recombinant dkk1  (R&D Systems)

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: Decreased MPO + , CD11b + , and MHC class II + neutrophils in MyD88 (PKO) - and DKK1 (PKO) -infected mice (A–G) Six-week-old female WT-BALB/c, MyD88 (PKO) and DKK1 (PKO) mice were challenged with infective metacyclic promastigote (2 ×10 6 parasites, n = 5 per group) of L . major via the footpad. Non-infected BALB/c mice ( n = 10/2 feet per mouse) were given 0.9% NaCl saline. Cells were isolated from the footpads of all infected and non-infected mice at days 3 and 14 PI. Samples were analyzed by flow cytometry for MPO + , CD11b + , and MHC class II + neutrophils. A representative flow cytometry dot plot of MPO + , CD11b + , and MHC class II + neutrophils on day 3 is presented in . The percentage of MPO + , CD11b + , and MHC class II + cells in the different experimental groups is shown in column graphs (A, B, C, D, E, and F), while the absolute number of neutrophils obtained on day 3 PI is indicated (G). In all experiments, infected and non-infected BALB/c mice served as positive and negative controls, respectively. Results are presented as mean (±SEM) and are representative of two independent experiments. One-way ANOVA followed by Bonferroni’s post hoc test was used to analyze the data ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ns, non-significant ( p > 0.05).

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Infection, Saline, Isolation, Flow Cytometry

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: Increased CD38 + macrophages and CD8α + dendritic cells in MyD88 (PKO) - and DKK1 (PKO) -infected mice (A–D) Six-week-old female WT-BALB/c, MyD88 (PKO) and DKK1 (PKO) mice were challenged with infective metacyclic promastigote (2 ×10 6 parasites, n = 5 per group) of L . major via the footpad. One week and two weeks post-infection, cells from the infected foot of each mouse in BALB/c, MyD88 (PKO) , DKK1 (PKO) , and non-infected BALB/c mice ( n = 10/2 feet per mouse) were harvested and counted for macrophages (A and B) and dendritic cells (C and D) by flow cytometry. (E–H) Two weeks post-infection, the infected foot from each mouse in BALB/c, MyD88 (PKO) , DKK1 (PKO) ( n = 5 per group), and non-infected BALB/c mice ( n = 10/2 feet per mouse) were examined for CD206 + and CD38 + macrophages (E and F), as well as CD11b + and CD8α + dendritic cells (G and H) by flow cytometry. Representative flow cytometry dot plots showing the analysis of CD206 + , CD38 + macrophages and CD11b + , CD8α + dendritic cells and a dot plot of each sample in all the experimental groups performed on day 14 PI are presented in . Results are presented as mean ± SEM and are representative of two independent experiments. Data analysis was done using one-way ANOVA followed by Bonferroni’s post hoc test ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and “ns” indicates non-significant.

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Infection, Flow Cytometry

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: DKK1 promotes IL-10 induction in BALB/c-infected mice (A–F) Six-week-old female BALB/c, MyD88 (PKO) and DKK1 (PKO) mice were challenged with infective metacyclic promastigote (2 ×10 6 parasites, n = 5 per group) of L . major via the footpad. Non-infected BALB/c mice ( n = 5) were given 0.9% NaCl saline. Two or fifteen weeks post-infection, cells from draining lymph nodes were isolated. Lymph node cells were incubated with SLAG (50 μg/mL derived from WT parasites). Cell culture supernatant samples obtained were analyzed by ELISA for cytokine production, as shown in the column graphs (A–F). In all the experiments, BALB/c infected and non-infected mice served as positive and negative controls, respectively. Results are presented as mean ± SEM and are representative of two independent experiments. One-way ANOVA followed by Bonferroni’s post hoc test was used to analyze the data ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, “ns” indicates not significant ( p > 0.05).

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Infection, Saline, Isolation, Incubation, Derivative Assay, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: Impaired CD8 + IL-10 + IFN-γ − , CD4 + IL-10 + IFN-γ − , and CD4 + IL-10 + IFNg + T cells in MyD88 (PKO) - and DKK1 (PKO) -infected mice on day 14 PI (A–M) Six-week-old female BALB/c, MyD88 (PKO) , and DKK1 (PKO) mice were challenged with infective metacyclic promastigote (2 ×10 6 parasites, n = 5 per group) of L . major via the footpad. Non-infected BALB/c mice ( n = 5) were given 0.9% NaCl saline. Two weeks post-infection, the draining and non-draining lymph node cells were isolated. Lymph node cells were incubated with a cell stimulation cocktail for 5 h. After a further 3 h in BFA, cells were stained for intracellular IL-10 and IFN-γ. The stained lymph node cells from each mouse in BALB/c, MyD88 (PKO) - and DKK1 (PKO) -infected mice were determined for the percentage and frequency of CD4 + and CD8 + T cells (A, B, C, and B), percentage and MFI of CD8 + IFN-γ + or CD8 + IL-10 + T cells (E, F, G, and H), percentage and MFI of CD4 + IFN-γ + or CD4 + IL-10 + T cells (I, J, K, and L), and percentage of CD4 + IFN-γ + IL-10 + T cells (M) by flow cytometry. Representative flow cytometry dot plots showing the analyses CD4 + and CD8 + T cells, CD8 + IFN-γ + or CD8 + IL-10 + T cells, CD4 + IFN-γ + or CD4 + IL-10 + T cells, and CD4 + IFN-γ + IL-10 + T cells performed on day 14 PI is indicated ( A). Results are presented as mean ± SEM. and are representative of triplicate experiments. Data analysis was done using one-way ANOVA followed by Bonferroni’s post hoc test ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and “ns” indicates non-significant.

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Infection, Saline, Isolation, Incubation, Cell Stimulation, Staining, Flow Cytometry

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: Minimal expression and percentage of MHC II + , CD86 + , and CD80 + dendritic cells in the presence of recombinant DKK1 (A) Monocyte-derived dendritic cells were incubated in rDKK1(100 ng/mL), rIL-10 (20 ng/mL) and rTNF-α (10 ng/mL). Cells harvested at 24 and 48 h post-incubation were used to determine the expression and percentage of MHC II + , CD86 + , and CD80 + cells by flow cytometry. Representative flow cytometry dot plots generated 24 h post-incubation showed the analysis of MHC II + , CD86 + and CD80 + as indicated (A). (B–G) The percentage (B, D, and F) and MFI (C, E, and G) of MHC II + , CD86 + and CD80 + in the different experimental conditions are shown in the bar graphs. Results are presented as mean (±SEM) and are representative of triplicate experiments. One-way ANOVA followed by Bonferroni’s post hoc test was used to analyze the data (∗ p < 0.05, ∗∗ p < 0.01; ∗∗∗ p < 0.001; and “ns” indicates non-significant.

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Expressing, Recombinant, Derivative Assay, Incubation, Flow Cytometry, Generated

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: r-DKK1 blocked TNF-α-induced MHC II + and CD86 + expression of dendritic cells (A) Monocyte-derived dendritic cells were incubated in rDKK1(100 ng/mL), rTNF-α (10 ng/mL), and (rDKK1 + rTNF-α). Cells harvested at 48 h post-incubation were used to determine the percentage of MHC II + and CD86 + cells by flow cytometry. Representative flow cytometry dot plots generated 48 h post-incubation showed the analysis of MHC II + and CD86 + as indicated (A). (B and C) The percentage (B and C) of MHC II + and CD86 + in the different experimental conditions are shown in the bar graphs. Results are presented as mean (±SEM) and are representative of triplicate experiments. One-way ANOVA followed by Bonferroni’s post hoc test was used to analyze the data (∗ p < 0.05, ∗∗ p < 0.01; and “ns” indicates non-significant.

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Expressing, Derivative Assay, Incubation, Flow Cytometry, Generated

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: Elevated IL-10 and reduced IL-12 production in rDKK1-treated dendritic cells (A and B) Monocyte-derived dendritic cells were incubated in r-DKK1(100 ng/mL), r-IL-10 (20 ng/mL), r-TNF-α (10 ng/mL) or r-TNF-α + r-DKK1. Cell culture supernatants harvested 24 and 48 h post-incubation were used to determine IL-10 and IL-12 production by ELISA, as shown in the bar graphs (A and B). Results are presented as mean ± SEM and are representative of triplicate experiments. One-way ANOVA followed by Bonferroni’s post hoc test was used to analyze the data (∗ p < 0.05, ∗∗ p < 0.01). “ns” indicates not significant.

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Derivative Assay, Incubation, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: rDKK1-treated dendritic cells significantly increased the percentage of IL-10-producing CD4 T cells (A and B) Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 or 10 ng/mL of recombinant TNF-α for 24 h. The rDKK1 and r-TNF-α-treated dendritic cells were incubated for 4 h in a 24-well plate (2 × 10 5 per well) in 200 μL of medium in the presence of SLAG (100 ng/ml). SLAG-loaded rDKK1 and r-TNF-α-treated dendritic cells (2 ×10 5 ) were then washed and co-cultured with infected WT-BALB/c lymph node T cells (1 ×10 6 ) in complete RPMI 1640 medium in a 1:5 ratio for 72 h. The cells were surface-stained with appropriate antibodies before intracellular IL-10 staining with a cell stimulation cocktail and BD Golgi plug. The stained lymph node cells from the experimental condition were determined for the percentage of CD4 + IL-10 + T cells by flow cytometry. Representative flow cytometry dot plots showed the analysis of CD4 + IL-10 + T cells (A). The percentage of CD4 + IL-10 + T cells in the different experimental conditions is indicated (B). The non-treated and r-TNF-α-treated dendritic cells serve as controls. Results are presented as mean ± SEM and are representative of three replicate experiments. One-way ANOVA followed by Bonferroni’s post hoc test was used to analyze the data ∗∗∗ p < 0.001; ns indicates not significant ( p > 0.05).

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Recombinant, Incubation, Cell Culture, Infection, Staining, Cell Stimulation, Flow Cytometry

Journal: iScience

Article Title: Platelet DKK1 promotes tolerogenic dendritic cells and non-healing responses in cutaneous leishmaniasis

doi: 10.1016/j.isci.2026.115090

Figure Lengend Snippet: Lesion size and parasite burden decreased in MyD88 (PKO) - and DKK1 (PKO) -infected mice (A–C) Six-week-old female WT-BALB/c, MyD88 (PKO) and DKK1 (PKO) mice were challenged with infective metacyclic promastigote (2 ×10 6 parasites) of L . major via the footpad. The infected foot from each mouse in BALB/c, MyD88 (PKO) - and DKK1 (PKO) -infected mice ( n = 5 per group) were measured for lesion size weekly using a vernier caliper (A), and parasite burden (at day week 6 and 15PI) was determined by limiting dilution assay (B and C). Results are presented as mean ± SEM. For Figure (A), mice in each infected group were compared with the non-infected group and data analysis was done using one-way ANOVA followed by Bonferroni’s post hoc test ∗ p < 0.05; ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: Rested immature dendritic cells were stimulated with 100 ng/mL of recombinant DKK1 (R & D Systems), 20 ng/mL of recombinant IL-10 (Thermo Fisher Scientific), or 10 ng/mL of recombinant TNF-α (Thermo Fisher Scientific).

Techniques: Infection, Limiting Dilution Assay

Journal: The FASEB Journal

Article Title: MECP2 Insufficiency Attenuates RUNX2 ‐Dependent Osteoblast Differentiation via miR ‐126‐3p/ DKK1 ‐Mediated Canonical Wnt Signaling Inhibition in Rett Syndrome

doi: 10.1096/fj.202503014RR

Figure Lengend Snippet: MECP2 knockdown inhibits canonical Wnt signaling by upregulating DKK1. (A) Active β‐catenin levels were measured using western blotting. The mean ± standard error of the mean (SEM) was obtained from three independent experiments. (B) mRNA levels of DKK1 were measured using qRT‐PCR. DKK1 protein levels in culture media were measured using ELISA. The mean ± standard error of the mean (SEM) was obtained from three independent experiments. * p < 0.05; ** p < 0.01.

Article Snippet: Extracellular DKK1 levels were measured using a Human DKK1 Quantikine ELISA Kit (R&D systems, Minneapolis, MN, USA, #DKK100B) according to the manufacturer's instructions.

Techniques: Knockdown, Western Blot, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: The FASEB Journal

Article Title: MECP2 Insufficiency Attenuates RUNX2 ‐Dependent Osteoblast Differentiation via miR ‐126‐3p/ DKK1 ‐Mediated Canonical Wnt Signaling Inhibition in Rett Syndrome

doi: 10.1096/fj.202503014RR

Figure Lengend Snippet: DKK1 knockdown promotes the canonical Wnt signaling in MECP2‐insufficient osteoblasts. (A) mRNA and protein levels of DKK1 were measured using qRT‐PCR and ELISA, respectively. The mean ± standard error of the mean (SEM) was obtained from three independent experiments for qRT‐PCR and four independent experiments for ELISA. (B) Active β‐catenin levels were measured using western blotting. The mean ± standard error of the mean (SEM) was obtained from three independent experiments. * p < 0.05; ** p < 0.01; n.s., not significant.

Article Snippet: Extracellular DKK1 levels were measured using a Human DKK1 Quantikine ELISA Kit (R&D systems, Minneapolis, MN, USA, #DKK100B) according to the manufacturer's instructions.

Techniques: Knockdown, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: The FASEB Journal

Article Title: MECP2 Insufficiency Attenuates RUNX2 ‐Dependent Osteoblast Differentiation via miR ‐126‐3p/ DKK1 ‐Mediated Canonical Wnt Signaling Inhibition in Rett Syndrome

doi: 10.1096/fj.202503014RR

Figure Lengend Snippet: DKK1 knockdown improves RUNX2‐dependent differentiation and mineralization in MECP2‐insufficient osteoblasts. (A) mRNA levels of RUNX2 , ALP , and OCN were measured using qRT‐PCR. The mean ± standard error of the mean (SEM) was obtained from four independent experiments for RUNX2 and three independent experiments for ALP and OCN . (B) Mineralization was measured using Alizarin Red S staining. The staining intensity was quantified and statistically analyzed using ImageJ software. The mean ± standard error of the mean (SEM) was obtained from three independent experiments. * p < 0.05; ** p < 0.01; n.s., not significant.

Article Snippet: Extracellular DKK1 levels were measured using a Human DKK1 Quantikine ELISA Kit (R&D systems, Minneapolis, MN, USA, #DKK100B) according to the manufacturer's instructions.

Techniques: Knockdown, Quantitative RT-PCR, Staining, Software

Journal: The FASEB Journal

Article Title: MECP2 Insufficiency Attenuates RUNX2 ‐Dependent Osteoblast Differentiation via miR ‐126‐3p/ DKK1 ‐Mediated Canonical Wnt Signaling Inhibition in Rett Syndrome

doi: 10.1096/fj.202503014RR

Figure Lengend Snippet: miR‐126‐3p upregulates DKK1 in MECP2‐insufficient osteoblasts. (A) Expression levels of miR‐126‐3p were measured using qRT‐PCR. The mean ± standard error of the mean (SEM) was obtained from three independent experiments. (B) Expression levels of miR‐126‐3p and DKK1 were measured using qRT‐PCR. DKK1 protein levels in culture media were measured using ELISA. The mean ± standard error of the mean (SEM) was obtained from three independent experiments. * p < 0.05; ** p < 0.01; n.s., not significant.

Article Snippet: Extracellular DKK1 levels were measured using a Human DKK1 Quantikine ELISA Kit (R&D systems, Minneapolis, MN, USA, #DKK100B) according to the manufacturer's instructions.

Techniques: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: The FASEB Journal

Article Title: MECP2 Insufficiency Attenuates RUNX2 ‐Dependent Osteoblast Differentiation via miR ‐126‐3p/ DKK1 ‐Mediated Canonical Wnt Signaling Inhibition in Rett Syndrome

doi: 10.1096/fj.202503014RR

Figure Lengend Snippet: Schematic representation of the proposed and potential pathways through which MECP2 insufficiency disrupts osteoblast differentiation. The proposed pathway is shown as black solid lines. MECP2 insufficiency leads to upregulation of miR‐126‐3p, which in turn enhances the endogenous Wnt antagonist DKK1. DKK1 inhibits the canonical Wnt signaling pathway, thereby impairing RUNX2‐dependent osteoblast differentiation. (A–F) Considering the broad functions of MECP2 and miRNAs, the potential pathways discussed in the study limitations section of the Discussion are depicted as gray dotted lines.

Article Snippet: Extracellular DKK1 levels were measured using a Human DKK1 Quantikine ELISA Kit (R&D systems, Minneapolis, MN, USA, #DKK100B) according to the manufacturer's instructions.

Techniques: