| Abstract No.: | A-D1150 |
| Country: | Canada |
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| Title: | PHOSPHATASE AND TENSIN HOMOLOGUE DELETED ON CHROMOSOME TEN (PTEN): A POSSIBLE ROLE IN PERIPHERAL NEURON INJURY AND REGENERATION |
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| Authors/Affiliations: | 1 Kim Vanneste*; 1 Christine Webber; 1 Jose Martinez; 1 Douglas Zochodne;
1 University of Calgary, AB, Canada
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| Content: | Background: The phosphoinositide 3-kinase (PI3-K) pathway is involved in cell growth and survival, and it also promotes neurite extension in embryonic dorsal root ganglia (DRG) and hippocampal neuron cultures. Following PI3-K activation, an intracellular signalling cascade is initiated activating downstream molecules such as phosphatase and tensin homologue deleted on chromosome ten (PTEN) and Akt. PTEN inhibits the PI3-K pathway by hydrolyzing phosphatidylinositol 3,4,5 triphosphate (PIP3) thereby decreasing Akt activation.
Objective: To determine the expression of PTEN and Akt in the sciatic nerve and DRG both in vivo and in vitro in normal and injured adult rats and to establish the impact of PTEN inhibition in vitro.
Materials and Methods: The sciatic nerve of adult male 200-250g Sprague-Dawley rats were cut at mid-thigh level for preconditioning and the DRG neurons and their regenerating nerve stumps were analyzed at three or seven days and compared to intact age-matched control animals. For expression experiments qRT-PCR, western blot and immunohistochemistry techniques were used to measure mRNA and protein levels. For in vitro experiments, rats were injured as above or sham injured and the DRGs were harvested and placed into culture three days after the nerve injury. At the time of plating, 0, 10, 50 or 200 nM of the PTEN pharmacological inhibitor, bpv(pic), was added to the culture medium. The cells were grown overnight and then fixed and processed for immunocytochemistry. Neurite extension was analyzed by MetaXpress software.
Results: Western blot and immunohistochemistry demonstrated expression of both total and phosphorylated PTEN and Akt in both DRGs and sciatic nerves. Adult sensory neurons and associated glial cells studied in vitro also expressed PTEN. Interestingly, three days following a sciatic nerve transection, PTEN and Akt mRNA levels decreased in DRG neurons. Immunohistochemistry identified that the proportion of DRG neurons expressing total and phosphorylated PTEN also decreased following a three day nerve injury. Despite its down-regulation, this pathway remained active in adult sensory neurons since the PTEN inhibitor was associated with rises in neurite outgrowth, observed both in injured and non-preconditioned neurons.
Conclusions: Overall, our findings confirm the presence of major players in the PI3-K pathway for neurite extension in peripheral neurons. In particular, PTEN, not previously identified in these neurons may be important by inhibiting this pathway and attenuating regeneration. Interrupting its action may enhance the growth of regenerating axons in vitro.
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