917. GM-CSF Gene Transfer to Acute Limb Ischemic Mouse for Limb Recuperation

Abstract

IntroductionPeripheral arterial diseases may be caused by any occlusive leg artery lesion that interferes with blood flow. When these conditions occur, there is an increase in vessel resistance that can lead to a reduction in distal perfusion pressure and blood flow. This process forces collateral vessel remodeling and it is known as arteriogenesis. Inflammation is the main triggering factor of arteriogenesis and several molecular factors are involved. One way to prolong the arteriogenic activity is extending monocytes half-life by apoptosis inhibition with GM-CSF (granulocyte and macrophage colony stimulating factor). This factor is also responsible for proliferation and differentiation of inflammatory cells, which are important growth factors producers and chemoattractants for arteriogenesis.The main objective of this study was to assess the capacity of reverting surgery-induced acute limb ischemia in mice by in vivo transference of plasmid vector with GM-CSF gene and to compare with the VEGF gene transfer.Materials and MethodsTwo plasmid vectors were used in this study: uP-GM and uP- VEGF to express GM-CSF and VEGF, respectively. To induce acute limb ischemia in mice BALB/c, the femoral artery was excised from its origin of the external iliac artery to the distal point, where it bifurcates into saphenous and popliteal arteries, without damaging the vein and femoral nerve. All branches of femoral artery were ligated. Gene therapy was performed by injecting 100 μg of plasmid vector at the thigh soon after the ischemia surgery.Results and DiscussionAbout 30 % of animals that underwent ischemic surgery lost their limbs after 1 week, meanwhile none of uP-GM treated animals lost their limbs, even after 4 weeks of the surgery. Functional tests of the GM-CSF gene treated muscles revealed about 70 % of recovery of force, meanwhile the VEGF gene treated animals and the non- treated animals recovered only 25 % and 5 % of force respectively after 4 weeks of the gene transfer. Weight of gastrocnemius reached 100 % of recovery with ischemic animals treated with uP-GM after 4 weeks in comparison to the non-ischemic animals, but the VEGF gene treated or non-treated animals showed recovery of only 60 %. Capillary density had a significant increase in the VEGF treated animals after 1 week of surgery, but those treated with GM-CSF did not change in comparison to the non-treated ischemic animals, and this profile continued for 4 weeks. Histological data from the animals treated with GM-CSF for 4 weeks revealed that the muscle organization returned to the normal form.Therefore, we conclude that GM-CSF gene transference therapy in the ischemic muscle promoted good recovery of physical and physiological parameters in the long term meanwhile the use of VEGF gene promoted rapid recovery but it was not sustainable for 4 weeks.