A four-year-old female leopard gecko E. macularius suffered from a subcutaneous tumor in the occipito-pterional portion behind its right eye. The subject weighed 35 g at the date of surgery and was fed daily with commercial mealworms supplemented with calcium and vitamin D powder and housed in a 60 (width) x 30 (depth) x 35 (height) cm3 acryl cage, equipped with a floor panel heater, calcium-containing sand bedding, and a 75 W night-light lamp throughout the observation. Room temperature was maintained at 27–30 °C, with the humidity at 60–80 %.
Surgical operation
Upon first inspection, the tumor mass (T in Fig. 1a and b) was palpable, motile, and smooth but had already developed to an ulcerated lesion. The appetite, activity and consciousness level were normal. The wound was bleeding, and the rear edge of the tumor was cracked and fused to the molting skin (Fig. 1a). Thus, an immediate surgical resection was performed under general anesthesia. A mixture of the following anesthetics was used: 8.0 mL midazolam (Dormicam, Astellas Pharma Inc., Tokyo, Japan), 1.5 mL medetomidine hydrochloride and 0.4 mL butorphanol tartrate (Domitol and Vetorphale, Meiji Seika Pharma Co., Ltd., Japan) was adjusted to 30 mL with normal saline. Prior to the operation, the patient was calmed in a dark box, and 100 μL of the anesthetic mixture was slowly injected subcutaneously twice within ten minutes (200 μL total) into the lateral part of the right femur using a 30-G needle (Terumo, Japan).
The incision was made from the crack at the outer perimeter of the ulcerated skin at the occipito-pterional area (Fig. 1b). The skin flap was partially trimmed with scissors to preserve five pigmented regions in the margin (red arrows; Fig. 1c). The isolated mass was easily enucleated. Note that no communicating fistula was found towards the ocular region (Fig. 1c). The wound was rinsed twice with saline and sutured using a 14 mm curved needle with a 7–0 nylon string (Nazme Seisakusho Co., Ltd., Japan).
The exposed sub-dermal area (E in Fig. 1c) was about 46 mm2 large, and the remaining skin flap (s in Fig. 1a) was about 6 mm2 large without tension. Consequently, the skin flap was sutured with a sufficient margin from the edge of the incision (Fig. 1d) because the shortened skin needed extensive stretching to cover as much lesion area as possible (Fig. 1e). The uncovered wound (about 2 mm2) was plastered with artificial ointment containing Vaseline, saline, and gentamicin (50 μg/mL) to avoid infection and dehydration (Fig. 1f).
A total of 100 μL gentamicin solution (100 μg/mL) was also orally administered twice a day for one post-operative week. The post-operative course was stable and uneventful. The dermal wound epithelium was rapidly regenerated by post-operative day 5 as previously reported [5], but the pigmented spots had not changed much (Fig. 2a). One month following the surgery, the subject underwent the first post-operative molting (Fig. 2b), and the pigmented spots had grown from the remaining pigments with a little bit blurred perimeter (Fig. 2c and d). In the four months since the surgery, the regenerated spots looked almost undistinguishable from the pre-existent spots with a rather vivid border, and the epidermal scales with yellow coloration were restored (Fig. 2e and f). The delayed yellowish coloration should be a general recovery course of the epidermis in E. macularius [4].
Followed up for three more months, there were no more changes in the wound healing process in the operated area (Fig. 3a). The newly pigmented spots were distributed evenly and the appearance looked rather natural compared to the un-operated side (Fig. 3b). The regenerated area surrounded by the imaginary borders of the surgical margin (yellow dotted line: Fig. 3c, d and g) was about 20 mm2. On higher magnification, the skin in E. macularius is tessellated with two different types of scales (Fig. 3e and f): small polygonal scales and cone-like large interspersing scales with round perimeters (defined as tubercles). The tubercles are rather flattened at the occipito-pterional region (pink asterisks in Fig.ure 3e) compared to those on the trunk (pink asterisks in Fig. 3f). The regenerated skin comprised of the small mosaic scales with no obvious scar as reported previously [3–5]. However, much of the large cone-like tubercles were not regenerated but only sparsely located in the operated side (pink asterisks in Fig. 3g), whereas such tubercles were evenly distributed in the intact side (pink asterisks in Fig. 3h). But, it is noteworthy that two tubercles were newly formed in the dorsal area of the operated region (green asterisks in Fig. 3d and g: compare with Fig. 1c).
