Scar tissue was removed and the gap between the rostral and caudal stumps was filled with pieces of respiratory (2WDC and 4WDC groups) or olfactory (2WDT and 4WDT groups) lamina propria (Fig. 7A, right). A piece of hemostatic sponge (Hemospon, Technew, São Paulo—SP, Brazil) was placed over the transplantation site to ensure blood homeostasis. Again, muscle and skin layers were sutured and post-operatory care was maintained

as previously described. Approximately 18 weeks after spinal cord injury, the viability of grafted tissue was demonstrated http://www.selleckchem.com/products/Everolimus(RAD001).html by the presence of fusiform-shaped OECs immunoreactive for p75NTR, S-100 and GFAP at the site of spinal cord transection (Fig. 8C). RLP control grafts continued to be devoid of OECs, as confirmed by the lack of cells expressing the three markers used in the lesion area (Fig. 8D). C59 wnt concentration Hindlimb motor function was assessed using the BBB locomotor rating (Basso et al., 1996).

This scale is qualitative, widely used and designated to assess the functional recovery of hindlimbs after lesions in thoracic spinal cord. The score of this scale ranges from 0 (no hindlimb movement) to 21 (normal movement of the hindlimbs). In this study, BBB assessment was accomplished preoperatively (naive) and postoperatively after the SCI (at days 5, 20, 35, 50, 65, 80 post-injury for the AC and AT groups; at days 5, 20, 35, 50, 65, 80, 95 post-injury for the 2WDC and 2WDT groups; and at days 5, 20, 35, 50, 65, 80, 95, 110 post-injury for the 4WDC and 4WDT groups). For each test, rats were placed in an open-field (60 × 30 × 40 cm) for 5 min. The test session was recorded with a video camera (Sony Handycam DCR-SR88, São Paulo—SP, Brazil) to allow later analysis by a blinded lambrolizumab observer. The scores of the left and right hindlimbs were averaged and taken as the BBB score of each animal. At the end of behavioral analysis, rats were anesthetized as described above. An

incision was made at the T12 vertebrae level to expose the spinal cord below the SCI site. After a laminectomy, FG retrograde tracer (2% dextran tetramethylenerhodamine, Biotium Inc., Hayward—CA, USA) was injected using a stereotaxic apparatus (Insight, Ribeirão Preto—SP, Brazil) coupled to a 1 μL Hamilton syringe (Hamilton Company, Reno—NV, USA). Three injections of FG (0.05 μL, 1 min duration each) were made at midline (0.5, 0.8 and 1.5 mm deep) and 1 mm laterally (0.5, 0.8 and 1.2 mm deep) in each side of this spinal cord level (Steward et al., 2006). Post-operatory care was done as previously described. One week after the retrograde tracing injections, rats received an overdose of pentobarbital (100 mg/kg body weight, i.p., Cristália, São Paulo—SP, Brazil) and were transcardially perfused with saline solution and buffered 4% paraformaldehyde (pH 7.4) using a peristaltic pump (30 mL/min, Milan Equipamentos Científicos, Colombo—PR, Brazil).