Effects of endostatin-vascular endothelial growth inhibitor chimeric recombinant adenoviruses on antiangiogenesis.
OBJECTIVE
To investigate the inhibitory effects of endostatin-vascular endothelial growth inhibitor (VEGI151) recombinant adenoviruses on neovascularization.


METHODS
We used recombinant adenoviruses to treat human vascular endothelial cell line ECV304, human hepatocellular carcinoma cell line HepG2, and murine fibroblast cell line L929, in order to study the chimeric gene expression in these cell lines. Chick choriallantic membrane (CAM) model, rabbit inflammatory corneal neovascularization (CNV) model, and liver cancer-bearing nude mice model were employed to investigate the negative biological effect of fusion molecules on neovascularization in vivo.


RESULTS
Western blot showed that the molecular weight of fusion protein was about 41 kD after infection of ECV304, HepG2 and L929 cells with supernatant of AdhENDO-VEGI151. The fusion protein showed a specific inhibitory effect on the proliferation of ECV304 cells, but no inhibitory effect on the growth of HepG2 and L929 cells (F=13112.13, P=0.0001). In the chick choriallantic membrane (CAM) assay, the expressed fusion protein significantly inhibited neovascularization. Rabbit inflammatory corneal neovascularization (CNV) induced by intrastromal sutures resulted in a uniform neovascular response. In this model, direct subconjunctival injection of AdhENDO-VEGI151 expressed the fusion protein in vivo and suppressed the development of CNV. Topical application of AdhENDO-VEGI151 led to a significant suppression of CNV (F=1413.11, P=0.0001), as compared with the control group of AdLacZ. Immunohistochemical staining showed the fusion protein dominantly expressed in corneal epithelium. Compared with the control group of AdLacZ (4075.9+/-1849.9 mm(3)), the average tumor size of group AdhENDO-VEGI151 reduced in size (487.7+/-241.2 mm(3)) (F=14.80, P=0.0085), with an inhibition rate of 88.03%. Immunohistochemical staining showed the adenoviruses carried the fusion gene expressed on liver cancer cell membrane. MVD decreased more significantly in treated mice (30.75+/-3.31%) than in AdLacZ control (50.25+/-8.65%) (F=17.72, P=0.0056) with an inhibition rate of 39%.


CONCLUSIONS
Fusion protein expressed by recombinant adenoviruses has a significant inhibitory effect on neovascularization.
Pan
Xin
X
Department of Microbiology, Second Military Medical University, Shanghai 200433, China. panxinpx@yahoo.com
Wang
Yong
Y
Zhang
Min
M
Pan
Wei
W
Qi
Zhong-Tian
ZT
Cao
Guang-Wen
GW
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism*; D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D001681:Biological Assay; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D002642:Chick Embryo; D049033:Chorioallantoic Membrane / Q000098:blood supply; D003315:Cornea / Q000098:blood supply; D043169:Endostatins / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D053380:Tumor Necrosis Factor Ligand Superfamily Member 15; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*
D020533:Angiogenesis Inhibitors; D043169:Endostatins; D011993:Recombinant Fusion Proteins; C117026:TNFSF15 protein, human; D053380:Tumor Necrosis Factor Ligand Superfamily Member 15; D014409:Tumor Necrosis Factor-alpha