Histological evaluation

After tail harvest, the specimens of caudal vertebrae and discs were fixed with 10% buffered neutral formalin, then decalcified in Surgipath Decalcifier II solution, followed by embedding in paraffin. Slides with the dried paraffin-embedded sections were deparaffinized in xylene, and rehydrated with a series of ethanol washes. The sections were stained with hematoxylin and eosin and Masson's trichrome before they were examined under light microscopy. The histological assessment was evaluated by two different research fellows under the guidance of an experienced pathologist, in accordance with the grading system of Emery et al. [15]. The fusion status was graded using a histological score of 0–7: score 0, empty islets; score 1, fibrosis tissue only; score 2, fibrosis tissue more than fibrocartilage tissue; score 3, fibrocartilage tissue more than fibrosis tissue; score 4, fibrocartilage tissue only; score 5, fibrocartilage tissue more than bony tissue; score 6, bone tissue more than fibrocartilage tissue and fibrosis tissue; score 7, bone tissue only.