Limits...
Tumor-derived microvesicles mediate human breast cancer invasion through differentially glycosylated EMMPRIN.

Menck K, Scharf C, Bleckmann A, Dyck L, Rost U, Wenzel D, Dhople VM, Siam L, Pukrop T, Binder C, Klemm F - J Mol Cell Biol (2014)

Bottom Line: Uptake of T-MV is essential for the proinvasive effect.Interestingly, the effect of EMMPRIN-bearing MV is not mediated by matrix metalloproteinases but by activation of the p38/MAPK signaling pathway in the tumor cells.In conclusion, T-MV stimulate cancer cell invasion via a direct feedback mechanism dependent on highly glycosylated EMMPRIN.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology/Oncology, University Medical Center Göttingen, 37099 Göttingen, Germany.

No MeSH data available.


Related in: MedlinePlus

EMMPRIN is expressed on T-MV in vitro and in vivo and contributes to their proinvasive phenotype. (A and B) Western blots showing the expression of highly (HG) and lowly glycosylated (LG) EMMPRIN (EMP) in whole cell lysates (C) and T-MV (A) as well as the corresponding T-Exo (B) of both breast cancer cell lines. (C) Western blots showing stable knockdown of EMMPRIN via shRNA (shEMP) in SK-BR-3 cells and MV (ns ctl, non-sense control). (D) Microinvasion assay of MCF-7 cells exposed to T-MVS (1 µg/ml) from EMMPRIN knockdown cells (mean ± SD, n = 3, *P < 0.01). (E) Total MV from peripheral blood of metastatic breast cancer (BC) patients and matched controls (ctl). The percentage of EMMPRIN-positive MV was quantified by flow cytometry. (F) Double staining for EMMPRIN and MUC1 of MV from a BC and a control patient (representative density plots).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4401212&req=5

MJU047F3: EMMPRIN is expressed on T-MV in vitro and in vivo and contributes to their proinvasive phenotype. (A and B) Western blots showing the expression of highly (HG) and lowly glycosylated (LG) EMMPRIN (EMP) in whole cell lysates (C) and T-MV (A) as well as the corresponding T-Exo (B) of both breast cancer cell lines. (C) Western blots showing stable knockdown of EMMPRIN via shRNA (shEMP) in SK-BR-3 cells and MV (ns ctl, non-sense control). (D) Microinvasion assay of MCF-7 cells exposed to T-MVS (1 µg/ml) from EMMPRIN knockdown cells (mean ± SD, n = 3, *P < 0.01). (E) Total MV from peripheral blood of metastatic breast cancer (BC) patients and matched controls (ctl). The percentage of EMMPRIN-positive MV was quantified by flow cytometry. (F) Double staining for EMMPRIN and MUC1 of MV from a BC and a control patient (representative density plots).

Mentions: Previously, we have shown that EMMPRIN expression in human breast cancer cells is involved in reprogramming tumor stroma cells toward a proinvasive, tumor-supporting phenotype (Hagemann et al., 2005). To investigate whether EMMPRIN is present on both EV populations, i.e. MV and Exo, of breast cancer cells, we analyzed its expression on the individual populations in comparison with the whole cell lysate. In fact, we found an enrichment of EMMPRIN on MV of both MCF-7 and SK-BR-3 cells (Figure 3A), whereas it was absent on the respective Exo (Figure 3B). To further clarify whether EMMPRIN is suitable for discriminating MV and Exo, we comparably analyzed various known markers on both vesicle populations (Supplementary Figure S3A). Interestingly, Flotillin-2, which is often used as a marker for Exo, was found mainly in MV preparations, while the MVB protein TSG101 was indeed specifically expressed on Exo. Tubulin was found predominantly on MV. Next, MV and Exo were loaded separately on sucrose gradients. Based on the distribution of the different markers, both populations were localized in comparable density fractions ranging from 1.13 to 1.22 g/ml (Supplementary Figure S3B). Again, EMMPRIN was present exclusively in the MV fraction, confirming that it can be used to visualize MV in sucrose gradient preparations and to distinguish them from Exo.Figure 3


Tumor-derived microvesicles mediate human breast cancer invasion through differentially glycosylated EMMPRIN.

Menck K, Scharf C, Bleckmann A, Dyck L, Rost U, Wenzel D, Dhople VM, Siam L, Pukrop T, Binder C, Klemm F - J Mol Cell Biol (2014)

EMMPRIN is expressed on T-MV in vitro and in vivo and contributes to their proinvasive phenotype. (A and B) Western blots showing the expression of highly (HG) and lowly glycosylated (LG) EMMPRIN (EMP) in whole cell lysates (C) and T-MV (A) as well as the corresponding T-Exo (B) of both breast cancer cell lines. (C) Western blots showing stable knockdown of EMMPRIN via shRNA (shEMP) in SK-BR-3 cells and MV (ns ctl, non-sense control). (D) Microinvasion assay of MCF-7 cells exposed to T-MVS (1 µg/ml) from EMMPRIN knockdown cells (mean ± SD, n = 3, *P < 0.01). (E) Total MV from peripheral blood of metastatic breast cancer (BC) patients and matched controls (ctl). The percentage of EMMPRIN-positive MV was quantified by flow cytometry. (F) Double staining for EMMPRIN and MUC1 of MV from a BC and a control patient (representative density plots).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4401212&req=5

MJU047F3: EMMPRIN is expressed on T-MV in vitro and in vivo and contributes to their proinvasive phenotype. (A and B) Western blots showing the expression of highly (HG) and lowly glycosylated (LG) EMMPRIN (EMP) in whole cell lysates (C) and T-MV (A) as well as the corresponding T-Exo (B) of both breast cancer cell lines. (C) Western blots showing stable knockdown of EMMPRIN via shRNA (shEMP) in SK-BR-3 cells and MV (ns ctl, non-sense control). (D) Microinvasion assay of MCF-7 cells exposed to T-MVS (1 µg/ml) from EMMPRIN knockdown cells (mean ± SD, n = 3, *P < 0.01). (E) Total MV from peripheral blood of metastatic breast cancer (BC) patients and matched controls (ctl). The percentage of EMMPRIN-positive MV was quantified by flow cytometry. (F) Double staining for EMMPRIN and MUC1 of MV from a BC and a control patient (representative density plots).
Mentions: Previously, we have shown that EMMPRIN expression in human breast cancer cells is involved in reprogramming tumor stroma cells toward a proinvasive, tumor-supporting phenotype (Hagemann et al., 2005). To investigate whether EMMPRIN is present on both EV populations, i.e. MV and Exo, of breast cancer cells, we analyzed its expression on the individual populations in comparison with the whole cell lysate. In fact, we found an enrichment of EMMPRIN on MV of both MCF-7 and SK-BR-3 cells (Figure 3A), whereas it was absent on the respective Exo (Figure 3B). To further clarify whether EMMPRIN is suitable for discriminating MV and Exo, we comparably analyzed various known markers on both vesicle populations (Supplementary Figure S3A). Interestingly, Flotillin-2, which is often used as a marker for Exo, was found mainly in MV preparations, while the MVB protein TSG101 was indeed specifically expressed on Exo. Tubulin was found predominantly on MV. Next, MV and Exo were loaded separately on sucrose gradients. Based on the distribution of the different markers, both populations were localized in comparable density fractions ranging from 1.13 to 1.22 g/ml (Supplementary Figure S3B). Again, EMMPRIN was present exclusively in the MV fraction, confirming that it can be used to visualize MV in sucrose gradient preparations and to distinguish them from Exo.Figure 3

Bottom Line: Uptake of T-MV is essential for the proinvasive effect.Interestingly, the effect of EMMPRIN-bearing MV is not mediated by matrix metalloproteinases but by activation of the p38/MAPK signaling pathway in the tumor cells.In conclusion, T-MV stimulate cancer cell invasion via a direct feedback mechanism dependent on highly glycosylated EMMPRIN.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology/Oncology, University Medical Center Göttingen, 37099 Göttingen, Germany.

No MeSH data available.


Related in: MedlinePlus