Naked DNA expressing two isoforms of hepatocyte growth factor induces collateral artery augmentation in a rabbit model of limb ischemia.
Hepatocyte growth factor (HGF) has been shown to induce angiogenesis in vivo and has potential as a candidate gene for 'therapeutic angiogenesis'. In vivo, two isoforms of HGF, HGF₇₂₃ and HGF₇₂₈, consisting of 723 and 728 amino acids, are generated through alternative splicing between exons 4 and 5, but the biological effects of their coexpression have not yet been elucidated. In this study, we generated a series of genomic-complementary DNA (cDNA) hybrids of the HGF gene by inserting various truncated intron 4 into the junction of exons 4 and 5 of HGF cDNA and analyzed the biological activities of these hybrid constructs. We showed that: (1) the hybrid called HGF-X7, which contained 1502 base pairs of intron 4, could drive a higher level of HGF expression than other hybrid constructs and cDNAs of each isoform alone; (2) the pCK vector was most efficient for the gene expression of HGF-X7; (3) coexpression of both isoforms of HGF could more efficiently induce the migration of human umbilical vein endothelial cell (HUVEC) and of the mouse myoblast cell line C₂C₁₂ myoblasts than a single isoform of HGF and human vascular endothelial growth factor (VEGF)₁₆₅ at a given concentration; (4) intramuscular administration of pCK-HGF-X7 resulted in transient and localized HGF expression in the injected muscle without an increase in the HGF protein levels in other tissues including serum; and (5) intramuscular injection of pCK-HGF-X7 could more efficiently increase the number of angiographically recognizable collateral vessels, as well as improve an intra-arterial Doppler wire-measured blood flow in the rabbit model of hindlimb ischemia when compared with the identical vector encoding VEGF₁₆₅ gene. These results showed that transfer of the genomic-cDNA hybrid of the HGF gene could be used as a potential therapeutic approach to human vascular diseases.
Pyun
W-B
WB
Department of Cardiology, Medical College of Ewha Womans University, Mokdong Hospital, Seoul, Korea.
Hahn
W
W
Kim
D-S
DS
Yoo
W-S
WS
Lee
S-D
SD
Won
J-H
JH
Rho
B-S
BS
Park
Z-Y
ZY
Kim
J-M
JM
Kim
S
S
D000818:Animals; D001158:Arteries / Q000254:growth & development / Q000378:metabolism; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D003097:Collateral Circulation / Q000187:drug effects*; D004247:DNA / Q000235:genetics / Q000627:therapeutic use*; D018076:DNA, Complementary / Q000235:genetics; D004195:Disease Models, Animal; D005121:Extremities / Q000098:blood supply; D005260:Female; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005818:Genetic Engineering; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics; D017228:Hepatocyte Growth Factor / Q000235:genetics* / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D007438:Introns / Q000235:genetics; D007511:Ischemia / Q000503:physiopathology / Q000628:therapy*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D012039:Regional Blood Flow / Q000187:drug effects
D018076:DNA, Complementary; D020033:Protein Isoforms; D017228:Hepatocyte Growth Factor; D004247:DNA