Effects Of Adenovirus-mediated BDNF Gene Transfer Into Spinal Cord To Treat Nerve Root Injury And Its Mechanism

Nerve root injury (NRI) is generally devastating which can result in disability of limb.The treatment of nerve root injury has not been satisfactorying.The bad prognosis of nerve root injury due to two main factors:1. Proximal stump of the injury root which close to the soma contain less schwann cells and side branches. There is a disruption in the supply of neurotrophic factors coming from Schwann cell and target-organ ,leading to mass death of the motor neurons and lack of regeneration. 2. The central processes of dorsal root ganglion within the dorsal roots fail to regenerate through the dorsal root entry zone and into the spinal cord.Axonal growth is apparently inhibited in this region. This appears in part due to the presence of neurite growth-inhibiting molecules secreted by glial cell. From this two points we know that the key reason of bad prognosis after nerve root injury is in the relevant spinal cord segment, so the fundamentality of treat should be in the relevant spinal cord segment .With the better understanding of regeneration from neurons and further refinement of instruments ,nerve root repair at the level of spinal cord appeared as a new but challenging technique.Brain-derived neurotrophic factor(BDNF) is an important neurotrophic factors which has motor and sensory activity. However, the application of exogenic BDNF on clinic was limited due to their short plasmic half-life, the block of blood nerve barrier and the transient expression within the host cells.In order to maintain a suitable supply for a sufficient length of time,treatment strategies should be based on gene transfer which would deliver critical exogenous gene to neural cells through the vector of recombinant adenovirus.It is well know that large diameter afferent neurons whose central processes normally end on motor neurons in the grey matter of the ventral horn. We project the delivery of an adenovirus-mediated BDNF gene into the grey matter of the ventral horn,hoping this project can not only prevend neurons from death but also promote the regeneration of dorsal root axons into the spinal cord and restore the spinal reflex arc.Nerve root transection and reanastomosis was used as the experiment model. Base on which, the vector of recombinant adenovirus-mediated brain-derived neurotrophic factor was transfered into spinal ventral horn through microinjection in a stereotaxic frame. Histopathologic observation, electrophsiologic test, immunocytochemistry.in situ hybridization,Tunnel reaction, sciatic nerve function index were used to investigate the effects and mechanisms of nerve root reconstruction combining BDNF gene therapy to treat nerve root injury.The Major results and conclusions are showed as follow:1. It has been confirmed that exogenous BDNF gene existed in the genome of the RAD we got, and the RAD holds bioactivity in vitro experiments.2. Adenovirus-mediated BDNF gene transfer into spinal cord could infect the spinal cord tissue, significantly increasing their BDNF gene expression level, and inducing trk-B receptor expression upwards.3. Treatment with AxCA-BDNF significantly prevented the loss of lesioned ventral horn motoneurons after nerve root injury.4. Nerve root injury induced the expression of caspaseS and NOS that triggers the procedure of apoptosis. It was showed that RAD mediated exogenous BDNF gene transfer not only inhibited caspase3 and NOS expression, but also increased bcl-2 expression, companing with the decreased apoptotic cells. RAD-mediated exogenous BDNF gene transfer protects neurons from delayed death by the mechanisms of blocking neuronal apoptosis.5. The activities of acetylcholinesterase in AxCA-BDNF group was highter while acid phosphatase was lower than injury group. AxCA-BDNF decreased the change of enzyme, protecting neurons from damage.6. Over-expression of BDNF could induces the injured axon regeneration into the ventral roots and enhanced the regeneration of dorsal root into spianl cord.7. Treatment with AxCA-BDNF cou...