The inventors down regulated Smad 2/3 (an inhibitor of neuro-regeneration) in vivo via delivery of siRNA to the spinal cord using catheters.
Inhibiting smad signaling promotes neuron regeneration.
Inventors: Fan Wang, Zhigang He
USPTO Application #: 20080031911
Inhibition of Smad2/3 Signaling Promotes Axonal Regeneration after Spinal Injury in Rats-Gene Expression Knockdown in vivo!
One hour after the spinal cord is lesioned, the rats in the SB-505124 group receive a bolus injection of SB-505124 (30 mg/kg) in 0.9% saline administered via a tail vein. The treatment is repeated every 24 hours on days 1 through 7 post-lesion. Vehicle only control rats undergo the same treatment but are injected with an equal volume of 0.09% saline in a tail vein. At the same treatment time points as the SB-505124 group, the Smad2/3 siRNA group and corresponding controls receive 10 .mu.l rat Smad2/3 siRNA (Dharmacon, Lafayette, Colo.), mismatch siRNA, or transfection reagent only delivered to the spinal cord via the catheters. The siRNA (or mismatch siRNA control) complexes are prepared immediately prior to administration by mixing the RNA solution (200 .mu.M in annealing buffer) with a transfection reagent, i-Fect ™ . (Neuromics, Edina, Minn.), in a ratio of 1:4 (w:v). At this ratio, the final concentration of RNA as an RNA/lipid complex is 2 .mu.g in 10 .mu.l.