Prostatic radiation therapy increases neuronal apoptosis, decreases nitrergic neurons and markedly impairs neuritogenesis in the major pelvic ganglion
Powers, SA1; Odom, MR1; Moomaw, MA1; Pak, ES1; Ashcraft, KA2; Koontz, BF2; Hannan, JL1
1: East Carolina University, United States; 2: Duke University, United States
Objective: Prostatic radiation therapy (RT) is presumed to cause erectile dysfunction (ED) through damage to the vasculature and the nerves supplying the penis. This study examined the impact of in vivo prostatic RT on 1) vascular function in the internal pudendal artery (IPA) and penis, 2) ex vivo survival and growth of major pelvic ganglia (MPG) neurons, and 3) neuronal markers of inflammation, regeneration and injury.
Materials and Methods: Adult male Sprague-Dawley rats underwent conformal single fraction 22Gy RT using small animal microirradiator to the prostate or sham treatment. Erections (ICP/MAP) were assessed by electrical stimulation of the cavernous nerve at 2 or 10 weeks post-RT (n=10/grp). MPGs were excised and neurons were dissociated and cultured. Axon length and branching, and neuronal nitric oxide synthase (nNOS), tyrosine hydroxylase (TH), ninjurin-1 (Ninj1, neuroinflammation marker), Rac-1 (role in dendritic spine elongation) and TUNEL assay expression were measured at 72h. IPA and penises were mounted in myograph. Concentration response curves to phenylephrine (PE), acetylcholine (ACh), and electrical field stimulated (EFS) contraction and NANC relaxation were assessed. MPG gene expression of nNOS, TH, beta-tubulin, GFAP (Schwann cell marker), GAP43 (regeneration marker), ATF3 (injury marker), Ninj1 and Rac-1 was measured.
Results: Overall, there was no change in vascular reactivity in the penises or IPA post-RT. Additionally, erectile function was not impaired at either time point. In neuron cultures, there was an early increase in length, while branching and nNOS positive neurons decreased (p<0.05). At 10 weeks post-RT, there was a 20% decrease in neuron length, decreased neuron branching, and 20-30% less nNOS and TH positive neurons (p<0.05). Rac-1 expression was decreased and Ninj1 expression was increased 10 weeks post-RT. Additionally at both time points, there was a 2-2.5 fold increase in TUNEL positive apoptotic neurons (p<0.05). Gene expression of Ninj1 and GFAP were increased early and late post-RT, while nNOS and GAP43 were decreased at 10 weeks post-RT.
Conclusions: While vascular changes were not evident in these animals, pelvic neurons displayed markedly impaired neuritogenesis, neuron survival and decreased nNOS. Schwann cells and neuroinflammatory protein Ninj1 were elevated post-RT and Rac-1 which plays a role in neuritogenesis and branching was decreased. These are potential novel targets to prevent pelvic nerve damage that contributes to RT-induced ED.
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