Journal: bioRxiv

Article Title: HIV broadly neutralizing antibodies with enhanced Fc-CD16 affinity increase NK cell ADCC even with limited envelope binding

doi: 10.64898/2026.04.27.721006

Figure Lengend Snippet: Cytokine stimulation enhances NK cell activation while downregulating CD16 expression through ADAM17-dependent shedding. ( A ) Representative flow plots of CD69 induction of NK cells stimulated with IL-15, IL-12/IL-15, or IL-12/IL-15/IL-18. ( B ) CD69 induction across donors (n=10). ( C ) Viability upon different treatments. ( D ) Changes in CD16 expression across multiple donors upon cytokine treatment. ( E ) Spearman correlation analysis between CD16 expression and CD69 induction. ( F ) Representative histogram of CD16 expression upon triple cytokine treatment in the presence of TAPI-I (right panel) or DMSO control (left panel). ( G ) CD16 expression in donors (n=13) stimulated with cytokines in the absence or presence of TAPI-1. ( H ) Effects of TAPI-I on NK cell activation. ( I ) Percentage remaining p24 + cells upon coculture of HIV-infected CD4 + T cells with autologous NK cells from donors (n=7) pre-stimulated with cytokines in the presence of TAPI-1 or DMSO control. ( J ) Percentage ADCC calculated as indicated in methods from data shown in ( I ). Statistical significance was calculated using one-way ANOVA with multiple comparison compared to the no cytokine condition for panel ( B-D ), two-way ANOVA with multiple comparison compared to the IL12/IL15/IL18 condition for panel ( I ), and two-way ANOVA for panel ( J ). Open symbols represent female donors and closed symbols male donors.

Article Snippet: Ten cytokines were measured using Quanterix SP-X Corplex Cytokine Panel (IFN-γ, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-22, TNF-α) (cat# 85-0329).

Techniques: Activation Assay, Expressing, Control, Infection, Comparison

Journal: bioRxiv

Article Title: HIV broadly neutralizing antibodies with enhanced Fc-CD16 affinity increase NK cell ADCC even with limited envelope binding

doi: 10.64898/2026.04.27.721006

Figure Lengend Snippet: Functional activity, cytokine secretion, and Fc binding of VRC07-523-LS and Fc variants. ( A ) Table summarizing VRC07-523LS antibody mutants with optimized Fc biding including specific mutations and effects on CD16 binding. ( B ) CD16 reporter activity in Jurkat-Lucia NFAT-CD16 cells exposed to antibody variants. ( C ) CD69 induction across donors (n=7). ( D ) Viability upon different conditions. ( E-O ) Supernatants from NK cells cultured overnight with WT VRC07-523-LS or Fc mutant antibodies were analyzed for cytokine secretion (fg/mL). Levels of ( E ) IFN-γ (Limit of detection (LOD) 1fg/mL), ( F ) TNF-α (LOD 16.7fg/mL), ( G ) IL-1β (LOD 16.3fg/mL), ( H ) IL-4 (LOD 339.1fg/mL), ( I ) IL-5 (LOD 8.7fg/ml), ( J ) IL-6 (LOD 21.1fg/mL), ( K ) IL-8 (LOD 41.7fg/mL), ( M ) IL-10 (LOD 16.8fg/mL), ( N ) IL-12p70 (LOD 24.7 fg/mL), ( O ) IL-22 (LOD 4.5 fg/mL) (n=6). ( P ) Representative flow cytometry histograms showing VRC07-523LS binding to surface Env on uninfected (blue), uninfected exposed (red), or HIV-infected cells (black). A FMO control is included (grey). Quantification of Env binding for the different antibodies (n=7). ( Q ) Surface expression of CD107a upon co-culture of NK cells alone (no CD4 + ), or NK cells co-cultured with uninfected autologous CD4 + or NL4-3 infected CD4 + T cells in the presence of the indicated antibody (n=2). Statistical significance was assessed using a Friedman test with multiple comparisons compared to unstimulated condition for panels ( C-O ), and two-way ANOVA with multiple comparisons for panel ( P ). Open symbols represent female and closed symbols male donors.

Article Snippet: Ten cytokines were measured using Quanterix SP-X Corplex Cytokine Panel (IFN-γ, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-22, TNF-α) (cat# 85-0329).

Techniques: Functional Assay, Activity Assay, Binding Assay, Cell Culture, Mutagenesis, Flow Cytometry, Infection, Control, Expressing, Co-Culture Assay

Journal: bioRxiv

Article Title: HIV broadly neutralizing antibodies with enhanced Fc-CD16 affinity increase NK cell ADCC even with limited envelope binding

doi: 10.64898/2026.04.27.721006

Figure Lengend Snippet: Enhancing antibody affinity to CD16 improves NK cell-mediated ADCC against HIV-infected cells independently of cytokine activation or ADAM17 inhibition. Percent of remaining HIV p24⁺ infected CD4⁺ T cells following co-culture with autologous NK cells from donors pre-treated with or without cytokines (IL-12/IL-15/IL-18) (n=16) ( A ) and ADAM17 inhibitor TAPI-1 (n=7) ( B ), in the presence of the indicated antibody. ( C ) Hierarchical cluster analysis of percentage of remaining p24⁺ cells in different conditions shown in A-B. ADCC activity under the same treatment conditions in the absence ( D ) or the presence ( E ) of the ADAM17 inhibitor TAPI-1. Statistical significance was determined using two-way ANOVA with multiple comparison compared to WT. Open symbols represent female and closed symbols male donors. ( F ) Hierarchical cluster analysis of ADCC in different conditions shown in D-E . Clusters were done using ClustVis.

Article Snippet: Ten cytokines were measured using Quanterix SP-X Corplex Cytokine Panel (IFN-γ, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-22, TNF-α) (cat# 85-0329).

Techniques: Infection, Activation Assay, Inhibition, Co-Culture Assay, Activity Assay, Comparison

Journal: bioRxiv

Article Title: HIV broadly neutralizing antibodies with enhanced Fc-CD16 affinity increase NK cell ADCC even with limited envelope binding

doi: 10.64898/2026.04.27.721006

Figure Lengend Snippet: Effects of Fc-optimized VRC07-523LS mutants across HIV-1 subtypes. ( A ) Neutralization curves of VRC07-523LS against indicated HIV-1 viral strain. ( B ) Env binding of the different antibodies to HIV-infected CD4T cells with the indicated viral strain (n=3-4). ( C ) Spearman correlation between Env binding and neutralization IC 50 values. ( D ) Percentage of remaining p24 + cells following natural cytotoxicity by autologous NK cells pretreated with a combination of cytokines or no cytokine control (n= 6-16). ( E-I ) Percentage of remaining p24+ cells for each of the indicated viruses in the presence of the different antibodies (n= 7-8). Statistical significance was assessed using a one-sample t test with a hypothetical value set to 1 for panel ( B ), wilcoxon signed rank test with hypothetical value set to 100 for panel ( E ), and two -way ANOVA with multiple comparison compared to WT for panels ( E–I ). Open symbols represent female and closed symbols male donors. ( J ) Hierarchical cluster analysis of the percentage remaining p24 + cells. Clusters were done using ClustVis.

Article Snippet: Ten cytokines were measured using Quanterix SP-X Corplex Cytokine Panel (IFN-γ, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-22, TNF-α) (cat# 85-0329).

Techniques: Neutralization, Binding Assay, Infection, Control, Comparison

Journal: bioRxiv

Article Title: HIV broadly neutralizing antibodies with enhanced Fc-CD16 affinity increase NK cell ADCC even with limited envelope binding

doi: 10.64898/2026.04.27.721006

Figure Lengend Snippet: Cytokine activation of NK cells requires higher levels of surface Env binding to promote ADCC. ( A ) Percentage of ADCC mediated by VRC07-523LS for each of the viral strains upon pretreatment of NK cells with the combination of cytokines or no cytokine control (n=6-16). Statistical significance between no cytokine and IL12/IL15/IL18 was determined by two-way ANOVA and Wilcoxon signed rank test with hypothetical value set to 0 for each condition. Open symbols represent female and closed symbols male donors. ( B ) Spearman correlation between the percentage of ADCC and surface Env binding for the indicated antibodies and NK cells pretreated with no cytokine (top) or IL12/IL15/IL18 (bottom). Each virus is indicated with a different color (n=4-6).

Article Snippet: Ten cytokines were measured using Quanterix SP-X Corplex Cytokine Panel (IFN-γ, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-22, TNF-α) (cat# 85-0329).

Techniques: Activation Assay, Binding Assay, Control, Virus

Journal: Clinical and Experimental Immunology

Article Title: Multi-nucleated giant cell formation from human cord blood monocytes in vitro , in comparison with adult peripheral blood monocytes

doi: 10.1111/j.1365-2249.2009.03990.x

Figure Lengend Snippet: Morphology of multi-nucleated giant cells (MGCs) induced by interleukin (IL)-4, IL-4 with macrophage colony-stimulating factor (M-CSF) and IL-4 with granulocyte–macrophage colony-stimulating factor (GM-CSF) in vitro. Adult peripheral blood and cord blood monocytes were cultured in RPMI-1640 medium supplemented with 10% heat-inactivated fetal bovine serum (FBS), 100 U/ml penicillin and 100 µg/ml streptomycin, at 1 × 106 cells/ml/well in 24-well tissue culture plates with or without cytokines. After 14 days' incubation without changing the medium, the cells were stained with a May–Grünwald–Giemsa stain. (a) No added cytokine: adult blood; (b) no added cytokine: cord blood; (c) IL-4 (20 ng/ml) alone: adult blood; (d) IL-4 (20 ng/ml) alone: cord blood; (e) IL-4 (20 ng/ml)+M-CSF (100 ng/ml): adult blood; (f) IL-4 (20 ng/ml)+M-CSF (100 ng/ml): cord blood; (g) IL-4 (20 ng/ml)+GM-CSF (20 ng/ml): adult blood; (h) IL-4 (20 ng/ml)+GM-CSF (20 ng/ml): cord blood.

Article Snippet: The human cytokines (IL-4, M-CSF, GM-CSF, IFN-γ and TNF-α) were from Wako Pure Chemical Industries (Osaka, Japan).

Techniques: In Vitro, Cell Culture, Incubation, Staining, Giemsa Stain

Journal: Clinical and Experimental Immunology

Article Title: Multi-nucleated giant cell formation from human cord blood monocytes in vitro , in comparison with adult peripheral blood monocytes

doi: 10.1111/j.1365-2249.2009.03990.x

Figure Lengend Snippet: Effect of cytokines on multi-nucleated giant cell (MGC) formation; adult blood versus cord blood monocytes. Adult peripheral blood (solid bar) and cord blood (open bar) monocytes were cultured as described in the legend to Fig. 1. The results were calculated from seven independent experiments as the fusion index. Significantly different from control or between cord blood and adult blood: *P < 0·0001, **P < 0·01, †P < 0·05.

Article Snippet: The human cytokines (IL-4, M-CSF, GM-CSF, IFN-γ and TNF-α) were from Wako Pure Chemical Industries (Osaka, Japan).

Techniques: Cell Culture

Journal: Clinical and Experimental Immunology

Article Title: Multi-nucleated giant cell formation from human cord blood monocytes in vitro , in comparison with adult peripheral blood monocytes

doi: 10.1111/j.1365-2249.2009.03990.x

Figure Lengend Snippet: Nuclei per multi-nucleated giant cells (MGCs) induced by cytokines; adult blood versus cord blood monocytes. Adult peripheral blood (solid box) and cord blood (open box) monocytes were cultured as described in the legend to Fig. 1. The number of nuclei per MGC (20 cells acquired randomly in each experiment) was counted. Results from seven independent experiments are shown in the box-and-whisker plots. Line in the box shows median point, box shows the point from first to third quartile, whiskers show the point from smallest to largest number. Significantly different between cord blood and adult blood: *P < 0·0001.

Article Snippet: The human cytokines (IL-4, M-CSF, GM-CSF, IFN-γ and TNF-α) were from Wako Pure Chemical Industries (Osaka, Japan).

Techniques: Cell Culture, Whisker Assay

Journal: Clinical and Experimental Immunology

Article Title: Multi-nucleated giant cell formation from human cord blood monocytes in vitro , in comparison with adult peripheral blood monocytes

doi: 10.1111/j.1365-2249.2009.03990.x

Figure Lengend Snippet: Concentration of interferon (IFN)-γ, interleukin (IL)-1β, IL-4 and tumour necrosis factor (TNF)-α in the supernatants of adult peripheral blood and cord blood monocytes after 14 days' culture with the cytokines listed below.

Article Snippet: The human cytokines (IL-4, M-CSF, GM-CSF, IFN-γ and TNF-α) were from Wako Pure Chemical Industries (Osaka, Japan).

Techniques: Concentration Assay

Journal: Experimental Dermatology

Article Title: Interferon‐γ downregulates tight junction function, which is rescued by interleukin‐17A

doi: 10.1111/exd.14425

Figure Lengend Snippet: Dose‐ and time‐dependent inhibition of CLDN1 expression by IFN‐γ in normal human epidermal keratinocytes (NHEKs). (A) Effects of IL‐4, IL‐17A, IFN‐γ and TNF‐α (30 ng ml −1 ) on CLDN1 expression in NHEK cultures incubated with the cytokines in a 24‐well plate for 48 h. Samples were prepared from another three wells ( n = 3). (B) Dose‐dependent inhibition of CLDN1 expression by IFN‐γ (3–100 ng ml −1 ) in NHEKs cultured with or without IFN‐γ for 48 h ( n = 2). (C) Time‐dependent inhibition of CLDN1 expression in NHEKs treated with 30 ng ml −1 IFN‐γ for 0–48 h ( n = 1). Western blotting performed with anti‐CLDN1 and anti‐GAPDH antibodies. Examples of typical blots in duplicate are illustrated. The expression level of CLDN1 was quantified by densitometry and normalized to GAPDH; the ratio is shown directly under each blot. Non‐treated control (0) is 1.00. The statistical analysis is shown in Figure . The results are representative of more than three independent experiments. (D) Changes in the relative CLDN1 mRNA expression in NHEKs cultured in a 24‐well plate with or without IFN‐γ (30 ng ml −1 ) for 3 or 6 h. (E) Changes in the relative CLDN1 mRNA expression in NHEKs treated with 2–50 ng ml −1 IFN‐γ for 6 h. mRNA expression was measured using qRT‐PCR. Data are presented as the mean ± SD ( n = 4). Data were analysed using the Dunnett's test (* p < 0.05; ** p < 0.01 vs. control (0 ng ml −1 ))

Article Snippet: Human recombinant cytokines (IL‐4, IL‐17A, IFN‐γ and TNF‐α) were purchased from FUJIFILM Wako Pure Chemical (Osaka, Japan).

Techniques: Inhibition, Expressing, Incubation, Cell Culture, Western Blot, Control, Quantitative RT-PCR