Tiam Mana Saffari

72 CHAPTER 4 vascular tree, a closer-meshed network of smaller vessels, contrast enhanced micro CT can become a powerful tool in quantifying angiogenesis 13 . Information collected from conventional photography may complement the micro CT. Photographs could be analyzed by measuring the ratio of vessel area and total nerve area digitally 23 . More conventional methods such as manual vessel counts per sub- segment of the nerve, providing a vessel density per mm 2 , have also been described 24 . These techniques have never been verified and are questionably representative for the entire quantification of the nerve vasculature when used solely. However, with evolving technology, the quality of conventional photography has improved and there are numerous photo editing software available providing a cost-effective way to measure vessel and nerve surface areas. The purpose of this study was to describe three-dimensional visualization of the vascular system in the rat sciatic nerve and to quantify angiogenesis of nerve reconstruction. The micro CT and conventional photography were used to objectively quantify vascular volume and vascular surface area, respectively, as measurements of angiogenesis in rat nerve. These methods will allow us to advance angiogenesis related research by improving the tools for studying and understanding vascular development and the mechanisms of neoangiogenesis. MATERIALS AND METHODS Animals Animal experiments were approved by our Institutional Animal Care and Use Committee (IACUC A3348-18). For this study, a total of 12 male Lewis rats (Envigo, USA) were used, weighing between 250 – 300 grams, in which 10 mm unilateral sciatic nerve gaps were repaired with a reverse autologous graft. All animals were housed with ad libitum access to food and water, with a 12-hour light-dark cycle after surgery. Experimental design A unilateral 10 mm sciatic nerve gap was repaired with an ipsilateral reversed autologous nerve graft to create a mismatch in the alignment of the nerve gap 25,26 .