Journal: Cell

Article Title: RAB11FIP5 Expression and Altered Natural Killer Cell Function Are Associated with Induction of HIV Broadly Neutralizing Antibody Responses

doi: 10.1016/j.cell.2018.08.064

Figure Lengend Snippet:

Article Snippet: Human KIR3DL2/CD158k APC-conjugated Antibody, Clone # 539304 , R and D Systems , Cat# FAB2878A.

Techniques: Recombinant, Staining, Cell Isolation, Enzyme-linked Immunosorbent Assay, Marker, Library Quantification, Gene Expression, Sequencing, Software

Journal: Breast cancer research and treatment

Article Title: Viral transduction of the HER2-extracellular domain expands trastuzumab-based photoimmunotherapy for HER2-negative breast cancer cells.

doi: 10.1007/s10549-015-3265-y

Figure Lengend Snippet: Fig. 1 HER2-extracellular domain (ECD) expression induced by Ad- HER2-ECD. a Western blot analysis showing expression of the 100-kDa HER2-ECD protein in Ad-HER2-ECD-infected cells but not in Ad-GFP-infected or parental cells. Actin was used as a loading control. b Flowcytometric analysis showing HER2-ECD expression on the cell membrane. Cells were labeled with APC-conjugated

Article Snippet: In experiments with replication-deficient adenoviral vector, cells were infected with Ad-HER2-ECD or Ad-GFP at a multiplicity of infection (MOI) of 50 for 48 h. Flowcytometric analysis To measure the expression of HER2-ECD in cells infected with Ad-HER2-ECD, cells were labeled with APC-conjugated mouse monoclonal anti-HER2-ECD antibody (R&D Systems Inc.) or APC-conjugated IgG2b as control (Miltenyi Biotec, Inc.) on ice for 45 min and were then analyzed using a FACS instrument (BD Biosciences).

Techniques: Expressing, Western Blot, Infection, Control, Membrane, Labeling

Journal: Breast cancer research and treatment

Article Title: Viral transduction of the HER2-extracellular domain expands trastuzumab-based photoimmunotherapy for HER2-negative breast cancer cells.

doi: 10.1007/s10549-015-3265-y

Figure Lengend Snippet: Fig. 2 Trastzumab-IR700 binds to the transduced cell surface HER2- ECD Immunofluorescent analysis of Trastzumab-IR700 (Tra-IR700) binding to HER2-ECD-transduced by Ad-HER2-ECD on the cell surface of breast cancer cells. Scale bars 50 lm

Article Snippet: In experiments with replication-deficient adenoviral vector, cells were infected with Ad-HER2-ECD or Ad-GFP at a multiplicity of infection (MOI) of 50 for 48 h. Flowcytometric analysis To measure the expression of HER2-ECD in cells infected with Ad-HER2-ECD, cells were labeled with APC-conjugated mouse monoclonal anti-HER2-ECD antibody (R&D Systems Inc.) or APC-conjugated IgG2b as control (Miltenyi Biotec, Inc.) on ice for 45 min and were then analyzed using a FACS instrument (BD Biosciences).

Techniques: Binding Assay

Journal: Breast cancer research and treatment

Article Title: Viral transduction of the HER2-extracellular domain expands trastuzumab-based photoimmunotherapy for HER2-negative breast cancer cells.

doi: 10.1007/s10549-015-3265-y

Figure Lengend Snippet: Fig. 3 Microscopic analysis of the effect of Tra-IR700-mediated PIT on the cell morphology and cell death of HER2-ECD-transduced HER2-negative cells. a Phase contrast analysis of the morphology of the breast cancer cells MCF-7 and MDA-MB-231 immediately following PIT using 0, 6, or 18 J. Scale bars 50 lm. b Phase contrast analysis of control, Ad-GFP-infected or Ad-HER2-ECD-infected MCF-7 and MDA-MB-231 cells before and after 72 h treatment with

Article Snippet: In experiments with replication-deficient adenoviral vector, cells were infected with Ad-HER2-ECD or Ad-GFP at a multiplicity of infection (MOI) of 50 for 48 h. Flowcytometric analysis To measure the expression of HER2-ECD in cells infected with Ad-HER2-ECD, cells were labeled with APC-conjugated mouse monoclonal anti-HER2-ECD antibody (R&D Systems Inc.) or APC-conjugated IgG2b as control (Miltenyi Biotec, Inc.) on ice for 45 min and were then analyzed using a FACS instrument (BD Biosciences).

Techniques: Control, Infection

Journal: Breast cancer research and treatment

Article Title: Viral transduction of the HER2-extracellular domain expands trastuzumab-based photoimmunotherapy for HER2-negative breast cancer cells.

doi: 10.1007/s10549-015-3265-y

Figure Lengend Snippet: Fig. 4 Effect of Tra-IR700 mediated PIT treatment of HER2-ECD transduced HER2-negative cells on cell viability. Tra-IR700-medi- ated PIT was applied to the indicated cells along with seven control conditions and cell viability was quantified 72 h after PIT (24 J for MDA-MB-231 and 36 J for MCF-7) using the XTT assay. Only the

Article Snippet: In experiments with replication-deficient adenoviral vector, cells were infected with Ad-HER2-ECD or Ad-GFP at a multiplicity of infection (MOI) of 50 for 48 h. Flowcytometric analysis To measure the expression of HER2-ECD in cells infected with Ad-HER2-ECD, cells were labeled with APC-conjugated mouse monoclonal anti-HER2-ECD antibody (R&D Systems Inc.) or APC-conjugated IgG2b as control (Miltenyi Biotec, Inc.) on ice for 45 min and were then analyzed using a FACS instrument (BD Biosciences).

Techniques: Control, XTT Assay

Journal: Cancer Communications

Article Title: Unfolded protein response kinase PERK supports survival and metastasis of circulating tumor cell clusters via SAM synthesis and H3K4me3‐dependent PDGFB signaling

doi: 10.1002/cac2.70072

Figure Lengend Snippet: Activated UPR in CTC clusters protects cells from cell death. (A) Schematic diagram of the experimental design for isolating CTCs from mice for RNA sequencing at week 4 following fat pad inoculation with MDA‐MB‐231/LM2 cells. (B) GO pathway enrichment analysis of upregulated gene sets in CTC clusters compared to single CTCs ( n = 3). (C) GSVA enrichment scores of single CTCs and CTC clusters from breast cancer patients and PDX‐Br16 mouse model. The single‐cell RNA sequencing data were obtained from the GEO database ( GSE111065 ). (D) Relative mRNA expression levels of UPR‐related genes in single CTCs and CTC clusters from the NCG‐MDA‐MB‐231/LM2 mouse model ( n = 3). (E) Representative flow cytometry plots showing the gating strategy for identifying ER tracker intensity in single CTCs and CTC clusters from the blood of NCG‐MDA‐MB‐231/LM2 mice. (F) MFI of TPE‐MI in single CTCs and CTC clusters, as determined by flow cytometry ( n = 5). G. Percentage of Annexin V + single and clustered tumor cells pretreated with thapsigargin ( n = 3). (H) Cell viability of single and clustered tumor cells under thapsigargin treatment ( n = 3). (I) Schematic diagram of the experimental design. Clustered tumor cells were pretreated with vehicle or azoramide for 6 h prior to tail vein injection into NCG mice, followed by assessment of disseminated tumor cells in the lung at 24 h. (J‐K) Bioluminescence images (left) and quantification of fluorescence signal intensity (right) of mice (J) and the lungs (K) to assess lung metastasis at 24 h after tail vein injection of clustered MDA‐MB‐231/LM2‐CTC cells ( n = 5). Results represent mean ± SD. Student's t‐test in C, D, F, J, and K, two‐way ANOVA test in G and H. * P < 0.05; ** P < 0.01; *** P < 0.001; ns, not significant. Abbreviations: CTCs, circulating tumor cells; RNA‐seq, RNA sequencing; GO, gene ontology; GSVA, gene set variation analysis; PDX‐Br16, patient‐derived xenograft‐Br16; GEO, ​gene expression omnibus​; NCG, NOD/ShiLtJGpt‐Prkdc em26Cd52 Il2rg em26Cd22 /Gpt; TPE‐MI, tetraphenylethene‐maleimide; MDA‐MB‐231/LM2​, ​MDA‐MB‐231 lung metastasis 2​; UPR, unfolded protein response; HSPA5, heat shock protein family A member 5; XBP1, x‐box binding protein 1; DDIT3, DNA damage inducible transcript 3; DNAJB1, dnaJ heat shock protein family member B1; ATF3, activating transcription factor 3; ATF4, activating transcription factor 4; PPP1R15A, protein phosphatase 1 regulatory subunit 15A; ER Tracker, endoplasmic reticulum tracker; MFI, mean fluorescence intensity; Azo, azoramide; IV, intravenous; ANOVA, analysis of variance.

Article Snippet: For annexin V detection analysis, Annexin V‐Elab Fluor 647/propidium iodide (PI) Apoptosis Kit (E‐CK‐A213, Elabscience, Wuhan, Hubei, China) or Annexin V‐allophycocyanin (APC)/4',6‐diamidino‐2‐phenylindole (DAPI) Apoptosis Kit (E‐CK‐A258, Elabscience) was employed to detect Annexin V + cells.

Techniques: RNA Sequencing, Expressing, Flow Cytometry, Injection, Fluorescence, Derivative Assay, Binding Assay

Journal: Cancer Communications

Article Title: Unfolded protein response kinase PERK supports survival and metastasis of circulating tumor cell clusters via SAM synthesis and H3K4me3‐dependent PDGFB signaling

doi: 10.1002/cac2.70072

Figure Lengend Snippet: PERK is a key factor in promoting CTC cluster survival. (A) Relative mRNA expression levels of UPR‐related genes in single CTCs and CTC clusters isolated from NCG‐MDA‐MB‐231/LM2 (left) and C57BL/6‐B16F10 (right) mouse models ( n = 3). (B) Percentage of Annexin V + single and clustered MDA‐MB‐231/LM2‐CTC (left) and B16F10 (right) cells transfected with siRNAs against PERK and IRE1α and treated with or without thapsigargin ( n = 3). (C) Percentage of Annexin V + clustered MDA‐MB‐231/LM2‐CTC (left) and B16F10 (right) cells treated with AMG44 prior to thapsigargin ( n = 3). (D) Relative PERK expression in single CTCs and CTC clusters isolated from BC patients and PDX mouse models (PDX‐1, PDX‐2, and PDX‐3) ( n = 15 for patients; n = 10 for PDX‐1; n = 8 for PDX‐2 and PDX‐3). (E) Schematic design of the experimental design. MDA‐MB‐231/LM2 cells were orthotopically implanted into mice, and lung metastases were analyzed via IHC at week 4. Representative IHC images of pPERK in lung metastases. The arrow indicates single CTCs; the dashed circle indicates CTC cluster. (F‐I) Representative immunofluorescence images of single CTCs and CTC clusters from a patient and the NCG‐MDA‐MB‐231/LM2 mouse model. Cells were stained for pPERK (red), EGFR (green) (F) or TUNEL (purple) (H), GFP (green) (G) or TUNEL (purple) (I), and DAPI (blue). (J) Schematic design of the experiment (left). MDA‐MB‐231/LM2‐CTC cells were pretreated with Vehicle, MK‐28 or AMG44 and then cultured in suspension for 24 h. Percentage of Annexin V + tumor cell clusters was analyzed by flow cytometry (right) ( n = 5). Results represent mean ± SD. Student's t‐test in A and D, one‐way ANOVA test in B, C, and J. * P < 0.05; ** P < 0.01; *** P < 0.001; ns, not significant. Abbreviations: UPR, unfolded protein response; CTCs, circulating tumor cells; NCG, NOD/ShiLtJGpt‐Prkdc em26Cd52 Il2rg em26Cd22 /Gpt; MDA‐MB‐231/LM2​, ​MDA‐MB‐231 lung metastasis 2​; ATF6, activating transcription factor 6; PERK, protein kinase R (PKR)‐like endoplasmic reticulum kinase; IRE1α, inositol‐requiring enzyme 1 alpha; BC, breast cancer; PDX, patient‐derived xenograft; IHC, immunohistochemistry; pPERK, phosphorylated PERK; DAPI, 4′,6‐diamidino‐2‐phenylindole; EGFR, epidermal growth factor receptor; GFP, green fluorescent protein; TUNEL, terminal deoxynucleotidyl transferase dUTP nick‐end labeling; siRNA, small interfering RNA;ANOVA, analysis of variance.

Article Snippet: For annexin V detection analysis, Annexin V‐Elab Fluor 647/propidium iodide (PI) Apoptosis Kit (E‐CK‐A213, Elabscience, Wuhan, Hubei, China) or Annexin V‐allophycocyanin (APC)/4',6‐diamidino‐2‐phenylindole (DAPI) Apoptosis Kit (E‐CK‐A258, Elabscience) was employed to detect Annexin V + cells.

Techniques: Expressing, Isolation, Transfection, Immunofluorescence, Staining, TUNEL Assay, Cell Culture, Suspension, Flow Cytometry, Derivative Assay, Immunohistochemistry, Small Interfering RNA

Journal: Cancer Communications

Article Title: Unfolded protein response kinase PERK supports survival and metastasis of circulating tumor cell clusters via SAM synthesis and H3K4me3‐dependent PDGFB signaling

doi: 10.1002/cac2.70072

Figure Lengend Snippet: PERK signaling regulates MAT2A via ATF4 to support enhanced methionine metabolism. (A) KEGG enrichment analysis of upregulated genes in scramble control tumor cell clusters compared to PERK‐KO clusters ( n = 3). (B) Volcano plot showing PERK‐related metabolites identified through targeted metabolomics ( n = 6). (C) Heatmap showing metabolites involved in the methionine cycle in scramble and PERK‐KO tumor cell clusters ( n = 6). (D) Intracellular metabolites in NC‐oe and PERK‐oe tumor cell clusters ( n = 6). (E) The schematic diagram for the conversion of [ 13 C 5 ]‐methionine into various metabolites (left) and LC‐MS quantification of M+5 methionine, M+5 SAM, and M+4 SAH following a 16 h incubation with [ 13 C 5 ]‐methionine in scramble and PERK‐KO tumor cell clusters (right) ( n = 4). (F) Percentage of Annexin V + MDA‐MB‐231/LM2‐CTC cells cultured in CM or medium lacking Ser, Gly, Met, or Cys for 24 h ( n = 5). (G‐I) Western blot analysis of metabolic enzyme expression in MDA‐MB‐231/LM2‐CTC and B16F10 tumor cell clusters with or without PERK (G), ATF4 expression in MDA‐MB‐231/LM2‐CTC with or without PERK (H), and ATF4 and MAT2A expression in MDA‐MB‐231/LM2‐CTC with or without ATF4 (I). (J) Relative MAT2A expression in MDA‐MB‐231/LM2‐CTC transfected with shNC or shATF4 #2 ( n = 3). (K) Percentage of Annexin V + MDA‐MB‐231/LM2‐CTC transduced with shNC or shMAT2A #3 and treated with or without SAM for 24 h (left) or pretreated with PF9366 prior to SAM (right) ( n = 3). (L) Schematic of the in vivo experiment design. (M) Bioluminescence imaging (left) and fluorescence intensity quantification (right) of lung metastases at 24 h after tail vein injection ( n = 5). (N) Representative H&E staining images (left) and quantification of pulmonary nodules (right) in NCG mice at 2 weeks after tail vein injection of MDA‐MB‐231/LM2‐CTC clusters (shNC and shMAT2A #3) ( n = 5). (O) Schematic representation of the experimental design. MDA‐MB‐231/LM2‐CTC clusters were cultured in CM or MRM for 24 h before tail vein injection into NCG mice, with lung metastases assessed at 24 h. (P) Bioluminescence imaging (left) and fluorescence intensity quantification (right) of lung metastases at 24 h after tail vein injection ( n = 5). Results represent mean ± SD. Student's t‐test in D, E, J, M, and P; one‐way ANOVA test in F and K. * P < 0.05; ** P < 0.01; *** P < 0.001; ns, not significant. Abbreviations: KEGG, Kyoto Encyclopedia of Genes and Genomes; FC, fold change; Sc, scramble; PERK, Protein kinase RNA‐like endoplasmic reticulum kinase; PERK‐KO, PRKR‐knockout; sgPERK, single‐guide RNA targeting PERK; LC‐MS, liquid chromatography‐mass spectrometry; Met, methionine; SAM; S‐adenosylmethionine; SAH, S‐adenosylhomocysteine; CM, control medium; Ser, serine; Gly, glycine; Cys, cysteine; PSAT1, phosphoserine aminotransferase 1; PHGDH, phosphoglycerate dehydrogenase; MTHFR, methylenetetrahydrofolate reductase; MAT2A, methionine adenosyltransferase 2A; ATF4, activating transcription factor 4; shNC, negative control short hairpin RNA; NC‐oe, negative control‐overexpression; PERK‐oe, PERK‐overexpression; shATF4, short hairpin RNA of ATF4; shMAT2A, short hairpin RNA of MAT2A; MRM, methionine‐restricted medium; NCG, NOD/ShiLtJGpt‐Prkdc em26Cd52 Il2rg em26Cd22 /Gpt; MDA‐MB‐231/LM2​, ​MDA‐MB‐231 lung metastasis 2​; CTCs, circulating tumor cells; BLI, lung Bioluminescence imaging; H&E, hematoxylin and eosin; ANOVA, analysis of variance.

Article Snippet: For annexin V detection analysis, Annexin V‐Elab Fluor 647/propidium iodide (PI) Apoptosis Kit (E‐CK‐A213, Elabscience, Wuhan, Hubei, China) or Annexin V‐allophycocyanin (APC)/4',6‐diamidino‐2‐phenylindole (DAPI) Apoptosis Kit (E‐CK‐A258, Elabscience) was employed to detect Annexin V + cells.

Techniques: Control, Liquid Chromatography with Mass Spectroscopy, Incubation, Cell Culture, Western Blot, Expressing, Transfection, Transduction, In Vivo, Imaging, Fluorescence, Injection, Staining, Knock-Out, Liquid Chromatography, Mass Spectrometry, Negative Control, shRNA, Over Expression

Journal: Cancer Communications

Article Title: Unfolded protein response kinase PERK supports survival and metastasis of circulating tumor cell clusters via SAM synthesis and H3K4me3‐dependent PDGFB signaling

doi: 10.1002/cac2.70072

Figure Lengend Snippet: Methionine metabolism promotes H3K4me3 methylation modification to upregulate PDGFB expression in CTC clusters. (A) Western blot analysis of PERK, MAT2A, and H3K4me3 levels in control and PERK‐KO MDA‐MB‐231/LM2‐CTC clusters treated with or without PF9366 or SAM. (B) The volcano plot depicting differentially expressed genes affected by methionine ( n = 3). (C) Metaplot comparing H3K4me3 enrichment profiles in MDA‐MB‐231/LM2‐CTC clusters cultured in CM or MRM. The plot is centered on TSS (±3.0 Kb) ( n = 3). (D) Bioinformatics analysis filtered out 10 survival‐related genes as downstream targets of H3K4me3. (E) Heatmap showing survival‐associated gene expression in MDA‐MB‐231/LM2‐CTC clusters under CM or MRM conditions ( n = 3). (F) Genome browser tracks of H3K4me3 at the PDGFB gene locus by ChIP‐seq ( n = 3). (G) TEM images of MDA‐MB‐231/LM2‐CTC cluster. (H‐J) Representative immunofluorescence images (left) and quantification (right) of PDGFB in CTC clusters from NCG‐MDA‐MB‐231/LM2 mouse model (H, n = 20); or in the complete or dispersed CTC clusters (I, n = 15); or in control or PDGFB‐knockdown CTC clusters (J, n = 15). Arrows indicate cell‐cell border; circle indicates intracellular regions. (K) Representative images of flow cytometry (left) and quantification of AnnexinV + MDA‐MB‐231/LM2‐CTC clusters (right) in control (shNC) and PDGFB‐knockdown (shP#1/2/3) groups ( n = 5). (L) Schematic representation of the experimental design. MDA‐MB‐231/LM2‐CTC clusters (shNC and shPDGFB #3) were injected into NCG mice via the tail vein, and metastatic burden was assessed by bioluminescence imaging at 24 h. (M‐N) Bioluminescence images (left) and quantification of fluorescence intensity (right) showing lung metastasis after tail vein injection at 24 h ( n = 5). Results represent mean ± SD. One‐way ANOVA test in A and K; student's t‐test in H, I, J, M, and N. * P < 0.05; ** P < 0.01; *** P < 0.001; ns, not significant. Abbreviations: MDA‐MB‐231/LM2​, ​MDA‐MB‐231 lung metastasis 2​; CTCs, circulating tumor cells; FC, fold change; PERK, PRKR‐like endoplasmic reticulum kinase; ERK‐KO, PRKR‐knockout; sgPERK, single‐guide RNA targeting PERK; MAT2A, methionine adenosyltransferase 2A; H3K4me3, trimethylation of histone H3 at lysine 4; SAM, S‐adenosylmethionine; CM, control medium; MRM, methionine‐restricted medium; No., number; TSS, transcription start site; ChIP‐seq, chromatin immunoprecipitation sequencing; RNA‐seq, RNA sequencing; PDGFB, platelet‐derived growth factor subunit B; IGF2BP3, insulin‐like growth factor 2 mRNA‐binding protein 3; PHGDH, phosphoglycerate dehydrogenase; ANXA11, annexin A11; ITGA6, integrin subunit alpha 6; ITGB4, integrin subunit beta 4; LOX, lysyl oxidase; PIK3C2B, phosphatidylinositol‐4‐phosphate 3‐kinase catalytic subunit type 2 beta; IRF9, interferon regulatory factor 9; ATP2A3, ATPase sarcoplasmic/endoplasmic reticulum Ca 2+ transporting 3; TEM, transmission electron microscopy; DAPI, 4′,6‐diamidino‐2‐phenylindole; shNC, negative control short hairpin RNA; shP#, short hairpin RNA targeting PDGFB; NCG, NOD/ShiLtJGpt‐Prkdc em26Cd52 Il2rg em26Cd22 /Gpt; BLI, lung Bioluminescence imaging; ANOVA, analysis of variance.

Article Snippet: For annexin V detection analysis, Annexin V‐Elab Fluor 647/propidium iodide (PI) Apoptosis Kit (E‐CK‐A213, Elabscience, Wuhan, Hubei, China) or Annexin V‐allophycocyanin (APC)/4',6‐diamidino‐2‐phenylindole (DAPI) Apoptosis Kit (E‐CK‐A258, Elabscience) was employed to detect Annexin V + cells.

Techniques: Methylation, Modification, Expressing, Western Blot, Control, Cell Culture, Gene Expression, ChIP-sequencing, Immunofluorescence, Knockdown, Flow Cytometry, Injection, Imaging, Fluorescence, Knock-Out, RNA Sequencing, Derivative Assay, Binding Assay, Transmission Assay, Electron Microscopy, Negative Control, shRNA

Journal: Chemical Engineering Journal

Article Title: Cryogenically 3D printed biomimetic scaffolds containing decellularized small intestinal submucosa and Sr2+/Fe3+ co-substituted hydroxyapatite for bone tissue engineering

doi: 10.1016/j.cej.2021.133459

Figure Lengend Snippet: Fig. 5. In vitro RAW cells polarization modulation after cultured on scaffolds for 3 days. (A) Calcein-AM staining of RAW; (B) RAW cells polarization examined by double marked immunofluorescence of NOS2/Arg-1; (C) RAW polarization quantified by double marked flow cytometry of CD86/CD206; (D) Quantitative CD206 and CD86 expression ratios; (E) RT-qPCR analysis of relative IL6, IL1-β, TNF-α, Arg-1 and IL10 gene expressions; (F) RT-qPCR analysis of relative VEGF, PDGF-BB, TGF-β1 and BMP2 gene expressions. Data represent the mean ± SD. *p < 0.05 versus SIS; #p < 0.05 versus SIS/FeHA.

Article Snippet: Briefly, RAW cells were harvested and incubated with CD86 antibody (553692, BD Pharmingen, USA) under 4 °C for 30 min. Then, cells were washed, resuspended, and incubated with CD206 antibody (FAB25351A, R&D Systems, USA) again to be analyzed using the flow cytometer (Beckman Coulter, CytoFLEX).

Techniques: In Vitro, Cell Culture, Staining, Immunofluorescence, Flow Cytometry, Expressing, Quantitative RT-PCR

Journal: Chemical Engineering Journal

Article Title: Cryogenically 3D printed biomimetic scaffolds containing decellularized small intestinal submucosa and Sr2+/Fe3+ co-substituted hydroxyapatite for bone tissue engineering

doi: 10.1016/j.cej.2021.133459

Figure Lengend Snippet: Fig. 7. In vivo immunomodulation and vascularization of scaffolds following 7 days subcutaneous implantation. (A) The histological graphs of implanted scaffolds : from the general view of horizontal HE sections to internal sites stained by HE, NOS2, CD206, and CD31 IHC in similar positions; (B). Quantitation of CD206 and NOS2 positive stained areas; (C) Quantitative CD206/NOS2 ratios; (D) Quantitation of new vessels density. Red arrows indicate neo-vessels stained by CD31. Data represent the mean ± SD. *p < 0.05 versus SIS; &p < 0.05 versus SIS/FeHA; #p < 0.05 versus SIS/SrHA.

Article Snippet: Briefly, RAW cells were harvested and incubated with CD86 antibody (553692, BD Pharmingen, USA) under 4 °C for 30 min. Then, cells were washed, resuspended, and incubated with CD206 antibody (FAB25351A, R&D Systems, USA) again to be analyzed using the flow cytometer (Beckman Coulter, CytoFLEX).

Techniques: In Vivo, Staining, Quantitation Assay