168 Targeted Depletion of the Enzyme Fidgetin-like 2 Promotes Axon Regeneration and Improves Erectile Function Outcomes in a Rodent Model of Radical Prostatectomy

Abstract

The majority of men undergoing radical prostatectomy (RP) suffer from erectile dysfunction (ED) after the surgery due to damage to the cavernous nerves (CN). We recently discovered that the enzyme fidgetin-like 2 (FL2), a negative regulator of the microtubule (MT) cytoskeleton, can be targeted in neurons to enhance axon regeneration. Here, we used a rodent model of RP to test the hypothesis that reducing levels of FL2 after CN injury will promote axon regeneration and recovery of erectile function. To better understand the mechanism by which FL2 regulates axon growth, we characterized changes in neurite outgrowth and the axonal microtubule array of adult rodent dorsal root ganglion (DRG) neurons with FL2 depletion in vitro. Three rat models of CN injury were used: mild (a smooth clamp applied for 2 minutes to the CN), moderate (a serrated clamp applied for 4 minutes to the CN) and severe (CN transection). Immediately after injury, two formulations (nanoparticle or liposomal) for delivery of FL2-siRNA or control-siRNA were applied. Erectile function was assessed at several time points after injury by measuring the intracorporal pressure/blood pressure (ICP/BP) ratio following electro-stimulation of the CN. To further understand the mechanism by which FL2 depletion promotes axon regeneration, dissociated adult rodent dorsal root ganglion (DRG) neurons were transduced with AAV5 containing a plasmid encoding FL2 or scrambled small hairpin RNA (shRNA) and a GFP reporter. 5 days after treatment, neurons were replated for live monitoring of neurite growth, then fixed and immunostained for microtubules.