Scatter factor/hepatocyte growth factor gene transfer enhances glioma growth and angiogenesis in vivo

Scatter factor (SF), also known as hepatocyte growth factor, is angiogenic in systemic tissues, and SF titers correlate with the malignancy and metastatic phenotype of certain systemic cancers. Human gliomas express SF and its receptor c-met, but their role in the malignant progression of these tumors has not been defined. To examine this, 9L glioma cells that express c-met but not SF were transfected with human SF cDNA, and their behavior in vitro and in vivo was examined. SF gene expression was detected in conditioned medium of 9L-SF but not in control 9L-neo-transfected cell lines, by reverse transcriptase-PCR, immunoblot, ELISA, and scatter activity assays. Gliomas derived from 9L-SF and control 9L-neo cell lines implanted in the caudate/putamen of Fisher 344 rats (intracranially) and in the flanks of SCID/Beige mice (subcutaneously) were examined. Extracts from intracranial (i.c.) gliomas contained elevated levels of SF protein as determined by ELISA (1 to 5.5 ng SF/mg protein), whereas no SF was detected in control tumors. Reverse transcriptase-PCR of RNA from i.c. gliomas revealed that only 9L-SF gliomas expressed SF and both 9L-neo and 9L-SF gliomas expressed the c-met SF receptor. By postimplantation Day 14, 9L-SF i.c. gliomas were approximately 5-fold larger than 9L-neo control tumors (p < 0.001). Subcutaneous 9L-SF glioma growth was also greater than that in controls, although the differences were more variable. SF-producing i.c. gliomas contained elevated levels of 48-kd urokinase (3.5-fold) and 92-kd type IV collagenase (2.8- fold), both enzymes that correlate with the malignant progression of human gliomas (p < 0.001). SF-producing and control 9L cell lines did not differ in rates of proliferation, thymidine incorporation, or adhesion-independent growth in vitro. Conditioned medium from 9L-SF cells stimulated thymidine incorporation into microvessel brain endothelial cells 3- to 4-fold higher than did CM from 9L-neo controls (p < 0.001). Intracranial 9L-SF gliomas were more angiogenic than controls based on elevated peak (2,25-fold; p < 0.005) and mean (1.7-fold; p < 0.008) blood vessel densities. These results suggest that SF production by glioma cells enhances glioma malignancy in vivo, in part, by paracrine mechanisms involving glioma-associated angiogenesis.