Controlled release of small interfering RNA targeting midkine attenuates intimal hyperplasia in vein grafts.
OBJECTIVE
Intimal hyperplasia is a major obstacle to patency after vein grafting. Despite of a diverse array of trials to prevent it, a satisfactory therapeutic strategy for clinical use has not been established. However, sufficient inhibition of early stages of intimal hyperplasia may prevent this long-term progressive disease. Midkine (MK) is a heparin-binding growth factor that was originally discovered as the product of a retinoic acid-responsive gene. We previously demonstrated that MK-deficient mice exhibit a striking reduction of neointima formation in a restenosis model, which is reversed on systemic MK administration. In this study, we evaluated a strategy of using small interfering RNA (siRNA) targeting MK as a therapy for vein graft failure.


METHODS
We first made a highly effective siRNA to rabbit MK. Jugular vein-to-carotid artery interposition vein grafts, which are applied to a low flow condition, were made in Japanese white rabbits. Small interfering RNA mixed with atelocollagen was administrated to the external wall of grafted veins. Cy3-conjugated stabilized siRNA was used to confirm its stability and successful transfer into the vein graft wall. Neointimal hyperplasia was evaluated 4 weeks after the operation. The proliferation index and leukocyte infiltration were determined.


RESULTS
MK expression was induced and reached the maximum level 7 days after operation. Fluorescence of Cy3-labeled siRNA could be detected in the graft wall even 7 days after operation. Knockdown of the gradually increasing expression was achieved by perivascular application of siRNA using atelocollagen. The intima-media ratio and the intima thickness at 28 days after grafting were both reduced >90% by this treatment compared with controls. This phenomenon was preceded by significant reductions of inflammatory cell recruitment to the vessel walls and subsequent cell proliferation in MK siRNA-treated grafts.


CONCLUSIONS
These results suggest that midkine is a candidate molecular target for preventing vein graft failure. Furthermore, for clinical applications of siRNA, a single intraoperative atelocollagen-based nonviral delivery method could be a reliable approach to achieve maximal function of siRNA in vivo. This strategy may be a useful and practical form of gene therapy against human vein graft failure.
Banno
Hiroshi
H
Department of Biochemistry, Nagoya University Graduate School of Medicine, Nagoya, Japan.
Takei
Yoshifumi
Y
Muramatsu
Takashi
T
Komori
Kimihiro
K
Kadomatsu
Kenji
K
D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology; D016207:Cytokines / Q000378:metabolism*; D004337:Drug Carriers; D006084:Graft Rejection; D006965:Hyperplasia; D000078792:Midkine; D009414:Nerve Growth Factors / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000008:administration & dosage*; D011817:Rabbits; D014162:Transfection; D017539:Tunica Intima / Q000473:pathology*; D014680:Veins / Q000637:transplantation*
D016207:Cytokines; D004337:Drug Carriers; D009414:Nerve Growth Factors; D034741:RNA, Small Interfering; C059821:atelocollagen; D000078792:Midkine; D003094:Collagen