Shigeyuki Nagato

Introduction of wild-type p53 enhances thrombospondin-1 expression in human glioma cells

Malignant gliomas are distinguished from low-grade gliomas by their intense angiogenesis. In gliomas, p53 is the most frequently altered gene and is involved in the early phase of glioma development. In contrast, homozygous p16 gene deletion is more common in high-grade gliomas. In order to understand the mechanism by which gliomas become more angiogenic during the malignant transformation, we examined the relationship between thrombospondin-1, a negative regulator in angiogenesis, and these tumor suppressor genes in malignant gliomas. Human glioma cell line U-251 MG, which has mutated p53 and deleted p16, was transduced with recombinant replication-defective adenovirus vectors containing the cDNA of wild-type p53, p16, and p21. Only the induction of wild-type p53 enhanced expression of thrombospondin-1 mRNA and the protein in U-251 MG cells. Furthermore, thrombospondin-1 that was secreted in the culture medium was significantly increased (3.8-fold) as compared with that of the viral control 36 h after infection with Ad5CMV-p53. In the presence of wild-type p53 plasmid DNA, the promoter activity was increased 7.4-fold as compared with an empty expression vector control. These studies may suggest that mutation of p53 gene endows gliomas with an angiogenic phenotype by reducing thrombospondin-1 production as well as enhancing the angiogenesis inducers in the early phase of malignant progression.

Downregulation of laminin α4 chain expression inhibits glioma invasion in vitro and in vivo

The laminin family is a structural constituent of the extracellular matrix that plays an essential role in promoting the motility of infiltrative tumor cells. We investigated the role of laminin α4 chain, a subset of laminin‐8, ‐9 and ‐14, in the motile and invasive activities of human glioma cells. All malignant glioma cell lines examined expressed more mRNA for the laminin α4 and β1 chains than for the β2 chain, indicating that these cells predominantly express the laminin‐8 isoform. Introducing an antisense oligonucleotide for laminin α4 chain (AS‐Ln‐α4) into the glioma cells resulted in downregulation of laminin α4 expression. AS‐Ln‐α4 also significantly suppressed glioma cell adhesion and migration. Furthermore, invasiveness was significantly reduced in cells transfected with AS‐Ln‐α4 compared to those transfected with the sense oligonucleotide (S‐Ln‐α4). Indeed, when glioma spheroids were implanted into rat brain slices, AS‐Ln‐α4‐transfected cells failed to invade surrounding normal brain tissues. In addition, intracerebral injection of glioma cells transfected with AS‐Ln‐α4 into nude mice resulted in the formation of a noninvasive tumor, whereas injection of cells transfected with S‐Ln‐α4 resulted in diffuse invasion of brain tissue. These results suggest that mainly laminin‐8 is essential for the invasive activity of human glioma cells; thus, a novel therapeutic strategy could target this molecule to treat patients with malignant glioma.

Downregulation of laminin alpha4 chain expression inhibits glioma invasion in vitro and in vivo.

The laminin family is a structural constituent of the extracellular matrix that plays an essential role in promoting the motility of infiltrative tumor cells. We investigated the role of laminin α4 chain, a subset of laminin‐8, ‐9 and ‐14, in the motile and invasive activities of human glioma cells. All malignant glioma cell lines examined expressed more mRNA for the laminin α4 and β1 chains than for the β2 chain, indicating that these cells predominantly express the laminin‐8 isoform. Introducing an antisense oligonucleotide for laminin α4 chain (AS‐Ln‐α4) into the glioma cells resulted in downregulation of laminin α4 expression. AS‐Ln‐α4 also significantly suppressed glioma cell adhesion and migration. Furthermore, invasiveness was significantly reduced in cells transfected with AS‐Ln‐α4 compared to those transfected with the sense oligonucleotide (S‐Ln‐α4). Indeed, when glioma spheroids were implanted into rat brain slices, AS‐Ln‐α4‐transfected cells failed to invade surrounding normal brain tissues. In addition, intracerebral injection of glioma cells transfected with AS‐Ln‐α4 into nude mice resulted in the formation of a noninvasive tumor, whereas injection of cells transfected with S‐Ln‐α4 resulted in diffuse invasion of brain tissue. These results suggest that mainly laminin‐8 is essential for the invasive activity of human glioma cells; thus, a novel therapeutic strategy could target this molecule to treat patients with malignant glioma.

Downregulation of laminin α4 chain expression inhibits glioma invasionin vitro andin vivo

The laminin family is a structural constituent of the extracellular matrix that plays an essential role in promoting the motility of infiltrative tumor cells. We investigated the role of laminin α4 chain, a subset of laminin‐8, ‐9 and ‐14, in the motile and invasive activities of human glioma cells. All malignant glioma cell lines examined expressed more mRNA for the laminin α4 and β1 chains than for the β2 chain, indicating that these cells predominantly express the laminin‐8 isoform. Introducing an antisense oligonucleotide for laminin α4 chain (AS‐Ln‐α4) into the glioma cells resulted in downregulation of laminin α4 expression. AS‐Ln‐α4 also significantly suppressed glioma cell adhesion and migration. Furthermore, invasiveness was significantly reduced in cells transfected with AS‐Ln‐α4 compared to those transfected with the sense oligonucleotide (S‐Ln‐α4). Indeed, when glioma spheroids were implanted into rat brain slices, AS‐Ln‐α4‐transfected cells failed to invade surrounding normal brain tissues. In addition, intracerebral injection of glioma cells transfected with AS‐Ln‐α4 into nude mice resulted in the formation of a noninvasive tumor, whereas injection of cells transfected with S‐Ln‐α4 resulted in diffuse invasion of brain tissue. These results suggest that mainly laminin‐8 is essential for the invasive activity of human glioma cells; thus, a novel therapeutic strategy could target this molecule to treat patients with malignant glioma.