Abstract
Background
Impaired wound healing is a frequent phenomenon in diabetes mellitus. However, little is known of the fundamental cause of this pathology. The present study examined the effect of human insulin‐like growth factor (hIGF)‐1 overexpression in combination with autologous cell transplantation to diabetic wounds in a preclinical large‐animal model.
Methods
Diabetes was induced in Yorkshire pigs with streptozotocin. Keratinocytes were cultured and transfected with hIGF‐1 or LacZ transgene. Plasmids were lipoplexed with either Lipofectin or Lipofectamin 2000. Transgene expression was assessed by enzyme‐linked immunosorbent assay or X‐gal staining. For in vivo studies, full‐thickness wounds were created and dressed with a sealed chamber. Transfected cells were transplanted into the wounds. Wound contraction was monitored and biopsies were obtained for measurement of re‐epithelialization. Wound fluid was collected and analysed for IGF‐1 concentrations.
Results
Quantification showed up to 740 ng/ml IGF‐1 in vitro and significantly higher concentrations over 14 days compared to controls for the Lipofectamin 2000 group. Lipofectin‐mediated gene transfer showed peak expression on day 2 with 68.5 ng/ml. In vivo, transfected cells showed peak expression of 457 ng/ml at day 1, followed by subsequent decline to 5 ng/ml on day 12 with Lipofectamin 2000. For Lipofectin, no significant IGF‐1 expression could be detected. Gene therapy caused significantly faster wound closure (83%) than both controls (native‐cell therapy = 57%; control wounds = 32%).
Conclusions
The present study demonstrates that optimized nonviral gene transfer increased IGF‐1 expression in diabetic wounds by up to 900‐fold. This high IGF‐1 concentration in combination with cell therapy improved diabetic wound healing significantly. Copyright © 2008 John Wiley & Sons, Ltd.