Sodium pentobarbitone

After seven months of exposure, mice from both experimental groups were sacrificed by intraperitoneal injection of sodium pentobarbitone, 100 mg/kg body wt (Abbot Laboratories, Lake Forest, IL, USA). The lung tissue was removed from the cardiopulmonary block and the lungs were fixed by filling them with 4% buffered formalin solution at a constant airway pressure of 25 cmH₂O for 24 h. The fixed lungs were kept in the formalin solution until they were embedded in paraffin and sections 5-μm-thick were cut using a rotary microtome HM325 (Microm International GmbH). The gastrocnemius muscles were collected from the hind limbs using standardized dissection methods and cut into 5 μm of the samples embedded in paraffin and fixed with formalin.

At the time of humane killing, groups of four pigs were moved in an animal transport trailer a short distance from the home pen building to holding pens located within a nearby surgical and post-mortem facility. The pigs were weighed individually to determine the required drug dosages. For sedation, the pigs received an intramuscular injection in the neck of ketamine (5 mg/kg, Vetoquinol, Buckingham, UK), midazolam (0.5 mg/kg, Hameln, Gloucester, UK) and medetomidine (10 μg/kg, Vetoquinol) and were left undisturbed under dimmed light conditions for 10–15 min. Once each pig was sedated (i.e. immobile, absence of reaction to touch and human presence) the ear vein was catheterized. Pigs were killed humanely by injection of sodium pentobarbitone (150 mg/kg intravenously; Abbott Laboratories, Abbott Park, Illinois, USA). Death was confirmed by respiratory arrest and loss of corneal reflex. All pigs were exsanguinated by cutting the jugular and carotid arteries prior to post-mortem tissue collection.