mouse monoclonal antihuman cd163  (Novus Biologicals)

Journal: World Journal of Surgical Oncology

Article Title: Hypoxic lung cancer cell-derived exosomal miR-21 mediates macrophage M2 polarization and promotes cancer cell proliferation through targeting IRF1

doi: 10.1186/s12957-022-02706-y

Figure Lengend Snippet: Hypoxic lung cancer cells promote macrophage M2 polarization. A IHC was performed to quantify the CD163 protein expression in clinical tissues. Scale bar = 400 μm. B Flow cytometry was used to detect the quantity of M2 macrophage through labeling the protein CD206 and CD163. GW4869 was used as the inhibitor of exosome secretion. C The quantitative analysis of flow cytometry results. D RT-qPCR was for the marker genes examining in mTHP-1 that co-cultured with the normoxia and hypoxia lung cancer cell H1299. Data was shown as mean ± SD. n = 3. * P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: After multiple steps, including dewaxed and rehydrated, the sections were stained with mouse monoclonal antihuman CD163 antibody (1:1000, Abcam, USA) at 4 °C overnight, then incubated with the secondary antibody (1:2000, Abcam, USA) for 30 min at 37 °C, and finally counterstained with hematoxylin.

Techniques: Expressing, Flow Cytometry, Labeling, Quantitative RT-PCR, Marker, Cell Culture

Journal: World Journal of Surgical Oncology

Article Title: Hypoxic lung cancer cell-derived exosomal miR-21 mediates macrophage M2 polarization and promotes cancer cell proliferation through targeting IRF1

doi: 10.1186/s12957-022-02706-y

Figure Lengend Snippet: Exosomes from hypoxic lung cancer cells promote macrophage M2 polarization. A Exosome morphological characteristics were determined by transmission electron microscope. Scale bar = 200 nm. B Dynamic light scattering was used to detect the exosome size. C The expression of exosome protein biomarkers CD9, CD81, and TS101 was measured by western blot. D The immunofluorescence image shows the internalization of PKH67-labeled exosomes by macrophage. Scale bar = 20 μm. E Flow cytometry was used to detect the quantity of CD163 + CD206 + M2 macrophage after treated with normoxia or hypoxia exosome for 48 h. F RT-qPCR was for the marker genes examining in macrophages that co-cultured with the normoxia and hypoxia exosomes, including TNF, Arg1, IL-10, and TGF-β. Data was expressed as mean ± SD. n = 3. * P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: After multiple steps, including dewaxed and rehydrated, the sections were stained with mouse monoclonal antihuman CD163 antibody (1:1000, Abcam, USA) at 4 °C overnight, then incubated with the secondary antibody (1:2000, Abcam, USA) for 30 min at 37 °C, and finally counterstained with hematoxylin.

Techniques: Transmission Assay, Microscopy, Expressing, Western Blot, Immunofluorescence, Labeling, Flow Cytometry, Quantitative RT-PCR, Marker, Cell Culture

Journal: World Journal of Surgical Oncology

Article Title: Hypoxic lung cancer cell-derived exosomal miR-21 mediates macrophage M2 polarization and promotes cancer cell proliferation through targeting IRF1

doi: 10.1186/s12957-022-02706-y

Figure Lengend Snippet: MiR-21 enriched in exosomes promotes macrophage M2 polarization. A RNA sequence was performed to analyze the different expression miRNAs between the exosomes secreted by H1299 cultured in normoxia (N) and hypoxia (H). B The top 10 differentially expressed miRNAs was validated by RT-qPCR in clinical tissues of lung cancer patients. The relative expressions were averaged and plotted. C The expression of miR-21 in mTHP-1 after transfected mimic or inhibitor and corresponding control was detected by RT-qPCR. D The quantity of CD163 + CD206 + M2 macrophage after overexpress or knockdown miR-21 in mTHP-1 was by using flow cytometry. E RT-qPCR was for the marker genes examining in macrophages that co-cultured with the normoxia and hypoxia exosomes, including TNF, Arg1, IL-10, and TGF-β. F Clone formation assay of H1299 was assayed after co-culture with miR-21 knockdown or overexpression of mTHP-1. Data was expressed as mean ± SD. n = 3. * P < 0.05,** P < 0.01, *** P < 0.001

Article Snippet: After multiple steps, including dewaxed and rehydrated, the sections were stained with mouse monoclonal antihuman CD163 antibody (1:1000, Abcam, USA) at 4 °C overnight, then incubated with the secondary antibody (1:2000, Abcam, USA) for 30 min at 37 °C, and finally counterstained with hematoxylin.

Techniques: Sequencing, Expressing, Cell Culture, Quantitative RT-PCR, Transfection, Control, Knockdown, Flow Cytometry, Marker, Tube Formation Assay, Co-Culture Assay, Over Expression

Journal: World Journal of Surgical Oncology

Article Title: Hypoxic lung cancer cell-derived exosomal miR-21 mediates macrophage M2 polarization and promotes cancer cell proliferation through targeting IRF1

doi: 10.1186/s12957-022-02706-y

Figure Lengend Snippet: MiR-21 regulates M2 polarization by targeting IRF1. A The sequence of miR-21 binding sites in 3′UTR of IRF1. B The binding relationship between miR-21 and IRF1 was verified through luciferase reporter assay. ** P < 0.01, compared with the mimic NC group. C Western blot was for IRF1 protein detecting. ** P < 0.01, compared with the mimic NC group; ## P < 0.01, compared with the inhibitor NC group. D Western blot was for IRF1 protein detecting. E RT-qPCR detected the expression of TNF, Arg1, IL-10, and TGF-β in mTHP-1 after overexpression of IRF1 or both IRF1 and miR-21. F Flow cytometry was for the CD163 + CD206 + M2 macrophage quantiting after overexpression of IRF1 or both IRF1 and miR-21. G Clone formation assay of H1299 was assayed after co-culture with mTHP-1 with overexpressed IRF1 or both IRF1 and miR-21. Data were expressed as mean ± SD. n = 3. * P < 0.05,** P < 0.01, *** P < 0.001

Article Snippet: After multiple steps, including dewaxed and rehydrated, the sections were stained with mouse monoclonal antihuman CD163 antibody (1:1000, Abcam, USA) at 4 °C overnight, then incubated with the secondary antibody (1:2000, Abcam, USA) for 30 min at 37 °C, and finally counterstained with hematoxylin.

Techniques: Sequencing, Binding Assay, Luciferase, Reporter Assay, Western Blot, Quantitative RT-PCR, Expressing, Over Expression, Flow Cytometry, Tube Formation Assay, Co-Culture Assay

Journal: World Journal of Surgical Oncology

Article Title: Hypoxic lung cancer cell-derived exosomal miR-21 mediates macrophage M2 polarization and promotes cancer cell proliferation through targeting IRF1

doi: 10.1186/s12957-022-02706-y

Figure Lengend Snippet: Exosomal miR-21 facilitates cancer cell proliferation and induces macrophage M2 polarization in vivo. H1299 cells mixed with or without the conditioned mTHP-1 stimulated by normoxia or hypoxia H1299 exosomes or transfected miR-21 mimic or NC mimic respectively. The mixed cells solution was injected into mice subcutaneous. A Photographs of tumors. B Growth curve of tumors. C The level of miR-21 in lung cancer tissues was detected by RT-qPCR assay. D IHC was performed to detect the CD163 expression in tumor tissues. Scale bar = 100 μm. E RT-qPCR assay was used to assess M2 polarization-related factors (Arg1, IL-10, and TGF-β) expression.* P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: After multiple steps, including dewaxed and rehydrated, the sections were stained with mouse monoclonal antihuman CD163 antibody (1:1000, Abcam, USA) at 4 °C overnight, then incubated with the secondary antibody (1:2000, Abcam, USA) for 30 min at 37 °C, and finally counterstained with hematoxylin.

Techniques: In Vivo, Transfection, Injection, Quantitative RT-PCR, Expressing

Journal: Journal of Innate Immunity

Article Title: The Gene Signature of Activated M-CSF-Primed Human Monocyte-Derived Macrophages Is IL-10-Dependent

doi: 10.1159/000519305

Figure Lengend Snippet: In vivo co-expression of Gene set 1. a, b Summary of GSEA on the ranked list of genes from the comparison of the transcriptome of M-MØ + LPS and GM-MØ + LPS using previously defined gene sets for (a) COVID-19 BALF macrophages [54, 55, 56] or (b) COVID-19 pathogenic monocyte-derived macrophage subsets [10, 11, 16]. c Gene ontology of Gene set 1 on the “COVID-19-related gene sets” database using Enrichr. d GSEA on the ranked list of the comparison of the transcriptome of PBMC from COVID-19 patients versus control PBMC [57], using Gene set 1 genes. The identity of the genes within the leading edge is shown. e Correlation of the expression on Gene set 1 genes with the expression of CD163 or SPI1 in breast carcinoma, as calculated using TIMER (http://timer.cistrome.org), with indication of the percentage of Gene set 1 genes whose positive correlation with the indicated gene is p < 0.05. LPS, lipopolysaccharide; GSEA, gene set enrichment analysis; BALF, bronchoalveolar lavage; PBMC, peripheral blood mononuclear cells; M-MØ, monocyte-derived human macrophages.

Article Snippet: After deparaffinization, rehydration, and antigen-retrieval using the automated PT Link system (Dako, Agilent Technologies), tissue microarray slides were incubated with a mixture of a mouse monoclonal antihuman CD163 (NB110-59935; Novus Biologicals) and a rabbit polyclonal antihuman CCL19 (NBP2-56275; Novus Biologicals) overnight at 4°C.

Techniques: In Vivo, Expressing, Comparison, Derivative Assay, Control

Journal: The Journal of Infectious Diseases

Article Title: Macrophage Activation Marker Soluble CD163 Is a Dynamic Marker of Liver Fibrogenesis in Human Immunodeficiency Virus/Hepatitis C Virus Coinfection

doi: 10.1093/infdis/jiy331

Figure Lengend Snippet: Human immunodeficiency virus (HIV) and hepatitis C virus (HCV) coinfection cooperatively elevate soluble CD163 (sCD163) levels. sCD163 levels in treatment-naive subjects with chronic HCV, HIV, or HIV/HCV coinfection; antiretroviral therapy (ART)–experienced subjects with HIV/HCV coinfection; and uninfected controls. Mann–Whitney U test.

Article Snippet: Slides were incubated with antihuman mouse-CD163 monoclonal antibody (LifeSpan Biosciences) and rabbit-CD68 polyclonal antibody (Abcam), followed by incubation with antimouse (AlexaFluor 488) and antirabbit (AlexaFluor 568) donkey secondary antibodies (Life Technologies).

Techniques: MANN-WHITNEY

Journal: The Journal of Infectious Diseases

Article Title: Macrophage Activation Marker Soluble CD163 Is a Dynamic Marker of Liver Fibrogenesis in Human Immunodeficiency Virus/Hepatitis C Virus Coinfection

doi: 10.1093/infdis/jiy331

Figure Lengend Snippet: Soluble CD163 (sCD163) decreases upon successful antiretroviral therapy (ART) (A) and only in sustained virologic responders (SVR) to anti-HCV therapy (B). A, sCD163 levels in human immunodeficiency virus (HIV) and HIV/hepatitis C virus (HCV)–infected subjects, prior to ART and 6 months post-ART. Patients were HCV and HIV treatment-naive at baseline. B, sCD163 levels before and following completion of interferon (IFN)–based therapy in SVR and nonresponders (NR). Patients were ART-treated for >3 months, and HCV treatment-naive at baseline. Dashed lines are median sCD163 levels in uninfected controls. The box plots represent the median and interquartile ranges, and the whiskers depict the minimum and maximum of the data set. Wilcoxon signed-rank test (paired) and Mann–Whitney U test (unpaired). *P < .05.

Article Snippet: Slides were incubated with antihuman mouse-CD163 monoclonal antibody (LifeSpan Biosciences) and rabbit-CD68 polyclonal antibody (Abcam), followed by incubation with antimouse (AlexaFluor 488) and antirabbit (AlexaFluor 568) donkey secondary antibodies (Life Technologies).

Techniques: Infection, MANN-WHITNEY

Journal: The Journal of Infectious Diseases

Article Title: Macrophage Activation Marker Soluble CD163 Is a Dynamic Marker of Liver Fibrogenesis in Human Immunodeficiency Virus/Hepatitis C Virus Coinfection

doi: 10.1093/infdis/jiy331

Figure Lengend Snippet: Soluble CD163 (sCD163) levels correlate with noninvasive assessment (A) and histological assessment (B) of liver fibrosis. Baseline sCD163 in 80 human immunodeficiency virus/hepatitis C virus (HCV) (antiretroviral therapy–treated, HCV treatment-naive) and 47 HCV (HCV treatment-naive). Histological liver fibrosis and necroinflammation assessed using Ishak scoring system [26]. Spearman correlation test. Abbreviations: APRI, aspartate aminotransferase–to-platelet ratio index; FIB-4, fibrosis index; HCV, hepatitis C virus; HIV, human immunodeficiency virus.

Article Snippet: Slides were incubated with antihuman mouse-CD163 monoclonal antibody (LifeSpan Biosciences) and rabbit-CD68 polyclonal antibody (Abcam), followed by incubation with antimouse (AlexaFluor 488) and antirabbit (AlexaFluor 568) donkey secondary antibodies (Life Technologies).

Techniques:

Journal: The Journal of Infectious Diseases

Article Title: Macrophage Activation Marker Soluble CD163 Is a Dynamic Marker of Liver Fibrogenesis in Human Immunodeficiency Virus/Hepatitis C Virus Coinfection

doi: 10.1093/infdis/jiy331

Figure Lengend Snippet: Soluble CD163 (sCD163) levels rise with increasing fibrosis stage, peaking prior to cirrhosis establishment. Baseline sCD163 per Ishak fibrosis stage in human immunodeficiency virus (HIV)/hepatitis C virus (HCV)–coinfected patients (antiretroviral therapy–treated, HCV treatment-naive). Light bars: stages 0–3 (mild to moderate fibrosis), dark bars: stages 4 (bridging fibrosis) and 5–6 (cirrhosis). The box plots represent the median and interquartile ranges, and the whiskers depict the minimum and maximum of the data set. Mann–Whitney U test. *P < .05.

Article Snippet: Slides were incubated with antihuman mouse-CD163 monoclonal antibody (LifeSpan Biosciences) and rabbit-CD68 polyclonal antibody (Abcam), followed by incubation with antimouse (AlexaFluor 488) and antirabbit (AlexaFluor 568) donkey secondary antibodies (Life Technologies).

Techniques: MANN-WHITNEY

Journal: The Journal of Infectious Diseases

Article Title: Macrophage Activation Marker Soluble CD163 Is a Dynamic Marker of Liver Fibrogenesis in Human Immunodeficiency Virus/Hepatitis C Virus Coinfection

doi: 10.1093/infdis/jiy331

Figure Lengend Snippet: Soluble CD163 (sCD163) levels are higher in human immunodeficiency virus (HIV)/hepatitis C virus (HCV) coinfection than in HCV monoinfection in mild to moderate fibrosis but not in advanced fibrosis. Baseline sCD163 levels in HIV/HCV-coinfected (antiretroviral therapy–treated, HCV treatment-naive) and HCV-monoinfected (HCV treatment-naive) patients stratified by liver fibrosis stage: mild to moderate fibrosis (Ishak stage 0–3) and advanced fibrosis (bridging fibrosis/cirrhosis, Ishak stage 4–6). The box plots represent the median and interquartile ranges, and the whiskers depict the minimum and maximum of the data set. Mann–Whitney U test.

Article Snippet: Slides were incubated with antihuman mouse-CD163 monoclonal antibody (LifeSpan Biosciences) and rabbit-CD68 polyclonal antibody (Abcam), followed by incubation with antimouse (AlexaFluor 488) and antirabbit (AlexaFluor 568) donkey secondary antibodies (Life Technologies).

Techniques: MANN-WHITNEY

Journal: The Journal of Infectious Diseases

Article Title: Macrophage Activation Marker Soluble CD163 Is a Dynamic Marker of Liver Fibrogenesis in Human Immunodeficiency Virus/Hepatitis C Virus Coinfection

doi: 10.1093/infdis/jiy331

Figure Lengend Snippet: Periportal proportion of CD163+ macrophages is enriched with incremental liver fibrosis and higher in human immunodeficiency virus (HIV)/hepatitis C virus (HCV) coinfection compared with HCV monoinfection. A, Liver tissues from 16 HIV/HCV-infected (antiretroviral therapy–treated, HCV treatment-naive) and 13 HCV-infected (treatment-naive) patients were co-stained for CD163/CD68. While the proportion of CD163+ macrophages enriched in periportal regions with increasing liver fibrosis in both HIV/HCV (r = 0.55, P = .04) and HCV liver tissue (r = 0.60, P = .03) (not shown), this enrichment was stronger in HIV/HCV liver tissue prior to cirrhosis (Ishak stage 0–4). Spearman correlation test. B, Proportion of periportal CD163+ macrophages in 14 HIV/HCV-infected and 11 HCV-infected patients without established cirrhosis (Ishak stage 0–4). Mann–Whitney U test.

Article Snippet: Slides were incubated with antihuman mouse-CD163 monoclonal antibody (LifeSpan Biosciences) and rabbit-CD68 polyclonal antibody (Abcam), followed by incubation with antimouse (AlexaFluor 488) and antirabbit (AlexaFluor 568) donkey secondary antibodies (Life Technologies).

Techniques: Infection, Staining, MANN-WHITNEY