S100b solution

The intraocular S100B injections were carried out as described previously [7 (link), 8 ]. 0.4 μg S100B solution was injected (Sigma Aldrich, St. Louis, MO, USA) in one eye (43 eyes in total). The control group received 2 μl of phosphate-buffered saline (PBS; Biochrome, Berlin, Germany, n = 14 eyes). The contralateral eyes were used as naïve controls (n = 14 eyes; Fig. 1a).
Fig. 1

Study timeline. In this study, two different concentrations of minocycline (25 mg/kg body weight; 50 mg/kg body weight) were intraperitoneally injected at day -1. S100B or its solvent PBS were intravitreally applied at day 0. The untreated eyes were used as naïve controls. Minocycline was then administered daily (13.5 mg/kg body weight; 25 mg/kg body weight) from day 1 to 14. Five groups were generated in this study: naïve: untreated (n = 14); PBS: intravitreal PBS injection (n = 14); S100B: intravitreal S100B injection (n = 13); 13.5 mg/kg mino: low minocycline dose of 13.5 mg/kg + S100B (n = 15); 25 mg/kg mino: high minocycline dose of 25 mg/kg + S100B (n = 15). After 14 days, Electroretinogramm (ERG) measurements were carried out and the eyes and optic nerves were obtained for further analysis, including (immuno-)histology and proteomics

Wistar rats received an intraocular S100B injection (n = 6–7/group). Before intraocular treatment, animals were anesthetized with a mixture of ketamine (50 mg/mL, Ratiopharm, Ulm, Germany) and xylazine (2%, Bayer Health Care, Leverkusen, Germany) followed by a topical anesthetic (Conjuncain, 4 mg/mL, Bausch & Lomb, Rochester, NY, USA) and a mydriatic to dilate the pupil (Tropicamide, 5 mg/mL, Pharma Stulln, Stulln, Germany). In the S100B group, 0.4 µg/µL S100B solution (Sigma-Aldrich) was injected in the vitreous of one eye with a 32-gauge needle (Hamilton, VWR, Radnor, PA, USA) under a stereomicroscope (Zeiss, Jena, Germany) [29 (link)]. Controls received 2 µL phosphate-buffered saline (PBS; Biochrome), which also functioned as a solvent for S100B. After the injection, the eyes were treated with an antibiotic ointment (Floxal, Bausch & Lomb). Animals were examined 2 h after injection and on the next day. Once a week, the general behavior and look of the animals was examined Eye bleeding or cataracts were considered as early endpoints in this study.