Transfer of the sFLT-1 gene in Morris hepatoma results in decreased growth and perfusion and induction of genes associated with stress response.
OBJECTIVE
Inhibition of tumor angiogenesis is emerging as a promising target in the treatment of malignancies. Therefore, monitoring of antiangiogenic approaches with functional imaging and histomorphometrical analyses are desirable to evaluate the biological effects caused by this treatment modality.


METHODS
Using a bicistronic retroviral vector for transfer of the soluble receptor for the vascular endothelial growth factor (sFLT) hepatoma (MH3924A) cell lines with sFLT expression were generated. In human umbilical vein endothelial cells cultured with conditioned medium of sFLT-expressing hepatoma cells, the inhibitory action of secreted sFLT was determined using a Coulter counter and a thymidine incorporation assay. Furthermore, in vivo experiments were done to measure the effects on tumor growth and perfusion. Finally, the tumors were examined by immunohistochemistry (including computer-assisted morphometry) and DNA chip analysis.


RESULTS
Stable sFLT-expressing hepatoma cells inhibited endothelial cell proliferation in vitro. In vivo, growth and perfusion, as measured by H(2)(15)O positron emission tomography, were reduced in genetically modified tumors. However, the immunohistochemically quantified microvascularization and macrovascularization, as indicated by CD31- and alpha-actin-positive area, revealed no significant changes, whereas the number of apoptotic cells was increased in sFLT-expressing tumors, although not significantly. DNA chip analysis of tumors with gene transfer showed an increase of genes related to apoptosis, signal transduction, and oxidative stress.


CONCLUSIONS
Our results suggest that sFLT expression inhibits tumor growth and perfusion and enhances expression of apoptosis-related genes in this model. Enhanced expression of genes for signal transduction, stress, and metabolism indicates tumor defense reactions.
Schmidt
Kerstin
K
Department of Anatomy and Cell Biology III, University of Heidelberg, Im Neuenheimer Feld 400, Heidelberg 69120, Germany.
Hoffend
Johannes
J
Altmann
Annette
A
Strauss
Ludwig G
LG
Dimitrakopoulou-Strauss
Antonia
A
Engelhardt
Britta
B
Koczan
Dirk
D
Peter
Jörg
J
Vorwald
Silke
S
Eskerski
Helmut
H
Eisenhut
Michael
M
Metz
Jürgen
J
Kinscherf
Ralf
R
Haberkorn
Uwe
U
D000818:Animals; D049109:Cell Proliferation; D017077:Culture Media, Conditioned; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D018014:Gene Transfer Techniques; D006801:Humans; D008114:Liver Neoplasms, Experimental / Q000235:genetics* / Q000473:pathology / Q000517:prevention & control*; D018384:Oxidative Stress / Q000235:genetics*; D049268:Positron-Emission Tomography; D011506:Proteins / Q000235:genetics*; D051381:Rats; D011923:Rats, Nude; D012190:Retroviridae / Q000235:genetics; D013936:Thymidine / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014471:Umbilical Veins / Q000378:metabolism / Q000473:pathology
D017077:Culture Media, Conditioned; D011506:Proteins; D013936:Thymidine