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Membrane-type 1 matrix metalloprotease (MT1-MMP) enables invasive migration of glioma cells in central nervous system white matter.

Beliën AT, Paganetti PA, Schwab ME - J. Cell Biol. (1999)

Bottom Line: Plasma membranes of both MT1-MMP-transfected fibroblasts and C6 glioma cells inactivated inhibitory myelin extracts, and this activity was sensitive to the same protease inhibitors.Interestingly, pretreatment of CNS myelin with gelatinase A/MMP-2 could not inactivate its inhibitory property.These data imply an important role of MT1-MMP in spreading and migration of glioma cells on white matter constituents in vitro and point to a function of MT1-MMP in the invasive behavior of malignant gliomas in the CNS in vivo.

View Article: PubMed Central - PubMed

Affiliation: Brain Research Institute, University of Zurich and Swiss Federal Institute of Technology, 8057 Zurich, Switzerland. belien@hifo.unizh.ch

ABSTRACT
Invasive glioma cells migrate preferentially along central nervous system (CNS) white matter fiber tracts irrespective of the fact that CNS myelin contains proteins that inhibit cell migration and neurite outgrowth. Previous work has demonstrated that to migrate on a myelin substrate and to overcome its inhibitory effect, rat C6 and human glioblastoma cells require a membrane-bound metalloproteolytic activity (C6-MP) which shares several biochemical and pharmacological characteristics with MT1-MMP. We show now that MT1-MMP is expressed on the surface of rat C6 glioblastoma cells and is coenriched with C6-MP activity. Immunodepletion of C6-MP activity is achieved with an anti-MT1-MMP antibody. These data suggest that MT1-MMP and the C6-MP are closely related or identical. When mouse 3T3 fibroblasts were transfected with MT1-MMP they acquired the ability to spread and migrate on the nonpermissive myelin substrate and to infiltrate into adult rat optic nerve explants. MT1-MMP-transfected fibroblasts and C6 glioma cells were able to digest bNI-220, one of the most potent CNS myelin inhibitory proteins. Plasma membranes of both MT1-MMP-transfected fibroblasts and C6 glioma cells inactivated inhibitory myelin extracts, and this activity was sensitive to the same protease inhibitors. Interestingly, pretreatment of CNS myelin with gelatinase A/MMP-2 could not inactivate its inhibitory property. These data imply an important role of MT1-MMP in spreading and migration of glioma cells on white matter constituents in vitro and point to a function of MT1-MMP in the invasive behavior of malignant gliomas in the CNS in vivo.

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Gelatin zymography. ProMMP-2 (lane 1) is partially  converted to mature MMP-2 by C6 plasma membranes (PM C6)  (lane 2), or MT1-MMP–transfected fibroblast plasma membranes (PM MT) (lane 4), but not by mock-transfected fibroblast  plasma membranes (PM Mo) (lane 6). Plasma membranes alone  (PM C6, lane 3; PM MT, lane 5; PM Mo, lane 7) neither contain  mature nor proMMP-2. Conditioned media of C6 cells (lane 8),  MT1-MMP–transfected fibroblasts (lane 9) and mock-transfected fibroblasts (lane 10) contain active and proMMP-2. Effects  of protease inhibitors: MMP-2 is strongly inhibited by 100 nM  TIMP2 (lane 12 versus lane 11) and 1 μM BB94 (lane 14), but not  by 100 nM TIMP1 (lane 13). Conversion of proMMP-2 to mature  MMP-2 by MT1-MMP–transfected fibroblast plasma membranes  is decreased by TIMP2 (100 nM, lane 16 versus lane 15), but not  by TIMP1 (100 nM, lane 17) or BB94 (1 μM).
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Figure 3: Gelatin zymography. ProMMP-2 (lane 1) is partially converted to mature MMP-2 by C6 plasma membranes (PM C6) (lane 2), or MT1-MMP–transfected fibroblast plasma membranes (PM MT) (lane 4), but not by mock-transfected fibroblast plasma membranes (PM Mo) (lane 6). Plasma membranes alone (PM C6, lane 3; PM MT, lane 5; PM Mo, lane 7) neither contain mature nor proMMP-2. Conditioned media of C6 cells (lane 8), MT1-MMP–transfected fibroblasts (lane 9) and mock-transfected fibroblasts (lane 10) contain active and proMMP-2. Effects of protease inhibitors: MMP-2 is strongly inhibited by 100 nM TIMP2 (lane 12 versus lane 11) and 1 μM BB94 (lane 14), but not by 100 nM TIMP1 (lane 13). Conversion of proMMP-2 to mature MMP-2 by MT1-MMP–transfected fibroblast plasma membranes is decreased by TIMP2 (100 nM, lane 16 versus lane 15), but not by TIMP1 (100 nM, lane 17) or BB94 (1 μM).

Mentions: C6-MP activity, as determined in a cell spreading assay, was also highest in the C6 plasma membrane fraction and lower in the C6 cell homogenate (Fig. 2 b) (Paganetti et al., 1988, Amberger et al., 1994). The presence of active MT1-MMP was confirmed by zymography where plasma membranes of C6 cells as well as of MT1-MMP–transfected fibroblasts showed conversion of progelatinase A/proMMP-2 to mature MMP-2 (Fig. 3, lanes 1–7). These results show the presence of active MT1-MMP on the plasma membranes of C6 glioblastoma cells.


Membrane-type 1 matrix metalloprotease (MT1-MMP) enables invasive migration of glioma cells in central nervous system white matter.

Beliën AT, Paganetti PA, Schwab ME - J. Cell Biol. (1999)

Gelatin zymography. ProMMP-2 (lane 1) is partially  converted to mature MMP-2 by C6 plasma membranes (PM C6)  (lane 2), or MT1-MMP–transfected fibroblast plasma membranes (PM MT) (lane 4), but not by mock-transfected fibroblast  plasma membranes (PM Mo) (lane 6). Plasma membranes alone  (PM C6, lane 3; PM MT, lane 5; PM Mo, lane 7) neither contain  mature nor proMMP-2. Conditioned media of C6 cells (lane 8),  MT1-MMP–transfected fibroblasts (lane 9) and mock-transfected fibroblasts (lane 10) contain active and proMMP-2. Effects  of protease inhibitors: MMP-2 is strongly inhibited by 100 nM  TIMP2 (lane 12 versus lane 11) and 1 μM BB94 (lane 14), but not  by 100 nM TIMP1 (lane 13). Conversion of proMMP-2 to mature  MMP-2 by MT1-MMP–transfected fibroblast plasma membranes  is decreased by TIMP2 (100 nM, lane 16 versus lane 15), but not  by TIMP1 (100 nM, lane 17) or BB94 (1 μM).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2132902&req=5

Figure 3: Gelatin zymography. ProMMP-2 (lane 1) is partially converted to mature MMP-2 by C6 plasma membranes (PM C6) (lane 2), or MT1-MMP–transfected fibroblast plasma membranes (PM MT) (lane 4), but not by mock-transfected fibroblast plasma membranes (PM Mo) (lane 6). Plasma membranes alone (PM C6, lane 3; PM MT, lane 5; PM Mo, lane 7) neither contain mature nor proMMP-2. Conditioned media of C6 cells (lane 8), MT1-MMP–transfected fibroblasts (lane 9) and mock-transfected fibroblasts (lane 10) contain active and proMMP-2. Effects of protease inhibitors: MMP-2 is strongly inhibited by 100 nM TIMP2 (lane 12 versus lane 11) and 1 μM BB94 (lane 14), but not by 100 nM TIMP1 (lane 13). Conversion of proMMP-2 to mature MMP-2 by MT1-MMP–transfected fibroblast plasma membranes is decreased by TIMP2 (100 nM, lane 16 versus lane 15), but not by TIMP1 (100 nM, lane 17) or BB94 (1 μM).
Mentions: C6-MP activity, as determined in a cell spreading assay, was also highest in the C6 plasma membrane fraction and lower in the C6 cell homogenate (Fig. 2 b) (Paganetti et al., 1988, Amberger et al., 1994). The presence of active MT1-MMP was confirmed by zymography where plasma membranes of C6 cells as well as of MT1-MMP–transfected fibroblasts showed conversion of progelatinase A/proMMP-2 to mature MMP-2 (Fig. 3, lanes 1–7). These results show the presence of active MT1-MMP on the plasma membranes of C6 glioblastoma cells.

Bottom Line: Plasma membranes of both MT1-MMP-transfected fibroblasts and C6 glioma cells inactivated inhibitory myelin extracts, and this activity was sensitive to the same protease inhibitors.Interestingly, pretreatment of CNS myelin with gelatinase A/MMP-2 could not inactivate its inhibitory property.These data imply an important role of MT1-MMP in spreading and migration of glioma cells on white matter constituents in vitro and point to a function of MT1-MMP in the invasive behavior of malignant gliomas in the CNS in vivo.

View Article: PubMed Central - PubMed

Affiliation: Brain Research Institute, University of Zurich and Swiss Federal Institute of Technology, 8057 Zurich, Switzerland. belien@hifo.unizh.ch

ABSTRACT
Invasive glioma cells migrate preferentially along central nervous system (CNS) white matter fiber tracts irrespective of the fact that CNS myelin contains proteins that inhibit cell migration and neurite outgrowth. Previous work has demonstrated that to migrate on a myelin substrate and to overcome its inhibitory effect, rat C6 and human glioblastoma cells require a membrane-bound metalloproteolytic activity (C6-MP) which shares several biochemical and pharmacological characteristics with MT1-MMP. We show now that MT1-MMP is expressed on the surface of rat C6 glioblastoma cells and is coenriched with C6-MP activity. Immunodepletion of C6-MP activity is achieved with an anti-MT1-MMP antibody. These data suggest that MT1-MMP and the C6-MP are closely related or identical. When mouse 3T3 fibroblasts were transfected with MT1-MMP they acquired the ability to spread and migrate on the nonpermissive myelin substrate and to infiltrate into adult rat optic nerve explants. MT1-MMP-transfected fibroblasts and C6 glioma cells were able to digest bNI-220, one of the most potent CNS myelin inhibitory proteins. Plasma membranes of both MT1-MMP-transfected fibroblasts and C6 glioma cells inactivated inhibitory myelin extracts, and this activity was sensitive to the same protease inhibitors. Interestingly, pretreatment of CNS myelin with gelatinase A/MMP-2 could not inactivate its inhibitory property. These data imply an important role of MT1-MMP in spreading and migration of glioma cells on white matter constituents in vitro and point to a function of MT1-MMP in the invasive behavior of malignant gliomas in the CNS in vivo.

Show MeSH
Related in: MedlinePlus