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SDF-1alpha concentration dependent modulation of RhoA and Rac1 modifies breast cancer and stromal cells interaction.

Pasquier J, Abu-Kaoud N, Abdesselem H, Madani A, Hoarau-Véchot J, Thawadi HA, Vidal F, Couderc B, Favre G, Rafii A - BMC Cancer (2015)

Bottom Line: This interaction increases migration from primary sites as well as homing at distant sites.Increased migration was obtained at 50 and 100 ng/ml of SDF-1α; however migration was reduced at 200 ng/ml.The adhesion between breast cancer cells and BMHC was significantly increased by SDF-1α treatment at 200 ng/ml and reduced using a blocking monoclonal antibody against CXCR4.

View Article: PubMed Central - PubMed

Affiliation: Stem Cell and Microenvironment Laboratory, Weill Cornell Medical College in Qatar, Education City, Qatar Foundation, Doha, Qatar. jep2026@qatar-med.cornell.edu.

ABSTRACT

Background: The interaction of SDF-1alpha with its receptor CXCR4 plays a role in the occurrence of distant metastasis in many solid tumors. This interaction increases migration from primary sites as well as homing at distant sites.

Methods: Here we investigated how SDF-1α could modulate both migration and adhesion of cancer cells through the modulation of RhoGTPases.

Results: We show that different concentrations of SDF-1α modulate the balance of adhesion and migration in cancer cells. Increased migration was obtained at 50 and 100 ng/ml of SDF-1α; however migration was reduced at 200 ng/ml. The adhesion between breast cancer cells and BMHC was significantly increased by SDF-1α treatment at 200 ng/ml and reduced using a blocking monoclonal antibody against CXCR4. We showed that at low SDF-1α concentration, RhoA was activated and overexpressed, while at high concentration Rac1 was promoting SDF-1α mediating-cell adhesion.

Conclusion: We conclude that SDF-1α concentration modulates migration and adhesion of breast cancer cells, by controlling expression and activation of RhoGTPases.

No MeSH data available.


Related in: MedlinePlus

SDF-1alpha regulates interaction between MDA-MB231 and BMHC. a Flow cytometry cell sorting chart. MDA-MB231 (green) and BMHC (purple) were gated through GFP fluorescence intensity. b Flow cytometry analysis of CXCR4 expression. After 5 days of co-culture with BMHC, MDA-MB231 were cell sorted and stained for CXCR4. MDA-MB231 display an increase of the receptor after the co-culture. c-d MDA-MB231 after co-culture with BMHC. CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC in western blot (C) or real-time qPCR (D). Relative transcript levels are represented as the ratios between the 2 subpopulations of their 2–ΔΔCp real-time PCR values. These are data representative of three different experiments. e-f BMHC after co-culture with MDA-MB231. SDF-1α is increased in BMHC after 2 or 5 days co-culture with MDA-MB231 in western blot (E) or real-time qPCR (F). CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC (right panel). g Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 eGFP MDA-MB231 were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α or CXCR4 monoclonal antibody. Plastic was used as negative control. SDF-1α is involved in MDA-MB231 adhesion. h Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 MCF7, T47D or MDA-MB361 (stained with Calcein-Am) were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α monoclonal antibody
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Fig2: SDF-1alpha regulates interaction between MDA-MB231 and BMHC. a Flow cytometry cell sorting chart. MDA-MB231 (green) and BMHC (purple) were gated through GFP fluorescence intensity. b Flow cytometry analysis of CXCR4 expression. After 5 days of co-culture with BMHC, MDA-MB231 were cell sorted and stained for CXCR4. MDA-MB231 display an increase of the receptor after the co-culture. c-d MDA-MB231 after co-culture with BMHC. CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC in western blot (C) or real-time qPCR (D). Relative transcript levels are represented as the ratios between the 2 subpopulations of their 2–ΔΔCp real-time PCR values. These are data representative of three different experiments. e-f BMHC after co-culture with MDA-MB231. SDF-1α is increased in BMHC after 2 or 5 days co-culture with MDA-MB231 in western blot (E) or real-time qPCR (F). CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC (right panel). g Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 eGFP MDA-MB231 were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α or CXCR4 monoclonal antibody. Plastic was used as negative control. SDF-1α is involved in MDA-MB231 adhesion. h Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 MCF7, T47D or MDA-MB361 (stained with Calcein-Am) were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α monoclonal antibody

Mentions: SDF-1α/CXCR4 interactions regulate chemotaxis and homing of BCC to the BHMC [16]. To investigate whether SDF-1α/CXCR4 plays a role in the interaction between BMHC and MDA-MB231, we performed cell sorting after 2 and 5 days of co-culture (Fig. 2a) and showed an increase of CXCR4 in the MDA-MB231 (Fig. 2b-d). Concurrently, an increase of SDF-1α production by BMHC could be observed after co-culture (Fig. 2e-f). Western blot and Flow Cytometry analysis revealed the same profile in 3 other breast cancer cell lines (MDA-MB361, MCF7 and T47D) and an absence of expression of CXCR4 or an absence of increase of this receptor upon co-culture with BMHC in two other one (Hs578T and SK-BR-3; Additional file 1: Figure S1B-C).Fig. 2


SDF-1alpha concentration dependent modulation of RhoA and Rac1 modifies breast cancer and stromal cells interaction.

Pasquier J, Abu-Kaoud N, Abdesselem H, Madani A, Hoarau-Véchot J, Thawadi HA, Vidal F, Couderc B, Favre G, Rafii A - BMC Cancer (2015)

SDF-1alpha regulates interaction between MDA-MB231 and BMHC. a Flow cytometry cell sorting chart. MDA-MB231 (green) and BMHC (purple) were gated through GFP fluorescence intensity. b Flow cytometry analysis of CXCR4 expression. After 5 days of co-culture with BMHC, MDA-MB231 were cell sorted and stained for CXCR4. MDA-MB231 display an increase of the receptor after the co-culture. c-d MDA-MB231 after co-culture with BMHC. CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC in western blot (C) or real-time qPCR (D). Relative transcript levels are represented as the ratios between the 2 subpopulations of their 2–ΔΔCp real-time PCR values. These are data representative of three different experiments. e-f BMHC after co-culture with MDA-MB231. SDF-1α is increased in BMHC after 2 or 5 days co-culture with MDA-MB231 in western blot (E) or real-time qPCR (F). CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC (right panel). g Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 eGFP MDA-MB231 were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α or CXCR4 monoclonal antibody. Plastic was used as negative control. SDF-1α is involved in MDA-MB231 adhesion. h Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 MCF7, T47D or MDA-MB361 (stained with Calcein-Am) were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α monoclonal antibody
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Fig2: SDF-1alpha regulates interaction between MDA-MB231 and BMHC. a Flow cytometry cell sorting chart. MDA-MB231 (green) and BMHC (purple) were gated through GFP fluorescence intensity. b Flow cytometry analysis of CXCR4 expression. After 5 days of co-culture with BMHC, MDA-MB231 were cell sorted and stained for CXCR4. MDA-MB231 display an increase of the receptor after the co-culture. c-d MDA-MB231 after co-culture with BMHC. CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC in western blot (C) or real-time qPCR (D). Relative transcript levels are represented as the ratios between the 2 subpopulations of their 2–ΔΔCp real-time PCR values. These are data representative of three different experiments. e-f BMHC after co-culture with MDA-MB231. SDF-1α is increased in BMHC after 2 or 5 days co-culture with MDA-MB231 in western blot (E) or real-time qPCR (F). CXCR4 is increased in MDA-MB231 after 2 or 5 days of co-culture with BMHC (right panel). g Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 eGFP MDA-MB231 were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α or CXCR4 monoclonal antibody. Plastic was used as negative control. SDF-1α is involved in MDA-MB231 adhesion. h Adhesion assay. BMHC were plated up to 60 % confluency, 50,000 MCF7, T47D or MDA-MB361 (stained with Calcein-Am) were allowed to adhere for 1 h in presence or not of SDF-1α and a SDF-1α monoclonal antibody
Mentions: SDF-1α/CXCR4 interactions regulate chemotaxis and homing of BCC to the BHMC [16]. To investigate whether SDF-1α/CXCR4 plays a role in the interaction between BMHC and MDA-MB231, we performed cell sorting after 2 and 5 days of co-culture (Fig. 2a) and showed an increase of CXCR4 in the MDA-MB231 (Fig. 2b-d). Concurrently, an increase of SDF-1α production by BMHC could be observed after co-culture (Fig. 2e-f). Western blot and Flow Cytometry analysis revealed the same profile in 3 other breast cancer cell lines (MDA-MB361, MCF7 and T47D) and an absence of expression of CXCR4 or an absence of increase of this receptor upon co-culture with BMHC in two other one (Hs578T and SK-BR-3; Additional file 1: Figure S1B-C).Fig. 2

Bottom Line: This interaction increases migration from primary sites as well as homing at distant sites.Increased migration was obtained at 50 and 100 ng/ml of SDF-1α; however migration was reduced at 200 ng/ml.The adhesion between breast cancer cells and BMHC was significantly increased by SDF-1α treatment at 200 ng/ml and reduced using a blocking monoclonal antibody against CXCR4.

View Article: PubMed Central - PubMed

Affiliation: Stem Cell and Microenvironment Laboratory, Weill Cornell Medical College in Qatar, Education City, Qatar Foundation, Doha, Qatar. jep2026@qatar-med.cornell.edu.

ABSTRACT

Background: The interaction of SDF-1alpha with its receptor CXCR4 plays a role in the occurrence of distant metastasis in many solid tumors. This interaction increases migration from primary sites as well as homing at distant sites.

Methods: Here we investigated how SDF-1α could modulate both migration and adhesion of cancer cells through the modulation of RhoGTPases.

Results: We show that different concentrations of SDF-1α modulate the balance of adhesion and migration in cancer cells. Increased migration was obtained at 50 and 100 ng/ml of SDF-1α; however migration was reduced at 200 ng/ml. The adhesion between breast cancer cells and BMHC was significantly increased by SDF-1α treatment at 200 ng/ml and reduced using a blocking monoclonal antibody against CXCR4. We showed that at low SDF-1α concentration, RhoA was activated and overexpressed, while at high concentration Rac1 was promoting SDF-1α mediating-cell adhesion.

Conclusion: We conclude that SDF-1α concentration modulates migration and adhesion of breast cancer cells, by controlling expression and activation of RhoGTPases.

No MeSH data available.


Related in: MedlinePlus