Potent inhibition of angiogenesis and liver tumor growth by administration of an aerosol containing a transferrin-liposome-endostatin complex.
OBJECTIVE
To obtain an efficient delivery system for transporting endostatin gene to mouse liver tumor xenografts by administration of aerosol.


METHODS
Recombinant plasmid pcDNA3.0/endostatin containing human endostatin gene together with signal peptide from alkaline phosphatase were transferred into human umbilical vein endothelial cell (HUVEC) by transferrin(TF)-liposome-endostatin complex. Western blot was used to detect the expression of human endostatin in transfected HUVEC cells and its medium. After the tumor-bearing mice were administrated with TF-liposome-endostatin complex, the lung tissue was analyzed by immunohistochemical method for expression of endostatin and the tumors were treated with CD-31 antibody to detect the density of microvessels in tumor tissues. The inhibition of tumor growth was estimated by the weight of tumors from groups treated with different doses of TF-liposome-endostatin complex. DNA fragmentation assay was used to detect the apoptosis of the cells from primary liver tumor.


RESULTS
Western blot analysis and immunohistochemical method confirmed the expression of endostatin protein in vitro and in vivo. After the tumor sections were treated with CD-31 antibody, the positive reaction cells appeared brown while the negative cells were colorless. The positively stained area of the TF-liposome-endostatin treated group was significantly smaller (P<0.01, 645.8+/-55.2 microm(2)) than that of the control group (1 325.4+/-198.5 microm(2)). The data showed a significant inhibition of angiogenesis. After administration of TF-liposome-endostatin, comparing with the control group administrated with TF-liposome-pcDNA3.0, liver tumor growth in the mice treated with 50, 250 and 500 mg DNA/kg was inhibited by 36.6 %, 40.8 %, and 72.8 %, respectively (P<0.01). And a typical DNA fragmentation of apoptosis was found in the cells from tumor tissues of the mice treated with TF-liposome-endostatin but none in the control group.


CONCLUSIONS
Endostatin gene could be efficiently transported into the mice with TF-liposome-DNA delivery system by administration of aerosol. TF-liposome-mediated endostatin gene therapy strongly inhibited angiogenesis and the growth of mouse xenograft liver tumors. It also could promote the development of apoptosis of tumors without direct influence on tumor cells.
Li
Xi
X
School of Life Science, Nanjing University, 22 Hankou Road, Nanjing 210093, Jiangsu Province, China.
Fu
Geng-Feng
GF
Fan
Yan-Rong
YR
Shi
Chan-Fu
CF
Liu
Xin-Juan
XJ
Xu
Gen-Xing
GX
Wang
Jian-Jun
JJ
D000336:Aerosols; D000818:Animals; D003094:Collagen / Q000235:genetics*; D043169:Endostatins; D005260:Female; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D008081:Liposomes / Q000008:administration & dosage; D008114:Liver Neoplasms, Experimental / Q000098:blood supply* / Q000473:pathology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009389:Neovascularization, Pathologic / Q000517:prevention & control*; D010446:Peptide Fragments / Q000235:genetics*; D014168:Transferrin / Q000008:administration & dosage
D000336:Aerosols; D043169:Endostatins; D008081:Liposomes; D010446:Peptide Fragments; D014168:Transferrin; D003094:Collagen