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R&D Systems goat polyclonal anti cxcl12
Goat Polyclonal Anti Cxcl12, supplied by R&D Systems, used in various techniques. Bioz Stars score: 94/100, based on 34 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 94 stars, based on 34 article reviews

goat polyclonal anti cxcl12 - by Bioz Stars, 2026-03

94/100 stars

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Serial sections of normal and diseased pancreatic tissue were stained for CXCL12, CXCR4, CXCR7, and CK19. CXCL12 staining apparent in normal exocrine ducts was diminished in PDAC tissue. CXCR4 staining increased in PanIN and PDAC relative to normal ductal epithelium. CXCR7 expression was variable in normal epithelium, PanIN lesions, and PDAC. (A)Normal tissue from a single patient with healthy pancreas represents observations from 25 different normal tissues from 21 individual patients. (B)PDAC tissue from one patient represents observations from 82 different tissues from 29 different patients. 1000× magnification represents inset box at 200×. (C) Staining was quantified by blinded scoring of serial sections in relation to CK19 staining. (***) denotes P ≤0.001.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: Serial sections of normal and diseased pancreatic tissue were stained for CXCL12, CXCR4, CXCR7, and CK19. CXCL12 staining apparent in normal exocrine ducts was diminished in PDAC tissue. CXCR4 staining increased in PanIN and PDAC relative to normal ductal epithelium. CXCR7 expression was variable in normal epithelium, PanIN lesions, and PDAC. (A)Normal tissue from a single patient with healthy pancreas represents observations from 25 different normal tissues from 21 individual patients. (B)PDAC tissue from one patient represents observations from 82 different tissues from 29 different patients. 1000× magnification represents inset box at 200×. (C) Staining was quantified by blinded scoring of serial sections in relation to CK19 staining. (***) denotes P ≤0.001.

Article Snippet: Secreted CXCL12 protein from supernatant of pancreatic cancer cells cultured in serum-free media was detected by ourpreviously established sandwich ELISA method using antibodies from R&D Systems (monoclonal mouse and human CXCL12 (MAB350) and goat anti-human CXCL12 (BAF310) .Cell surface CXCR4 or CXCR7was detected using the aforementioned antibodies (Abcam) along withFITC-conjugated secondary antibodies using our previously established method .

Techniques: Staining, Expressing

Representative 200× images of the same (A) normal or (B) pancreatic ductal adenocarcinoma (PDAC) tissues shown at 1000× magnification in . Representative serial section images of a pancreatic intestinal neoplasm (PanIN) lesion at (C) 200× or (D) 1000× magnification. Serial tissue sections were immunostained with antibodies to CK19, CXCL12, CXCR4, and CXCR7, or the isotype controls. n = 25 different normal tissues from 21 individual patients, 82 different tissues from 29 different PDAC patients, or 19 pathologically confirmed PanIN lesions.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: Representative 200× images of the same (A) normal or (B) pancreatic ductal adenocarcinoma (PDAC) tissues shown at 1000× magnification in . Representative serial section images of a pancreatic intestinal neoplasm (PanIN) lesion at (C) 200× or (D) 1000× magnification. Serial tissue sections were immunostained with antibodies to CK19, CXCL12, CXCR4, and CXCR7, or the isotype controls. n = 25 different normal tissues from 21 individual patients, 82 different tissues from 29 different PDAC patients, or 19 pathologically confirmed PanIN lesions.

Techniques:

RT-PCR analysis revealed that (A) patient-derived pancreatic cancer cell lines (#1, #2, #3, #4) or (B) established cell lines lacked expression of CXCL12 and maintained expression of CXCR4. CXCR7 mRNA was present in 3 of 8 PDAC lines.Flow cytometric detection(C)of surface CXCR4 or CXCR7 protein expression. Cell lines treatedseven days with a concentration curve of (D) 5-aza-2-deoxycytidine (5-aza) restored expression of CXCL12.Linestreated 4 days with a concentration curve of (E) Trichostatin-A (TSA)restored CXCL12 mRNA expression in Capan2 cells.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: RT-PCR analysis revealed that (A) patient-derived pancreatic cancer cell lines (#1, #2, #3, #4) or (B) established cell lines lacked expression of CXCL12 and maintained expression of CXCR4. CXCR7 mRNA was present in 3 of 8 PDAC lines.Flow cytometric detection(C)of surface CXCR4 or CXCR7 protein expression. Cell lines treatedseven days with a concentration curve of (D) 5-aza-2-deoxycytidine (5-aza) restored expression of CXCL12.Linestreated 4 days with a concentration curve of (E) Trichostatin-A (TSA)restored CXCL12 mRNA expression in Capan2 cells.

Techniques: Reverse Transcription Polymerase Chain Reaction, Derivative Assay, Expressing, Concentration Assay

(A) Panc1 and MiaPaCa2 cells migrated towards exogenous gradients of CXCL12 in a range from 1 nM to 1000 nM under serum-free conditions(*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001, respectively, in comparison to unstimulated cells (NS). Receptor-null HPAFII cells did not migrate in response to CXCL12. (B) CXCL12 directs Panc1 cell chemoinvasion into a three-dimensional Matrigel plug. The positive control (+) was 10% serum-containing medium. (*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001, respectively, in comparison to unstimulated cells (NS).

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: (A) Panc1 and MiaPaCa2 cells migrated towards exogenous gradients of CXCL12 in a range from 1 nM to 1000 nM under serum-free conditions(*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001, respectively, in comparison to unstimulated cells (NS). Receptor-null HPAFII cells did not migrate in response to CXCL12. (B) CXCL12 directs Panc1 cell chemoinvasion into a three-dimensional Matrigel plug. The positive control (+) was 10% serum-containing medium. (*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001, respectively, in comparison to unstimulated cells (NS).

Techniques: Positive Control

Luciferase levels in control GFP or three different (31, #2, #3) CXCL12-expressing MiaPaCa2 clones as measured by spectrophotometer (A) or IVIS-100 biophotonic imager (B). Levels of CXCL12 were measured by ELISA (C) in established PDAC cell lines as well as GFP- and CXCL12-expressing MiaPaCa2-luciferase clones. (D) CXCL12-secreted by transfected MiaPaCa2-luciferase clones #1 and #2 stimulated U937 chemotaxis. Cells treated with neutralizing antibody to CXCL12 (αL12) confirmed the specificity of U937 chemotaxis. Values in A, C, and D are mean±SEM, n = 2–3.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: Luciferase levels in control GFP or three different (31, #2, #3) CXCL12-expressing MiaPaCa2 clones as measured by spectrophotometer (A) or IVIS-100 biophotonic imager (B). Levels of CXCL12 were measured by ELISA (C) in established PDAC cell lines as well as GFP- and CXCL12-expressing MiaPaCa2-luciferase clones. (D) CXCL12-secreted by transfected MiaPaCa2-luciferase clones #1 and #2 stimulated U937 chemotaxis. Cells treated with neutralizing antibody to CXCL12 (αL12) confirmed the specificity of U937 chemotaxis. Values in A, C, and D are mean±SEM, n = 2–3.

Techniques: Luciferase, Expressing, Clone Assay, Spectrophotometry, Enzyme-linked Immunosorbent Assay, Transfection, Chemotaxis Assay

(A) Transwell migration assays revealed significantly reduced chemotaxis of CXCL12-expressing clones (#1 and #2) compared to ligand null (GFP) cells. Attractants were serum-free media (--), 10% serum (+), or 10 nM CXCL12 (L12) in serum-free media. denote P ≤0.01 and P ≤0.001, respectively, compared to 10 nM CXCL12-stimulated GFP cells, n = 5. (B) CXCL12 re-expression diminished TGF-β (5 ng/mL)-induced chemotaxis relative to the CXCL12-null cells. (##) denotes P ≤0.01 compared to TGF-β-stimulated GFP cells, n = 4. (C) & (D) Representative images of experiments in (A) and (B) respectively. (E) CXCL12-expressing cells were significantly more adherent to tissue culture plastic compared to CXCL12-null cells. Untreated cells = (--), neutralizing antibody for CXCL12 activity = (αL12), and a positive control = 1 ng/mL of EGF (+). (##) denotes P ≤0.01 compared to 10% serum-stimulated control cells. (*), (**), and (***) denote P ≤0.05, P ≤0.01 and P ≤0.001, respectively, compared to unstimulated GFP cells, n = 7.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: (A) Transwell migration assays revealed significantly reduced chemotaxis of CXCL12-expressing clones (#1 and #2) compared to ligand null (GFP) cells. Attractants were serum-free media (--), 10% serum (+), or 10 nM CXCL12 (L12) in serum-free media. denote P ≤0.01 and P ≤0.001, respectively, compared to 10 nM CXCL12-stimulated GFP cells, n = 5. (B) CXCL12 re-expression diminished TGF-β (5 ng/mL)-induced chemotaxis relative to the CXCL12-null cells. (##) denotes P ≤0.01 compared to TGF-β-stimulated GFP cells, n = 4. (C) & (D) Representative images of experiments in (A) and (B) respectively. (E) CXCL12-expressing cells were significantly more adherent to tissue culture plastic compared to CXCL12-null cells. Untreated cells = (--), neutralizing antibody for CXCL12 activity = (αL12), and a positive control = 1 ng/mL of EGF (+). (##) denotes P ≤0.01 compared to 10% serum-stimulated control cells. (*), (**), and (***) denote P ≤0.05, P ≤0.01 and P ≤0.001, respectively, compared to unstimulated GFP cells, n = 7.

Techniques: Migration, Chemotaxis Assay, Expressing, Clone Assay, Activity Assay, Positive Control

(A) Representative bioluminescence images of mice xenografted with GFP- or CXCL12-expressing cells at implantation (Day 0) or study endpoint (Day 28). (B) Whole-body in vivo radiance over time of GFP- and CXCL12-mice. (C) Ex vivo radiance of excised spleen (C), reflecting tumor cells at the site of injection, or the metastatic destination (D), reflecting decreased hepatic metastasis of CXCL12-expressing cells relative to GFP-cells. (**) denotes P ≤0.01, n = 4–5. (E) Representative H&E images showing pronounced tumor mass in the liver of control (GFP), relative to experimental (CXCL12) xenografted mice.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: (A) Representative bioluminescence images of mice xenografted with GFP- or CXCL12-expressing cells at implantation (Day 0) or study endpoint (Day 28). (B) Whole-body in vivo radiance over time of GFP- and CXCL12-mice. (C) Ex vivo radiance of excised spleen (C), reflecting tumor cells at the site of injection, or the metastatic destination (D), reflecting decreased hepatic metastasis of CXCL12-expressing cells relative to GFP-cells. (**) denotes P ≤0.01, n = 4–5. (E) Representative H&E images showing pronounced tumor mass in the liver of control (GFP), relative to experimental (CXCL12) xenografted mice.

Techniques: Expressing, In Vivo, Ex Vivo, Injection

Apoptosis of GFP and CXCL12-expressing MiaPaCa2 cells were assessed using the caspase 3/7 glo assay.Cells were starved for 24 hours and cultured in 0% serum (A, C) or 1% serum (B, D) containing medium. (A–B) Apoptosis in adherent CXCL12-expressing cells was elevated compared to either GFP-expressing clones in serum-free conditions with no change seen in 1% serum. GFP-cells werestimulated with 100 µM gemcitabine as a control. (C–D) To measure cell number and detachment based apoptosis of cells in suspension, MiaPaCa2-luciferase cells were cultured on poly-HEMA. Using the Viacount reagent and flow cytometric cell counting, there was no difference in live cell number observed in non-adherent CXCL12-null (GFP) or expressing cells when cultured either in 0% (C) or 1% serum (D). Apoptosis of poly-HEMA cultured cells revealed an in increase in active caspase-3/7 restricted to cells cultured in serum-free conditions only (C) with no change observed in 1% serum (D). Gemcitabine (GEM) was used as a control for decreased cell count and increased apoptosis.(*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001 respectively in comparison to control cells (GFP). Values are mean±SEM, n = 4–5.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: Apoptosis of GFP and CXCL12-expressing MiaPaCa2 cells were assessed using the caspase 3/7 glo assay.Cells were starved for 24 hours and cultured in 0% serum (A, C) or 1% serum (B, D) containing medium. (A–B) Apoptosis in adherent CXCL12-expressing cells was elevated compared to either GFP-expressing clones in serum-free conditions with no change seen in 1% serum. GFP-cells werestimulated with 100 µM gemcitabine as a control. (C–D) To measure cell number and detachment based apoptosis of cells in suspension, MiaPaCa2-luciferase cells were cultured on poly-HEMA. Using the Viacount reagent and flow cytometric cell counting, there was no difference in live cell number observed in non-adherent CXCL12-null (GFP) or expressing cells when cultured either in 0% (C) or 1% serum (D). Apoptosis of poly-HEMA cultured cells revealed an in increase in active caspase-3/7 restricted to cells cultured in serum-free conditions only (C) with no change observed in 1% serum (D). Gemcitabine (GEM) was used as a control for decreased cell count and increased apoptosis.(*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001 respectively in comparison to control cells (GFP). Values are mean±SEM, n = 4–5.

Techniques: Expressing, Glo Assay, Cell Culture, Clone Assay, Luciferase, Cell Counting

Population growth of adherent GFP and CXCL12-expressing MiaPaCa2 cells was assessed using the Viacount reagent and flow cytometric cell counting (A–B). CXCL12-expressing cells starved for 24 hours and cultured in both 0% serum (A) or 1% serum (B) containing medium were found to have decreased population growth.(B) Two CXCL12-expressing clones (#1, #2) were compared to GFP alone or GFP + gemcitabine (GEM) controls in 1% serum containing medium. Doubling time of adherent clones (T 2 ) was calculated using a linear regression of the data to determine slope and the intercept at y = 10 6 , with increased T 2 observed in both CXCL12-expressing clones. (C–D) Propidium Iodide cell cycle analysis revealed a decrease in percentage of cells in the G 2 phase in CXCL12-expressing cells compared to GFP controls. (*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001 respectively in comparison to control cells (GFP). Values are mean±SEM, n = 4–5.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: Population growth of adherent GFP and CXCL12-expressing MiaPaCa2 cells was assessed using the Viacount reagent and flow cytometric cell counting (A–B). CXCL12-expressing cells starved for 24 hours and cultured in both 0% serum (A) or 1% serum (B) containing medium were found to have decreased population growth.(B) Two CXCL12-expressing clones (#1, #2) were compared to GFP alone or GFP + gemcitabine (GEM) controls in 1% serum containing medium. Doubling time of adherent clones (T 2 ) was calculated using a linear regression of the data to determine slope and the intercept at y = 10 6 , with increased T 2 observed in both CXCL12-expressing clones. (C–D) Propidium Iodide cell cycle analysis revealed a decrease in percentage of cells in the G 2 phase in CXCL12-expressing cells compared to GFP controls. (*), (**), and (***) denote P ≤0.05, P ≤0.01, and P ≤0.001 respectively in comparison to control cells (GFP). Values are mean±SEM, n = 4–5.

Techniques: Expressing, Cell Counting, Cell Culture, Clone Assay, Cell Cycle Assay

(A) Kaplan-Meier survival curves for GFP- and CXCL12-expressing cell groups. Three experimental CXCL12 PDAC injected mice were removed for non-study reasons (tick marks). (B) Percent change in bioluminescence from baseline-level measured at day 7 for both GFP and CXCL12 engrafted mice. Dotted lines represent individual mice. Solid lines are quadratic regression fitted curves of each group. Statistical comparison was done between both groups independent of time. (C–D) Representative bioluminescence images of mice from each groupat days 7, 49, and endpoint for GFP (98) or CXCL12 (106). (E) Tumor wet weight was significantly reduced in CXCL12-expressing tumors relative GFP-tumors. Representative photomicrographs are shown in lower panels. (F) CXCL12-producing tumors had significantly fewer Ki-67 positive cells compared to GFP-expressing tumors, as counted in a cross-section of each tumor normalized to the total cross-sectional area of each tumor. (G) Representative images of Ki-67immunostaining and rabbit isotype control (inset, Rab IgG) are shown.n = 8–10. (**) and (***) denote P ≤0.01 and P ≤0.001 respectively, between CXCL12-expressing and control xenografted mice.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: (A) Kaplan-Meier survival curves for GFP- and CXCL12-expressing cell groups. Three experimental CXCL12 PDAC injected mice were removed for non-study reasons (tick marks). (B) Percent change in bioluminescence from baseline-level measured at day 7 for both GFP and CXCL12 engrafted mice. Dotted lines represent individual mice. Solid lines are quadratic regression fitted curves of each group. Statistical comparison was done between both groups independent of time. (C–D) Representative bioluminescence images of mice from each groupat days 7, 49, and endpoint for GFP (98) or CXCL12 (106). (E) Tumor wet weight was significantly reduced in CXCL12-expressing tumors relative GFP-tumors. Representative photomicrographs are shown in lower panels. (F) CXCL12-producing tumors had significantly fewer Ki-67 positive cells compared to GFP-expressing tumors, as counted in a cross-section of each tumor normalized to the total cross-sectional area of each tumor. (G) Representative images of Ki-67immunostaining and rabbit isotype control (inset, Rab IgG) are shown.n = 8–10. (**) and (***) denote P ≤0.01 and P ≤0.001 respectively, between CXCL12-expressing and control xenografted mice.

Techniques: Expressing, Injection

Ex vivo bioluminescence analysis revealed significantly decreased metastasis to the liver (A), lung (C), and mesenteric lymph nodes (D) of CXCL12-expressing cells compared to GFP-controls. Representative biophotonic images of hepatic metastases are shown in panel B. (**) and (***) denote statistically significant P ≤0.01 and P ≤0.001, respectively, differences between CXCL12-expressing and control tumor engrafted mice. n = 8–10 mice in each group.

Journal: PLoS ONE

Article Title: CXCL12 Chemokine Expression Suppresses Human Pancreatic Cancer Growth and Metastasis

doi: 10.1371/journal.pone.0090400

Figure Lengend Snippet: Ex vivo bioluminescence analysis revealed significantly decreased metastasis to the liver (A), lung (C), and mesenteric lymph nodes (D) of CXCL12-expressing cells compared to GFP-controls. Representative biophotonic images of hepatic metastases are shown in panel B. (**) and (***) denote statistically significant P ≤0.01 and P ≤0.001, respectively, differences between CXCL12-expressing and control tumor engrafted mice. n = 8–10 mice in each group.

Techniques: Ex Vivo, Expressing

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: Sequences of primers used in this study

Article Snippet: A goat polyclonal antibody against human CXCL12 (R&D Systems AF-310-NA), rabbit polyclonal antibody against CXCR4 (Abcam Inc. ab2074), mouse monoclonal antibody against human CXCR7/RDC1 (R&D Systems clone 11G8; MAB42273), a rabbit polyclonal antibody against COUP-TFI (Abcam Inc. ab11954) and a rabbit polyclonal antibody against HA epitope (Santa Cruz sc-805) were used for the immunofluorescence and western blot assays.

Techniques:

COUP-TFI modifies the expression of the CXCL12 signaling axis in MCF-7 cells. (A) Characterization of the control and COUP clones. An immunofluorescence cytochemistry assay was used to detect the relative expression of HA/COUP-TFI or COUP-TFI proteins in the control (Cont.) and COUP clones. The cells were fixed and processed for immunofluorescence as described in Methods; the nuclei were stained with DAPI. (B) CXCL12 , CXCR4 , and CXCR7 mRNAs were quantified by a real-time PCR analysis from two independent MCF-7 control and COUP clones. The results were normalized to GAPDH mRNA used as an internal control. The results were expressed as the relative mRNA expression level of CXCL12 , CXCR4 , or CXCR7 . Data are the mean values ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the control and COUP clones. (C) The amount of intracellular HA/COUP-TFI, COUP-TFI, CXCL12, CXCR4, and CXCR7 protein was determined from whole-cell extracts of the different MCF-7 clones and compared to total ERK. A representative western blot is shown. (D) The control and COUP clones were fixed, and an immunofluorescence cytochemistry assay was used to detect the relative expression of CXCL12, CXCR4, and CXCR7 proteins. Staining with DAPI is also presented to visualize the nucleus of the cells.

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: COUP-TFI modifies the expression of the CXCL12 signaling axis in MCF-7 cells. (A) Characterization of the control and COUP clones. An immunofluorescence cytochemistry assay was used to detect the relative expression of HA/COUP-TFI or COUP-TFI proteins in the control (Cont.) and COUP clones. The cells were fixed and processed for immunofluorescence as described in Methods; the nuclei were stained with DAPI. (B) CXCL12 , CXCR4 , and CXCR7 mRNAs were quantified by a real-time PCR analysis from two independent MCF-7 control and COUP clones. The results were normalized to GAPDH mRNA used as an internal control. The results were expressed as the relative mRNA expression level of CXCL12 , CXCR4 , or CXCR7 . Data are the mean values ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the control and COUP clones. (C) The amount of intracellular HA/COUP-TFI, COUP-TFI, CXCL12, CXCR4, and CXCR7 protein was determined from whole-cell extracts of the different MCF-7 clones and compared to total ERK. A representative western blot is shown. (D) The control and COUP clones were fixed, and an immunofluorescence cytochemistry assay was used to detect the relative expression of CXCL12, CXCR4, and CXCR7 proteins. Staining with DAPI is also presented to visualize the nucleus of the cells.

Techniques: Expressing, Control, Clone Assay, Immunofluorescence, Staining, Real-time Polymerase Chain Reaction, Western Blot

COUP-TFI modulates the chromatin structure of the CXCL12 and CXCR4 gene promoters. The FAIRE assay was performed as described in Methods. Real-time PCR was performed to monitor the enrichment of DNA corresponding to the proximal promoter of the CXCL12 (A) , the CXCR4 (B) and the CXCR7 (C) genes relative to the input chromatin from the control (Cont.) and COUP clones. The data are from triplicate samples and are representative of three separate experiments. The asterisk indicates significant differences ( p < 0.05) between the control and COUP clones.

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: COUP-TFI modulates the chromatin structure of the CXCL12 and CXCR4 gene promoters. The FAIRE assay was performed as described in Methods. Real-time PCR was performed to monitor the enrichment of DNA corresponding to the proximal promoter of the CXCL12 (A) , the CXCR4 (B) and the CXCR7 (C) genes relative to the input chromatin from the control (Cont.) and COUP clones. The data are from triplicate samples and are representative of three separate experiments. The asterisk indicates significant differences ( p < 0.05) between the control and COUP clones.

Techniques: Real-time Polymerase Chain Reaction, Control, Clone Assay

Estrogenic regulation of CXCL12 and CXCR4 in control and COUP clones. Control (Cont.) and COUP clones were treated with ethanol (EtOH) as the vehicle or E2 10 −8 M and ICI 10 −6 M alone or both together for 48 h. The CXCL12 (A) and CXCR4 (B) relative mRNA levels were monitored by a real-time PCR analysis, normalized to GAPDH mRNA as the internal control, and were expressed as the relative mRNA expression of CXCL12 or CXCR4 . Data are the mean ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the untreated and treated control clones. The pound sign indicates significant differences ( p < 0.05) between the untreated and treated COUP clones.

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: Estrogenic regulation of CXCL12 and CXCR4 in control and COUP clones. Control (Cont.) and COUP clones were treated with ethanol (EtOH) as the vehicle or E2 10 −8 M and ICI 10 −6 M alone or both together for 48 h. The CXCL12 (A) and CXCR4 (B) relative mRNA levels were monitored by a real-time PCR analysis, normalized to GAPDH mRNA as the internal control, and were expressed as the relative mRNA expression of CXCL12 or CXCR4 . Data are the mean ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the untreated and treated control clones. The pound sign indicates significant differences ( p < 0.05) between the untreated and treated COUP clones.

Techniques: Control, Clone Assay, Real-time Polymerase Chain Reaction, Expressing

The effect of COUP-TFI on the CXCL12/CXCR4 axis is mediated by EGF/EGFR activation. (A) The relative expression of EGF and EGFR mRNA was monitored by a real-time PCR analysis using MCF-7 control (Cont.) and COUP clones. The results were normalized against GAPDH as the internal control and are expressed as the mean EGF or EGFR mRNA/GAPDH mRNA ratio ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the control and COUP clones. (B) ERK activation was examined in the MCF-7 control (Cont.) and COUP clones after a 5- or 10-min stimulation with EGF (10 −9 M) or CXCL12 (200 ng/mL). Western blots were performed using antibodies against phospho-ERK (P-ERK) and total ERK (ERK1/2); a representative western blot is presented. The importance of EGFR-specific signaling and general ERK signaling on CXCL12 (C) and CXCR4 (D) regulation was assayed by treating the cells with EGF (10 −9 M), AG1478 (25 μM), or U0126 (25 μM) for 48 h. The CXCL12 and CXCR4 relative mRNA levels were monitored by the real-time PCR analysis, normalized to GAPDH mRNA as the internal control, and were expressed as the relative mRNA expression of CXCL12 or CXCR4 . Data are the mean ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the untreated and treated control clones. The pound sign indicates significant differences ( p < 0.05) between the untreated and treated COUP clones.

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: The effect of COUP-TFI on the CXCL12/CXCR4 axis is mediated by EGF/EGFR activation. (A) The relative expression of EGF and EGFR mRNA was monitored by a real-time PCR analysis using MCF-7 control (Cont.) and COUP clones. The results were normalized against GAPDH as the internal control and are expressed as the mean EGF or EGFR mRNA/GAPDH mRNA ratio ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the control and COUP clones. (B) ERK activation was examined in the MCF-7 control (Cont.) and COUP clones after a 5- or 10-min stimulation with EGF (10 −9 M) or CXCL12 (200 ng/mL). Western blots were performed using antibodies against phospho-ERK (P-ERK) and total ERK (ERK1/2); a representative western blot is presented. The importance of EGFR-specific signaling and general ERK signaling on CXCL12 (C) and CXCR4 (D) regulation was assayed by treating the cells with EGF (10 −9 M), AG1478 (25 μM), or U0126 (25 μM) for 48 h. The CXCL12 and CXCR4 relative mRNA levels were monitored by the real-time PCR analysis, normalized to GAPDH mRNA as the internal control, and were expressed as the relative mRNA expression of CXCL12 or CXCR4 . Data are the mean ± SEM of at least three independent experiments. The asterisks indicate significant differences ( p < 0.05) between the untreated and treated control clones. The pound sign indicates significant differences ( p < 0.05) between the untreated and treated COUP clones.

Techniques: Activation Assay, Expressing, Real-time Polymerase Chain Reaction, Control, Clone Assay, Western Blot

COUP-TFI overexpression influences cellular responses to CXCL12. (A) The relative growth of the control (Cont.) and COUP clones was assayed with or without CXCL12 treatment for 7 days. The basal and CXCL12-induced cell growth were evaluated by MTT assays (n = 6) and determined in three independent experiments. The results are expressed as the relative cell number obtained when the control cells were treated with the vehicle control. Significant differences between the unstimulated control cells and the other conditions ( p < 0.05) are indicated with an asterisk. Significant differences between stimulated control cells and stimulated COUP clones ( p < 0.05 ) are indicated with a pound sign. (B) The migratory capacity of control (Cont.) and COUP clones was analyzed. The cells were maintained in phenol red-free DMEM/2.5% dsFBS for 48 h and then seeded in phenol red-free DMEM/0.5% dsFBS in the upper chamber of a PET 8-μm pore insert. The cells were allowed to migrate for 24 h toward the phenol red-free DMEM/2.5% dsFBS medium complemented or not with CXCL12 (200 ng/mL) and AMD3100 (50 μM). (C) CXCL12 was also added to the culture medium in the upper chamber prior to migration. The results are expressed as the mean ± SEM of the relative number of migratory cells compared to the basal migration of the control cells measured in three independent experiments. The asterisks indicate significant differences ( p < 0.05) from the basal migration of the control clones. The pound sign indicates significant differences ( p < 0.05) between two conditions linked by black lines.

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: COUP-TFI overexpression influences cellular responses to CXCL12. (A) The relative growth of the control (Cont.) and COUP clones was assayed with or without CXCL12 treatment for 7 days. The basal and CXCL12-induced cell growth were evaluated by MTT assays (n = 6) and determined in three independent experiments. The results are expressed as the relative cell number obtained when the control cells were treated with the vehicle control. Significant differences between the unstimulated control cells and the other conditions ( p < 0.05) are indicated with an asterisk. Significant differences between stimulated control cells and stimulated COUP clones ( p < 0.05 ) are indicated with a pound sign. (B) The migratory capacity of control (Cont.) and COUP clones was analyzed. The cells were maintained in phenol red-free DMEM/2.5% dsFBS for 48 h and then seeded in phenol red-free DMEM/0.5% dsFBS in the upper chamber of a PET 8-μm pore insert. The cells were allowed to migrate for 24 h toward the phenol red-free DMEM/2.5% dsFBS medium complemented or not with CXCL12 (200 ng/mL) and AMD3100 (50 μM). (C) CXCL12 was also added to the culture medium in the upper chamber prior to migration. The results are expressed as the mean ± SEM of the relative number of migratory cells compared to the basal migration of the control cells measured in three independent experiments. The asterisks indicate significant differences ( p < 0.05) from the basal migration of the control clones. The pound sign indicates significant differences ( p < 0.05) between two conditions linked by black lines.

Techniques: Over Expression, Control, Clone Assay, Migration

Box plots of CXCR4, CXCR7, CXCL12, and COUP-TFI mRNA expression in breast cancer and normal tissue. CXCR4 (A) , CXCR7 (B) , CXCL12 (C) , and COUP-TFI (D) mRNA expression was measured by real-time PCR in 23 normal breast tissue samples (NT), in 20 SBR grades 1 and 2, and in 19 SBR grades 3. The expression level was normalized by 18S RNA expression and analyzed with IQ5 software (Bio-Rad). The data are presented as whisker plots in which the horizontal bar represents the median, the grey boxes are the 25 th and 75 th percentiles, the vertical bar is the standard deviation, and the plus signs are the extreme points. All the Mann-Whitney tests were performed with Minitab 16 software, and the p value is indicated on the different graphs (ns denotes non-significant).

Journal: BMC Cancer

Article Title: COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration

doi: 10.1186/1471-2407-14-407

Figure Lengend Snippet: Box plots of CXCR4, CXCR7, CXCL12, and COUP-TFI mRNA expression in breast cancer and normal tissue. CXCR4 (A) , CXCR7 (B) , CXCL12 (C) , and COUP-TFI (D) mRNA expression was measured by real-time PCR in 23 normal breast tissue samples (NT), in 20 SBR grades 1 and 2, and in 19 SBR grades 3. The expression level was normalized by 18S RNA expression and analyzed with IQ5 software (Bio-Rad). The data are presented as whisker plots in which the horizontal bar represents the median, the grey boxes are the 25 th and 75 th percentiles, the vertical bar is the standard deviation, and the plus signs are the extreme points. All the Mann-Whitney tests were performed with Minitab 16 software, and the p value is indicated on the different graphs (ns denotes non-significant).

Techniques: Expressing, Real-time Polymerase Chain Reaction, RNA Expression, Software, Whisker Assay, Standard Deviation, MANN-WHITNEY

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