Artiss

To ensure a spatial localization of the transplanted cells, we decided to use a clinically approved fibrin hydrogel (ARTISS®, Baxter, Deerfield, IL, USA), as preliminary results showed a high biocompatibility and metabolic activity over a long culture period in vitro (data not shown). Cell-loaded conduits with a length of 25 mm and a 2 mm wall thickness were prepared by mixing 3 × 10⁶ cells in standard medium and thrombin solution at a 1:10 ratio. Afterwards the fibrinogen solution was allowed to polymerize in a sterile syringe with a centered metal rod to create a 2 mm lumen. Initial polymerization was carried out for 30 minutes, and after the removal from the syringe, the fibrin conduit was hardened for two hours in complete medium, before gently opening it longitudinally with micro scissors in order to be put around the nerve autograft (Figure 1G). Pre-operative anesthesia was initiated by intramuscular injection of 0.02 mg/kg fentanyl (Janssen, Germany), 1.0 mg/kg midozilam (Ratiopharm, Ulm, Germany) and 0.2 mg/kg medetomidin (Orion, Espoo, Finnland). Anesthesia was post operatively antagonized with 0.03 mg/kg naloxone (Bristol myers Squibb, New York, NY, USA), 0.1 mg/kg flumazenile (Roche, Basel, Switzerland) and 1.0 mg/kg atipamezole (cp-pharma, Burgdorf, Germany).

To create cell seeded fibrin conduits (25 mm length, 2 mm wall thickness, 2 mm inner diameter), we used a clinically approved and commercially available two-component fibrin sealant (ARTISS, Baxter, Deerfield, IL, USA) as previously described (Saller et al., 2018). In brief, cells were trypsinized (0.5% trypsin, Merck, Darmstadt, Germany) and counted, and 3 × 10⁶ were resuspended in standard culture medium and mixed in a 1:10 ratio with thrombin. The conduit was prepared in sterile syringes with a metal wire in the middle. After a polymerization period of 30 minutes at 37°C in an incubator, the cell seeded conduits were applied during surgery as described above.