Artikel
Antagonization of PTPzeta by siRNA expression inhibits glioblastoma growth in vivo
Die Antagonisierung der PTPz Expression mittels siRNA inhibiert das Glioblastomwachstum In-vivo
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Autoren
Veröffentlicht: | 4. Mai 2005 |
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Gliederung
Text
Objective
We previously identified the protein tyrosine phosphatase zeta (PTPξ) and its ligand pleiotrophin (PTN) as overexpressed in glioblastomas using cDNA arrays (Müller et al., Oncogene 2003). Both molecules have been implicated in neuronal migration during central nervous system development. We confirmed PTPξ and PTN overexpression in glioblastomas at the protein level and showed that matrix-immobilized PTN stimulates glioma cell migration (Ulbricht et al. J Neuropathol Exp Neurol 2003). In the present study we analyzed the effect of PTPξ expression on glioma growth in vitro and in vivo using siRNA technology.
Methods
PTPξ expression was downregulated in U251-MG glioblastoma cells by stable siRNA transfection. Downregulation was detected by northern and western blotting. Colorimetric proliferation assays and modified Boyden chamber haptotactic migration assays were performed in vitro. Tumor growth in vivo was studied using a subcutaneous nude mouse model. Tumors were analyzed immunohistochemically to assess cell proliferation, apoptosis and vascularization.
Results
Two siRNA-transfected clones with strong downregulation of PTPξ expression were obtained (PTPξ-), and these clones were subsequently compared with 2 mock transfected control clones (PTPξ+). Proliferation of PTPξ- clones was inhibited by 56%-90% compared with PTPξ+ clones. Haptotaxis (migration towards substrate-bound molecules) induced by PTN was inhibited by 52-76% in PTPξ- clones compared with controls. Collagen I and fibronectin were also haptotactic, however, no differences were observed between PTPξ- and PTPξ+ clones, suggesting that the inhibition of haptotaxis towards PTN is specifically mediated through PTPξ. In vivo, growth of RPTPβ- clones was inhibited by 93-98% compared with controls. The fraction of proliferating tumor cells was reduced by 49-74% in tumors derived from PTPξ- clones, whereas no differences were observed for apoptosis or microvessel density.
Conclusions
Glioma growth in vivo can be inhibited by downregulation of PTPξ expression. In vitro, PTPξ contributes to glioma cell proliferation and mediates PTN-induced haptotactic migration. PTN was demonstrated earlier to be secreted by U251-MG cells, therefore our findings suggest that upregulated expression of PTN and PTPξ in human astrocytoma cells can create an autocrine loop that contributes to tumor growth in vivo.