Journal: Inflammation Research

Article Title: Quantitative temporal analysis of pancreatic islet T lymphocyte and macrophage infiltration heralded by serum IgE in congenic BioBreeding (BB) Gimap5 − / − rats at risk for insulitis and acute onset diabetes

doi: 10.1007/s00011-025-02101-9

Figure Lengend Snippet: Spatial infiltration of CD3 + and ED1 + immune cells in pancreatic islets. A Representative image of CD3+ T cell infiltration in a sBBM Gimap5-DP rat. The perimeter of the analyzed islet is marked in green, defined using a convolutional neural network (CNN)-based segmentation algorithm. B Schematic illustrating the spatial analysis of infiltrating immune cells. The green line indicates the islet boundary. Analysis was performed in 10 μm intervals extending up to 100 μm into the islets interior (negative values) and 100 μm into the surrounding exocrine tissue (positive values), reltive to the islet perimeter (0 μm). C Quantification of infiltrating CD3+ T cells (top row) and ED1+ macrophages (bottom row) shown as cell density (cells/mm 2 ) in 10 μm segments. Data are presented for sBBM Gimap5 + / + (Gimap5-DR) rats (n = 5; red bars), Gimap5 − / − (Gimap5-DP) prior to diabetes onset (n = 8; green bars), and Gimap5 − / − (Gimap5-DP) rats at the time of clinical onset of diabetes (n = 4; blue bars). Data are presented as mean ± SD

Article Snippet: Sections at depths of 15, 40, 65, 225, 250, 275, 435, 460, and 485 μm were stained for CD3, a marker of T lymphocytes (CD3 antibody (clone SP7), diluted 1:500, product no ab 16,669, Abcam, Cambridge, UK) and at 20, 45, 70, 230, 255, 280, 440, 465, and 490 μm were stained for ED1, a macrophage marker (mouse anti-rat CD68, diluted 1:100, MCA341R, Bio-Rad, Hercules, CA, USA).

Techniques:

Journal: The FASEB Journal

Article Title: ATF6 Alleviates Endothelial Inflammation Following Extended Hepatectomy Through Inhibition of TRIM10 / NF ‐ κB Signaling

doi: 10.1096/fj.202402197RRR

Figure Lengend Snippet: Expression of unfolded protein response (UPR) genes following extended hepatectomy (PH80) and major hepatectomy (PH70). (A) Transcriptomic analysis of the expression of UPR genes (atf6, atf4, xbp1, and hspa5/GRP78) at 0 h (control), 1 h, 6 h, 12 h, 1 day, and 3 days after PH80 and PH70 in rats (* p < 0.05 between PH80 and PH70, N = 3 at each time point); (B) the expression of ATF6 and cleaved ATF6 (cATF6) following PH80 and PH70 in mice determined by western blot analysis and semiquantitative analysis ( N = 5). (C) Western blot analysis of the expression of XBP1, spliced XBP1 (XBP1s) and ATF4 following PH80 and PH70 in mice and semiquantitative analysis ( N = 5); (D) western blot analysis of GRP78 and GRP94 expression following PH80 and PH70 treatment in mice and semiquantitative analysis of GRP78 and GRP94 expression ( N = 5); (E) immunohistochemical analysis of hepatic ATF6 expression following PH80 and PH70 treatment in mice (original magnification ×400, scale bars: 50 μm) and semiquantitative analysis of the results ( N = 5); (F) assessment of hepatic ATF6 expression following marginal hepatectomy ( N = 8) and minor hepatectomy ( N = 6) in humans by immunohistochemistry (original magnification ×400, scale bars: 50 μm) and semiquantitative analysis; (G) hepatic ATF6 expression (green) in liver sinusoidal endothelial cells (CD31, purple), macrophages (CD68, orange) and Th cells (CD4, white) in the early stage after extended hepatectomy in mice by immunofluorescence (original magnification ×400, scale bars: 20 μm).

Article Snippet: Immunostaining was conducted with primary antibodies against ATF6 (1:100, ab122897, Abcam, Cambridge, UK), Ki‐67 (1:200, ab15580, Abcam, Cambridge, UK), LSEC marker CD31 (1:100, 11265‐1‐AP, Proteintech, Wuhan, China), macrophage marker CD68 (1:100, 25747‐1‐AP, Proteintech, Wuhan, China), T helper cell marker CD4 (1:100, 65104‐1‐AP, Proteintech, Wuhan, China), and TRIM10 (1:200, bs‐9409R, Bioss, Beijing, China).

Techniques: Expressing, Control, Western Blot, Immunohistochemical staining, Immunohistochemistry, Immunofluorescence

Journal: Frontiers in Immunology

Article Title: CRP deposition in human abdominal aortic aneurysm is associated with transcriptome alterations toward aneurysmal pathogenesis: insights from in situ spatial whole transcriptomic analysis

doi: 10.3389/fimmu.2024.1475051

Figure Lengend Snippet: Morphologic marker staining and area of illumination selection in aortic tissue analysis. (A) Whole TMA slide scan view images for a normal aorta, AAA with low CRP, and AAA with high CRP. Each TMA section was prepared by creating 3-mm-diameter cores from the aortic wall, focusing on areas in contact with atheroma if present [AAA–low-CRP (n = 3), AAA–high-CRP (n = 7), and control group (n = 3)]. (B) Representative sections of a normal aorta, AAA–low CRP, and AAA–high CRP. The normal aorta primarily consists of αSMA-positive vascular smooth muscle cells with minimal inflammatory cell presence. In AAA–low CRP, the αSMA-positive vascular wall is thinner with fibrotic changes (white arrow), and CD68-positive macrophage infiltration is observed between fibrotic regions (red arrow). In AAA–high CRP, an extensively stained CD68-positive atherosclerotic plaque (magenta arrow) accumulates on a significantly thinned αSMA-positive vascular smooth muscle cell layer (yellow arrow). (C) Representative image of AOI segmentation using IF morphologic markers. The atheromatous plaque and vascular smooth muscle interface were stained simultaneously with CD45, αSMA, and CD68 markers. Vascular smooth muscle segmentation is highlighted (yellow arrow).

Article Snippet: To perform transcriptomic analysis of specific cell types in aortic aneurysm, we used four fluorochromes (AF488, AF532, AF594, and AF647) to label the following immunofluorescence (IF) markers: αSMA (#ab202368, Abcam, Cambridge, UK), CD45 (#NBP2-34528AF532, Novus, CO, USA), and CD68 (#sc-20060AF647, Santa Cruz, TX, USA) ( ).

Techniques: Marker, Staining, Selection, Control

Journal: Frontiers in Immunology

Article Title: CRP deposition in human abdominal aortic aneurysm is associated with transcriptome alterations toward aneurysmal pathogenesis: insights from in situ spatial whole transcriptomic analysis

doi: 10.3389/fimmu.2024.1475051

Figure Lengend Snippet: Differentially expressed genes (DEGs) in CD68-positive macrophages from spatial whole transcriptomic analysis [AAA–high CRP (n = 7) versus AAA–low CRP (n = 3)]. (A) Volcano plot displaying highly expressed DEGs in CD68-positive macrophages based on the degree of CRP deposition in AAA, using log 10 fold changes. (B) Clustered heatmap of the selected ROIs showing whole transcriptomic expression fold changes (average gene expression, z-score) in relation to CRP levels in AAA. Approximately 1,100 genes were differentially expressed according to the CRP deposition level. (C) Key DEGs between the high-CRP and low-CRP groups from (A) . The asterisk symbol (*) indicates statistical significance at p < 0.05.

Article Snippet: To perform transcriptomic analysis of specific cell types in aortic aneurysm, we used four fluorochromes (AF488, AF532, AF594, and AF647) to label the following immunofluorescence (IF) markers: αSMA (#ab202368, Abcam, Cambridge, UK), CD45 (#NBP2-34528AF532, Novus, CO, USA), and CD68 (#sc-20060AF647, Santa Cruz, TX, USA) ( ).

Techniques: Expressing, Gene Expression

Journal: Frontiers in Immunology

Article Title: CRP deposition in human abdominal aortic aneurysm is associated with transcriptome alterations toward aneurysmal pathogenesis: insights from in situ spatial whole transcriptomic analysis

doi: 10.3389/fimmu.2024.1475051

Figure Lengend Snippet: Functional enrichment and protein–protein interaction analysis of GO terms related to CRP deposition in AAA tissues: in CD68-positive macrophages [AAA–high CRP (n = 7) versus AAA–low CRP (n = 3)]. (A) Scatter plot from REVIGO illustrating cluster representatives after redundancy reduction in a two-dimensional space. Multidimensional scaling was applied to a matrix of the 34 GO terms’ semantic similarities. Notable GO terms identified include inflammatory response and blood vessel diameter maintenance. The bubble size indicates the frequency of the GO terms in the underlying GOA database, with color shading representing user-supplied p-values (legend in the lower right corner). (B) Potential enriched GO pathways in CD68+/CRP (high/low) regions. A total of 34 statistically significant biological process GO terms were observed. Further analysis highlighted representative GO terms such as inflammatory responses and MAPK cascade. Protein–protein interaction analysis using the STRING program with high confidence revealed that “ STAT3 ” connects two other clusters associated with apoptosis and immune responses within the inflammation cluster. (C) GO analysis results of genes associated with “blood vessel diameter maintenance,” and “inflammation response.” DEG, differentially expressed gene; Down, downregulated; Up, upregulated; AAA–high CRP, abdominal aortic aneurysm with high CRP; AAA–low CRP, abdominal aortic aneurysm with low CRP. The asterisk symbol (*) indicates statistical significance at p < 0.05.

Article Snippet: To perform transcriptomic analysis of specific cell types in aortic aneurysm, we used four fluorochromes (AF488, AF532, AF594, and AF647) to label the following immunofluorescence (IF) markers: αSMA (#ab202368, Abcam, Cambridge, UK), CD45 (#NBP2-34528AF532, Novus, CO, USA), and CD68 (#sc-20060AF647, Santa Cruz, TX, USA) ( ).

Techniques: Functional Assay

Journal: Gut

Article Title: DYRK1B blockade promotes tumoricidal macrophage activity in pancreatic cancer

doi: 10.1136/gutjnl-2023-331854

Figure Lengend Snippet: Intratumoral macrophage abundance correlates with DYRK1B levels in human PDAC. (A) Immunohistochemical CD68 staining (brown) of human PDAC tissue microarrays (Marburg cohort). DYRK1B levels were determined by bulk RNAseq. (B) Quantification of CD68 immunohistochemistry intensity in patients with PDAC, which were split into DYRK1B -low and high subgroups (n=15 each). Each dot represents one patient tumour (mean±SD). (C) CD24 mRNA levels in patients with PDAC as assessed by bulk RNA sequencing. Patients were split into DYRK1B -low/high subgroups (n=15 each). Each dot represents one patient tumour (mean±SD). (D) Correlation between CD68 and DYRK1B bulk mRNA expression in patients with PDAC of the TCGA cohort. (E) Correlation between MSR1 and DYRK1B bulk mRNA expression in patients with PDAC of the TCGA cohort. (F) Correlation between ITGAM (encoding CD11B) and DYRK1B bulk mRNA expression in patients with PDAC of the TCGA cohort. DYRK1B, dual specificity and tyrosine phosphorylation-regulated kinase 1B; PDAC, pancreatic ductal adenocarcinoma; mRNA, messenger RNA; TCGA, The Cancer Genome Atlas.

Article Snippet: To this end, we first made use of the Marburg cohort of patients with PDAC, whose tissue was stained by immunohistochemistry for the pan-macrophage marker CD68 (n=46).

Techniques: Immunohistochemical staining, Staining, Immunohistochemistry, RNA Sequencing, Expressing, Phospho-proteomics

Journal: BMC Veterinary Research

Article Title: Characterization of giant endocrine cells in the fundic stomach of African catfish (Clarias gariepinus) demonstrated by histochemical, immunohistochemical and ultrastructure microscopy methods suggesting their role in immunity

doi: 10.1186/s12917-024-04237-y

Figure Lengend Snippet: Identity, sources, and working dilution of antibodies used in the present immunohistochemical analysis

Article Snippet: CD68 (Macrophage Marker) Ab-3 (Clone KP1) , Mouse Anti-CD68 (Thermo Fisher Scientific Lab Vision Corporation, Fremont, USA) , Mouse Monoclonal Antibody Cat. #MS-397-R7 , 1:100 , Over night , boiling in citrate buffer (pH 6.0), 20 min , Goat anti-rabbit secondary antibody (Cat. no. K4003, EN Vision + TM System Horseradish Peroxidase Labelled Polymer; Dako). Ready to use (30 min at room temperature).

Techniques: Immunohistochemical staining, Incubation, Marker, Polymer

Journal: BMC Veterinary Research

Article Title: Characterization of giant endocrine cells in the fundic stomach of African catfish (Clarias gariepinus) demonstrated by histochemical, immunohistochemical and ultrastructure microscopy methods suggesting their role in immunity

doi: 10.1186/s12917-024-04237-y

Figure Lengend Snippet: Shows the immune reactivity of the endocrine cells with CD68. The fundic stomach paraffin section was immune stained with CD68. A : Endocrine cells positive for CD68 (shown by arrows) are seen in the gastric gland (gl), lamina propria (lp), and submucosa (sb). B : Endocrine cells positive for CD68 (shown by arrows) formed clusters within the sub-epithelial lymphatic space (ls). C , D : CD68-positive endocrine cells (shown by arrows) are situated in the submucosa (sb)

Article Snippet: CD68 (Macrophage Marker) Ab-3 (Clone KP1) , Mouse Anti-CD68 (Thermo Fisher Scientific Lab Vision Corporation, Fremont, USA) , Mouse Monoclonal Antibody Cat. #MS-397-R7 , 1:100 , Over night , boiling in citrate buffer (pH 6.0), 20 min , Goat anti-rabbit secondary antibody (Cat. no. K4003, EN Vision + TM System Horseradish Peroxidase Labelled Polymer; Dako). Ready to use (30 min at room temperature).

Techniques: Paraffin Section, Staining