mouse il 13rα2  (R&D Systems)

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 1. Expression of IL-13Rα2 in breast cancer tissue. (a) RT-PCR analysis of IL-13Rα2 mRNA expression in cDNA array samples (TissueScan™, Origene) derived from breast cancer (number of samples (n = 60)) and non-malignant (n = 7) tissue. Shown is the fold-change relative to mean expression in non-malignant samples (mean fold-change, cancer versus non-malignant; *** p ≤0.001). (b) Stratification analysis of IL-13Rα2 mRNA expression data comparing TNBC-type (n = 19) versus non-TNBC (n = 41) tumors (mean fold-change, TNBC versus non-TNBC, *** p ≤0.001). (c) Im- munohistochemical analysis of IL-13Rα2 protein expression in breast cancer tissue array samples (US Biomax, BR1009). Shown images (at 40× magnification) are representative examples of specific staining with an anti-IL-13Rα2 antibody (sc-134363) versus a mouse IgG2a isotype control: (left panel), non-malignant tissue; (middle panel) non-TNBC tumor (ER+, stage IIa); (right panel) TNBC tumor (stage IIb). (Right Figure), scoring analysis of anti-IL-13Rα2 reactivity in non-malignant (n = 9), non-TNBC (n = 8), and TNBC (n = 21) tissue array sections (mean score, TNBC versus non-malignant, *** p ≤0.001).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Derivative Assay, Staining, Control

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 2. Expression of IL-13Rα2 in breast cancer cell lines. (a) RT-PCR analysis of IL-13Rα2 mRNA expression in non-malignant (MCF-10A), non-TNBC (MCF-7), and TNBC (MDA-MB-231, LM2) cells. (b) Quantification of IL-13Rα2 protein expression in representative cell lines by Western blot. For densitometry, individual band density was normalized against α-tubulin expression. The uncropped bolts are shown in Supplementary Materials File S1. (c) Cell-surface expression of IL-13Rα2 protein was measured in non-permeabilized cells by cell-based ELISA. All graphs show fold-change relative to mean expression in MCF-10A cells. Data = mean values ± SEM of three independent experiments (*** p ≤0.001; **** p ≤0.0001).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, In-Cell ELISA

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 3. Evaluation of Pep-1-Phor21 activity. (a) Protein samples from transfected COS-7 cells expressing the red fluorescent protein (RFP) mCherry (27kDa) or IL-13Rα2-mCherry (75 kDa) were immunoblotted (IB) using either an anti-RFP or an anti-IL-13Rα2 antibody with α-tubulin detection as a loading control. The uncropped bolts are shown in Supplementary Materials File S1. (b) COS- 7 cells transfected with IL-13Rα2-mCherry plasmid or empty mCherry vector were treated with Pep-1-Phor21 (O) or Phor21 (S) at 0–10 µM for 3 h and their viability was determined by Alamar Blue assay (IC50 for Pep-1-Phor21 against IL-13Rα2-transfected cells = 0.133 µM ± 0.04). (c) CellTox assay was used to determine the cytotoxic activity of Pep-1-Phor21 against IL-13Rα2-transfected cells (IC50 = 0.130 µM ± 0.03). (d) IL-13Rα2- or LHCGR-transfected COS-7 cells were treated with 0.5 µM Pep-1-Phor21 or βCG-Phor21 for 3 h and their relative cytotoxicity was determined (versus empty vector-transfected cells). Only IL-13Rα2-transfected cells exhibited susceptibility to the cytotoxic effects of Pep-1-Phor21, whilst remaining resistant to βCG-Phor21. Data = mean value ± SEM of three independent experiments (ns = not significant).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Activity Assay, Transfection, Expressing, Control, Plasmid Preparation, Alamar Blue Assay, CellTox Assay

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 4. Treatment of representative breast cancer cell lines with Pep-1-Phor21. (a) Dose-dependent effect of Pep-1-Phor21 (O), Pep-1 (S), or Phor21 (∆) on the viability of non-tumorigenic (MCF-10A), non- TNBC (MCF-7), and TNBC cells (MDA-MB 231, LM2). Cells were treated with different concentrations of Pep-1-Phor21 (effective concentration range = 0–120 µM, a 5-fold serial dilution) and their viability was assessed after 3 h, 6 h, or 24 h of the treatment by Alamar Blue assay. (b) The IC50 of peptides for various cell lines for different incubation times. (c) The cytotoxic effect of individual peptides on cell lines was also assessed by CellTox assay. MCF-10A and MCF-7 cells were treated for 3 h with 120 µM Pep-1-Phor21 (maximum concentration used in the dose-response analysis, Alamar Blue, Figure 4a). MDA-MB 231 and LM2 cells were treated with Pep-1-Phor21 at 24 µM (maximal effective concentration against LM2, as determined in dose-response analysis, Alamar Blue, Figure 4a). Pep-1- Phor21 had a significant cytotoxic effect only against IL-13Rα2-expressing TNBC cells (MDA-MB-231, LM2; relative cytotoxicity = 85.2% ± 5.4 and 96.9% ± 0.38, respectively, versus non-treated cells). Data = mean value ± SEM of three independent experiments (* p ≤0.05; ** p ≤0.01; *** p ≤0.001; **** p ≤0.0001).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Concentration Assay, Serial Dilution, Alamar Blue Assay, Incubation, CellTox Assay, Expressing

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 6. Epigenetic modulation of IL-13Rα2 expression in breast cancer cells. Non-malignant breast epithelial (MCF-10A), non-TNBC (MCF-7), and TNBC (MDA-MB 231, LM2) cell lines were treated with an HDAC inhibitor (TSA) or with a DNMT inhibitor (5-aza-dC) for 24 h and IL-13Rα2 expression at the mRNA, total protein, and cell-surface levels were subsequently measured by RT-PCR (a), Western blot (b), and cell-based ELISA (c), respectively. Treatment with either epigenetically active compound induced detectable levels (all assays) of IL-13Rα2 expression in MCF-7 cells, whereas expression was not altered in MCF-10A cells (remaining IL-13Rα2-negative). In IL-13Rα2-positive cells, 10 µM TSA significantly upregulated IL-13Rα2 expression in MDA-MB-231 cells, as measured in all assays. Data = mean value ± SEM of three independent experiments (* p ≤0.05; ** p ≤0.01; *** p ≤0.001; ns = non-significant; relative to untreated cells).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, In-Cell ELISA

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 8. IL-13Rα2 expression in breast cancer spheroids and targeting with Pep-1-Phor21. (a) IL- 13Rα2 mRNA expression was determined by RT-PCR in the indicated cell lines cultured as 3-D spheroids for 48 h (fold-change relative to MCF-10A cells). (b) Established spheroids (48 h) were treated for 3 h with the indicated peptides (dose-response range = 0–120 µM) and their cytotoxicity was assessed (CellTox assay). Only IL-13Rα2-positive, TNBC spheroids exhibited susceptibility to the cytotoxic effect of Pep-1-Phor21 (MDA-MB-231, LM2; IC50 = 22.98 µM ± 1.5, 12.22 µM ± 2.5, respectively). Data = mean value ± SEM of three independent experiments (* p ≤0.05; *** p ≤0.001).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Cell Culture, CellTox Assay

Journal: Cancers

Article Title: Targeting and Sensitization of Breast Cancer Cells to Killing with a Novel Interleukin-13 Receptor α2-Specific Hybrid Cytolytic Peptide.

doi: 10.3390/cancers15102772

Figure Lengend Snippet: Figure 10. Epigenetic modulation of IL-13Rα2 expression in spheroids and targeting with Pep-1- Phor21. (a) Established spheroids (48 h) were treated with either 5-aza-dC or TSA (−10 µM, 24 h), and relative IL-13Rα2 mRNA expression was determined by RT-PCR (fold-change relative to expression in untreated spheroid cells). Whilst non-malignant MCF-10A spheroids remained refractory to treatment, both compounds significantly upregulated IL-13Rα2 expression in non-TNBC MCF-7 spheroids. Similarly, 10µM 5-aza-dC/TSA significantly upregulated IL-13Rα2 expression in TNBC MDA-MB-231 spheroids. (b) Spheroids pre-treated with 5-aza-dC or TSA (dose-response range 0–10 µM) were subsequently treated with Pep-1-Phor21 (MCF-10A, MCF-7, MDA-MB-231, and LM2 were treated with 100 µM, 50 µM, 25 µM, and 10 µM, respectively, based on pre-determined IC50 values for each spheroid type, Figure 8b) for 3 h, and relative cytotoxicity was evaluated (CellTox assay, relative to untreated spheroids). Data = mean value ± SEM of three experiments (* p ≤0.05; ** p ≤0.01; ns = not significant).

Article Snippet: IL-13Rα2 protein was detected with an anti-IL-13Rα2 mouse monoclonal antibody (sc-134363, Santa Cruz Biotechnology) diluted 1:500 in blocking buffer (TBS, 0.1% Tween-20 containing 5% milk powder) followed by incubation with an HRP-conjugated anti-mouse secondary antibody.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, CellTox Assay

Journal: Frontiers in Endocrinology

Article Title: Vertical sleeve gastrectomy associates with airway hyperresponsiveness in a murine model of allergic airway disease and obesity

doi: 10.3389/fendo.2023.1092277

Figure Lengend Snippet: Summary of Findings.

Article Snippet: Leptin (DY498), IL-13Rα2 (DY539), total (EMIGHE) and HDM-specific (3037) IgE, IL-13 (DY413), IL-5 (DY405), and total and active TGF-β1 (DY1679), were measured in serum, BAL fluid or homogenized lung tissue by ELISA using kits purchased from R&D systems or ThermoFisher Scientific.

Techniques: