anti mouse ccr7 neutralizing antibody  (R&D Systems)

Journal: Hepatology Communications

Article Title: Single-cell profiling reveals hepatic monocyte-derived macrophages heterogeneity during steatotic liver disease

doi: 10.1097/HC9.0000000000000928

Figure Lengend Snippet: CCR7 + (c4) MoMFs drive MASLD progression through enhanced antigen presentation and are recruited via the CCL19/CCL21 axis. (A) Feature plots displaying the expression of H2-Eb2 , Ccr7 , Cd209a , and Ciita . (B) Flow cytometry analysis showing CCR7 + MoMFs proportions among total MoMFs across NCD-fed, CDHFD-fed, MCD-fed, and HFD-fed mice, with representative plots (left) and quantitative data (right). (C) Correlation between the proportion of CCR7 + MoMFs and the NAFLD activity score in the combined cohort of NCD-fed and MCD-fed mice. (D, E) Dot plots show the enriched GO (D) and KEGG (E) terms of DEGs in CCR7 + MoMFs between the NCD and MCD groups. Dot size denotes the enriched gene counts. The color bar denotes the adjusted p -value of enrichment. (F) Dot plots show the average expression levels of selected DEGs. Dot size denotes the percentage of expressed cells, and the color bar denotes scaled average expression. (G) Flow cytometry analysis showing MHC-II expression levels in CCR7 + MoMFs across NCD-fed, CDHFD-fed, MCD-fed, and HFD-fed mice, with representative plots (left) and quantitative data (right). (H) Percentage of CCR7 + cells among total intrahepatic MoMFs in mice. (I, J) Plasma ALT and AST levels in each group. (K–M) Representative images (K) and quantification of H&E staining (L) and Sirius red staining (M) in liver paraffin sections. Scale bars, 200 μm. (N) Relative mRNA levels of Ccl19 (left) and Ccl21 (right) in liver tissues of NCD-fed, CDHFD-fed, MCD-fed, and HFD-fed mice. (O) Feature plots displaying the expression of Top2a , Mki67 , and Stmn1 . (P) Flow cytometry analysis showing Ki67 + MoMFs proportions among total MoMFs across NCD-fed, CDHFD-fed, MCD-fed, and HFD-fed mice, with representative plots (left) and quantitative data (right). (Q) Dot plots show the enriched GO terms of DEGs in Ki67 + MoMFs between the NCD and MCD groups. Dot size denotes the enriched gene counts. The color bar denotes the adjusted p -value of enrichment. (R) Feature plots displaying the expression of Cd3d , Gzma , and Gzmb . (S) Flow cytometry analysis showing CD3 + MoMFs proportions among total MoMFs across NCD-fed, CDHFD-fed, MCD-fed, and HFD-fed mice, with representative plots (left) and quantitative data (right). (T) Dot plots show the enriched GO terms of DEGs in CD3 + MoMFs between the NCD and MCD groups. Dot size denotes the enriched gene counts. The color bar denotes the adjusted p -value of enrichment. Abbreviations: ALT, alanine transaminase; AST, aspartate aminotransferase; CDHFD, choline-deficient high-fat diet; HFD, high-fat diet; H&E, Hematoxylin and Eosin; MASLD, metabolic dysfunction–associated steatotic liver disease; MCD, methionine/choline-deficient diet; MoMFs, monocyte-derived macrophages; NAFLD, nonalcoholic fatty liver disease; NCD, normal control diet; UMAP, uniform manifold approximation and projection.

Article Snippet: To target CCR7 + MoMFs, mice received twice-weekly intravenous (i.v.) injections of an anti-mouse CCR7 neutralizing antibody (MAB3477, R&D Systems) at 10 μg per mouse or an IgG2A isotype control (clone 4B12, R&D Systems)., To target CCR2 + MoMFs, mice were fed an MCD diet for 4 weeks.

Techniques: Immunopeptidomics, Expressing, Flow Cytometry, Activity Assay, Clinical Proteomics, Staining, Derivative Assay, Control

Journal: Hepatology Communications

Article Title: Single-cell profiling reveals hepatic monocyte-derived macrophages heterogeneity during steatotic liver disease

doi: 10.1097/HC9.0000000000000928

Figure Lengend Snippet: The basal cluster 1 (c1) is depleted during MASLD progression and acquires CD14 + and CCR7 + phenotypes under inflammatory stimulation in vitro. (A) Feature plots displaying the expression of Uba52 , Rpl27 , Rpl15 , and Ifi27l2a . (B) The typical flow cytometry gating strategy and statistical analysis of cluster 1 relative to total MoMFs in NCD-fed, CDHFD-fed, MCD-fed, and HFD-fed mice. (C) Correlation between the proportion of cluster 1 and the NAFLD activity score in the combined cohort of NCD-fed and MCD-fed mice. (D) UMAP plots displaying the major cellular subsets identified in the NCD and MCD groups. (E) Trajectory analysis of clusters 0, 1, 4, and 5 using Monocle 2. (F) Heatmap displaying gene expression dynamics across pseudotime for selected marker genes in clusters 0, 1, 4, and 5. (G–I) Flow cytometry analysis of CD14 (G), CCR7 (H), and CCR3 (I) expression in cluster 1 cells isolated from NCD-fed mouse liver, with or without LPS stimulation. (J) Dot plots depicting the expression of genes in liver macrophages derived from healthy and MASLD samples ( GSE212837 ). Abbreviations: CDHFD, choline-deficient high-fat diet; HFD, high-fat diet; LPS, lipopolysaccharide; MASLD, metabolic dysfunction–associated steatotic liver disease; MCD, methionine/choline-deficient diet; MoMFs, monocyte-derived macrophages; NCD, normal control diet; UMAP, uniform manifold approximation and projection.

Article Snippet: To target CCR7 + MoMFs, mice received twice-weekly intravenous (i.v.) injections of an anti-mouse CCR7 neutralizing antibody (MAB3477, R&D Systems) at 10 μg per mouse or an IgG2A isotype control (clone 4B12, R&D Systems)., To target CCR2 + MoMFs, mice were fed an MCD diet for 4 weeks.

Techniques: In Vitro, Expressing, Flow Cytometry, Activity Assay, Gene Expression, Marker, Isolation, Derivative Assay, Control

Journal: Hepatology Communications

Article Title: Single-cell profiling reveals hepatic monocyte-derived macrophages heterogeneity during steatotic liver disease

doi: 10.1097/HC9.0000000000000928

Figure Lengend Snippet: Heterogeneity and dynamics of intrahepatic MoMFs during MASLD progression. This scRNA-seq analysis delineated 7 distinct MoMFs clusters (c0–c6) in the liver with dynamic changes during steatohepatitis progression. CD14 + (c0) MoMFs, the predominant CCR2 + population, exhibited multifaceted functions beyond inflammation, including roles in fibrosis, lipid uptake, and antigen presentation. The basal cluster (c1) is depleted during MASLD progression and acquires CD14 + and CCR7⁺ phenotypes under inflammatory stimulation in vitro. Lipid overload drives mitochondrial dysfunction and apoptosis in CD206 + (c2) MoMFs in MASLD. CCR3 + (c3) MoMFs drive MASLD progression through lipid accumulation, oxidative stress, and CCL5-mediated recruitment. CCL19/CCL21-recruited CCR7 + (c4) MoMFs exacerbate MASLD pathogenesis through enhanced antigen presentation. In addition, Ki67 + (c5) proliferative and CD3 + (c6) TCR-related MoMFs maintained stable proportions but exhibited functional alterations during MASLD. Abbreviations: MASLD, metabolic dysfunction–associated steatotic liver disease; MoMFs, monocyte-derived macrophages; scRNA-seq, single-cell RNA sequencing.

Article Snippet: To target CCR7 + MoMFs, mice received twice-weekly intravenous (i.v.) injections of an anti-mouse CCR7 neutralizing antibody (MAB3477, R&D Systems) at 10 μg per mouse or an IgG2A isotype control (clone 4B12, R&D Systems)., To target CCR2 + MoMFs, mice were fed an MCD diet for 4 weeks.

Techniques: Immunopeptidomics, In Vitro, Functional Assay, Derivative Assay, Single Cell, RNA Sequencing

Journal: Journal of Biomedical Research

Article Title: CC-chemokine receptor 7 and its ligand CCL19 promote mitral valve interstitial cell migration and repair

doi: 10.7555/JBR.29.20150031

Figure Lengend Snippet: (A) CCR7 was not found in normal mitral valves. (B) CCR7 was expressed by hMVICs in mitral valves of RMS (arrow). (C) CCL19 was not found in normal mitral valves. (D) CCL19 was also observed in the endothelium of vessels in mitral valves of RMS. (E) CCL19 was expressed by mononuclear cells in mitral valves of RMS (arrow). The scale bar is 50 µM.

Article Snippet: Immediately after wounding, cells were treated with 100 or 200 ng/mL CCL19 (R&D, 361-MI-025), with or without 15 μ g/mL anti-CCR7 neutralizing antibody (R&D, MAB197).

Techniques:

Journal: Journal of Biomedical Research

Article Title: CC-chemokine receptor 7 and its ligand CCL19 promote mitral valve interstitial cell migration and repair

doi: 10.7555/JBR.29.20150031

Figure Lengend Snippet: (A) Representative photomicrograph of rheumatic hMVICs. Immnofluorescent staining showed that hMVICs consisted of vimentin positive (green) fibroblastic cells and vimentin and a-SMA double positive (red) myofibroblasts. (B) CCL19 stimulation was required before CCR7 was observed in normal hMVICs, while rheumatic hMVICs showed positive CCR7 staining without additional CCL19 stimulation. (C) The CCR7 mRNA level of rheumatic hMVICs was higher than that of normal hMVICs, but the concentration of CCL19 did not alter the level of CCR7 expression. * P < 0.05. (D-F) CCR7 expression was confirmed by immunofluorescence in rheumatic hMVICs and 100 ng/ml CCL19 treated normal hMVICs. The more CCR7 positive cells were observed in rheumatic group even without CCL19 stimulation. * P < 0.05. The imaging photos are captured at the same exposure time. The scale bar is 100 µM.

Article Snippet: Immediately after wounding, cells were treated with 100 or 200 ng/mL CCL19 (R&D, 361-MI-025), with or without 15 μ g/mL anti-CCR7 neutralizing antibody (R&D, MAB197).

Techniques: Staining, Concentration Assay, Expressing, Immunofluorescence, Imaging

Journal: Journal of Biomedical Research

Article Title: CC-chemokine receptor 7 and its ligand CCL19 promote mitral valve interstitial cell migration and repair

doi: 10.7555/JBR.29.20150031

Figure Lengend Snippet: (A–H) Representative photomicrographs of hMVICs 24 h after wounding of the hMVICs monolayer. (A–D, I) In rheumatic groups, the migration rate of CCL19-treated hMVICs significantly increased compared with hMVICs cultured in media alone. There were no statistical differences between the 100 ng/ml and 200 ng/ml CCL19 groups. (E–I) In normal groups, the migration rate of CCL19-treated hMVICs significantly increased when compared with hMVICs cultured in media alone; however, the migration rate of cells treated with 200 ng/ml CCL19 was longer than cells treated with 100 ng/ml CCL19, indicating a dose response. The effect of CCL19 on migration was blocked with an anti-CCR7 neutralizing antibody. *15ug/ml anti-CCR7, * P < 0.05, ** P < 0.01. The scale bar is 200 µM.

Article Snippet: Immediately after wounding, cells were treated with 100 or 200 ng/mL CCL19 (R&D, 361-MI-025), with or without 15 μ g/mL anti-CCR7 neutralizing antibody (R&D, MAB197).

Techniques: Migration, Cell Culture