anti integrin αvβ3  (Santa Cruz Biotechnology)

Journal: Journal of Cellular and Molecular Medicine

Article Title: CD44 Participates to Extramedullary Haematopoiesis Onset by Mediating the Interplay Between Monocytes and Haematopoietic Stem Cells in Myelofibrosis

doi: 10.1111/jcmm.70720

Figure Lengend Snippet: CD44 is highly expressed in MF monocytes and HSPCs and its expression is increased in the spleen of MF mice. (a) Immunophenotypic evaluation of α4β1, αvβ3 and CD44 in circulating CD14 + monocytes in HDs ( n = 15) and MF patients ( n = 8). (b) Immunophenotypic evaluation of α4β1, αvβ3 and CD44 in circulating CD34 + cells in HDs ( n = 15) and MF patients ( n = 8). (c, d) Flow cytometry evaluation of CD44 , α4β1 and αvβ3 expression in macrophages (c) and LSK cells (d) in the spleen of NT ( n = 2) and TPO ‐ RA ( n = 4) treated mice. Each dot represents a mouse. (e, f) Flow cytometry evaluation of the frequency of CD44 + macrophages (e) and LSK cells (f) in the spleen of NT ( n = 2), TPO ‐ RA ( n = 4) and TPO ‐ RA + Ruxolitinib ( n = 5) treated mice. Each dot represents a mouse. In graphs, each dot represents a patient or a mouse, and each column represents group mean ± standard deviation. Comparisons were performed by means of unpaired T ‐test for panels (a–d) and one‐way ANOVA for panels (e, f). p ‐values are reported only when < 0.05.

Article Snippet: For in vitro extravasation assay the following antibodies and corresponding isotype controls were used: monoclonal anti‐CD44 antibody (clone IM7, cat. #14‐0441‐82), rat IgG2a kappa Isotype Control as control IgG for the monoclonal anti‐CD44 antibody (clone eBR2a, cat. #14‐4321‐82; both from eBioscience, Invitrogen, ThermoFisher, Waltham, Massachussets, USA); Polyclonal anti‐CD44 antibody (clone HCAM DF1485, cat. #sc‐7297), anti‐αvβ3 antibody (clone 23C6, cat. #sc‐7312), normal mouse IgG as control IgG for the polyclonal anti‐CD44 and anti‐αvβ3 antibodies (cat. #sc‐2025, all from Santa Cruz Biotechnology Inc., Dallas, Texas, USA); anti‐α4β1 antibody, InVivoMAb anti‐mouse/human VLA‐4 (CD49d) (clone PS/2, cat. #BE0071), InVivoMAb rat IgG2b isotype control as control IgG for the anti‐α4β1 antibody (clone LTF‐2, cat. #BE0090, all from BioXCell, Lebanon, New Hampshire, USA); anti‐human OPN Antibody (cat. AF1433, Biotechne, R&D systems, Minnesota, USA); Hyaluronan inhibitor (cat. #AS‐62622, Anaspec, California, USA).

Techniques: Expressing, Flow Cytometry, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: CD44 Participates to Extramedullary Haematopoiesis Onset by Mediating the Interplay Between Monocytes and Haematopoietic Stem Cells in Myelofibrosis

doi: 10.1111/jcmm.70720

Figure Lengend Snippet: CD44 regulates HD and MF CD14 + monocyte migration. Panel (a) shows the effect of anti‐αvβ3 antibody on migration of HD CD14 + monocyte in vitro in presence of HUVEC coating while panel (b) displays results for the anti‐α4β1 antibody treatment. Panel (c) shows the effect of the polyclonal anti‐CD44 antibody on migration of HD CD14 + monocyte in vitro in presence of HUVEC endothelium while panel (d) displays results for the monoclonal anti‐CD44 antibodies. (e) Graph displays the difference in migration between HD ( n = 6) and MF ( n = 3) CD14 + monocytes in the established in vitro extravasation model. (f) The effect of anti‐CD44 antibody on migration of MF CD14 + monocyte in vitro ( n = 3). (g) Bar plot represents results of HD CD14 + monocyte migration assays with and without HUVEC for the monoclonal anti‐CD44 antibody; Transwell samples with HUVEC are represented with purple bars while Transwell samples without HUVEC are illustrated with pink bars; the percentage of migration is normalised considering control IgG samples, not shown in this graph. In all bar plots the red dashed line represents 100% migration. On X axis are reported antibodies doses, Y axis reports the normalised cell count (see Section ). In graphs each dot represents a replicate, each column represents group mean ± standard deviation. Comparisons between samples treated with the specific antibody or control IgG were performed by means of paired T ‐test. p ‐values are reported only if < 0.05.

Article Snippet: For in vitro extravasation assay the following antibodies and corresponding isotype controls were used: monoclonal anti‐CD44 antibody (clone IM7, cat. #14‐0441‐82), rat IgG2a kappa Isotype Control as control IgG for the monoclonal anti‐CD44 antibody (clone eBR2a, cat. #14‐4321‐82; both from eBioscience, Invitrogen, ThermoFisher, Waltham, Massachussets, USA); Polyclonal anti‐CD44 antibody (clone HCAM DF1485, cat. #sc‐7297), anti‐αvβ3 antibody (clone 23C6, cat. #sc‐7312), normal mouse IgG as control IgG for the polyclonal anti‐CD44 and anti‐αvβ3 antibodies (cat. #sc‐2025, all from Santa Cruz Biotechnology Inc., Dallas, Texas, USA); anti‐α4β1 antibody, InVivoMAb anti‐mouse/human VLA‐4 (CD49d) (clone PS/2, cat. #BE0071), InVivoMAb rat IgG2b isotype control as control IgG for the anti‐α4β1 antibody (clone LTF‐2, cat. #BE0090, all from BioXCell, Lebanon, New Hampshire, USA); anti‐human OPN Antibody (cat. AF1433, Biotechne, R&D systems, Minnesota, USA); Hyaluronan inhibitor (cat. #AS‐62622, Anaspec, California, USA).

Techniques: Migration, In Vitro, Control, Cell Counting, Standard Deviation

Journal: International Journal of Biological Sciences

Article Title: Apelin facilitates integrin αvβ3 production and enhances metastasis in prostate cancer by activating STAT3 and inhibiting miR-8070

doi: 10.7150/ijbs.113161

Figure Lengend Snippet: Figure 2. Apelin enhances integrin αvβ3-dependent prostate cancer motility. (A) Cells were treated with apelin for 24 hours, the indicated integrin expression was examined by qPCR (n=3). (B&C) Cells were treated with integrin αvβ3 or α5β1 antibody then with apelin, the wound healing and cell migration was examined (n=3). (D) Integrin αv and β3 gene levels in normal and prostate cancer patients retrieved from the GEO database. (E&F) Representative images showing the results of IHC staining for integrin αvβ3 in tissue samples from healthy individuals (n=3) and prostate cancer patients (n=3). (G&H) Cells were co-transfected with integrin αv and β3 siRNA then with apelin, the wound healing and cell migration was examined (n=3). * p < 0.05 compared with the control group. # p < 0.05 compared with the apelin-treated group.

Article Snippet: JNK (SC-474), p38 (SC-4972), ERK (SC-1647), STAT3 (SC-482), p-JNK (SC-6254), p-p38 (SC-166182), p-ERK (SC-7383) and integrin αvβ3 (SC-7312) antibodies as well as JNK (sc-29380), p38 (sc-29433), STAT3 (sc-29493), integrin αv (sc-29373), integrin β3 (sc-29375) and apelin (sc-44741) siRNA were purchased from Santa Cruz Biotechnology (CA, USA).

Techniques: Expressing, Migration, Immunohistochemistry, Transfection, Control

Journal: International Journal of Biological Sciences

Article Title: Apelin facilitates integrin αvβ3 production and enhances metastasis in prostate cancer by activating STAT3 and inhibiting miR-8070

doi: 10.7150/ijbs.113161

Figure Lengend Snippet: Figure 3. MAPK pathway is regulated apelin-induced integrin expression and prostate cancer migration. (A&B) IPA pathway enrichment figure showing pathways that were changed in the GSE7930 dataset (Orange color indicated upregulated gene profile; blue color indicated downregulated gene profile). (C-H) Cells were treated with ERK (ERK II inhibitor; 10 μM), p38 (SB203580; 10 μM) and JNK (SP600125; 10 μM) inhibitors or transfected with ERK, p38 and JNK siRNA for 24 hours then with apelin, the wound healing, cell migration and integrin mRNA expression was examined (n=3). (I) PC3 cells were stimulated with apelin and ERK, p38 and JNK phosphorylation was examined by Western blotting (n=3). * p < 0.05 compared with the control group. # p < 0.05 compared with the apelin-treated group.

Article Snippet: JNK (SC-474), p38 (SC-4972), ERK (SC-1647), STAT3 (SC-482), p-JNK (SC-6254), p-p38 (SC-166182), p-ERK (SC-7383) and integrin αvβ3 (SC-7312) antibodies as well as JNK (sc-29380), p38 (sc-29433), STAT3 (sc-29493), integrin αv (sc-29373), integrin β3 (sc-29375) and apelin (sc-44741) siRNA were purchased from Santa Cruz Biotechnology (CA, USA).

Techniques: Expressing, Migration, Transfection, Phospho-proteomics, Western Blot, Control

Journal: International Journal of Biological Sciences

Article Title: Apelin facilitates integrin αvβ3 production and enhances metastasis in prostate cancer by activating STAT3 and inhibiting miR-8070

doi: 10.7150/ijbs.113161

Figure Lengend Snippet: Figure 4. Apelin promotes integrin production and cell motility via the STAT3 pathway. (A-F) Cells were treated with STAT3 inhibitor (10 μM) or transfected with STAT3 siRNA for 24 hours then with apelin, the wound healing, cell migration and integrin mRNA expression was examined (n=3). (G&H) PC3 cells were treated with apelin or pretreated with ERK, p38 and JNK inhibitor then with apelin, the STAT3 phosphorylation was examined by Western blotting (n=3). * p < 0.05 compared with the control group. # p < 0.05 compared with the apelin-treated group.

Article Snippet: JNK (SC-474), p38 (SC-4972), ERK (SC-1647), STAT3 (SC-482), p-JNK (SC-6254), p-p38 (SC-166182), p-ERK (SC-7383) and integrin αvβ3 (SC-7312) antibodies as well as JNK (sc-29380), p38 (sc-29433), STAT3 (sc-29493), integrin αv (sc-29373), integrin β3 (sc-29375) and apelin (sc-44741) siRNA were purchased from Santa Cruz Biotechnology (CA, USA).

Techniques: Transfection, Migration, Expressing, Phospho-proteomics, Western Blot, Control

Journal: International Journal of Biological Sciences

Article Title: Apelin facilitates integrin αvβ3 production and enhances metastasis in prostate cancer by activating STAT3 and inhibiting miR-8070

doi: 10.7150/ijbs.113161

Figure Lengend Snippet: Figure 5. Apelin enhances integrin expression and promotes cell migration by inhibiting miR-8070 expression. (A) The diagrams depict the selection of miRNA candidates targeting integrin αv and β3. (B) PC3 cells were treated with apelin, the miRNAs expression was examined by qPCR (n=3). (C) Cells were treated with apelin for 24 hours, the miR-8070 expression was examined by qPCR (n=3). (D-F) Cells were transfected with miR-8070 mimic for 24 hours then with apelin, the wound healing, cell migration and integrin mRNA expression was examined (n=3). (G-I) Cells were treated with ERK, p38 and JNK inhibitor or siRNA then with apelin, the 3’UTR activity and miRNA expression was examined by luciferase activity and qPCR (n=3). * p < 0.05 compared with the control group. # p < 0.05 compared with the apelin-treated group.

Article Snippet: JNK (SC-474), p38 (SC-4972), ERK (SC-1647), STAT3 (SC-482), p-JNK (SC-6254), p-p38 (SC-166182), p-ERK (SC-7383) and integrin αvβ3 (SC-7312) antibodies as well as JNK (sc-29380), p38 (sc-29433), STAT3 (sc-29493), integrin αv (sc-29373), integrin β3 (sc-29375) and apelin (sc-44741) siRNA were purchased from Santa Cruz Biotechnology (CA, USA).

Techniques: Expressing, Migration, Selection, Transfection, Activity Assay, Luciferase, Control

Journal: International Journal of Biological Sciences

Article Title: Apelin facilitates integrin αvβ3 production and enhances metastasis in prostate cancer by activating STAT3 and inhibiting miR-8070

doi: 10.7150/ijbs.113161

Figure Lengend Snippet: Figure 6. Apelin blockade inhibits prostate cancer metastasis in vivo. (A-C) Cells were transfected with apelin siRNA, the integrin expression, wound healing and cell migration was examined (n=3). (D&E) IHC analysis of prostate cancer tissue samples stained with integrin αvβ3 antibody (n=3). (F) IHC analysis of leg bone (n=3), liver (n=3), and lung (n=3) sections stained with apelin and integrin αvβ3 antibodies. * p < 0.05 compared with the control group.

Article Snippet: JNK (SC-474), p38 (SC-4972), ERK (SC-1647), STAT3 (SC-482), p-JNK (SC-6254), p-p38 (SC-166182), p-ERK (SC-7383) and integrin αvβ3 (SC-7312) antibodies as well as JNK (sc-29380), p38 (sc-29433), STAT3 (sc-29493), integrin αv (sc-29373), integrin β3 (sc-29375) and apelin (sc-44741) siRNA were purchased from Santa Cruz Biotechnology (CA, USA).

Techniques: In Vivo, Transfection, Expressing, Migration, Staining, Control

Journal: International Journal of Biological Sciences

Article Title: Apelin facilitates integrin αvβ3 production and enhances metastasis in prostate cancer by activating STAT3 and inhibiting miR-8070

doi: 10.7150/ijbs.113161

Figure Lengend Snippet: Figure 7. Schematic diagram illustrating the mechanism underlying the effects of apelin in prostate cancer metastasis. Apelin stimulation enhances integrin αvβ3-dependent prostate cancer migration and metastasis. The activation of STAT3 and inhibition of miR-8070 via the ERK, p38 and JNK pathways mediate apelin-facilitated integrin synthesis and cell motility.

Article Snippet: JNK (SC-474), p38 (SC-4972), ERK (SC-1647), STAT3 (SC-482), p-JNK (SC-6254), p-p38 (SC-166182), p-ERK (SC-7383) and integrin αvβ3 (SC-7312) antibodies as well as JNK (sc-29380), p38 (sc-29433), STAT3 (sc-29493), integrin αv (sc-29373), integrin β3 (sc-29375) and apelin (sc-44741) siRNA were purchased from Santa Cruz Biotechnology (CA, USA).

Techniques: Migration, Activation Assay, Inhibition

Journal: EJVES Vascular Forum

Article Title: Overexpression of αVβ3 Integrin in an Aortic Aneurysm Murine Model: Confirmation in Ex Vivo Human Abdominal Aortic Aneurysm Tissue and Its Potential Application for Molecular Imaging Characterisation ☆

doi: 10.1016/j.ejvsvf.2025.08.003

Figure Lengend Snippet: Immunofluorescence analysis with the primary αVβ3 integrin in Wistar rats aortic tissue demonstrating significant differences when comparing A. aneurysmal lesions with B. control tissue. PAX8 (A, small box) was used as a negative control, a transcription factor expressed during embryonic development of Müllerian organs, kidney, and thyroid (PAX8 mouse monoclonal antibody (MRQ-50), Cell Marque 363M, San Diego, CA, USA), which was simultaneously processed with αVβ3 integrin. C. Graphic demonstrating differences in mean values of immunofluorescence intensity (II) in the aneurysmal wall (II mean 20.1) compared with the control tissue (II mean 9.1) ( p < 0.001).

Article Snippet: Primary antibody αVβ3 integrin Alexa Fluor 546 (1:200, Cat. No. sc-7312 AF 546, Santa Cruz Biotechnology, Dallas, TX, USA) was applied to slides incubated in a humid chamber for one hour in the dark.

Techniques: Immunofluorescence, Control, Negative Control

Journal: Translational Neuroscience

Article Title: Thyroid hormone T3 induces Fyn modification and modulates palmitoyltransferase gene expression through αvβ3 integrin receptor in PC12 cells during hypoxia

doi: 10.1515/tnsci-2022-0347

Figure Lengend Snippet: JAK/STAT pathway’s protein phosphorylation in differentiated PC-12 cells in hypoxia. Densitometric analysis (arbitrary units of spot signal densities normalized to the positive control signals) showing the effect of 10 nM T3 and αvβ3-Ab integrin (µg/ml). Results represent the mean ± SEM of duplicate samples from two independent experiments. A significant level is defined as * p < 0.05 vs control and # p < 0.05 to T3.

Article Snippet: Differentiated cells were incubated with 10 nM T3 and αvβ3 blocking antibody (αvβ3-Ab, 1 μg/ml; sc-7312, Santa Cruz Biotechnology).

Techniques: Phospho-proteomics, Positive Control, Control

Journal: Translational Neuroscience

Article Title: Thyroid hormone T3 induces Fyn modification and modulates palmitoyltransferase gene expression through αvβ3 integrin receptor in PC12 cells during hypoxia

doi: 10.1515/tnsci-2022-0347

Figure Lengend Snippet: The action of T3 on the phosphorylation and palmitoylation of Fyn A. The ratio of palmitoylated and non-palmitoylated Fyn and p-Fyn forms. The palm-Fyn/Fyn ratio in differentiated PC-12 cells exposed to 10 nM T3 and µg/ml αvβ3-Ab integrin during 1 h of hypoxia. (a) Immunoblotting image of palm-Fyn and total Fyn; the palm-p-Fyn/p-Fyn ratio in differentiated PC-12 cells exposed to 10 nM T3 and αvβ3-Ab integrin (µg/ml) during 1 h of hypoxia. (b) Immunoblotting image of palm-p-Fyn and total p-Fyn; plot of the ratio of palm-p-Fyn to total p-Fyn. Plot of the ratio of palm-Fyn to total Fyn. Results represent the mean ± SEM of duplicate samples from two independent experiments. A significant level was defined as * p < 0.05 vs control and # p < 0.05 to T3.

Article Snippet: Differentiated cells were incubated with 10 nM T3 and αvβ3 blocking antibody (αvβ3-Ab, 1 μg/ml; sc-7312, Santa Cruz Biotechnology).

Techniques: Phospho-proteomics, Western Blot, Control

Journal: Translational Neuroscience

Article Title: Thyroid hormone T3 induces Fyn modification and modulates palmitoyltransferase gene expression through αvβ3 integrin receptor in PC12 cells during hypoxia

doi: 10.1515/tnsci-2022-0347

Figure Lengend Snippet: TH supplementation of hypoxic cells increases the proportion of phosphorylated to non-phosphorylated forms of palmitoylated Fyn kinase in differentiated PC-12 cells exposed to 10 nM T3 and αvβ3-Ab (µg/ml) integrin during 1 h hypoxia. Results represent the mean ± SEM of duplicate samples from two independent experiments. A significant level was defined as * p < 0.05. vs control and # p < 0.05. to T3.

Article Snippet: Differentiated cells were incubated with 10 nM T3 and αvβ3 blocking antibody (αvβ3-Ab, 1 μg/ml; sc-7312, Santa Cruz Biotechnology).

Techniques: Control

Journal: Translational Neuroscience

Article Title: Thyroid hormone T3 induces Fyn modification and modulates palmitoyltransferase gene expression through αvβ3 integrin receptor in PC12 cells during hypoxia

doi: 10.1515/tnsci-2022-0347

Figure Lengend Snippet: RT-qPCR analysis of genes ZDHHC2, ZDHHC3, ZDHHC8, ZDHHC9, and ZDHHC16 showing the effect of 10 nM T3 and αvβ3-Ab integrin (µg/ml) in differentiated PC-12 cells exposed to 1 h hypoxia

Article Snippet: Differentiated cells were incubated with 10 nM T3 and αvβ3 blocking antibody (αvβ3-Ab, 1 μg/ml; sc-7312, Santa Cruz Biotechnology).

Techniques: Control

Journal: Translational Neuroscience

Article Title: Thyroid hormone T3 induces Fyn modification and modulates palmitoyltransferase gene expression through αvβ3 integrin receptor in PC12 cells during hypoxia

doi: 10.1515/tnsci-2022-0347

Figure Lengend Snippet: The palmitoyltransferase gene expression levels of (a) ZDHHC2 and (b) ZDHHC9It were determined using the RT-PCR method in differentiated PC-12 cells exposed to 10 nM T3 and αvβ3-Ab integrin (µg/ml) during 1 h hypoxia. Results represent the mean ± SEM of duplicate samples from three independent experiments. A significant level was defined as * p < 0.05 vs control and # p < 0.05 to T3.

Article Snippet: Differentiated cells were incubated with 10 nM T3 and αvβ3 blocking antibody (αvβ3-Ab, 1 μg/ml; sc-7312, Santa Cruz Biotechnology).

Techniques: Gene Expression, Reverse Transcription Polymerase Chain Reaction, Control

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: P 4 regulates gene expression in PR-positive MCF-7 and PR-negative MDA-MB-231 breast cancer cells. (A, B) Serum-starved cells were left unstimulated (━) or stimulated with different concentrations of P 4 (10 −7 and 10 −5 M) for 24 h. The cells were lysed and the mRNAs extracted from cell lysates were subjected to the reverse transcription reaction. The mRNA expression of PCNA , CCND1 , MMP-9 , PD-L1 , integrin αv , integrin β3 , integrin β5 , and β-actin , as a loading control, was quantified by qRT-PCR. The mRNA expression of these genes was normalized to that of β-actin . The quantitative values were expressed as relative mRNA levels by defining the amounts of gene expression in unstimulated group as 1. Data are represented as the mean ± standard deviation of triplicate cultures in three independent experiments. *P < 0.05 , **P < 0.01 , ***P < 0.001 compared to the unstimulated group.

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Gene Expression, Reverse Transcription, Expressing, Control, Quantitative RT-PCR, Standard Deviation

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: Predicted docking poses of P 4 bound at the cRGD-binding site of integrin αvβ3. (A, B) Docking models 1 and models 2 of P 4 are respectively colored in blue and white, and the free binding energies are anticipated to be −6.7 -and −6.5 kcal/mol, respectively. (C) Superimpositions of binding models for modes 1 (blue) and 2 (white) mapped into cRGD peptide (purple) of αvβ3 integrin subunits. (D, E) Binding mode 1 and mode 2 of P 4 are illustrated within integrin αvβ3 and their corresponding 2D interaction plots by the BIOVIA Discovery Studio Visualizer ( http://accelrys.com ). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Binding Assay

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: Blocking of integrin αvβ3 activity down-regulates P 4 -induced integrin β3 expression in MDA-MB-231 cells. Serum-starved cells were left unpretreated or pretreated with anti-integrin αvβ3 antibody (2 μg/ml) or the RGD peptide (500 nM) for 1 h and then were left unstimulated or stimulated with10 −5 M P 4 for 72 h. The cells were then lysed and cell lysates were subjected to Western blotting for the detection of the indicated integrin β3, integrin β5, and GAPDH, as a loading control (These original blot images are provided in the Supplementary file). Similar results were obtained in three independent experiments. The quantitative results were expressed as fold increase by defining the amounts of the indicated detected proteins in untreated cells, where the absence of P 4 stimulation was considered as 1. Data are represented as the mean ± SD of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 compared to untreated cells, where the absence of P 4 stimulation; # P < 0.05, ## P < 0.01 compared to the P 4 -stimulated unpretreated cells.

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Blocking Assay, Activity Assay, Expressing, Western Blot, Control

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: Blocking of Integrin αvβ3 activity affects P 4 -induced gene expressions in MDA-MB-231 cells. (A–F) Serum-starved cells were left unpretreated or pretreated with anti-integrin αvβ3 antibody (2 μg/ml) or the RGD peptide (500 nM) for 1 h and then were left unstimulated or stimulated with10 −5 M P 4 for 72 h. The cells were lysed and the mRNAs extracted from cell lysates were subjected to the reverse transcription reaction. The mRNA expression of integrin αv , integrin β3 , integrin β5 , CCND1 , p21 , PCNA , and β-actin , as a loading control, was quantified by qRT-PCR. The mRNA expression of these genes was normalized to that of β-actin . The quantitative values were expressed as relative mRNA levels by defining the amounts of gene expression in untreated cells, where the absence of P 4 stimulation as 1. Data are represented as the mean ± standard deviation of triplicate cultures in three‐independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 compared to untreated cells, where the absence of P 4 stimulation; ## P < 0.01, ### P < 0.001 compared to the P 4 -stimulated unpretreated cells.

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Blocking Assay, Activity Assay, Reverse Transcription, Expressing, Control, Quantitative RT-PCR, Gene Expression, Standard Deviation

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: P 4 -induced ERK1/2 activation is integrin αvβ3-dependent in MDA-MB-231 breast cancer cells. (A, B) Cells were seeded on a cover glass and starved for 48 h. Different combinations of treatment were described in the Confocal microscopy section of Materials and Methods. Cells then were fixed for confocal microscopy. The cells were fixed, permeabilized and immunostained with antibodies against integrin αvβ3 (red color) and p -ERK1/2 (green color). The merge image shows colocalization (yellow color) of these two proteins. Nuclei were counterstained with DAPI (blue color). Accumulation of p -ERK1/2 in the nucleus was showed as white arrows. The right panel (B) shows a zoom-in image of the left panel (A) to present cells in a more focused manner. (C) Quantification of the number of p -ERK1/2 accumulation in the nucleus. Data are represented as the mean ± standard deviation of triplicate cultures in three‐independent experiments. *P < 0.05 , ** P < 0.0 1 , ***P < 0.001 compared to untreated cells, where the absence of P 4 stimulation; # P < 0.01 compared to the P 4 -stimulated unpretreated cells. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Activation Assay, Confocal Microscopy, Standard Deviation

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: P 4 regulates signal transduction protein profiles in MDA-MB-231 cells. Serum-starved cells were left unpretreated or pretreated with anti-integrin αvβ3 antibody (2 μg/ml) or the RGD peptide (500 nM) for 1 h and then were left unstimulated or stimulated with 10 −5 M P 4 for 72 h. The cells were then lysed and cell lysates were subjected to Western blotting for the detection of the indicated p -FAK (Y397), FAK, p -ERK1/2 (T202/Y204), ERK1/2, and GAPDH, as a loading control (These original blot images are provided in the Supplementary file). Similar results were obtained in three independent experiments. The quantitative results were expressed as fold increase by defining the amounts of the indicated detected proteins in untreated cells, where the absence of P 4 stimulation was considered as 1. Data are represented as the mean ± SD of three independent experiments. *P < 0.05, ***P < 0.001 compared to untreated cells, where the absence of P 4 stimulation; # P < 0.05 compared to the P 4 -stimulated unpretreated cells.

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Transduction, Western Blot, Control

Journal: Heliyon

Article Title: Progesterone modulates cell growth via integrin αvβ3-dependent pathway in progesterone receptor-negative MDA-MB-231 cells

doi: 10.1016/j.heliyon.2024.e34006

Figure Lengend Snippet: Blocking of RGD binding site affects P 4 -induced cell growth in MDA-MB-231 cells. (A, B) Serum-starved cells were left unpretreated or pretreated with anti-integrin αvβ3 antibody (2 μg/ml) or the RGD peptide (500 nM) for 1 h and then were left unstimulated or stimulated with 10 −5 M P 4 for 72 h. (A) The cells were then subjected to the cell viability assay. Data are represented normalized to the untreated cells, where the absence of P 4 stimulation and presented as the mean ± standard deviation of triplicate cultures in three‐independent experiments. ***P < 0.001 compared to untreated cells, where the absence of P 4 stimulation; ### P < 0.001, compared to the P 4 -stimulated unpretreated cells. (B) The cells were then subjected to the flow cytometric analysis. The bar graphs represent the percentage of cell populations in each cell cycle phase, as measured by DNA content stained with PI. Similar results were obtained in three‐independent experiments.

Article Snippet: For integrin αvβ3 interfering studies, MDA-MB-231 cells were treated with 10 −5 M P 4 in the presence or absence of anti-integrin αvβ3 antibody (2 μg/ml, Cat. No.: sc-7312, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or the RGD peptide (500 nM, Cat. No.: HY-P0023, MedChemExpress, Monmouth Junction, NJ, USA) for 72 h. Cell viability was evaluated using the Cell Counting Kit-8 (Cat. No.: 96992, Sigma-Aldrich) according to the manufacturer's instructions.

Techniques: Blocking Assay, Binding Assay, Viability Assay, Standard Deviation, Staining