Xylapan

All of the animals were humanely euthanized 24 h after the last treatment by exsanguination under Xylapan/Narketan anaesthesia (Narketan, Vetoquinol UK Ltd., Towcester, UK, 80 mg/kg b. w.; Xylapan, Vetoquinol UK Ltd., 12 mg/kg b. w., i.p.). A licensed veterinarian at IMROH examined gross pathological changes of the internal organs in all of the animals.
Testes and epididymis were dissected, cleaned from adhering matters, and rinsed in cold PBS buffer (without Ca²⁺ and Mg²⁺). The tissues were symmetrically bisected and weighed. The left testis/epididymis was stored in cryo-tubes without additional media or cryoprotectants, while the right testis/epididymis was symmetrically halved and stored separately. One-half was used for the measurements of antioxidant status parameters and the other one for element analysis. The tubes were immediately frozen in liquid nitrogen and stored at −80 °C until use.

At the end of the experiment, all animals were humanely euthanized by exsanguination under intraperitoneal Xylapan/Narketan anesthesia (Xylapan, Vetoquinol UK Ltd., Towcester, UK, 12 mg/kg bw i. p./Narketan, Vetoquinol UK Ltd., 80 mg/kg bw). A licensed veterinarian at IMROH examined the animals for gross pathological changes of the internal organs. The blood samples were collected in heparinized vacutainers by a dissection of the carotid artery under general anesthesia and further processed. One portion of whole blood was immediately used to prepare agarose microgels for the alkaline comet assay while the portion used for biochemical analyses was centrifuged (980× g, 10 min, at +4 °C) to separate red blood cells and plasma that were stored at −20 °C until further processing. Livers, kidneys, and brains were dissected out and weighted. Based on the obtained values, relative liver, kidney, and brain weight were calculated using the following formula:
Tissues were rinsed in cold PBS buffer (without Ca²⁺ and Mg²⁺), weighed, and divided into two portions intended for biochemical analyses and the comet assay. The samples used for biochemical analyses were immediately frozen in liquid nitrogen and stored at −80 °C until further processing while the tissue samples used for comet assay were prepared as described later in the text.

Twenty New Zealand White rabbits underwent bilateral posterolateral lumbar fusion (L5-L6) as described^{34 (link),37 (link)}. Animals were premedicated with a mixture of Xylazine (Xylapan^®, Vetoquinol, Switzerland) 5 mg/kg and Ketamine (Ketaminol^® Vetoquinol, Switzerland) 35 mg/kg applied intramuscularly. General anesthesia was maintained using a mixture of isoflurane (1–1.5%) and oxygen delivered by mask. A dorsal midline skin incision extending from L4 to L7 was made, followed by a paramedian fascial incision. Paravertebral muscles were retracted laterally, facilitating exposure of L5–L6 transverse processes. Exposed transverse processes were then decorticated using a high-speed burr, and Osteogrow-C implants were placed bilaterally in the gutter between L5–L6 transverse processes (Fig. 8). The fascial and skin incisions were closed with 3‐0 synthetic glycolide/lactide copolymer absorbable sutures in a continuous fashion. There were no adverse effects during the follow-up period. Experimental animals were euthanized 14 or 27 weeks following Osteogrow-C implantation using premedication of 3 mg/kg xylazine and 20 mg/kg ketamine i.m. and administration of T61 (1 ml/kg) i.v. Rabbit lumbar spine was harvested to conduct analyses described in the following sections.