Journal: Journal of Immunology Research

Article Title: DKK2 Impairs Tumor Immunity Infiltration and Correlates with Poor Prognosis in Pancreatic Ductal Adenocarcinoma

doi: 10.1155/2019/8656282

Figure Lengend Snippet: Correlations of DKK2 with clinical characteristics in PDAC patients.

Article Snippet: Antibodies used in our work include DKK2 (21051-1-AP, Proteintech), CD8A (ab17147, Abcam), T-bet (sc-21763, Santa Cruz), GATA3 (ab199428, Abcam), and FOXP3 (22228-1-AP, Proteintech).

Techniques: Expressing

Journal: Journal of Immunology Research

Article Title: DKK2 Impairs Tumor Immunity Infiltration and Correlates with Poor Prognosis in Pancreatic Ductal Adenocarcinoma

doi: 10.1155/2019/8656282

Figure Lengend Snippet: DKK2 was highly expressed in PDAC and was associated with poor prognosis in PDAC patients. (a, b) The mRNA expression level between tumor tissue and adjacent normal tissue in GSE15471 dataset, TCGA data, Oncomine, GTEx data, and Renji cohort. (c) DKK2 expression plot of various types of cancer from TCGA database. (d) Representative scoring criterion images of immunohistochemical staining of DKK2 expression in 208 paired PADC tumor and adjacent nontumor tissues. Scale bar: 50 μ m. (e) DKK2 was upregulated in PDAC tumor tissues. (f) High DKK2 expression correlated with inferior survival time. Abbreviations: TCGA: The Cancer Genome Atlas; GTEx: Genotype-Tissue Expression. Significant differences are indicated as ∗ P < 0.05; ∗∗∗∗ P < 0.0001.

Article Snippet: Antibodies used in our work include DKK2 (21051-1-AP, Proteintech), CD8A (ab17147, Abcam), T-bet (sc-21763, Santa Cruz), GATA3 (ab199428, Abcam), and FOXP3 (22228-1-AP, Proteintech).

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Journal of Immunology Research

Article Title: DKK2 Impairs Tumor Immunity Infiltration and Correlates with Poor Prognosis in Pancreatic Ductal Adenocarcinoma

doi: 10.1155/2019/8656282

Figure Lengend Snippet: Prognostic significance of DKK2 assessed in PDAC patients classified by three conditions. The correlation between DKK2 expression and overall survival rate with patients grouped by clinical classification I-II stage (a), clinical classification III-IV stage (b), without node metastasis (c), with node metastasis (d), without vascular invasion (e), and with vascular invasion (f).

Article Snippet: Antibodies used in our work include DKK2 (21051-1-AP, Proteintech), CD8A (ab17147, Abcam), T-bet (sc-21763, Santa Cruz), GATA3 (ab199428, Abcam), and FOXP3 (22228-1-AP, Proteintech).

Techniques: Expressing

Journal: Journal of Immunology Research

Article Title: DKK2 Impairs Tumor Immunity Infiltration and Correlates with Poor Prognosis in Pancreatic Ductal Adenocarcinoma

doi: 10.1155/2019/8656282

Figure Lengend Snippet: Univariate and multivariate analyses of prognostic parameters for survival in PDAC patients.

Article Snippet: Antibodies used in our work include DKK2 (21051-1-AP, Proteintech), CD8A (ab17147, Abcam), T-bet (sc-21763, Santa Cruz), GATA3 (ab199428, Abcam), and FOXP3 (22228-1-AP, Proteintech).

Techniques:

Journal: Journal of Immunology Research

Article Title: DKK2 Impairs Tumor Immunity Infiltration and Correlates with Poor Prognosis in Pancreatic Ductal Adenocarcinoma

doi: 10.1155/2019/8656282

Figure Lengend Snippet: Knockdown of DKK2 promoted tumor cells' migration ability. (a) The enrichment plots in DKK2-highly expressed group compared to the low performed by gene set enrichment analysis. (b) Transwell assay showed that silencing DKK2 led to more migrated cells in AsPC-1 and CAPAN-1. For each transwell assay, the cell number was counted (c). (d) Movement of DKK2 knockout and control cells into the wound was shown in AsPC-1 and CAPAN-1 at 0 h and 12 h. The percentage of wound healing was assessed (e). Significant differences are indicated as ∗ P < 0.05 and ∗∗ P < 0.001.

Article Snippet: Antibodies used in our work include DKK2 (21051-1-AP, Proteintech), CD8A (ab17147, Abcam), T-bet (sc-21763, Santa Cruz), GATA3 (ab199428, Abcam), and FOXP3 (22228-1-AP, Proteintech).

Techniques: Knockdown, Migration, Transwell Assay, Knock-Out, Control

Journal: Journal of Immunology Research

Article Title: DKK2 Impairs Tumor Immunity Infiltration and Correlates with Poor Prognosis in Pancreatic Ductal Adenocarcinoma

doi: 10.1155/2019/8656282

Figure Lengend Snippet: DKK2 impacted the infiltration capability of immune cells. (a) Gene set enrichment analysis of DKK2 showed the potential relationship between DKK2 and PDAC immunity invasion. (b) Representative immunohistochemical samples of DKK2, CD8A, FOXP3, GATA3, and T-bet in PDAC tissue microarray. (c) Patients were divided into two groups according to the expression of DKK2, and the column bar graphs showed the correlation between CD8A, FOXP3, GATA3, T-bet, and DKK2. (d) PBMCs treated with siRNA were performed with migration assay and then were counted by flow cytometry, and the percent was displayed by the bar graph (e). (f) Representative flow cytometry profiles of T cell subtype were shown. CD4 + and CD8 + cells, which were gated on single cells and lymphocytes, were gated as depicted. Significant differences are indicated as ∗ P < 0.05, ∗∗ P < 0.001, and ∗∗∗ P < 0.0001.

Article Snippet: Antibodies used in our work include DKK2 (21051-1-AP, Proteintech), CD8A (ab17147, Abcam), T-bet (sc-21763, Santa Cruz), GATA3 (ab199428, Abcam), and FOXP3 (22228-1-AP, Proteintech).

Techniques: Immunohistochemical staining, Microarray, Expressing, Migration, Flow Cytometry

Journal: Journal of Ovarian Research

Article Title: LINC01094/miR-577 axis regulates the progression of ovarian cancer

doi: 10.1186/s13048-020-00721-9

Figure Lengend Snippet: Knockdown of LINC01094 inhibited the proliferation, migration, invasion and EMT of OC cells. a SKOV3 and 3AO cells were transfected with si-LINC01094 or si-NC and the inhibition efficiency was detected by qRT-PCR assay. b CCK-8 assay was used to detect the effect of LINC01094 on the proliferation of SKOV3 and 3AO cells. c - d The effects of LINC01094 on the migration and invasion of SKOV3 ( c ) and 3AO ( d ) cells were measured using Transwell assay. e Western blot was adopted to analyze the effect of LINC01094 on the expressions of EMT markers E-cadherin and Vimentin. All experiments were performed in triplicate. ** P < 0.01 and *** P < 0.001

Article Snippet: The primary antibodies used in this work included anti-E-cadherin antibody (Abcam, ab197751, 1:2000), anti-vimentin antibody (Abcam, ab92547, 1:1000), anti-β-catenin antibody (Abcam, ab16051, 1:1000), anti-c-Myc antibody (Abcam, ab32072, 1:2000), anti-cyclin D1 antibody (Abcam, ab16663, 1:2000) and anti-GAPDH antibody (Abcam, ab9484, 1:4000).

Techniques: Knockdown, Migration, Transfection, Inhibition, Quantitative RT-PCR, CCK-8 Assay, Transwell Assay, Western Blot

Journal: Journal of Ovarian Research

Article Title: LINC01094/miR-577 axis regulates the progression of ovarian cancer

doi: 10.1186/s13048-020-00721-9

Figure Lengend Snippet: Inhibition of miR-577 promoted the proliferation, migration and invasion of OC cells. a SKOV3 and 3AO cells were transfected with miR-577-in or miR-con-in, and the inhibition efficiency was detected by qRT-PCR assay. b CCK-8 assay was used to detect the effect of miR-577 on the proliferation of SKOV3 and 3AO cells. c - d The effects of miR-577 on the migration and invasion of SKOV3 ( c ) and 3AO ( d ) cells were measured using Transwell assay. e Western blot was adopted to analyze the effect of miR-577 on the expressions of EMT markers E-cadherin and Vimentin. All experiments were performed in triplicate. ** P < 0.01 and *** P < 0.001

Article Snippet: The primary antibodies used in this work included anti-E-cadherin antibody (Abcam, ab197751, 1:2000), anti-vimentin antibody (Abcam, ab92547, 1:1000), anti-β-catenin antibody (Abcam, ab16051, 1:1000), anti-c-Myc antibody (Abcam, ab32072, 1:2000), anti-cyclin D1 antibody (Abcam, ab16663, 1:2000) and anti-GAPDH antibody (Abcam, ab9484, 1:4000).

Techniques: Inhibition, Migration, Transfection, Quantitative RT-PCR, CCK-8 Assay, Transwell Assay, Western Blot

Journal: Scientific Reports

Article Title: Trypanosoma cruzi infection induces DNA double-strand breaks and activates DNA damage response pathway in host epithelial cells

doi: 10.1038/s41598-024-53589-w

Figure Lengend Snippet: Trypanosoma cruzi infection induces rapid H2AX phosphorylation in LLC-MK2 cells in response to the DNA damage. (A) Confocal microscopy images of non-infected (NI) or T. cruzi -infected LLC-MK2 cells (2–24 hpi) are stained with anti-γ-H2AX antibody (green), demonstrating DNA damage in host cell nuclei. Actin is stained with rhodamin-phalloidin (red), host and parasite nuclei and kinetoplasts (arrowheads) are labelled with DAPI (blue). Host cell nuclei were delimited by a dotted line. Merged images are shown as indicated. Scale bar: 10 µm. (B) Quantification of the nuclear γ-H2AX fluorescence intensity of images acquired in epifluorescence microscopy during parasite progression of infection (n > 100 cells/time points) and analysed by ImageJ. Data presented are mean ± SEM, representative of two independent experiments (n = 2). ***: (ρ < 0.001) obtained by the One-way ANOVA test and the comparison performed by Dunnett as a posthoc test in the GraphPad Prism v.8.0 software. a.u. arbitrary units. Bars represent a mean value from the dataset. (C) Western Blot of LLC-MK2 cells lysates highlighting active levels of phospho-H2AX at different times of infection (2–24 hpi) compared to non-infected (NI) cells revealed with anti-γ-H2AX antibody. GAPDH was labelled with an anti-GAPDH antibody as endogenous control. Data are representative of two independent experiments (n = 2). kDa kilodaltons. (D) The bar graph represents the relative intensity bands in ( C ) normalized with the GAPDH protein.

Article Snippet: The antibodies used in this work include the following commercial antibodies: anti-phospho-histone H2AX (Ser139) (#80312-mouse), anti-phospho-53BP1 (Ser1778) (#2675-rabbit), anti-phospho-ATR (Thr1989) (#30632-rabbit), anti-phospho-Rad50 (Ser635) (#14223-rabbit) and anti-GAPDH (#3683-rabbit-HRP conjugated) from Cell Signaling Technology, USA.

Techniques: Infection, Phospho-proteomics, Confocal Microscopy, Staining, Fluorescence, Epifluorescence Microscopy, Comparison, Software, Western Blot, Control

Journal: Scientific Reports

Article Title: Trypanosoma cruzi infection induces DNA double-strand breaks and activates DNA damage response pathway in host epithelial cells

doi: 10.1038/s41598-024-53589-w

Figure Lengend Snippet: Phospho-53BP1 protein and phospho-ATM kinase are recruited rapidly to the nuclear foci in response to T. cruzi infection. (A) Confocal microscopy images are showing the localisation of phospho-53BP1 (red) and phospho-H2AX (green) in T. cruzi -infected LLC-MK2 cells at different time points (2–24 hpi). Host and parasite nuclei and kinetoplasts are stained with DAPI (blue). Arrowheads indicate parasites. Host cell nuclei were delimited by a dotted line. Merged images are shown as indicated. Scale bar: 10 µm. (B) Quantification of phospho-53BP1 nuclear fluorescence intensity acquired by epifluorescence microscopy during the progression of parasitic infection (n > 50 cells/time point). Data are representative of three independent experiments (n = 3). ***: (ρ < 0.001) obtained by the Kruskal–Wallis test and Dunn's multiple comparisons as a posthoc test in GraphPad Prism v.8.0 software. a.u. arbitrary units. The black bar shows the median value for each time point. (C) Confocal microscopy images of T. cruzi -infected LLC-MK2 cells (2–24 hpi) and NI cells showing the localisation of phospho-ATM (red) and phospho-H2AX (green) proteins in the cell nuclei. Host cell nuclei, parasite nuclei and kinetoplasts (arrowheads) were stained with DAPI (blue). Host cell nuclei were delimited by a dotted line. Merged images are shown as indicated. Scale bar: 10 µm. (D) Quantification of nuclear phospho-ATM fluorescence intensity in T. cruzi -infected cells obtained by epifluorescence microscopy compared to NI cells (n > 50 cells/time point). Black bars indicate the median value of the dataset. a.u. arbitrary units. ***: (ρ < 0.001), *: (ρ < 0.05), ns: (ρ > 0.05) obtained by the Kruskal–Wallis test and Dunn's multiple comparisons as a posthoc test in GraphPad Prism v.8.0 software. Data are representative of three independent experiments (n = 3).

Article Snippet: The antibodies used in this work include the following commercial antibodies: anti-phospho-histone H2AX (Ser139) (#80312-mouse), anti-phospho-53BP1 (Ser1778) (#2675-rabbit), anti-phospho-ATR (Thr1989) (#30632-rabbit), anti-phospho-Rad50 (Ser635) (#14223-rabbit) and anti-GAPDH (#3683-rabbit-HRP conjugated) from Cell Signaling Technology, USA.

Techniques: Infection, Confocal Microscopy, Staining, Fluorescence, Epifluorescence Microscopy, Software

Journal: Scientific Reports

Article Title: Trypanosoma cruzi infection induces DNA double-strand breaks and activates DNA damage response pathway in host epithelial cells

doi: 10.1038/s41598-024-53589-w

Figure Lengend Snippet: DSB repair is dependent by DNA-PK kinase during T. cruzi infection. (A) Confocal microscopy of T. cruzi -infected LLC-MK2 cells (2–24 hpi) and NI cells showing the localisation of phospho-DNA-PK (red) and phospho-H2AX (green) proteins. Host and parasite nuclei and kinetoplasts were stained with DAPI (blue). Host cell nuclei were delimited by a dotted line. Arrowheads indicate intracellular parasites. Merged images are shown as indicated. Scale bar: 10 µm. (B) Quantification of nuclear phospho-DNA-PK fluorescence intensity of T. cruzi -infected cells obtained by epifluorescence microscopy (n > 50 cells/time point). Data are representative of three independent experiments (n = 3). ***: (ρ < 0.001) obtained using the Kruskal–Wallis test and Dunn's multiple comparisons as a posthoc test in GraphPad Prism v.8.0 software. Black bars indicate the median value of the dataset. (C) Quantification of DNA damage by comet assay in LLC-MK2 cells that were either infected (+) or not (−) with T. cruzi (12 and 24 hpi), treated with the specific DNA-PK inhibitor (AZD7648) or with DMSO vehicle. The median value of each group is represented by the black bar. ns: (ρ > 0.05), ***: (ρ < 0.001). Statistical analysis was performed using the Kruskal–Wallis test followed by Dunn's multiple comparisons post hoc test in GraphPad Prism v.8.0 software.

Article Snippet: The antibodies used in this work include the following commercial antibodies: anti-phospho-histone H2AX (Ser139) (#80312-mouse), anti-phospho-53BP1 (Ser1778) (#2675-rabbit), anti-phospho-ATR (Thr1989) (#30632-rabbit), anti-phospho-Rad50 (Ser635) (#14223-rabbit) and anti-GAPDH (#3683-rabbit-HRP conjugated) from Cell Signaling Technology, USA.

Techniques: Infection, Confocal Microscopy, Staining, Fluorescence, Epifluorescence Microscopy, Software, Single Cell Gel Electrophoresis