Review





Similar Products

86
Revvity Signals cx3cr1
Hepatic Ly6C low macrophages in Rnaset2 −/− mice are moKCs. (A) Cluster analyses of the genes expressed in Ly6C hi and Ly6C low macrophages from wild-type and Rnaset2 −/− mice ( n = 3–4). (B, C, and F) Bars show reads per million (RPM) of indicated genes in indicated macrophages ( n = 3–4). (D) Dot plots show expression of MerTK, Axl, and <t>CX3CR1</t> in hepatic Ly6C low macrophages from wild-type and Rnaset2 −/− mice. (E) Red and black histograms show staining with antibodies to indicated molecules and isotype-matched control, respectively. (G) The numbers of KCs and moKCs in wild-type mice and those that were irradiated and received BM cells from Rnaset2 −/− mice. Closed and open bars show the numbers of macrophages from wild-type (CD45.1) and Rnaset2 −/− mice (CD45.2). ****P < 0.0001.
Cx3cr1, supplied by Revvity Signals, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/cx3cr1/product/Revvity Signals
Average 86 stars, based on 1 article reviews
cx3cr1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Danaher Inc polyclonal rabbit anti cx3cr1
Hepatic Ly6C low macrophages in Rnaset2 −/− mice are moKCs. (A) Cluster analyses of the genes expressed in Ly6C hi and Ly6C low macrophages from wild-type and Rnaset2 −/− mice ( n = 3–4). (B, C, and F) Bars show reads per million (RPM) of indicated genes in indicated macrophages ( n = 3–4). (D) Dot plots show expression of MerTK, Axl, and <t>CX3CR1</t> in hepatic Ly6C low macrophages from wild-type and Rnaset2 −/− mice. (E) Red and black histograms show staining with antibodies to indicated molecules and isotype-matched control, respectively. (G) The numbers of KCs and moKCs in wild-type mice and those that were irradiated and received BM cells from Rnaset2 −/− mice. Closed and open bars show the numbers of macrophages from wild-type (CD45.1) and Rnaset2 −/− mice (CD45.2). ****P < 0.0001.
Polyclonal Rabbit Anti Cx3cr1, supplied by Danaher Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/polyclonal rabbit anti cx3cr1/product/Danaher Inc
Average 86 stars, based on 1 article reviews
polyclonal rabbit anti cx3cr1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Revvity Signals pe anti mouse cx3cr1
Hepatic Ly6C low macrophages in Rnaset2 −/− mice are moKCs. (A) Cluster analyses of the genes expressed in Ly6C hi and Ly6C low macrophages from wild-type and Rnaset2 −/− mice ( n = 3–4). (B, C, and F) Bars show reads per million (RPM) of indicated genes in indicated macrophages ( n = 3–4). (D) Dot plots show expression of MerTK, Axl, and <t>CX3CR1</t> in hepatic Ly6C low macrophages from wild-type and Rnaset2 −/− mice. (E) Red and black histograms show staining with antibodies to indicated molecules and isotype-matched control, respectively. (G) The numbers of KCs and moKCs in wild-type mice and those that were irradiated and received BM cells from Rnaset2 −/− mice. Closed and open bars show the numbers of macrophages from wild-type (CD45.1) and Rnaset2 −/− mice (CD45.2). ****P < 0.0001.
Pe Anti Mouse Cx3cr1, supplied by Revvity Signals, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/pe anti mouse cx3cr1/product/Revvity Signals
Average 86 stars, based on 1 article reviews
pe anti mouse cx3cr1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Revvity Signals cx3cr1 apc
Hepatic Ly6C low macrophages in Rnaset2 −/− mice are moKCs. (A) Cluster analyses of the genes expressed in Ly6C hi and Ly6C low macrophages from wild-type and Rnaset2 −/− mice ( n = 3–4). (B, C, and F) Bars show reads per million (RPM) of indicated genes in indicated macrophages ( n = 3–4). (D) Dot plots show expression of MerTK, Axl, and <t>CX3CR1</t> in hepatic Ly6C low macrophages from wild-type and Rnaset2 −/− mice. (E) Red and black histograms show staining with antibodies to indicated molecules and isotype-matched control, respectively. (G) The numbers of KCs and moKCs in wild-type mice and those that were irradiated and received BM cells from Rnaset2 −/− mice. Closed and open bars show the numbers of macrophages from wild-type (CD45.1) and Rnaset2 −/− mice (CD45.2). ****P < 0.0001.
Cx3cr1 Apc, supplied by Revvity Signals, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/cx3cr1 apc/product/Revvity Signals
Average 86 stars, based on 1 article reviews
cx3cr1 apc - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Becton Dickinson anti cx3cr1 2a9 1
a) Proportion of classical monocytes (CM), intermediate monocytes (IM), and nonclassical monocytes (NCM) in the study population. P values are indicated. b) Proportion of MCT1-positive cells in CM, IM, and NCM. P values are indicated. c) CCR5, CCR2, and <t>CX3CR1</t> expression in CM, IM, and NCM. P values are indicated. d) Proportion of CCR5 + and CX3CR1 + cells in CM, IM, and NCM. P values are indicated. e) Migration assay scheme and percentage of migrated monocytes in the study population. f) Volcano diagram of differentially abundant plasma proteins between PWH and PWoH using Olink proteomics data. Proteins with adjusted p<0.05 are considered as significantly expressed in PWH compared to PWoH. g) The heatmap shows the expression pattern of proteins significantly regulated between PWH and PWoH in CM, IM, and NCM, which were obtained from the human protein atlas. Row annotation shows expression direction as log2 fold change according to Olink proteomics data h) Results of pathway enrichment analysis between PWH and PWoH using Olink proteomics data. Y-axis represents the negative log10 scaled fitted p-values belonging to a specific directional class.
Anti Cx3cr1 2a9 1, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/anti cx3cr1 2a9 1/product/Becton Dickinson
Average 86 stars, based on 1 article reviews
anti cx3cr1 2a9 1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Revvity Signals bv650 anti mouse cx3cr1
(A) Schematic of tumor challenges in mice immunized with OVA/ poly(I:C). (B) The growth of B16-OVA tumors after tumor injection on day 7 after immunization. ( C ) The growth of B16-OVA (left flank) or B16F10 (top right flank) tumors injected on day 45 in mice that rejected the first tumor challenge at lower right flank on day 7 as in ( B ). The growth of B16-OVA or B16F10 tumors in naïve mice was used as control. (D) The growth of B16-OVA tumors after tumor injection on day 21 after immunization. ( E - G ) Frequency of CD44+CD62L+( E ), <t>CX3CR1+(</t> F ), CD107a degranulation ( G ) in splenic CD11a high CD8+ T cells from the baseline and weeks after immunization. CD11a was used to identify antigen-primed T cells. ( H-J ) the tumor growth ( H-I ) and survival ( J ) of mice with B16-OVA tumors after treatment with anti-PD-L1 (10B5) starting at day 7 after tumor injection for a total of five doses.
Bv650 Anti Mouse Cx3cr1, supplied by Revvity Signals, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/bv650 anti mouse cx3cr1/product/Revvity Signals
Average 86 stars, based on 1 article reviews
bv650 anti mouse cx3cr1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Danaher Inc antibodies against cx3cr1 ab8020
( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in <t>CX3CR1</t> + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.
Antibodies Against Cx3cr1 Ab8020, supplied by Danaher Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/antibodies against cx3cr1 ab8020/product/Danaher Inc
Average 86 stars, based on 1 article reviews
antibodies against cx3cr1 ab8020 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Revvity Signals mouse cx3cr1
( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in <t>CX3CR1</t> + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.
Mouse Cx3cr1, supplied by Revvity Signals, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/mouse cx3cr1/product/Revvity Signals
Average 86 stars, based on 1 article reviews
mouse cx3cr1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

86
Thermo Fisher human cx3cr1
( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in <t>CX3CR1</t> + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.
Human Cx3cr1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/human cx3cr1/product/Thermo Fisher
Average 86 stars, based on 1 article reviews
human cx3cr1 - by Bioz Stars, 2025-02
86/100 stars
  Buy from Supplier

Image Search Results


Hepatic Ly6C low macrophages in Rnaset2 −/− mice are moKCs. (A) Cluster analyses of the genes expressed in Ly6C hi and Ly6C low macrophages from wild-type and Rnaset2 −/− mice ( n = 3–4). (B, C, and F) Bars show reads per million (RPM) of indicated genes in indicated macrophages ( n = 3–4). (D) Dot plots show expression of MerTK, Axl, and CX3CR1 in hepatic Ly6C low macrophages from wild-type and Rnaset2 −/− mice. (E) Red and black histograms show staining with antibodies to indicated molecules and isotype-matched control, respectively. (G) The numbers of KCs and moKCs in wild-type mice and those that were irradiated and received BM cells from Rnaset2 −/− mice. Closed and open bars show the numbers of macrophages from wild-type (CD45.1) and Rnaset2 −/− mice (CD45.2). ****P < 0.0001.

Journal: The Journal of Experimental Medicine

Article Title: RNase T2 deficiency promotes TLR13-dependent replenishment of tissue-protective Kupffer cells

doi: 10.1084/jem.20230647

Figure Lengend Snippet: Hepatic Ly6C low macrophages in Rnaset2 −/− mice are moKCs. (A) Cluster analyses of the genes expressed in Ly6C hi and Ly6C low macrophages from wild-type and Rnaset2 −/− mice ( n = 3–4). (B, C, and F) Bars show reads per million (RPM) of indicated genes in indicated macrophages ( n = 3–4). (D) Dot plots show expression of MerTK, Axl, and CX3CR1 in hepatic Ly6C low macrophages from wild-type and Rnaset2 −/− mice. (E) Red and black histograms show staining with antibodies to indicated molecules and isotype-matched control, respectively. (G) The numbers of KCs and moKCs in wild-type mice and those that were irradiated and received BM cells from Rnaset2 −/− mice. Closed and open bars show the numbers of macrophages from wild-type (CD45.1) and Rnaset2 −/− mice (CD45.2). ****P < 0.0001.

Article Snippet: Monoclonal anti-mouse CD11b (clone M1/70), CX3CR1 (clone SA011F11), F4/80 (clone BM8), NK1.1 (clone PK136), CD16.2 (clone 9E9), CD3ε (clone 145-2c11), CD19 (clone 6D5), CD11c (clone N418), CD317 (clone 927), CD45.2 (clone 104), CD8 (clone 53-6.7), Ki67 (clone 16A8), Tim4 (clone F31-5G3), CD34 (clone HM34), CD16/32 (clone 93), CD135 (clone A2/F10), and Sca-1 (clone D7) antibodies were purchased from BioLegend.

Techniques: Expressing, Staining, Control, Irradiation

a) Proportion of classical monocytes (CM), intermediate monocytes (IM), and nonclassical monocytes (NCM) in the study population. P values are indicated. b) Proportion of MCT1-positive cells in CM, IM, and NCM. P values are indicated. c) CCR5, CCR2, and CX3CR1 expression in CM, IM, and NCM. P values are indicated. d) Proportion of CCR5 + and CX3CR1 + cells in CM, IM, and NCM. P values are indicated. e) Migration assay scheme and percentage of migrated monocytes in the study population. f) Volcano diagram of differentially abundant plasma proteins between PWH and PWoH using Olink proteomics data. Proteins with adjusted p<0.05 are considered as significantly expressed in PWH compared to PWoH. g) The heatmap shows the expression pattern of proteins significantly regulated between PWH and PWoH in CM, IM, and NCM, which were obtained from the human protein atlas. Row annotation shows expression direction as log2 fold change according to Olink proteomics data h) Results of pathway enrichment analysis between PWH and PWoH using Olink proteomics data. Y-axis represents the negative log10 scaled fitted p-values belonging to a specific directional class.

Journal: bioRxiv

Article Title: Disrupted α-ketoglutarate homeostasis trains monocyte-derived macrophages towards M2-like phenotype in long-term treated HIV-infection

doi: 10.1101/2025.01.15.633214

Figure Lengend Snippet: a) Proportion of classical monocytes (CM), intermediate monocytes (IM), and nonclassical monocytes (NCM) in the study population. P values are indicated. b) Proportion of MCT1-positive cells in CM, IM, and NCM. P values are indicated. c) CCR5, CCR2, and CX3CR1 expression in CM, IM, and NCM. P values are indicated. d) Proportion of CCR5 + and CX3CR1 + cells in CM, IM, and NCM. P values are indicated. e) Migration assay scheme and percentage of migrated monocytes in the study population. f) Volcano diagram of differentially abundant plasma proteins between PWH and PWoH using Olink proteomics data. Proteins with adjusted p<0.05 are considered as significantly expressed in PWH compared to PWoH. g) The heatmap shows the expression pattern of proteins significantly regulated between PWH and PWoH in CM, IM, and NCM, which were obtained from the human protein atlas. Row annotation shows expression direction as log2 fold change according to Olink proteomics data h) Results of pathway enrichment analysis between PWH and PWoH using Olink proteomics data. Y-axis represents the negative log10 scaled fitted p-values belonging to a specific directional class.

Article Snippet: PBMCs were stained using anti-CD3 (UCHT-1) (562280, BD Bioscience), anti-CD19 (HIB19) (562294, BD Bioscience), anti-CD56 (B159) (562289, BD Bioscience), anti-HLADR (L243) (335795, BD Bioscience), anti-CD11c (B-LY6) (741827, BD Bioscience), anti-CD11b (ICRF44) (563839, BD Bioscience), anti-CD14 (M5E2) (555398, BD Bioscience), anti-CD16 (3G8) (560195, BD Bioscience), anti-CCR2 (1D9) (747849, BD Bioscience), anti-CCR5 (2D7) (556889, BD Bioscience), and anti-CX3CR1 (2A9-1) (744488, BD Bioscience) for chemokine receptors and anti-CD3 (OKT3) (317328, Biolegend), anti-CD4 (SK3) (563552, BD Bioscience), anti-CD8 (RPA-T8) (301014, Biolegend), anti-CD14 (M5E2) (301842, Biolegend), anti-CD16 (3G8) (563690, BD Bioscience), anti-MCT1 (882616) (FAB8275V, R&D Systems), anti-GLUT1 (202915) (FAB1418F, R&D Systems), and anti-xCT (NB300-318AF594, Novus Biologicals) for metabolite receptors.

Techniques: Expressing, Migration

(A) Schematic of tumor challenges in mice immunized with OVA/ poly(I:C). (B) The growth of B16-OVA tumors after tumor injection on day 7 after immunization. ( C ) The growth of B16-OVA (left flank) or B16F10 (top right flank) tumors injected on day 45 in mice that rejected the first tumor challenge at lower right flank on day 7 as in ( B ). The growth of B16-OVA or B16F10 tumors in naïve mice was used as control. (D) The growth of B16-OVA tumors after tumor injection on day 21 after immunization. ( E - G ) Frequency of CD44+CD62L+( E ), CX3CR1+( F ), CD107a degranulation ( G ) in splenic CD11a high CD8+ T cells from the baseline and weeks after immunization. CD11a was used to identify antigen-primed T cells. ( H-J ) the tumor growth ( H-I ) and survival ( J ) of mice with B16-OVA tumors after treatment with anti-PD-L1 (10B5) starting at day 7 after tumor injection for a total of five doses.

Journal: bioRxiv

Article Title: PD-1 prelimits both the cytotoxic and exhaustion potential in thymic CD8+ T cells and impacts the maintenance of peripheral tumor immunity

doi: 10.1101/2025.01.18.631253

Figure Lengend Snippet: (A) Schematic of tumor challenges in mice immunized with OVA/ poly(I:C). (B) The growth of B16-OVA tumors after tumor injection on day 7 after immunization. ( C ) The growth of B16-OVA (left flank) or B16F10 (top right flank) tumors injected on day 45 in mice that rejected the first tumor challenge at lower right flank on day 7 as in ( B ). The growth of B16-OVA or B16F10 tumors in naïve mice was used as control. (D) The growth of B16-OVA tumors after tumor injection on day 21 after immunization. ( E - G ) Frequency of CD44+CD62L+( E ), CX3CR1+( F ), CD107a degranulation ( G ) in splenic CD11a high CD8+ T cells from the baseline and weeks after immunization. CD11a was used to identify antigen-primed T cells. ( H-J ) the tumor growth ( H-I ) and survival ( J ) of mice with B16-OVA tumors after treatment with anti-PD-L1 (10B5) starting at day 7 after tumor injection for a total of five doses.

Article Snippet: After that, cells were stained with cell surface mouse antibodies based on the needs of experiments including PerCP/Cyanine5.5 anti-mouse TCRβ chain Antibody (Biolegend, Cat#109227, Clone H57-597), BUV395 Rat Anti-Mouse CD4 (BD Biosciences, Cat#563790, clone GK1.5), BUV496 Rat Anti-Mouse CD8a (BD Biosciences, Cat#569181, Clone 53-6.7), Brilliant Ultra Violet™ 563 CD25 monoclonal antibody (Thermofisher, Cat# 365-0251-82, Clone PC61.5), BUV615 Rat Anti-Mouse CD24 (BD biosciences, Cat#751499, Clone M1/69), BV711 Rat Anti-Mouse CD11a (BD biosciences, Cat#740676, Clone M1/4), BV570 anti-mouse CD44 Antibody (Biolegend, Cat#103037, Clone IM7), PE/Cy7 anti-mouse CD62L Antibody (Biolegend, Cat#104418, Clone, MEL-14), APC/Cy7 anti-mouse PD-1 Antibody (Biolegend, Cat#135224, Clone 29F.1A12), BV785 anti-mouse antibody (Biolegend, Cat#104543, Clone H1.2F3), BV750 Rat Anti-Mouse CD117 (BD Biosciences, Cat#747412, Clone 2B8), BV650 anti-mouse CX3CR1 (Biolegend, Cat#149033, Clone SA011F11), BUV737 Rat Anti-Mouse CD127 (BD Biosciences, Cat#612841, Clone SB/199), PE/Fire 810 anti-mouse Tim-3 (Biolegend, Cat#149033, Clone RMT3-23) resuspended in the FACS buffer (PBS + 2% FBS + 2mM EDTA).

Techniques: Injection, Control

( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in CX3CR1 + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in CX3CR1 + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques: Marker, Expressing, Flow Cytometry, Comparison, Immunofluorescence, Staining, Control, Fluorescence

( A ) Experimental workflow of eliminating intimal CX3CR1 + macrophages to observe TAA development in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The average quantity of CX3CR1 + macrophages in the vehicle group was set as 100%, while that in the DT group was relative to that in the vehicle group. n = 9 for vehicle and n = 8 for DT. * P < 0.05 by unpaired Student’s t test. ( C ) En face immunofluorescence staining of intimal CX3CR1 + macrophages (red) and VE-cadherin (green) in ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in intima averaged from 4 randomly selected areas for each mouse. The average of intensities of CX3CR1 staining in vehicle group were set as 1, while the intensities in DT group were presented as the relative values. n = 4 mice. * P < 0.05 by unpaired Student’s t test. ( D ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice at different ages with or without DT-mediated depletion. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( E ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 10 for vehicle and n = 8 for DT. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( F ) EVG staining of the aortic roots in 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 8 for each group. * P < 0.05 by Mann-Whitney U test for elastin degradation grade and unpaired Student’s t test for aortic thickness.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) Experimental workflow of eliminating intimal CX3CR1 + macrophages to observe TAA development in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The average quantity of CX3CR1 + macrophages in the vehicle group was set as 100%, while that in the DT group was relative to that in the vehicle group. n = 9 for vehicle and n = 8 for DT. * P < 0.05 by unpaired Student’s t test. ( C ) En face immunofluorescence staining of intimal CX3CR1 + macrophages (red) and VE-cadherin (green) in ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in intima averaged from 4 randomly selected areas for each mouse. The average of intensities of CX3CR1 staining in vehicle group were set as 1, while the intensities in DT group were presented as the relative values. n = 4 mice. * P < 0.05 by unpaired Student’s t test. ( D ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice at different ages with or without DT-mediated depletion. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( E ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 10 for vehicle and n = 8 for DT. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( F ) EVG staining of the aortic roots in 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 8 for each group. * P < 0.05 by Mann-Whitney U test for elastin degradation grade and unpaired Student’s t test for aortic thickness.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques: Flow Cytometry, Immunofluorescence, Staining, Fluorescence, MANN-WHITNEY

( A ) The pseudotime path of the VSMC transcriptome derived from the scRNA-Seq data on WT and Fbn1 C1041G/+ mice. ( B ) KEGG pathways enriched using the top 100 genes upregulated in Fbn1 C1041G/+ VSMCs compared with WT cells. ( C ) Bioinformatic analysis of ligand-receptor interactions between CX3CR1 + macrophages and VSMCs based on scRNA-Seq data. ( D ) The workflow of CX3CR1 + and CX3CR1 – macrophages isolated from MFS patients by FACS. ( E ) ELISA measurements of predicted ligand proteins secreted from CX3CR1 + or CX3CR1 – macrophages in conditioned media. n = 6. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The pseudotime path of the VSMC transcriptome derived from the scRNA-Seq data on WT and Fbn1 C1041G/+ mice. ( B ) KEGG pathways enriched using the top 100 genes upregulated in Fbn1 C1041G/+ VSMCs compared with WT cells. ( C ) Bioinformatic analysis of ligand-receptor interactions between CX3CR1 + macrophages and VSMCs based on scRNA-Seq data. ( D ) The workflow of CX3CR1 + and CX3CR1 – macrophages isolated from MFS patients by FACS. ( E ) ELISA measurements of predicted ligand proteins secreted from CX3CR1 + or CX3CR1 – macrophages in conditioned media. n = 6. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques: Derivative Assay, Isolation, Enzyme-linked Immunosorbent Assay

( A ) The experimental workflow of the evaluation of gene expression in MFS patient-specific iPSC-derived VSMCs with 24-hour treatment of conditioned media produced from 48-hour cultures of MFS patient-derived CX3CR1 + and CX3CR1 – macrophages. ( B ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment. n = 4. * P < 0.05 by unpaired Student’s t test. ( C and D ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditional media treatment in the presence of adalimumab (10 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( E and F ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment in the presence of teprotumumab (200 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) Real-time PCR of gene expression in the aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 4. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The experimental workflow of the evaluation of gene expression in MFS patient-specific iPSC-derived VSMCs with 24-hour treatment of conditioned media produced from 48-hour cultures of MFS patient-derived CX3CR1 + and CX3CR1 – macrophages. ( B ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment. n = 4. * P < 0.05 by unpaired Student’s t test. ( C and D ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditional media treatment in the presence of adalimumab (10 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( E and F ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment in the presence of teprotumumab (200 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) Real-time PCR of gene expression in the aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 4. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques: Expressing, Derivative Assay, Produced, Real-time Polymerase Chain Reaction, Comparison

( A ) The experimental workflow of parabiosis of age-matched Fbn1 C1041G/+ mice and Cx3cr1 GFP/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of GFP-expressing CX3CR1 + macrophages in aortic root and ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. n = 5. Unpaired Student’s t test. ( C ) En face immunofluorescence staining of GFP-expressed intimal CX3CR1 + macrophages in ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. Scale bars: 20 μm. ( D ) The experimental workflow of bone marrow transplantation (BMT) with Fbn1 C1041G/+ bone marrow cells infected with lentivirus encoding shRNA targeting TNF-α or IGF1 or control shRNA in 12-week-old Fbn1 C1041G/+ mice. ( E ) Real-time PCR of gene expression in aortic root and ascending aortas from Fbn1 C1041G/+ mice after BMT. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Fbn1 C1041G/+ mice at different ages with control, Igf1, or Tnfα shRNA treatment. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 5. * ,# P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots in 24-week-old mice with control, Igf1, or Tnfα shRNA treatment. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The experimental workflow of parabiosis of age-matched Fbn1 C1041G/+ mice and Cx3cr1 GFP/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of GFP-expressing CX3CR1 + macrophages in aortic root and ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. n = 5. Unpaired Student’s t test. ( C ) En face immunofluorescence staining of GFP-expressed intimal CX3CR1 + macrophages in ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. Scale bars: 20 μm. ( D ) The experimental workflow of bone marrow transplantation (BMT) with Fbn1 C1041G/+ bone marrow cells infected with lentivirus encoding shRNA targeting TNF-α or IGF1 or control shRNA in 12-week-old Fbn1 C1041G/+ mice. ( E ) Real-time PCR of gene expression in aortic root and ascending aortas from Fbn1 C1041G/+ mice after BMT. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Fbn1 C1041G/+ mice at different ages with control, Igf1, or Tnfα shRNA treatment. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 5. * ,# P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots in 24-week-old mice with control, Igf1, or Tnfα shRNA treatment. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques: Flow Cytometry, Expressing, Immunofluorescence, Staining, Transplantation Assay, Infection, shRNA, Control, Real-time Polymerase Chain Reaction, Comparison

( A – D ) Six-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 26 weeks. Age-matched WT mice were used as a normal control. ( A ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 4. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( B ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( C ) Quantification of transthoracic echocardiography. n = 4 for WT and n = 8 for Fbn1 C1041G/+ mice. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( D ) EVG staining of the aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness. ( E – H ) Twenty-four-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 36 weeks. Age-matched WT mice were used as a normal control. ( E ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of transthoracic echocardiography. n = 5. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A – D ) Six-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 26 weeks. Age-matched WT mice were used as a normal control. ( A ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 4. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( B ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( C ) Quantification of transthoracic echocardiography. n = 4 for WT and n = 8 for Fbn1 C1041G/+ mice. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( D ) EVG staining of the aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness. ( E – H ) Twenty-four-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 36 weeks. Age-matched WT mice were used as a normal control. ( E ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of transthoracic echocardiography. n = 5. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques: Injection, Control, Flow Cytometry, Comparison, Staining

CX3CR1 + macrophages mainly located in the aortic intima and originated from circulating Ly6C + monocytes. Intimal CX3CR1 + macrophages upregulated inflammatory genes, including IL6 , MCP1 , and CXCL2 , in VSMCs by producing and secreting TNF-α and IGF1, thereby aggravating TAA development in MFS. Either elimination of intimal CX3CR1 + macrophages or administration of a CCR2 inhibitor to suppress monocyte recruitment efficiently alleviated TAA progression in MFS.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: CX3CR1 + macrophages mainly located in the aortic intima and originated from circulating Ly6C + monocytes. Intimal CX3CR1 + macrophages upregulated inflammatory genes, including IL6 , MCP1 , and CXCL2 , in VSMCs by producing and secreting TNF-α and IGF1, thereby aggravating TAA development in MFS. Either elimination of intimal CX3CR1 + macrophages or administration of a CCR2 inhibitor to suppress monocyte recruitment efficiently alleviated TAA progression in MFS.

Article Snippet: Antibodies against CX3CR1 (ab8020) and CD68 (ab955) were obtained from Abcam.

Techniques:

( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in CX3CR1 + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in CX3CR1 + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques: Marker, Expressing, Flow Cytometry, Comparison, Immunofluorescence, Staining, Control, Fluorescence

( A ) Experimental workflow of eliminating intimal CX3CR1 + macrophages to observe TAA development in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The average quantity of CX3CR1 + macrophages in the vehicle group was set as 100%, while that in the DT group was relative to that in the vehicle group. n = 9 for vehicle and n = 8 for DT. * P < 0.05 by unpaired Student’s t test. ( C ) En face immunofluorescence staining of intimal CX3CR1 + macrophages (red) and VE-cadherin (green) in ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in intima averaged from 4 randomly selected areas for each mouse. The average of intensities of CX3CR1 staining in vehicle group were set as 1, while the intensities in DT group were presented as the relative values. n = 4 mice. * P < 0.05 by unpaired Student’s t test. ( D ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice at different ages with or without DT-mediated depletion. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( E ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 10 for vehicle and n = 8 for DT. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( F ) EVG staining of the aortic roots in 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 8 for each group. * P < 0.05 by Mann-Whitney U test for elastin degradation grade and unpaired Student’s t test for aortic thickness.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) Experimental workflow of eliminating intimal CX3CR1 + macrophages to observe TAA development in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The average quantity of CX3CR1 + macrophages in the vehicle group was set as 100%, while that in the DT group was relative to that in the vehicle group. n = 9 for vehicle and n = 8 for DT. * P < 0.05 by unpaired Student’s t test. ( C ) En face immunofluorescence staining of intimal CX3CR1 + macrophages (red) and VE-cadherin (green) in ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in intima averaged from 4 randomly selected areas for each mouse. The average of intensities of CX3CR1 staining in vehicle group were set as 1, while the intensities in DT group were presented as the relative values. n = 4 mice. * P < 0.05 by unpaired Student’s t test. ( D ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice at different ages with or without DT-mediated depletion. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( E ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 10 for vehicle and n = 8 for DT. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( F ) EVG staining of the aortic roots in 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 8 for each group. * P < 0.05 by Mann-Whitney U test for elastin degradation grade and unpaired Student’s t test for aortic thickness.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques: Flow Cytometry, Immunofluorescence, Staining, Fluorescence, MANN-WHITNEY

( A ) The pseudotime path of the VSMC transcriptome derived from the scRNA-Seq data on WT and Fbn1 C1041G/+ mice. ( B ) KEGG pathways enriched using the top 100 genes upregulated in Fbn1 C1041G/+ VSMCs compared with WT cells. ( C ) Bioinformatic analysis of ligand-receptor interactions between CX3CR1 + macrophages and VSMCs based on scRNA-Seq data. ( D ) The workflow of CX3CR1 + and CX3CR1 – macrophages isolated from MFS patients by FACS. ( E ) ELISA measurements of predicted ligand proteins secreted from CX3CR1 + or CX3CR1 – macrophages in conditioned media. n = 6. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The pseudotime path of the VSMC transcriptome derived from the scRNA-Seq data on WT and Fbn1 C1041G/+ mice. ( B ) KEGG pathways enriched using the top 100 genes upregulated in Fbn1 C1041G/+ VSMCs compared with WT cells. ( C ) Bioinformatic analysis of ligand-receptor interactions between CX3CR1 + macrophages and VSMCs based on scRNA-Seq data. ( D ) The workflow of CX3CR1 + and CX3CR1 – macrophages isolated from MFS patients by FACS. ( E ) ELISA measurements of predicted ligand proteins secreted from CX3CR1 + or CX3CR1 – macrophages in conditioned media. n = 6. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques: Derivative Assay, Isolation, Enzyme-linked Immunosorbent Assay

( A ) The experimental workflow of the evaluation of gene expression in MFS patient-specific iPSC-derived VSMCs with 24-hour treatment of conditioned media produced from 48-hour cultures of MFS patient-derived CX3CR1 + and CX3CR1 – macrophages. ( B ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment. n = 4. * P < 0.05 by unpaired Student’s t test. ( C and D ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditional media treatment in the presence of adalimumab (10 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( E and F ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment in the presence of teprotumumab (200 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) Real-time PCR of gene expression in the aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 4. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The experimental workflow of the evaluation of gene expression in MFS patient-specific iPSC-derived VSMCs with 24-hour treatment of conditioned media produced from 48-hour cultures of MFS patient-derived CX3CR1 + and CX3CR1 – macrophages. ( B ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment. n = 4. * P < 0.05 by unpaired Student’s t test. ( C and D ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditional media treatment in the presence of adalimumab (10 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( E and F ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment in the presence of teprotumumab (200 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) Real-time PCR of gene expression in the aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 4. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques: Expressing, Derivative Assay, Produced, Real-time Polymerase Chain Reaction, Comparison

( A ) The experimental workflow of parabiosis of age-matched Fbn1 C1041G/+ mice and Cx3cr1 GFP/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of GFP-expressing CX3CR1 + macrophages in aortic root and ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. n = 5. Unpaired Student’s t test. ( C ) En face immunofluorescence staining of GFP-expressed intimal CX3CR1 + macrophages in ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. Scale bars: 20 μm. ( D ) The experimental workflow of bone marrow transplantation (BMT) with Fbn1 C1041G/+ bone marrow cells infected with lentivirus encoding shRNA targeting TNF-α or IGF1 or control shRNA in 12-week-old Fbn1 C1041G/+ mice. ( E ) Real-time PCR of gene expression in aortic root and ascending aortas from Fbn1 C1041G/+ mice after BMT. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Fbn1 C1041G/+ mice at different ages with control, Igf1, or Tnfα shRNA treatment. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 5. * ,# P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots in 24-week-old mice with control, Igf1, or Tnfα shRNA treatment. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The experimental workflow of parabiosis of age-matched Fbn1 C1041G/+ mice and Cx3cr1 GFP/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of GFP-expressing CX3CR1 + macrophages in aortic root and ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. n = 5. Unpaired Student’s t test. ( C ) En face immunofluorescence staining of GFP-expressed intimal CX3CR1 + macrophages in ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. Scale bars: 20 μm. ( D ) The experimental workflow of bone marrow transplantation (BMT) with Fbn1 C1041G/+ bone marrow cells infected with lentivirus encoding shRNA targeting TNF-α or IGF1 or control shRNA in 12-week-old Fbn1 C1041G/+ mice. ( E ) Real-time PCR of gene expression in aortic root and ascending aortas from Fbn1 C1041G/+ mice after BMT. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Fbn1 C1041G/+ mice at different ages with control, Igf1, or Tnfα shRNA treatment. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 5. * ,# P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots in 24-week-old mice with control, Igf1, or Tnfα shRNA treatment. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques: Flow Cytometry, Expressing, Immunofluorescence, Staining, Transplantation Assay, Infection, shRNA, Control, Real-time Polymerase Chain Reaction, Comparison

( A – D ) Six-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 26 weeks. Age-matched WT mice were used as a normal control. ( A ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 4. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( B ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( C ) Quantification of transthoracic echocardiography. n = 4 for WT and n = 8 for Fbn1 C1041G/+ mice. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( D ) EVG staining of the aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness. ( E – H ) Twenty-four-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 36 weeks. Age-matched WT mice were used as a normal control. ( E ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of transthoracic echocardiography. n = 5. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A – D ) Six-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 26 weeks. Age-matched WT mice were used as a normal control. ( A ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 4. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( B ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( C ) Quantification of transthoracic echocardiography. n = 4 for WT and n = 8 for Fbn1 C1041G/+ mice. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( D ) EVG staining of the aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness. ( E – H ) Twenty-four-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 36 weeks. Age-matched WT mice were used as a normal control. ( E ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of transthoracic echocardiography. n = 5. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques: Injection, Control, Flow Cytometry, Comparison, Staining

CX3CR1 + macrophages mainly located in the aortic intima and originated from circulating Ly6C + monocytes. Intimal CX3CR1 + macrophages upregulated inflammatory genes, including IL6 , MCP1 , and CXCL2 , in VSMCs by producing and secreting TNF-α and IGF1, thereby aggravating TAA development in MFS. Either elimination of intimal CX3CR1 + macrophages or administration of a CCR2 inhibitor to suppress monocyte recruitment efficiently alleviated TAA progression in MFS.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: CX3CR1 + macrophages mainly located in the aortic intima and originated from circulating Ly6C + monocytes. Intimal CX3CR1 + macrophages upregulated inflammatory genes, including IL6 , MCP1 , and CXCL2 , in VSMCs by producing and secreting TNF-α and IGF1, thereby aggravating TAA development in MFS. Either elimination of intimal CX3CR1 + macrophages or administration of a CCR2 inhibitor to suppress monocyte recruitment efficiently alleviated TAA progression in MFS.

Article Snippet: Antibodies against mouse CD45 (Pacific blue labeled, 103126), mouse CD11b (APC labeled, 101212), mouse F4/80 (FITC labeled, 123107), mouse CX3CR1 (PE labeled, 149005; PerCP-Cyanine5.5 labeled, 149009), mouse CD3 (FITC labeled, 100203), mouse CD19 (AF700 labeled, 152413), and mouse CD144/VE-cadherin (A647 labeled,138006) were obtained from BioLegend.

Techniques:

( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in CX3CR1 + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) UMAP visualization of leukocytes in aortic root and ascending aortas from 20-week-old WT and Fbn1 C1041G/+ male mice. ( B ) Selected marker gene expression in distinct leukocyte subtypes. ( C ) Cell counts in each leukocyte subtype. WT, n = 6,508 cells versus Fbn1 C1041G/+ , n = 3,827 cells. χ 2 test to compare the cell quantitative changes. ( D ) GO analysis of genes enriched in CX3CR1 + macrophages (cluster 0). ( E ) The expression of CX3CR1 in distinct leukocyte subtypes. ( F ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from WT and Fbn1 C1041G/+ mice at different ages. n = 5. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) En face immunofluorescence staining of VE-cadherin (green) and CX3CR1 (red) in the intima of ascending aortas from 6- or 20-week-old WT and Fbn1 C1041G/+ mice. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the percentages of CX3CR1 + cells (red cells) in total intima cells (the number of nuclei) averaged from 4 randomly selected areas of ascending aortas for each mouse. n = 4 mice. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( H ) Immunofluorescence staining of CX3CR1 (green) and CD31 (red) in cross-sections of normal aortic tissues from control individuals (controls) and aneurysmal tissues from MFS patients. The nuclei were stained blue with DAPI. Scale bars:100 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in CD31 intimal areas averaged from 4 randomly selected areas for each patient. The average of intensities of CX3CR1 staining in normal controls was set as 1, while the intensities in MFS samples were presented as the relative values. n = 4. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: Marker, Expressing, Flow Cytometry, Comparison, Immunofluorescence, Staining, Control, Fluorescence

( A ) Experimental workflow of eliminating intimal CX3CR1 + macrophages to observe TAA development in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The average quantity of CX3CR1 + macrophages in the vehicle group was set as 100%, while that in the DT group was relative to that in the vehicle group. n = 9 for vehicle and n = 8 for DT. * P < 0.05 by unpaired Student’s t test. ( C ) En face immunofluorescence staining of intimal CX3CR1 + macrophages (red) and VE-cadherin (green) in ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in intima averaged from 4 randomly selected areas for each mouse. The average of intensities of CX3CR1 staining in vehicle group were set as 1, while the intensities in DT group were presented as the relative values. n = 4 mice. * P < 0.05 by unpaired Student’s t test. ( D ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice at different ages with or without DT-mediated depletion. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( E ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 10 for vehicle and n = 8 for DT. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( F ) EVG staining of the aortic roots in 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 8 for each group. * P < 0.05 by Mann-Whitney U test for elastin degradation grade and unpaired Student’s t test for aortic thickness.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) Experimental workflow of eliminating intimal CX3CR1 + macrophages to observe TAA development in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of CX3CR1 + macrophages in aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The average quantity of CX3CR1 + macrophages in the vehicle group was set as 100%, while that in the DT group was relative to that in the vehicle group. n = 9 for vehicle and n = 8 for DT. * P < 0.05 by unpaired Student’s t test. ( C ) En face immunofluorescence staining of intimal CX3CR1 + macrophages (red) and VE-cadherin (green) in ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. The nuclei were stained blue with DAPI. Scale bars: 20 μm. Data were quantified as the relative fluorescence intensities of CX3CR1 staining in intima averaged from 4 randomly selected areas for each mouse. The average of intensities of CX3CR1 staining in vehicle group were set as 1, while the intensities in DT group were presented as the relative values. n = 4 mice. * P < 0.05 by unpaired Student’s t test. ( D ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice at different ages with or without DT-mediated depletion. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( E ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 10 for vehicle and n = 8 for DT. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( F ) EVG staining of the aortic roots in 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 8 for each group. * P < 0.05 by Mann-Whitney U test for elastin degradation grade and unpaired Student’s t test for aortic thickness.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: Flow Cytometry, Immunofluorescence, Staining, Fluorescence, MANN-WHITNEY

( A ) The pseudotime path of the VSMC transcriptome derived from the scRNA-Seq data on WT and Fbn1 C1041G/+ mice. ( B ) KEGG pathways enriched using the top 100 genes upregulated in Fbn1 C1041G/+ VSMCs compared with WT cells. ( C ) Bioinformatic analysis of ligand-receptor interactions between CX3CR1 + macrophages and VSMCs based on scRNA-Seq data. ( D ) The workflow of CX3CR1 + and CX3CR1 – macrophages isolated from MFS patients by FACS. ( E ) ELISA measurements of predicted ligand proteins secreted from CX3CR1 + or CX3CR1 – macrophages in conditioned media. n = 6. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The pseudotime path of the VSMC transcriptome derived from the scRNA-Seq data on WT and Fbn1 C1041G/+ mice. ( B ) KEGG pathways enriched using the top 100 genes upregulated in Fbn1 C1041G/+ VSMCs compared with WT cells. ( C ) Bioinformatic analysis of ligand-receptor interactions between CX3CR1 + macrophages and VSMCs based on scRNA-Seq data. ( D ) The workflow of CX3CR1 + and CX3CR1 – macrophages isolated from MFS patients by FACS. ( E ) ELISA measurements of predicted ligand proteins secreted from CX3CR1 + or CX3CR1 – macrophages in conditioned media. n = 6. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: Derivative Assay, Isolation, Enzyme-linked Immunosorbent Assay

( A ) The experimental workflow of the evaluation of gene expression in MFS patient-specific iPSC-derived VSMCs with 24-hour treatment of conditioned media produced from 48-hour cultures of MFS patient-derived CX3CR1 + and CX3CR1 – macrophages. ( B ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment. n = 4. * P < 0.05 by unpaired Student’s t test. ( C and D ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditional media treatment in the presence of adalimumab (10 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( E and F ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment in the presence of teprotumumab (200 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) Real-time PCR of gene expression in the aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 4. * P < 0.05 by unpaired Student’s t test.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The experimental workflow of the evaluation of gene expression in MFS patient-specific iPSC-derived VSMCs with 24-hour treatment of conditioned media produced from 48-hour cultures of MFS patient-derived CX3CR1 + and CX3CR1 – macrophages. ( B ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment. n = 4. * P < 0.05 by unpaired Student’s t test. ( C and D ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditional media treatment in the presence of adalimumab (10 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( E and F ) Real-time PCR of gene expression in MFS patient-specific iPSC-derived VSMCs with conditioned media treatment in the presence of teprotumumab (200 μg/mL) or IgG. n = 4. * P < 0.05 by 2-way ANOVA followed by Tukey’s test for post hoc comparison. ( G ) Real-time PCR of gene expression in the aortic root and ascending aortas from 21-week-old Cx3cr1 -CreER T2 iDTR F/+ Fbn1 C1041G/+ mice with or without DT-mediated depletion. n = 4. * P < 0.05 by unpaired Student’s t test.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: Expressing, Derivative Assay, Produced, Real-time Polymerase Chain Reaction, Comparison

( A ) The experimental workflow of parabiosis of age-matched Fbn1 C1041G/+ mice and Cx3cr1 GFP/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of GFP-expressing CX3CR1 + macrophages in aortic root and ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. n = 5. Unpaired Student’s t test. ( C ) En face immunofluorescence staining of GFP-expressed intimal CX3CR1 + macrophages in ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. Scale bars: 20 μm. ( D ) The experimental workflow of bone marrow transplantation (BMT) with Fbn1 C1041G/+ bone marrow cells infected with lentivirus encoding shRNA targeting TNF-α or IGF1 or control shRNA in 12-week-old Fbn1 C1041G/+ mice. ( E ) Real-time PCR of gene expression in aortic root and ascending aortas from Fbn1 C1041G/+ mice after BMT. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Fbn1 C1041G/+ mice at different ages with control, Igf1, or Tnfα shRNA treatment. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 5. * ,# P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots in 24-week-old mice with control, Igf1, or Tnfα shRNA treatment. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A ) The experimental workflow of parabiosis of age-matched Fbn1 C1041G/+ mice and Cx3cr1 GFP/+ Fbn1 C1041G/+ mice. ( B ) Flow cytometry analysis of GFP-expressing CX3CR1 + macrophages in aortic root and ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. n = 5. Unpaired Student’s t test. ( C ) En face immunofluorescence staining of GFP-expressed intimal CX3CR1 + macrophages in ascending aortas from Fbn1 C1041G/+ and Cx3cr1 GFP/+ Fbn1 C1041G/+ parabionts. Scale bars: 20 μm. ( D ) The experimental workflow of bone marrow transplantation (BMT) with Fbn1 C1041G/+ bone marrow cells infected with lentivirus encoding shRNA targeting TNF-α or IGF1 or control shRNA in 12-week-old Fbn1 C1041G/+ mice. ( E ) Real-time PCR of gene expression in aortic root and ascending aortas from Fbn1 C1041G/+ mice after BMT. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas in Fbn1 C1041G/+ mice at different ages with control, Igf1, or Tnfα shRNA treatment. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of aortic root and ascending aorta diameters measured by transthoracic echocardiography. n = 5. * ,# P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots in 24-week-old mice with control, Igf1, or Tnfα shRNA treatment. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: Flow Cytometry, Expressing, Immunofluorescence, Staining, Transplantation Assay, Infection, shRNA, Control, Real-time Polymerase Chain Reaction, Comparison

( A – D ) Six-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 26 weeks. Age-matched WT mice were used as a normal control. ( A ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 4. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( B ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( C ) Quantification of transthoracic echocardiography. n = 4 for WT and n = 8 for Fbn1 C1041G/+ mice. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( D ) EVG staining of the aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness. ( E – H ) Twenty-four-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 36 weeks. Age-matched WT mice were used as a normal control. ( E ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of transthoracic echocardiography. n = 5. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: ( A – D ) Six-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 26 weeks. Age-matched WT mice were used as a normal control. ( A ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 4. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( B ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( C ) Quantification of transthoracic echocardiography. n = 4 for WT and n = 8 for Fbn1 C1041G/+ mice. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( D ) EVG staining of the aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness. ( E – H ) Twenty-four-week-old Fbn1 C1041G/+ mice were intraperitoneally injected with RS504393 (4 mg/kg/d) until age 36 weeks. Age-matched WT mice were used as a normal control. ( E ) Flow cytometry analysis of Ly6C + monocytes and CX3CR1 + macrophages in aortic root and ascending aortas. n = 5. * P < 0.05 by 1-way ANOVA followed by Tukey’s test for post hoc comparison. ( F ) Representative transthoracic echocardiographic images of the aortic root and ascending aortas. Scale bars: 2 mm. White arrows depict the sinus of Valsalva measurements. Yellow arrows depict ascending aorta measurement. ( G ) Quantification of transthoracic echocardiography. n = 5. * P < 0.05 by repeated-measures ANOVA with the Greenhouse-Geisser adjustment followed by Bonferroni’s post hoc comparisons. ( H ) EVG staining of aortic roots. n = 5. * P < 0.05 by Kruskal-Wallis test followed by Dunn’s test for elastin degradation grade and 1-way ANOVA followed by Tukey’s test for post hoc comparison for aortic thickness.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: Injection, Control, Flow Cytometry, Comparison, Staining

CX3CR1 + macrophages mainly located in the aortic intima and originated from circulating Ly6C + monocytes. Intimal CX3CR1 + macrophages upregulated inflammatory genes, including IL6 , MCP1 , and CXCL2 , in VSMCs by producing and secreting TNF-α and IGF1, thereby aggravating TAA development in MFS. Either elimination of intimal CX3CR1 + macrophages or administration of a CCR2 inhibitor to suppress monocyte recruitment efficiently alleviated TAA progression in MFS.

Journal: The Journal of Clinical Investigation

Article Title: Inhibition of aortic CX3CR1 + macrophages mitigates thoracic aortic aneurysm progression in Marfan syndrome in mice

doi: 10.1172/JCI178198

Figure Lengend Snippet: CX3CR1 + macrophages mainly located in the aortic intima and originated from circulating Ly6C + monocytes. Intimal CX3CR1 + macrophages upregulated inflammatory genes, including IL6 , MCP1 , and CXCL2 , in VSMCs by producing and secreting TNF-α and IGF1, thereby aggravating TAA development in MFS. Either elimination of intimal CX3CR1 + macrophages or administration of a CCR2 inhibitor to suppress monocyte recruitment efficiently alleviated TAA progression in MFS.

Article Snippet: Antibodies against human CD45 (FITC labeled, 11-0459-41), human CD14 (PE Cyanine7 labeled, 25-0149-41), and human CX3CR1 (PE labeled, 12-6099-41) were obtained from Thermo Fisher.

Techniques: