The Anti–angiogenic Effect of Transplanted Amniotic Membrane on Alkalic Burn Models

Abstract

Abstract: : Purpose: To evaluate the anti–angiogenic effect of transplanted amniotic membrane(AM) on alkalic burn rabbit models Methods: Chemical burn model was established using 1N of NaOH for 60 seconds in 12 eyes of 12 rabbits. Four fresh AM, four cryopreserved AM or two lyophilized AM (AmniSite–Cornea®, Bioland, Korea) were transplanted into each cornea immediately after chemical burn. Transplanted AMs were maintained for 3 and 7 days before sacrifice. The length and the number of newly developed vessel were measured in total cornea using Image–Pro ® Plus (Version4.5, Media Cybernetics, USA). Neovacularization in each group was graded by multiplying of number of new vessels (1:0–10,2:11–15,3:16–30,4:>31) and length of new vessels (1:<3, 2:4–6.0, 3:7–9, 4:>10 mm) and compared it with that in the control. TIMP1, TIMP2 were measured using RT–PCR in a ratio of density of TIMP to density of betha actin with Image analyzer(Vilberlourmat) and densitometry(Tina 2.0, Raytest, Straubenhardt, Germany) in each transplanted AM for 3 and 7 days. Results: Neovascularization grading score was 4 in all transplanted group with fresh AM or cryopreserved AM or lyophilized AM and the grading was 16 in non–transplanted group after 3 days, respectively. It was significantly lower in transplanted group compare with that in control. The grading was the same between in 3 days– and in 7 days– transplantation group. Before transplantation, TIMP–1 and 2 were 1.56 and 0.55 in fresh AM, 1.46 and 0.92 in cryopreserved AM, 1.46 and 0.92 in lyophilized AM. 3 days after transplantation, TIMP–1 and 2 were 0.98 and 0.42 in fresh AM, 1.78 and 0.60 in cryopreserved AM, 0.32 and 0.61 in lyophilized AM. 7 days after transplantation, TIMP 1 and 2 were 0.29 and 0.11 in fresh AM, 0.56 and 0.54 in cryopreserved AM, 0.54 and 0.36 in lyophilized AM. The mean density of TIMP1 and 2 in 3 day–transplanted AM was not significantly different from that in pre–transplanted AM. It was significantly decreased in 7 day– transplanted AM compared with that in pre–transplanted AM Conclusions: Human amniotic membrane seemed to be effective on the suppression of neovascularization in burned rabbit eyes regardless of the preservation method. TIMP–1 might play a major role on anti–angiogenesis by maintaining effective concentration for the first 3 days in transplanted AM

Keywords: cornea: basic science • neovascularization • wound healing