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Exogenous expression of N-cadherin in breast cancer cells induces cell migration, invasion, and metastasis.

Hazan RB, Phillips GR, Qiao RF, Norton L, Aaronson SA - J. Cell Biol. (2000)

Bottom Line: To determine whether N-cadherin promotes invasion and metastasis, we transfected a weakly metastatic and E-cadherin-expressing breast cancer cell line, MCF-7, with N-cadherin and analyzed the effects on cell migration, invasion, and metastasis.These results demonstrate that N-cadherin promotes motility, invasion, and metastasis even in the presence of the normally suppressive E-cadherin.The increase in MMP-9 production by N-cadherin-expressing cells in response to a growth factor may endow them with a greater ability to penetrate matrix protein barriers, while the increase in their adherence to endothelium may improve their ability to enter and exit the vasculature, two properties that may be responsible for metastasis of N-cadherin-expressing cells.

View Article: PubMed Central - PubMed

Affiliation: The Derald H. Ruttenberg Cancer Center, Mount Sinai School of Medicine of New York University, New York, New York 10029, USA. rhazan@smtplink.mssm.edu

ABSTRACT
E- and N-cadherin are calcium-dependent cell adhesion molecules that mediate cell-cell adhesion and also modulate cell migration and tumor invasiveness. The loss of E-cadherin-mediated adhesion has been shown to play an important role in the transition of epithelial tumors from a benign to an invasive state. However, recent evidence indicates that another member of the cadherin family, N-cadherin, is expressed in highly invasive tumor cell lines that lacked E-cadherin expression. These findings have raised the possibility that N-cadherin contributes to the invasive phenotype. To determine whether N-cadherin promotes invasion and metastasis, we transfected a weakly metastatic and E-cadherin-expressing breast cancer cell line, MCF-7, with N-cadherin and analyzed the effects on cell migration, invasion, and metastasis. Transfected cells expressed both E- and N-cadherin and exhibited homotypic cell adhesion from both molecules. In vitro, N-cadherin-expressing cells migrated more efficiently, showed an increased invasion of Matrigel, and adhered more efficiently to monolayers of endothelial cells. All cells produced low levels of the matrix metalloproteinase MMP-9, which was dramatically upregulated by treatment with FGF-2 only in N-cadherin-expressing cells. Migration and invasion of Matrigel were also greatly enhanced by this treatment. When injected into the mammary fat pad of nude mice, N-cadherin-expressing cells, but not control MCF-7 cells, metastasized widely to the liver, pancreas, salivary gland, omentum, lung, lymph nodes, and lumbar spinal muscle. The expression of both E- and N-cadherin was maintained both in the primary tumors and metastatic lesions. These results demonstrate that N-cadherin promotes motility, invasion, and metastasis even in the presence of the normally suppressive E-cadherin. The increase in MMP-9 production by N-cadherin-expressing cells in response to a growth factor may endow them with a greater ability to penetrate matrix protein barriers, while the increase in their adherence to endothelium may improve their ability to enter and exit the vasculature, two properties that may be responsible for metastasis of N-cadherin-expressing cells.

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FGF-2 stimulates Matrigel invasion of N-cadherin–expressing cells and expression of MMP-9. (A) Invasion through Matrigel-coated filters of control and N-cadherin–expressing cells, pretreated for 24 h with 10 ng/ml of FGF-2 or left untreated, as measured 5 h after inoculation into Transwell chambers. The invading cells were stained (see Materials and Methods) and photographed using a digital microscope camera. Each panel illustrates a sample representing three to six filters. (B) Gelatinolytic activity of conditioned media of control (Neo-mass) (lanes 1 and 2) or N-cadherin–transfected MCF-7 cells (N-cad-mass, N-cad-5, and N-cad-17; lanes 3–8) that were either treated with FGF-2, or left untreated, as indicated. (C) Gelatinolytic activity of conditioned media of Neo-mass, N-cad-mass, N-cad-5, and N-cad-17 that were pretreated for 24 h either with 10 ng/ml of FGF-2 (lanes 1, 3, 5, and 7, respectively) or 5 μg/ml insulin (lanes 2, 4, 6, and 8, respectively).
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Figure 9: FGF-2 stimulates Matrigel invasion of N-cadherin–expressing cells and expression of MMP-9. (A) Invasion through Matrigel-coated filters of control and N-cadherin–expressing cells, pretreated for 24 h with 10 ng/ml of FGF-2 or left untreated, as measured 5 h after inoculation into Transwell chambers. The invading cells were stained (see Materials and Methods) and photographed using a digital microscope camera. Each panel illustrates a sample representing three to six filters. (B) Gelatinolytic activity of conditioned media of control (Neo-mass) (lanes 1 and 2) or N-cadherin–transfected MCF-7 cells (N-cad-mass, N-cad-5, and N-cad-17; lanes 3–8) that were either treated with FGF-2, or left untreated, as indicated. (C) Gelatinolytic activity of conditioned media of Neo-mass, N-cad-mass, N-cad-5, and N-cad-17 that were pretreated for 24 h either with 10 ng/ml of FGF-2 (lanes 1, 3, 5, and 7, respectively) or 5 μg/ml insulin (lanes 2, 4, 6, and 8, respectively).

Mentions: The effect of FGF-2 on the invasive activity of N-cadherin–expressing MCF-7 cells was next assessed in a Matrigel invasion assay after 5 h of incubation (Fig. 9 A). The low basal invasiveness of the N-cad-mass cells and the high invasiveness of the N-cad-5 and -17 clones were all further enhanced by FGF-2 treatment. In contrast, there was no effect on the Neo-mass cell invasion (Fig. 9 A). Of note, we found that the Matrigel-invading N-cadherin–transfected MCF-7 cells appeared on the underside of the filter as cell clusters rather than individual cells. This suggests that the gain of invasive properties by N-cadherin does not involve a switch from an adhesive to a more scattered, mesenchymal phenotype usually observed with invasive carcinomas (Oka et al. 1993; Sommers et al. 1994; Hazan et al. 1997).


Exogenous expression of N-cadherin in breast cancer cells induces cell migration, invasion, and metastasis.

Hazan RB, Phillips GR, Qiao RF, Norton L, Aaronson SA - J. Cell Biol. (2000)

FGF-2 stimulates Matrigel invasion of N-cadherin–expressing cells and expression of MMP-9. (A) Invasion through Matrigel-coated filters of control and N-cadherin–expressing cells, pretreated for 24 h with 10 ng/ml of FGF-2 or left untreated, as measured 5 h after inoculation into Transwell chambers. The invading cells were stained (see Materials and Methods) and photographed using a digital microscope camera. Each panel illustrates a sample representing three to six filters. (B) Gelatinolytic activity of conditioned media of control (Neo-mass) (lanes 1 and 2) or N-cadherin–transfected MCF-7 cells (N-cad-mass, N-cad-5, and N-cad-17; lanes 3–8) that were either treated with FGF-2, or left untreated, as indicated. (C) Gelatinolytic activity of conditioned media of Neo-mass, N-cad-mass, N-cad-5, and N-cad-17 that were pretreated for 24 h either with 10 ng/ml of FGF-2 (lanes 1, 3, 5, and 7, respectively) or 5 μg/ml insulin (lanes 2, 4, 6, and 8, respectively).
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Figure 9: FGF-2 stimulates Matrigel invasion of N-cadherin–expressing cells and expression of MMP-9. (A) Invasion through Matrigel-coated filters of control and N-cadherin–expressing cells, pretreated for 24 h with 10 ng/ml of FGF-2 or left untreated, as measured 5 h after inoculation into Transwell chambers. The invading cells were stained (see Materials and Methods) and photographed using a digital microscope camera. Each panel illustrates a sample representing three to six filters. (B) Gelatinolytic activity of conditioned media of control (Neo-mass) (lanes 1 and 2) or N-cadherin–transfected MCF-7 cells (N-cad-mass, N-cad-5, and N-cad-17; lanes 3–8) that were either treated with FGF-2, or left untreated, as indicated. (C) Gelatinolytic activity of conditioned media of Neo-mass, N-cad-mass, N-cad-5, and N-cad-17 that were pretreated for 24 h either with 10 ng/ml of FGF-2 (lanes 1, 3, 5, and 7, respectively) or 5 μg/ml insulin (lanes 2, 4, 6, and 8, respectively).
Mentions: The effect of FGF-2 on the invasive activity of N-cadherin–expressing MCF-7 cells was next assessed in a Matrigel invasion assay after 5 h of incubation (Fig. 9 A). The low basal invasiveness of the N-cad-mass cells and the high invasiveness of the N-cad-5 and -17 clones were all further enhanced by FGF-2 treatment. In contrast, there was no effect on the Neo-mass cell invasion (Fig. 9 A). Of note, we found that the Matrigel-invading N-cadherin–transfected MCF-7 cells appeared on the underside of the filter as cell clusters rather than individual cells. This suggests that the gain of invasive properties by N-cadherin does not involve a switch from an adhesive to a more scattered, mesenchymal phenotype usually observed with invasive carcinomas (Oka et al. 1993; Sommers et al. 1994; Hazan et al. 1997).

Bottom Line: To determine whether N-cadherin promotes invasion and metastasis, we transfected a weakly metastatic and E-cadherin-expressing breast cancer cell line, MCF-7, with N-cadherin and analyzed the effects on cell migration, invasion, and metastasis.These results demonstrate that N-cadherin promotes motility, invasion, and metastasis even in the presence of the normally suppressive E-cadherin.The increase in MMP-9 production by N-cadherin-expressing cells in response to a growth factor may endow them with a greater ability to penetrate matrix protein barriers, while the increase in their adherence to endothelium may improve their ability to enter and exit the vasculature, two properties that may be responsible for metastasis of N-cadherin-expressing cells.

View Article: PubMed Central - PubMed

Affiliation: The Derald H. Ruttenberg Cancer Center, Mount Sinai School of Medicine of New York University, New York, New York 10029, USA. rhazan@smtplink.mssm.edu

ABSTRACT
E- and N-cadherin are calcium-dependent cell adhesion molecules that mediate cell-cell adhesion and also modulate cell migration and tumor invasiveness. The loss of E-cadherin-mediated adhesion has been shown to play an important role in the transition of epithelial tumors from a benign to an invasive state. However, recent evidence indicates that another member of the cadherin family, N-cadherin, is expressed in highly invasive tumor cell lines that lacked E-cadherin expression. These findings have raised the possibility that N-cadherin contributes to the invasive phenotype. To determine whether N-cadherin promotes invasion and metastasis, we transfected a weakly metastatic and E-cadherin-expressing breast cancer cell line, MCF-7, with N-cadherin and analyzed the effects on cell migration, invasion, and metastasis. Transfected cells expressed both E- and N-cadherin and exhibited homotypic cell adhesion from both molecules. In vitro, N-cadherin-expressing cells migrated more efficiently, showed an increased invasion of Matrigel, and adhered more efficiently to monolayers of endothelial cells. All cells produced low levels of the matrix metalloproteinase MMP-9, which was dramatically upregulated by treatment with FGF-2 only in N-cadherin-expressing cells. Migration and invasion of Matrigel were also greatly enhanced by this treatment. When injected into the mammary fat pad of nude mice, N-cadherin-expressing cells, but not control MCF-7 cells, metastasized widely to the liver, pancreas, salivary gland, omentum, lung, lymph nodes, and lumbar spinal muscle. The expression of both E- and N-cadherin was maintained both in the primary tumors and metastatic lesions. These results demonstrate that N-cadherin promotes motility, invasion, and metastasis even in the presence of the normally suppressive E-cadherin. The increase in MMP-9 production by N-cadherin-expressing cells in response to a growth factor may endow them with a greater ability to penetrate matrix protein barriers, while the increase in their adherence to endothelium may improve their ability to enter and exit the vasculature, two properties that may be responsible for metastasis of N-cadherin-expressing cells.

Show MeSH
Related in: MedlinePlus