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InvivoGen nf κb seap irf luc reporter monocytes
Nf κb Seap Irf Luc Reporter Monocytes, supplied by InvivoGen, used in various techniques. Bioz Stars score: 98/100, based on 371 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/irf+luc+reporter+thp1/THP1-Dual+Cells/bio_rxiv__2025__04__22__650092-51-1-5
Average 98 stars, based on 371 article reviews

nf κb seap irf luc reporter monocytes - by Bioz Stars, 2026-10

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Article Title: Intradermal vaccination with a phytoglycogen nanoparticle and STING agonist induces cytotoxic T lymphocyte-mediated antitumor immunity
Article Snippet: Similarly, the NF-ĸB-SEAP and IRF-Luc reporter THP1-Dual cells (RRID: CVCL_X599; InvivoGen, San Diego, CA) were grown at a density of 1 × 10 5 cells per well in a 96-well plate in complete RPMI supplemented with 10% FBS, 100 μg/ml zeocin, 100 μg/ml normocin, and 10 μg/ml blasticidin.

Article Title: Intradermal vaccination with a phytoglycogen nanoparticle and STING agonist induces cytotoxic T lymphocyte-mediated antitumor immunity.
Article Snippet: Similarly, the NF-ĸB-SEAP and IRF-Luc reporter THP1-Dual cells (RRID: CVCL_X599; InvivoGen, San Diego, CA) were grown at a density of 1 × 105 cells per well in a 96-well plate in complete RPMI supplemented with 10% FBS, 100 μg/ml zeocin, 100 μg/ml normocin, and 10 μg/ml blasticidin.

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Article Title: Effective and Safe Stimulation of Humoral and Cell-Mediated Immunity by Intradermal Immunization with a Cyclic Dinucleotide/Nanoparticle Combination Adjuvant.
Article Snippet: Supernatants were analyzed for IL-1b by ELISA (eBioscience, San Diego, CA). .. The NF-kB–SEAP and IRF-Luc reporter THP1-Dual cells (InvivoGen) were cultured in complete RPMI supplemented with 10% FBS, 10 mg/ml blasticidin, 100 mg/ml zeocin, and 100 mg/ml normocin. ..

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( A-C ) Expression of macrophage anti-inflammatory markers ( A ), pro-inflammatory markers ( B ), and major cytokines ( C ) in primary human MDMs, 48h post-irradiation. MDMs were differentiated from PBMCs, isolated from 6 donors, by treatment with M-CSF and exposed to 5Gy γ IR; box plots of RT-qPCR measured expression of n = 6 is shown. * p <0.05, paired Wilcoxon test. ( D and E ) Protein concentrations of soluble macrophage activation markers CXCL10, CCL2 and sCD163 ( D ) and cytokines IL-6, TNF-α, and IL-10 ( E ) in culture medium of primary MDMs, 48h post-irradiation. Error bars indicate ±SD of three independent biological replicates; * p <0.05, ** p <0.01, panel D and below–paired t test. ( F and G ) Expression of macrophage pro-inflammatory ( F ) and anti-inflammatory ( G ) markers in human monocytic cells THP1 48h after exposure to γ IR. Error bars indicate ±SD of four independent biological replicates. ( H ) Expression of cytokines in THP1 cells, 48h after γ IR. Cells were exposed to indicated γ IR doses; RT-qPCR analysis was performed with primers specific for indicated cytokines. Error bars indicate ±SD of 4 independent biological replicates. ( I ) Quantitation of proinflammatory (IL-1β, IL-6, TNF-α, CCL2) and anti-inflammatory (IL-10) cytokines in culture media of γ IR-exposed THP1 cells using Luminex multiplex immunoassay, 48h after γ IR, PMA (20 nM) or poly(I:C) (2 μg/ml) treatment. Error bars indicate ±SD of three independent biological replicates; ** p <0.01 vs 0 Gy. For panels F to I: * p <0.05, ** p <0.01, NS non-significant.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: ( A-C ) Expression of macrophage anti-inflammatory markers ( A ), pro-inflammatory markers ( B ), and major cytokines ( C ) in primary human MDMs, 48h post-irradiation. MDMs were differentiated from PBMCs, isolated from 6 donors, by treatment with M-CSF and exposed to 5Gy γ IR; box plots of RT-qPCR measured expression of n = 6 is shown. * p <0.05, paired Wilcoxon test. ( D and E ) Protein concentrations of soluble macrophage activation markers CXCL10, CCL2 and sCD163 ( D ) and cytokines IL-6, TNF-α, and IL-10 ( E ) in culture medium of primary MDMs, 48h post-irradiation. Error bars indicate ±SD of three independent biological replicates; * p <0.05, ** p <0.01, panel D and below–paired t test. ( F and G ) Expression of macrophage pro-inflammatory ( F ) and anti-inflammatory ( G ) markers in human monocytic cells THP1 48h after exposure to γ IR. Error bars indicate ±SD of four independent biological replicates. ( H ) Expression of cytokines in THP1 cells, 48h after γ IR. Cells were exposed to indicated γ IR doses; RT-qPCR analysis was performed with primers specific for indicated cytokines. Error bars indicate ±SD of 4 independent biological replicates. ( I ) Quantitation of proinflammatory (IL-1β, IL-6, TNF-α, CCL2) and anti-inflammatory (IL-10) cytokines in culture media of γ IR-exposed THP1 cells using Luminex multiplex immunoassay, 48h after γ IR, PMA (20 nM) or poly(I:C) (2 μg/ml) treatment. Error bars indicate ±SD of three independent biological replicates; ** p <0.01 vs 0 Gy. For panels F to I: * p <0.05, ** p <0.01, NS non-significant.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Expressing, Irradiation, Isolation, Quantitative RT-PCR, Activation Assay, Quantitation Assay, Luminex, Multiplex Assay

( A ) Expression of intracellular senescence markers (CDKN1A and GLB1) in THP1 cells (blue columns) and primary MDMs (red columns) 48h after irradiation. Error bars: ±SD of three independent biological replicates. ( B ) Western blot analysis of senescence marker p21 Waf1 (CDKN1A) protein in THP1 lysates, 48 h after irradiation or treatment with PMA or poly(I:C). Forty μg of total protein were loaded. ( C ) Expression of indicated macrophage SASP markers in human monocytic cells THP1 (blue columns) and primary MDMs (red columns) 48h after irradiation. Error bars indicate ±SD of three independent biological replicates. In all panels: * p <0.05, ** p <0.01, NS non-significant. ( D ) Quantitation of indicated macrophage SASP markers in culture media of irradiated THP1 cells (n = 6) using Luminex multiplex immunoassay, 48h after γIR. Error bars indicate ±SD; * p <0.05, paired Wilcoxon test. ( E ) Microscope images of untreated, irradiated, and PMA-treated THP1 monocytes, 48h incubation. Cells were fixed with paraformaldehyde, labeled for F actin with phalloidin-rhodamine conjugate and for nuclear DNA with DAPI (upper panels) or vital images were obtained (lower panels). Scale bars: 75 μm. ( F ) Proportion of CD11b-positive THP1 cells in irradiated or control cells detected by flow cytometry of viable (DAPI - ) cells, 72h after γ IR (graphs represent median fluorescence intensity data of 4 independent experiments). ( G ) Quantitation (flow cytometry) of CD11b-positive populations in cells exposed to increasing doses of γ IR, or treated with PMA or poly(I:C), 72h post-exposure (only viable, DAPI - cells were analyzed; box plot of n = 4 is shown. ** p <0.01.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: ( A ) Expression of intracellular senescence markers (CDKN1A and GLB1) in THP1 cells (blue columns) and primary MDMs (red columns) 48h after irradiation. Error bars: ±SD of three independent biological replicates. ( B ) Western blot analysis of senescence marker p21 Waf1 (CDKN1A) protein in THP1 lysates, 48 h after irradiation or treatment with PMA or poly(I:C). Forty μg of total protein were loaded. ( C ) Expression of indicated macrophage SASP markers in human monocytic cells THP1 (blue columns) and primary MDMs (red columns) 48h after irradiation. Error bars indicate ±SD of three independent biological replicates. In all panels: * p <0.05, ** p <0.01, NS non-significant. ( D ) Quantitation of indicated macrophage SASP markers in culture media of irradiated THP1 cells (n = 6) using Luminex multiplex immunoassay, 48h after γIR. Error bars indicate ±SD; * p <0.05, paired Wilcoxon test. ( E ) Microscope images of untreated, irradiated, and PMA-treated THP1 monocytes, 48h incubation. Cells were fixed with paraformaldehyde, labeled for F actin with phalloidin-rhodamine conjugate and for nuclear DNA with DAPI (upper panels) or vital images were obtained (lower panels). Scale bars: 75 μm. ( F ) Proportion of CD11b-positive THP1 cells in irradiated or control cells detected by flow cytometry of viable (DAPI - ) cells, 72h after γ IR (graphs represent median fluorescence intensity data of 4 independent experiments). ( G ) Quantitation (flow cytometry) of CD11b-positive populations in cells exposed to increasing doses of γ IR, or treated with PMA or poly(I:C), 72h post-exposure (only viable, DAPI - cells were analyzed; box plot of n = 4 is shown. ** p <0.01.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Expressing, Irradiation, Western Blot, Marker, Quantitation Assay, Luminex, Multiplex Assay, Microscopy, Incubation, Labeling, Flow Cytometry, Fluorescence

( A and B ) Box plots showing expression, measured by RT-qPCR, of IFNα ( A ) and IFNβ ( B ) in MDMs, 48h after irradiation (n = 15). M-CSF-differentiated MDMs from PBMCs isolated from 5 donors were exposed to indicated doses of γ IR. Error bars: ±SD, * p <0.05, NS non-significant, paired Wilcoxon test. ( C ) Expression of IFNα and IFNβ in THP1 cells, 48h post-irradiation, measured by RT-qPCR. Cells were exposed to indicated doses of γ IR or treated with PMA or poly(I:C) with or without indicated transfection reagents. Error bars: ±SD of four independent biological replicates; * p <0.05, ** p <0.01. ( D ) Immunoblotting of phosphorylated STAT1 at Ser727 and Tyr701 (40 μg of total protein) in THP1 cells, 48h post-irradiation. ( E ) Volcano plot showing ISG expression in irradiated vs. control THP1 cells, measured by PCR array of total cellular RNA samples, 48h post-irradiation. Red dots indicate significant (above blue line) or insignificant (below blue line) increase in gene expression in irradiated cells ( p <0.05, fold change >2.0). ( F and G ) Activation of NF-κB-dependent transcription ( F ) and IFNAR signaling ( G ) by culture media from irradiated (5 Gy) or non-irradiated THP1 in the reporter THP1-Dual cells (pink columns) expressing SEAP gene driven by an IFN-β minimal promoter activated by NF-κB and secreted Lucia luciferase gene under the control of an IFNAR signaling-activated ISG54 minimal promoter, and THP1-Dual KO-IFNAR2 cells (blue columns), generated from THP1-Dual cells by stable knockout of the IFNAR2 receptor. Similarly, the culture supernatant from Poly(I:C)-transfected THP1 was used as a positive control. After 18 h of incubation and staining with SEAP-sensitive QUANTI-Luc dye, the absorbance (600 nm) (F) or luminescence (G) of each sample were measured and normalized to fresh RPMI media controls. Error bars indicate ±SD of a minimum of 3 independent biological replicates; p <0.01. ( H ) Immunoblotting to assess STAT1 Ser727 and Tyr701 phosphorylation (20 μg of total protein) in THP1 cells incubated for 24h with filtered culture media from irradiated or non-irradiated THP1, with or without interferon-α/β receptor inhibitor IFNAR-IN-1. ( I ) Expression ratio of IFNα and indicated cytokines (5 Gy-to-0 Gy ratio of RNA count) in THP1 cells transfected with indicated siRNA (24 h before irradiation), with or without exposure to 5 Gy γ IR for 48 hours. RNA was quantified by RT-qPCR. Error bars: ±SD of at least four independent biological replicates; * p <0.05. ( J ) Immunoblot of ssRNA sensor RIG-I and dsRNA sensors MDA-5 and TLR3 (40 μg of total protein) in THP1 cells, 48h post-irradiation. ( K ) Western blot of downstream markers of the activation of RIG-I/MDA-5 (MAVS, phospho-IKKε, and phospho-IRF3) and TLR3 (phospho-IKKε, and phospho-IRF3) pathways. THP1 lysates (40 μg total protein), harvested 48h post-exposure.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: ( A and B ) Box plots showing expression, measured by RT-qPCR, of IFNα ( A ) and IFNβ ( B ) in MDMs, 48h after irradiation (n = 15). M-CSF-differentiated MDMs from PBMCs isolated from 5 donors were exposed to indicated doses of γ IR. Error bars: ±SD, * p <0.05, NS non-significant, paired Wilcoxon test. ( C ) Expression of IFNα and IFNβ in THP1 cells, 48h post-irradiation, measured by RT-qPCR. Cells were exposed to indicated doses of γ IR or treated with PMA or poly(I:C) with or without indicated transfection reagents. Error bars: ±SD of four independent biological replicates; * p <0.05, ** p <0.01. ( D ) Immunoblotting of phosphorylated STAT1 at Ser727 and Tyr701 (40 μg of total protein) in THP1 cells, 48h post-irradiation. ( E ) Volcano plot showing ISG expression in irradiated vs. control THP1 cells, measured by PCR array of total cellular RNA samples, 48h post-irradiation. Red dots indicate significant (above blue line) or insignificant (below blue line) increase in gene expression in irradiated cells ( p <0.05, fold change >2.0). ( F and G ) Activation of NF-κB-dependent transcription ( F ) and IFNAR signaling ( G ) by culture media from irradiated (5 Gy) or non-irradiated THP1 in the reporter THP1-Dual cells (pink columns) expressing SEAP gene driven by an IFN-β minimal promoter activated by NF-κB and secreted Lucia luciferase gene under the control of an IFNAR signaling-activated ISG54 minimal promoter, and THP1-Dual KO-IFNAR2 cells (blue columns), generated from THP1-Dual cells by stable knockout of the IFNAR2 receptor. Similarly, the culture supernatant from Poly(I:C)-transfected THP1 was used as a positive control. After 18 h of incubation and staining with SEAP-sensitive QUANTI-Luc dye, the absorbance (600 nm) (F) or luminescence (G) of each sample were measured and normalized to fresh RPMI media controls. Error bars indicate ±SD of a minimum of 3 independent biological replicates; p <0.01. ( H ) Immunoblotting to assess STAT1 Ser727 and Tyr701 phosphorylation (20 μg of total protein) in THP1 cells incubated for 24h with filtered culture media from irradiated or non-irradiated THP1, with or without interferon-α/β receptor inhibitor IFNAR-IN-1. ( I ) Expression ratio of IFNα and indicated cytokines (5 Gy-to-0 Gy ratio of RNA count) in THP1 cells transfected with indicated siRNA (24 h before irradiation), with or without exposure to 5 Gy γ IR for 48 hours. RNA was quantified by RT-qPCR. Error bars: ±SD of at least four independent biological replicates; * p <0.05. ( J ) Immunoblot of ssRNA sensor RIG-I and dsRNA sensors MDA-5 and TLR3 (40 μg of total protein) in THP1 cells, 48h post-irradiation. ( K ) Western blot of downstream markers of the activation of RIG-I/MDA-5 (MAVS, phospho-IKKε, and phospho-IRF3) and TLR3 (phospho-IKKε, and phospho-IRF3) pathways. THP1 lysates (40 μg total protein), harvested 48h post-exposure.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Expressing, Quantitative RT-PCR, Irradiation, Isolation, Transfection, Western Blot, Activation Assay, Luciferase, Generated, Knock-Out, Positive Control, Incubation, Staining

( A ) Top : Venn diagram of differentially expressed retroelements and HERVs identified between irradiated vs. control THP1 cells or MDMs. Bottom : expression fold changes detected for the common set (626 features) shown on the upper Venn diagram. ( B ) Expression of six HERV clades, identified upregulated by RNA-Seq in both irradiated THP1 cells and MDMs (panel A, red circles), measured by RT-qPCR at 48h post-exposure to 5 Gy γ IR dose. Primers were designed for the alignments within the clusters identified across gag , pol or env genes of the sequences within the clades. Error bars: ±SD of three independent biological replicates. ( C ) Expression, measured by RT-qPCR, of randomly selected differentially expressed HERVK in THP1 cells (grey columns) and primary MDMs (red columns), 48h post-irradiation. Error bars: ±SD of three independent biological replicates. ( D ) Transcription of HERVK HML-2 env in response to 5 Gy γ IR dose, measured by RT-qPCR at 48h post-exposure in CD11b-positive and negative THP1 cells, separated by sorting flow cytometry. Only viable, DAPI-negative cells were analyzed; box plot of n = 3 is shown. Error bars: ±SD of three independent biological replicates. ( E ) Ratio of HERV RNA bound to anti-dsRNA antibodies to total RNA input: HML-2 (left) and HERV9NC Cl.1 pol (right) from 5 Gy γ IR exposed and unexposed THP1 cells. RT-qPCR of RNA-IP (RIP) complexes with rJ2 and 9D5 antibodies, 48h post-irradiation. Error bars: ±SD of five (HML-2) or four (HERV9NC) independent biological replicates; in panels B-E,* p <0.05, ** p<0.01. ( F ) Relative count of HERVK HML-2 antisense RNA identified with antisense strand-specific reverse transcription primers targeting regions in the env , pol and gag genes, and quantitated by qPCR. Error bars: ±SD of at least 6 independent biological replicates. ( G ) Relative count of the sense and antisense transcripts of HERVK HML-2 measured by RT-qPCR using env- specific primers, 48h after irradiation of MDMs. MDMs were differentiated with M-CSF from PBMCs isolated from 5 donors. Box plot of n = 15 is shown. In panels F and G, * p <0.05, NS non-significant, paired Wilcoxon test.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: ( A ) Top : Venn diagram of differentially expressed retroelements and HERVs identified between irradiated vs. control THP1 cells or MDMs. Bottom : expression fold changes detected for the common set (626 features) shown on the upper Venn diagram. ( B ) Expression of six HERV clades, identified upregulated by RNA-Seq in both irradiated THP1 cells and MDMs (panel A, red circles), measured by RT-qPCR at 48h post-exposure to 5 Gy γ IR dose. Primers were designed for the alignments within the clusters identified across gag , pol or env genes of the sequences within the clades. Error bars: ±SD of three independent biological replicates. ( C ) Expression, measured by RT-qPCR, of randomly selected differentially expressed HERVK in THP1 cells (grey columns) and primary MDMs (red columns), 48h post-irradiation. Error bars: ±SD of three independent biological replicates. ( D ) Transcription of HERVK HML-2 env in response to 5 Gy γ IR dose, measured by RT-qPCR at 48h post-exposure in CD11b-positive and negative THP1 cells, separated by sorting flow cytometry. Only viable, DAPI-negative cells were analyzed; box plot of n = 3 is shown. Error bars: ±SD of three independent biological replicates. ( E ) Ratio of HERV RNA bound to anti-dsRNA antibodies to total RNA input: HML-2 (left) and HERV9NC Cl.1 pol (right) from 5 Gy γ IR exposed and unexposed THP1 cells. RT-qPCR of RNA-IP (RIP) complexes with rJ2 and 9D5 antibodies, 48h post-irradiation. Error bars: ±SD of five (HML-2) or four (HERV9NC) independent biological replicates; in panels B-E,* p <0.05, ** p<0.01. ( F ) Relative count of HERVK HML-2 antisense RNA identified with antisense strand-specific reverse transcription primers targeting regions in the env , pol and gag genes, and quantitated by qPCR. Error bars: ±SD of at least 6 independent biological replicates. ( G ) Relative count of the sense and antisense transcripts of HERVK HML-2 measured by RT-qPCR using env- specific primers, 48h after irradiation of MDMs. MDMs were differentiated with M-CSF from PBMCs isolated from 5 donors. Box plot of n = 15 is shown. In panels F and G, * p <0.05, NS non-significant, paired Wilcoxon test.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Irradiation, Expressing, RNA Sequencing Assay, Quantitative RT-PCR, Flow Cytometry, Isolation

( A ) Transcription of HERVK HML-2 and five other HERV clades, identified upregulated by RNA-Seq in both irradiated THP1 cells and MDMs , measured by RT-qPCR in THP1 cells expressing control (grey) or shRNA-Env (red), 48h post-irradiation. Error bars: ±SD of three independent biological replicates. ( B ) Expression, measured by RT-qPCR, of macrophage activation markers in human monocytic THP1 cells, expressing shRNA-Env (red) or control shRNA (grey), 48h post-irradiation. Error bars: ±SD of three independent biological replicates. In panels A and B, * p <0.05, ** p <0.01; A and B, genes, whose expression changed significantly in response to shRNA-Env are indicated by red symbols. ( C ) Immunoblot against intracellular senescence marker p21 Waf1 (CDKN1A) in lysates (40 μg total protein) of THP1 cells, expressing either shRNA-Env or control shRNA, 48h after irradiation. ( D ) Phosphorylation of STAT1 Ser727 and Tyr701 (40 μg total protein) in THP1 cells, expressing control shRNA or shRNA-Env, 48h post-irradiation. ( E ) Volcano plot depicting ISG expression in THP1 cells expressing shRNA-Env or control shRNA, 48h post-IR: measured by PCR array of total cellular RNA samples. Green dots represent significant (above blue line) or non-significant decrease in expression of indicated genes in shRNA-Env vs. control shRNA expressing cells ( p <0.05, fold change >2.0). ( F and G ) Soluble macrophage activation markers ( F ) and key cytokines ( G ) in culture medium of THP1 cells, 48h post-irradiation, determined by multiplex immunoassay (Luminex). Error bars: ±SD of three independent biological replicates. * p <0.05, ** p <0.01, NS non-significant. ( H ) Macrophage SASP markers in culture media of irradiated THP1 cells (n = 6), expressing shRNA-Env (yellow) or control shRNA (blue), using multiplex immunoassay (Luminex), 48h after γ IR. Error bars indicate ±SD; p value of all differences is <0.05, unpaired Mann-Whitney test. ( I ) Transcription levels, measured by RT-qPCR, of HERVK HML-2 env , IFN-I (IFNα, IFNβ), inflammation (IL-1β, TNF-α) and anti-inflammatory (TGF-β, IL-10) markers in irradiated primary human MDMs, expressing control or shRNA-Env, 48h post-irradiation. M-CSF-differentiated MDMs from n = 6 donors. * p <0.05, NS non-significant, paired Wilcoxon test. ( J ) Soluble macrophage activation markers in culture medium of primary human MDMs, 48h post-irradiation assessed by multiplex immunoassay. Error bars: ±SD of three independent biological replicates; ** p <0.01, two-tailed paired t test.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: ( A ) Transcription of HERVK HML-2 and five other HERV clades, identified upregulated by RNA-Seq in both irradiated THP1 cells and MDMs , measured by RT-qPCR in THP1 cells expressing control (grey) or shRNA-Env (red), 48h post-irradiation. Error bars: ±SD of three independent biological replicates. ( B ) Expression, measured by RT-qPCR, of macrophage activation markers in human monocytic THP1 cells, expressing shRNA-Env (red) or control shRNA (grey), 48h post-irradiation. Error bars: ±SD of three independent biological replicates. In panels A and B, * p <0.05, ** p <0.01; A and B, genes, whose expression changed significantly in response to shRNA-Env are indicated by red symbols. ( C ) Immunoblot against intracellular senescence marker p21 Waf1 (CDKN1A) in lysates (40 μg total protein) of THP1 cells, expressing either shRNA-Env or control shRNA, 48h after irradiation. ( D ) Phosphorylation of STAT1 Ser727 and Tyr701 (40 μg total protein) in THP1 cells, expressing control shRNA or shRNA-Env, 48h post-irradiation. ( E ) Volcano plot depicting ISG expression in THP1 cells expressing shRNA-Env or control shRNA, 48h post-IR: measured by PCR array of total cellular RNA samples. Green dots represent significant (above blue line) or non-significant decrease in expression of indicated genes in shRNA-Env vs. control shRNA expressing cells ( p <0.05, fold change >2.0). ( F and G ) Soluble macrophage activation markers ( F ) and key cytokines ( G ) in culture medium of THP1 cells, 48h post-irradiation, determined by multiplex immunoassay (Luminex). Error bars: ±SD of three independent biological replicates. * p <0.05, ** p <0.01, NS non-significant. ( H ) Macrophage SASP markers in culture media of irradiated THP1 cells (n = 6), expressing shRNA-Env (yellow) or control shRNA (blue), using multiplex immunoassay (Luminex), 48h after γ IR. Error bars indicate ±SD; p value of all differences is <0.05, unpaired Mann-Whitney test. ( I ) Transcription levels, measured by RT-qPCR, of HERVK HML-2 env , IFN-I (IFNα, IFNβ), inflammation (IL-1β, TNF-α) and anti-inflammatory (TGF-β, IL-10) markers in irradiated primary human MDMs, expressing control or shRNA-Env, 48h post-irradiation. M-CSF-differentiated MDMs from n = 6 donors. * p <0.05, NS non-significant, paired Wilcoxon test. ( J ) Soluble macrophage activation markers in culture medium of primary human MDMs, 48h post-irradiation assessed by multiplex immunoassay. Error bars: ±SD of three independent biological replicates; ** p <0.01, two-tailed paired t test.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: RNA Sequencing Assay, Irradiation, Quantitative RT-PCR, Expressing, shRNA, Activation Assay, Western Blot, Marker, Multiplex Assay, Luminex, MANN-WHITNEY, Two Tailed Test

( A and B ) Immunoblot of MDA-5 ( A ) and TLR3 ( B ) with mouse monoclonal antibodies in immunocomplexes after IP of these proteins with rabbit polyclonal antibodies from THP1 lysates, exposed to indicated doses of γ IR or treated with PMA or poly(I:C); 48 h post-irradiation. β-Actin in cell lysates, utilized for IP, was used as a reference. ( C and D ) Relative abundance of sense and antisense HERVK HML-2 env RNA in immunocomplexes after RNP-immunoprecipitation (RIP) of MDA-5 ( C ) and TLR3 ( D ) from THP1 lysates, shown on panels A and B. The fold change RNA count (ΔΔCt) was calculated in relation to β-actin reference RNA, detected in the lysates used for RIP. Error bars: ±SD of three independent replicates; * p <0.05, ** p <0.01, NS non-significant. ( E ) PLA assay visualizing association of HML-2 env RNA with MDA-5 receptor in THP1 in irradiated or control cells; 48h post-IR. Nuclei–DAPI staining, MDA-5/RNA complexes–Texas Red. Scale bars: 30 μm.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: ( A and B ) Immunoblot of MDA-5 ( A ) and TLR3 ( B ) with mouse monoclonal antibodies in immunocomplexes after IP of these proteins with rabbit polyclonal antibodies from THP1 lysates, exposed to indicated doses of γ IR or treated with PMA or poly(I:C); 48 h post-irradiation. β-Actin in cell lysates, utilized for IP, was used as a reference. ( C and D ) Relative abundance of sense and antisense HERVK HML-2 env RNA in immunocomplexes after RNP-immunoprecipitation (RIP) of MDA-5 ( C ) and TLR3 ( D ) from THP1 lysates, shown on panels A and B. The fold change RNA count (ΔΔCt) was calculated in relation to β-actin reference RNA, detected in the lysates used for RIP. Error bars: ±SD of three independent replicates; * p <0.05, ** p <0.01, NS non-significant. ( E ) PLA assay visualizing association of HML-2 env RNA with MDA-5 receptor in THP1 in irradiated or control cells; 48h post-IR. Nuclei–DAPI staining, MDA-5/RNA complexes–Texas Red. Scale bars: 30 μm.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Western Blot, Irradiation, Immunoprecipitation, Staining

Expression and secretion of pro-inflammatory and anti-inflammatory macrophage markers in THP1 monocytes and primary monocyte-derived macrophages in response to gamma radiation.

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: Expression and secretion of pro-inflammatory and anti-inflammatory macrophage markers in THP1 monocytes and primary monocyte-derived macrophages in response to gamma radiation.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Expressing, Marker, Irradiation, shRNA

Journal: PLoS Pathogens

Article Title: Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses

doi: 10.1371/journal.ppat.1009305

Figure Lengend Snippet: Key resources.

Article Snippet: THP1 -dual NF-kB-SEAP IRF_Luc reporter Monocytes , , InvivoGen , thpd-nfis.

Techniques: Clone Assay, Virus, Plasmid Preparation, Recombinant, Blocking Assay, Membrane, Protease Inhibitor, Negative Control, Transfection, In Situ, Luciferase, Bicinchoninic Acid Protein Assay, SYBR Green Assay, Sequencing, Expressing, shRNA, Software

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