA).

Techniques: Marker, Expressing

Journal: Mediators of Inflammation

Article Title: Anti-Inflammatory Effect of Curcumin on the Mouse Model of Myocardial Infarction through Regulating Macrophage Polarization

doi: 10.1155/2021/9976912

Figure Lengend Snippet: Curcumin modulates M1/M2 macrophage polarization partly via AMPK. (a) The expression of p-AMPK and AMPK protein in homogenized hearts at 7 days after MI: representative protein bands (left) and quantification of protein gray values (right), 7 mice in each group. Data were shown as the mean ± SEM. ∗ was P < 0.05 vs. the sham group, and # was P < 0.05 vs. the MI group. And P value was calculated by Student's t -test. (b) After being stimulated with curcumin for different times, p-AMPK and AMPK proteins in BMMs were expressed: representative protein bands (left) and quantification of protein gray values (right). Representative protein bands (c) and quantification of the grayscale value of iNOS (d), CD86 (e), Arg1 (f), and CD163 (g) protein. Each experiment was repeated 3 times independently. Data were shown as the mean ± SEM. ∗ was P < 0.05 vs. the LPS+IFN γ stimulation group, and # was P < 0.05 vs. the LPS+IFN γ +Cur stimulation group. P value was calculated by post hoc comparisons.

Article Snippet: We performed intracellular staining after cells were fixed and permeabilized, i.e., incubation with following antibodies for 30 min on ice: anti-iNOS (PE, 14792, Cell Signaling Technology, USA) and anti-arginase 1 (FITC, 554001, BD, USA).

Techniques: Expressing