53-2069-42
antibody from Invitrogen Antibodies
Targeting: MRC1
bA541I19.1, CD206, CLEC13D, CLEC13DL, MRC1L1
Antibody data
- Antibody Data
- Antigen structure
- References [22]
- Comments [0]
- Validations
- Flow cytometry [1]
- Other assay [20]
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- Product number
- 53-2069-42 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- CD206 (MMR) Monoclonal Antibody (19.2), Alexa Fluor™ 488, eBioscience™
- Antibody type
- Monoclonal
- Antigen
- Other
- Description
- Description: This 19.2 monoclonal antibody reacts with human CD206, which is also known as the macrophage mannose receptor (MMR). CD206 is expressed on macrophages and dendritic cells. This type I transmembrane protein can also be detected on non-immune cells, including hepatic and lymphathic epithelia and kidney mesengial cells. CD206 binds to glycoproteins that terminate in D-mannose, L-fucose, or N-acetylglucosamine, as well as a variety of hormones. This receptor undergoes constitutive internalization and recycling between the plasma membrane and the endosomal compartment. CD206 is involved in antigen processing and presentation, cell migration, and intracellular signaling. Moreover, CD206 plays a key role in phagocytosis pathogens such as Candida albicans, Leishmania, and Mycobacterium tuberculosis. Applications Reported: This 19.2 antibody has been reported for use in flow cytometric analysis. Applications Tested: This 19.2 antibody has been pre-titrated and tested by flow cytometric analysis of culture normal human peripheral blood cells. This can be used at 5 µL (0.125 µg) per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. Excitation: 488 nm; Emission: 519 nm; Laser: Blue Laser. Filtration: 0.2 µm post-manufacturing filtered.
- Reactivity
- Human
- Host
- Mouse
- Conjugate
- Green dye
- Isotype
- IgG
- Antibody clone number
- 19.2
- Vial size
- 100 Tests
- Concentration
- 5 µL/Test
- Storage
- 4° C, store in dark, DO NOT FREEZE!
Submitted references M2-like tumor-associated macrophages transmit exosomal miR-27b-3p and maintain glioblastoma stem-like cell properties.
TcpC Inhibits M1 but Promotes M2 Macrophage Polarization via Regulation of the MAPK/NF-κB and Akt/STAT6 Pathways in Urinary Tract Infection.
Interleukin 6 regulates the expression of programmed cell death ligand 1 in thyroid cancer.
Red blood cell mannoses as phagocytic ligands mediating both sickle cell anaemia and malaria resistance.
The role of irreversible electroporation in promoting M1 macrophage polarization via regulating the HMGB1-RAGE-MAPK axis in pancreatic cancer.
Paliperidone Inhibits Glioblastoma Growth in Mouse Brain Tumor Model and Reduces PD-L1 Expression.
Annexin A1 drives macrophage skewing to accelerate muscle regeneration through AMPK activation.
Lung transplantation for patients with severe COVID-19.
CAIX Regulates GBM Motility and TAM Adhesion and Polarization through EGFR/STAT3 under Hypoxic Conditions.
Astragalus polysaccharides (PG2) Enhances the M1 Polarization of Macrophages, Functional Maturation of Dendritic Cells, and T Cell-Mediated Anticancer Immune Responses in Patients with Lung Cancer.
Ginsenoside Rg3 Mitigates Atherosclerosis Progression in Diabetic apoE-/- Mice by Skewing Macrophages to the M2 Phenotype.
Helicobacter pylori-induced exosomal MET educates tumour-associated macrophages to promote gastric cancer progression.
Similarities and differences between helminth parasites and cancer cell lines in shaping human monocytes: Insights into parallel mechanisms of immune evasion.
Metformin-treated cancer cells modulate macrophage polarization through AMPK-NF-κB signaling.
Mannose Receptor Mediates the Immune Response to Ganoderma atrum Polysaccharides in Macrophages.
Renal Sodium Gradient Orchestrates a Dynamic Antibacterial Defense Zone.
miR-142-5p and miR-130a-3p are regulated by IL-4 and IL-13 and control profibrogenic macrophage program.
Porphyromonas gingivalis evasion of autophagy and intracellular killing by human myeloid dendritic cells involves DC-SIGN-TLR2 crosstalk.
Doxycycline inhibits polarization of macrophages to the proangiogenic M2-type and subsequent neovascularization.
Influence of the mannose receptor in host immune responses.
Inflammatory and alternatively activated human macrophages attract vessel-associated stem cells, relying on separate HMGB1- and MMP-9-dependent pathways.
Expression of a mannosyl-fucosyl receptor for endocytosis on cultured primary macrophages and their hybrids.
Zhao G, Ding L, Yu H, Wang W, Wang H, Hu Y, Qin L, Deng G, Xie B, Li G, Qi L
Cell death discovery 2022 Aug 4;8(1):350
Cell death discovery 2022 Aug 4;8(1):350
TcpC Inhibits M1 but Promotes M2 Macrophage Polarization via Regulation of the MAPK/NF-κB and Akt/STAT6 Pathways in Urinary Tract Infection.
Fang J, Ou Q, Wu B, Li S, Wu M, Qiu J, Cen N, Hu K, Che Y, Ma Y, Pan J
Cells 2022 Aug 28;11(17)
Cells 2022 Aug 28;11(17)
Interleukin 6 regulates the expression of programmed cell death ligand 1 in thyroid cancer.
Zhang GQ, Jiao Q, Shen CT, Song HJ, Zhang HZ, Qiu ZL, Luo QY
Cancer science 2021 Mar;112(3):997-1010
Cancer science 2021 Mar;112(3):997-1010
Red blood cell mannoses as phagocytic ligands mediating both sickle cell anaemia and malaria resistance.
Cao H, Antonopoulos A, Henderson S, Wassall H, Brewin J, Masson A, Shepherd J, Konieczny G, Patel B, Williams ML, Davie A, Forrester MA, Hall L, Minter B, Tampakis D, Moss M, Lennon C, Pickford W, Erwig L, Robertson B, Dell A, Brown GD, Wilson HM, Rees DC, Haslam SM, Alexandra Rowe J, Barker RN, Vickers MA
Nature communications 2021 Mar 19;12(1):1792
Nature communications 2021 Mar 19;12(1):1792
The role of irreversible electroporation in promoting M1 macrophage polarization via regulating the HMGB1-RAGE-MAPK axis in pancreatic cancer.
He C, Sun S, Zhang Y, Xie F, Li S
Oncoimmunology 2021 Mar 11;10(1):1897295
Oncoimmunology 2021 Mar 11;10(1):1897295
Paliperidone Inhibits Glioblastoma Growth in Mouse Brain Tumor Model and Reduces PD-L1 Expression.
Liu YS, Huang BR, Lin CJ, Shen CK, Lai SW, Chen CW, Lin HJ, Lin CH, Hsieh YC, Lu DY
Cancers 2021 Aug 28;13(17)
Cancers 2021 Aug 28;13(17)
Annexin A1 drives macrophage skewing to accelerate muscle regeneration through AMPK activation.
McArthur S, Juban G, Gobbetti T, Desgeorges T, Theret M, Gondin J, Toller-Kawahisa JE, Reutelingsperger CP, Chazaud B, Perretti M, Mounier R
The Journal of clinical investigation 2020 Mar 2;130(3):1156-1167
The Journal of clinical investigation 2020 Mar 2;130(3):1156-1167
Lung transplantation for patients with severe COVID-19.
Bharat A, Querrey M, Markov NS, Kim S, Kurihara C, Garza-Castillon R, Manerikar A, Shilatifard A, Tomic R, Politanska Y, Abdala-Valencia H, Yeldandi AV, Lomasney JW, Misharin AV, Budinger GRS
Science translational medicine 2020 Dec 16;12(574)
Science translational medicine 2020 Dec 16;12(574)
CAIX Regulates GBM Motility and TAM Adhesion and Polarization through EGFR/STAT3 under Hypoxic Conditions.
Huang BR, Liu YS, Lai SW, Lin HJ, Shen CK, Yang LY, Lu DY
International journal of molecular sciences 2020 Aug 14;21(16)
International journal of molecular sciences 2020 Aug 14;21(16)
Astragalus polysaccharides (PG2) Enhances the M1 Polarization of Macrophages, Functional Maturation of Dendritic Cells, and T Cell-Mediated Anticancer Immune Responses in Patients with Lung Cancer.
Bamodu OA, Kuo KT, Wang CH, Huang WC, Wu ATH, Tsai JT, Lee KY, Yeh CT, Wang LS
Nutrients 2019 Sep 20;11(10)
Nutrients 2019 Sep 20;11(10)
Ginsenoside Rg3 Mitigates Atherosclerosis Progression in Diabetic apoE-/- Mice by Skewing Macrophages to the M2 Phenotype.
Guo M, Xiao J, Sheng X, Zhang X, Tie Y, Wang L, Zhao L, Ji X
Frontiers in pharmacology 2018;9:464
Frontiers in pharmacology 2018;9:464
Helicobacter pylori-induced exosomal MET educates tumour-associated macrophages to promote gastric cancer progression.
Che Y, Geng B, Xu Y, Miao X, Chen L, Mu X, Pan J, Zhang C, Zhao T, Wang C, Li X, Wen H, Liu Z, You Q
Journal of cellular and molecular medicine 2018 Nov;22(11):5708-5719
Journal of cellular and molecular medicine 2018 Nov;22(11):5708-5719
Similarities and differences between helminth parasites and cancer cell lines in shaping human monocytes: Insights into parallel mechanisms of immune evasion.
Narasimhan PB, Akabas L, Tariq S, Huda N, Bennuru S, Sabzevari H, Hofmeister R, Nutman TB, Tolouei Semnani R
PLoS neglected tropical diseases 2018 Apr;12(4):e0006404
PLoS neglected tropical diseases 2018 Apr;12(4):e0006404
Metformin-treated cancer cells modulate macrophage polarization through AMPK-NF-κB signaling.
Chiang CF, Chao TT, Su YF, Hsu CC, Chien CY, Chiu KC, Shiah SG, Lee CH, Liu SY, Shieh YS
Oncotarget 2017 Mar 28;8(13):20706-20718
Oncotarget 2017 Mar 28;8(13):20706-20718
Mannose Receptor Mediates the Immune Response to Ganoderma atrum Polysaccharides in Macrophages.
Li WJ, Tang XF, Shuai XX, Jiang CJ, Liu X, Wang LF, Yao YF, Nie SP, Xie MY
Journal of agricultural and food chemistry 2017 Jan 18;65(2):348-357
Journal of agricultural and food chemistry 2017 Jan 18;65(2):348-357
Renal Sodium Gradient Orchestrates a Dynamic Antibacterial Defense Zone.
Berry MR, Mathews RJ, Ferdinand JR, Jing C, Loudon KW, Wlodek E, Dennison TW, Kuper C, Neuhofer W, Clatworthy MR
Cell 2017 Aug 24;170(5):860-874.e19
Cell 2017 Aug 24;170(5):860-874.e19
miR-142-5p and miR-130a-3p are regulated by IL-4 and IL-13 and control profibrogenic macrophage program.
Su S, Zhao Q, He C, Huang D, Liu J, Chen F, Chen J, Liao JY, Cui X, Zeng Y, Yao H, Su F, Liu Q, Jiang S, Song E
Nature communications 2015 Oct 5;6:8523
Nature communications 2015 Oct 5;6:8523
Porphyromonas gingivalis evasion of autophagy and intracellular killing by human myeloid dendritic cells involves DC-SIGN-TLR2 crosstalk.
El-Awady AR, Miles B, Scisci E, Kurago ZB, Palani CD, Arce RM, Waller JL, Genco CA, Slocum C, Manning M, Schoenlein PV, Cutler CW
PLoS pathogens 2015 Feb;10(2):e1004647
PLoS pathogens 2015 Feb;10(2):e1004647
Doxycycline inhibits polarization of macrophages to the proangiogenic M2-type and subsequent neovascularization.
He L, Marneros AG
The Journal of biological chemistry 2014 Mar 21;289(12):8019-28
The Journal of biological chemistry 2014 Mar 21;289(12):8019-28
Influence of the mannose receptor in host immune responses.
Gazi U, Martinez-Pomares L
Immunobiology 2009;214(7):554-61
Immunobiology 2009;214(7):554-61
Inflammatory and alternatively activated human macrophages attract vessel-associated stem cells, relying on separate HMGB1- and MMP-9-dependent pathways.
Lolmede K, Campana L, Vezzoli M, Bosurgi L, Tonlorenzi R, Clementi E, Bianchi ME, Cossu G, Manfredi AA, Brunelli S, Rovere-Querini P
Journal of leukocyte biology 2009 May;85(5):779-87
Journal of leukocyte biology 2009 May;85(5):779-87
Expression of a mannosyl-fucosyl receptor for endocytosis on cultured primary macrophages and their hybrids.
Stahl P, Gordon S
The Journal of cell biology 1982 Apr;93(1):49-56
The Journal of cell biology 1982 Apr;93(1):49-56
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Supportive validation
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- Staining of 3-day GM-CSF-treated human peripheral blood monocytes with Mouse IgG1 K Isotype Control Alexa Fluor® 488 (Product # 53-4714-42) (blue histogram) or Anti-Human CD206 (MMR) Alexa Fluor® 488 (purple histogram). Total viable cells, as determined by Fixable Viability Dye eFluor® 450, were used for analysis.
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- Figure 2 miR-142-5p and miR-130a-3p regulate M2 activation of macrophages. ( a - d ) Human macrophages were transduced with nc, miR-142-5p ASO or miR-130a-3p mimics or both using lentiviral vectors. After 24 h, the cells were treated with IL-4 for 48 h. ( a ) The schematics of the approach. ( b ) Expression of CD206 and CD36 in macrophages as determined by flow-cytometry analysis. The representative histograms and quantitation of the MFI are shown (mean+-s.e.m., n =4 independent experiments; * P
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- Figure 9 Dyregulated Mph miR-142-5p and miR-130a-3p enhance liver fibrosis. ( a - f ) Mice were intravenously injected with LNA-modified miR-142-5p ASO, miR-130a-3p mimic or both every 3 days after CCL 4 challenge. Mice were sacrificed after 6 weeks (mean+-s.e.m., n =8 mice/group; * P
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- Figure 1 Metformin treated cancer cells polarized macrophage toward M1 phenotype THP-1 cells were stimulated with PMA (200 nM) for 24 h, then incubated with breast cancer (MDA-MB231/MDA-MB453) conditioned medium (CM) with or without metformin (60 muM) for 6 h, followed by flow cytometry analysis to quantify the amount of CD206, an M2 macrophage marker, and CD16, an M1 marker ( A, B ). Data are expressed as mean +- SD, * p < 0.05. DMSO: control; Met: metformin. Representative flow data shown are from experiments independently performed at least three times.
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- Figure 5 AMPK-NF-kappaB signaling participated in macrophage polarization Breast cancer cells (MDA-MB231) were treated with metformin 60 muM combined with an AMPK inhibitor (Compound C, CC) or NF-kappaB inhibitor (BAY-117082, BAY) for 6 h. The supernatant was collected to treat macrophages for 48 h, followed by flow cytometry analysis of CD206, M2 phenotype ( A ) and CD16, M1 phenotype ( B ). Data are expressed as mean +- SD, * p < 0.05. DMSO: control; CM: conditioned medium, Met: metformin. Representative flow data shown are from experiments independently performed at least three times.
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- 4 FIGURE M2-phenotype macrophages produce interleukin 6 (IL-6) in thyroid cancer. A, Flow cytometry showed that IL-6 was distributed in F4/80 staining-positive tumor-associated macrophages (TAMs). B, Flow cytometry showed that IL-6 was distributed in CD206-positive M2-phenotype macrophages. C, Double immunofluorescence analysis illustrated the colocalization of IL-6 and M2 phenotype within thyroid cancer
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- Figure 4 The polarizations of monocyte and GBM progression in a co-cultured model under hypoxic conditions. ( A ) THP-1 monocytes were co-cultured with U87-green fluorescent protein (GFP) or U251-GFP under hypoxic conditions for 48 h. The co-cultured cells were plotted on a side scatter versus FITC. The THP-1 monocytes (GFP-negative cells) were analyzed to assess the levels of cluster of differentiation (CD) 206, arginase 1 (Arg1), interleukin (IL)-10, interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha by flow cytometry. ( B ) The median fluorescence intensity (MFI) of CD206, Arg1, IL-10, IFN-gamma, and TNF-alpha in THP-1 monocytes in the co-cultured model under hypoxic conditions for 48 h. * p < 0.05 compared with the THP-1 monocyte group. Quantitative data are presented as the mean +- standard error (representative of n = 3). ( C ) U87-GFP and U251-GFP were treated with U104 (CAIX inhibitor) and subsequently co-cultured with THP-1 monocytes under hypoxic conditions for 48 h. THP-1 monocytes (GFP-negative cells) were analyzed to assess the levels of CD206, Arg1, and IL-10 by flow cytometry. ( D ) U87-GFP or U251-GFP was treated with U104 and subsequently co-cultured with THP-1 monocytes under hypoxic conditions for 48 h. The co-cultured cells were plotted on a side scatter versus FITC. The GFP-positive gated GBM was analyzed to assess the levels of programmed death ligand 1 (PD-L1) by flow cytometry. ( E ) The levels of CAIX, PD-L1, tumor growth factor-beta, and c
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- Figure 2. The effect of TSN on macrophage polarization. (a) The comparisons of mRNA levels of cytokines from THP-1 stimulated by TSN of Panc-1-treated with electric fields. The mRNA levels of M1 markers increased along with the increasing electric fields, while the mRNA levels of M2 markers did not show a continuous increase along with the increasing electric fields. (b) The expression of surface markers of THP-1 stimulated by TSN of Bxpc-3 treated with electric fields. Increased expression of HLA-DR on THP-1 along with the increasing electric fields was observed. (c) IRE could increase infiltration of M1 macrophage in pancreatic tumor tissue (IHC). Orthotopic or subcutaneous tumor tissue was used as control. (d) IRE could increase the expression of CD16/32 in infiltrated macrophages in pancreatic tumor tissue. Orthotopic or subcutaneous tumor tissue was used as control. F4/80 was used to indicate macrophage; CD16/32 was the marker of M1 macrophage; CD206 was the marker of M2 macrophage; DAPI was used to indicate the nuclear of cells. (e) The expression of CD16/32 in macrophage from tumor tissue, spleen, peripheral blood, and lymph node in an orthotopic model of mouse pancreatic cancer after IRE treatment. IRE increased the expression of CD16/32 on the surface of macrophages from these tissues in the orthotopic model of mouse pancreatic cancer. (f) The expression of CD16/32 in macrophages from tumor tissue, spleen, peripheral blood, and lymph node in the subcutaneous model of
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- Fig. 3 Display of membrane skeleton-associated mannose patches is induced by oxidative stress and recognized by the mannose receptor on macrophages. a Peanut agglutinin (PNA) and Galanthus nivalis Agglutinin (GNA) lectin binding to normal hemoglobin (HbAA) RBCs with or without oxidation. Mannan blockade for GNA lectin binding shown in blue. 2 tailed Wilcoxon, paired data, PNA, n = 7; GNA, n = 8 biologically independent RBC donors over two independent experiments. b Immunofluorescence microscopy of GNA lectin/streptavidin (red) staining of healthy HbAA RBCs (above) and after the oxidative insult (below). c Normalized geometric mean fluorescence (gMFI) for binding analyzed by flow cytometry of murine Fc fusions with C-type lectins or sub-domains applied to oxidized versus undamaged RBCs. Mannan blockade of mannose receptor-carbohydrate recognition domain (MR-CRD) binding is shown in blue. MAH, macrophage antigen H. MR-CR, mannose receptor cysteine-rich domain. 2 tailed Wilcoxon, paired data, n = 7 biologically independent RBC donors over two independent experiments. d Immunofluorescence microscopy image of human monocyte-derived macrophages (HMDM) stained with DAPI (blue) and for mannose receptor (green) after incubation with oxidized HbAA RBCs, shown in magenta. e Percentage phagocytosis of oxidized RBCs by HMDM treated with human MR specific or scrambled siRNA. 2 tailed Mann-Whitney, n = 4 biologically independent RBC donors over two independent experiments. f Percentage phag
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- Fig. 4 Single-cell RNA sequencing of lung tissue from patients with severe COVID-19. ( A , C , and E ) Uniform Manifold Approximation and Projection (UMAP) plots showing individual populations of epithelial cells (A), macrophages (C), and mesenchymal cells (E). ( B , D , and F ) Heatmaps illustrating expression of select marker genes in epithelial cells (B), macrophages (D), and mesenchymal cells (F). Gene expression for the pulmonary fibrosis dataset of Habermann et al. () is shown as an average per condition; gene expression for the end-stage COVID-19 dataset is shown per individual patient. Labels on heatmaps (B, D, and F) correspond to the following samples: Control (H), healthy controls; IPF, idiopathic pulmonary fibrosis samples; Other PF, samples from patients with other forms of pulmonary fibrosis, all from the Habermann et al. dataset. Donor 1, Donor 2, control donor lungs; Case 1, lung transplant case 1; PMB1, PMB2, postmortem lung biopsies from two patients with COVID-19, all from the end-stage COVID-19 dataset. ( G ) Immunofluorescence microscopy revealed KRT17 staining (magenta) of flat epithelial cells resembling alveolar type 1 cells in nonfibrotic lung tissue from a patient who died of COVID-19 (palliative COVID-19). ( H and I ) Immunofluorescence microscopy revealed KRT17 staining (magenta) of distal explanted lung tissue from a patient with COVID-19 undergoing lung transplantation (H) and lung tissue from a patient (PMB1) who died from late-stage severe COVI
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- Figure 3 DRD2 and PD-L1 in GBM are involved in the polarization of TAMs in the GBM-macrophage co-cultured system. ( A ) BMDMs were co-cultured with ALTS1C1-GFP cells and then treated with or without PAL (20 muM) for 48 h. CD80 and CD206 expressions were determined by flow cytometry analysis. ( B ) Establishment of stable PD-L1 expression in ALTS1C1-GFP cells. The expression of PD-L1 was verified by Western blot. ( C ) BMDMs were co-cultured with PD-L1 ALTS1C1-GFP cells for 48 h. CD80 and CD206 were determined by flow cytometry analysis. ( D ) ALTS1C1-GFP cells were transfected with DRD2, and the expression of DRD2 was verified by real time-PCR. * p < 0.05 compared with shLacz group. ( E ) BMDMs were co-cultured with DRD2 knockdown of ALTS1C1-GFP cells for 48 h. CD80 and CD206 were determined by flow cytometry analysis.
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- Fig. 2 M2-TAM-derived exosomes enhance the stemness of GSCs. A TEM images of M2-TAM-derived exosomes. The presence of CD63, CD206, and CD163 in M2-TAM-derived exosomes analyzed by flow cytometric ( B ) and Western blot ( C ) analyses. D Fluorescence images of GSCs co-cultured with Dil-labeled M2-TAM-derived exosomes with nuclei stained with DAPI (blue). E Representative images showing neurosphere formation of GSCs along with the statistics of GSC formation rate and diameter of spheres. F Expression of stem cell-related protein CD133, Nestin, Oct4, Sox2, and GFAP in GSCs measured by Western blot analysis. G HE staining images of xenograft tumors from mice. H The representative immunohistochemical images of Sox2 in xenograft tumors and the percentage of GSCs labeled by Sox2. I Kaplan-Meier survive curve of tumor-bearing mice. n = 10. * p < 0.05 vs. the control EXO group. Measurement data were depicted as mean +- standard deviation, comparison of that between two groups was conducted by unpaired t test. Cell experiments were repeated three times independently.
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- FIGURE 2 Flow cytometry analysis of M1 and M2 surface markers (A) The expression of CD86 (M1 surface marker) and CD206 (M2 surface marker) on RAW264.7 cells were examined by flow cytometry. (B) Quantification of mean fluorescence intensity (MFI) of the surface markers ( n = 3, respectively). (C) The expression of CD86 and CD206 on THP-1 cells were examined by flow cytometry. (D) Quantification of mean fluorescence intensity (MFI) of the surface markers ( n = 3, respectively). The results are expressed as the mean values +- S.E.M. * p < 0.05, ** p < 0.01, *** p < 0.001.
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- Figure 1 Differential macrophage response to different inflammatory cytokine stimuli. ( A ) Morphological and immunocytochemical images showing monocyte-derived macrophage (MDM) polarization to M1 or M2 functional phenotype using type 1 cytokine IFN-gamma and lipopolysaccharide (LPS) or type 2 cytokine IL-4 and IL-13, respectively. ( B ) Increases in CD80+ M1 and CD206+ M2 MDMs were seen after treatment with IFN-gamma /LPS or IL-4/IL-13, respectively, using flow-cytometry analysis. Green arrows point to the CD80+ M1 macrophage, while red arrows indicate CD206+ M2 macrophages. ( C ) Flow cytometry and morphology imaging of M1 and M2 cell sorting and isolation using the fluorescence-activated cell sorting (FACS) assay. ( D ) Compared with the H1299 cells treated with M1 conditioned medium, the H1299 cells cultured with M2 conditioned medium exhibited higher invasion ability at 18 h in matrigel study. APC: antigen-presenting cell.
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- Figure 2 Astragalus polysaccharide (PG2) enhances M1 polarization and down-regulates IL-4/IL-13-induced M2 polarization. Images from flow cytometric analyses showing ( A ) the differentiation of THP-1 monocyte into macrophages after 24 h incubation in phorbol 12-myristate 13-acetate (PMA), ( B ) incubation of MDM in IL-4 and IL-13 induced a CD206 high CD80 low M2 phenotype, while ( C ) incubation of MDM in PG2 16 mg/mL for 48 h induced a CD80 high CD206 low M1 phenotype. ( D ) A graphical representation of the differential effect of IL-4/IL-13 and low dose (8 mg/mL) or high dose (16 mg/mL) PG2 treatment on M1-M2 polarization. ** p < 0.01
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- Figure 3 The enhancement of M1 macrophage polarization by PG2 is akin to the effect of LPS/IFN-gamma stimulation of MDMs. Images from flow cytometric analyses showing ( A ) the differentiation of THP-1 monocyte into macrophages after 24 h incubation in PMA; ( B ) MDMs after exposure to IFN-gamma and LPS induced a CD80 high CD206 low M1 phenotype; ( C ) cancer cell culture medium (CCCM) induced 17.1% CD80+ and 61.9% CD206+ MDMs; ( D ) PG2-treatment of MDMs pre-incubated in CCCM induced a CD80 high CD206 low M1 phenotype, similar to IFN-gamma/LPS exposure; ( E ) a graphical representation of the differential effect of IFN-gamma/LPS, CCCM, and 16 mg/mL PG2 treatment on M1-M2 polarization. PG2 enhanced the M1 phenotype akin to IFN-gamma/LPS exposure effect. ** p < 0.01
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- Figure 6 PG2 modulated the CD80+ M1/CD206+ M2 macrophage population and increased the population of CD80+, CD103+, and CD86+ dendritic cells derived from peripheral blood mononuclear cells (PBMCs) of cancer patient's ex vivo. The effect of PMA, LPS+INF-gamma, or PG2 on the proportion of ( A ) CD80+ and ( B ) CD206+ cells, as shown by flow cytometry. Graphical representation of the effect of PG2 on the population of ( C ) CD80+, ( D ) CD103+, and ( E ) CD86+ dendritic cells derived from GM-CSF+IL-4-treated PBMCs of NSCLC patients. ( F ) Graphical representation of the effect of PG2 on the number of functional CD103+ dendritic cells derived from GM-CSF+IL-4-treated PBMCs of breast, colon, ovarian, or liver patients. 1: GM-CSF+IL4; 2: GM-CSF+IL4, followed by the treatment of PG2 (16 mg/mL); 3: GM-CSF+IL4, washed out, and followed by the treatment of PG2 (16 mg/mL); * p < 0.05, ** p < 0.01.
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- Figure 3 Exogenous hrANXA1 controls human and mouse macrophage polarization in vitro. Human PBMC-derived macrophages were incubated for 24 hours with lipopolysaccharide plus IFN-gamma to promote an M1-like phenotype, before addition of human recombinant ANXA1 (hrANXA1; 10 nM) for a further 6 hours. ( A and B ) Median fluorescence intensity (MFI) units measured by flow cytometry of MHCII proinflammatory ( A ) and CD206 antiinflammatory ( B ) markers. Shown are MFI quantification (left) and representative FACS plots (right). ( C - F ) Quantitative reverse transcriptase PCR analysis of Tnfa ( C ) and Nos-2 ( D ) proinflammatory genes, and Il-10 ( E ) and Tgfb1 ( F ) antiinflammatory genes. ( G and H ) MFI units as measured by flow cytometry of MHCII proinflammatory ( G ) and CD206 antiinflammatory ( H ) markers after treatment by hrANXA1 in the presence or absence of the FPR2/ALX antagonist WRW 4 (10 muM). Experiments were performed on PBMCs from 6 ( A - F ) or 3 ( G and H ) independent donors. Each color represents an independent PBMC donor. * P < 0.05 vs. vehicle; # P < 0.05 vs. control.
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