ANALYSIS OF MECHANISMS OF NEURONAL CELL DEATH USING ADENOVIRUS VECTOR SYSTEM

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Aims of this project were (I) to establish adenovirus vector system that is capable of introducing genes into neuronal cells of the chick embryo, and (II) to analyze effects of Bcl-2 and Crm A on neuronal cell death in vitro and in vivo. Major results are the following :(1) In the first trial, we failed to obtain recombinant adenovirus that express Bcl-2 under regulation of CAG promoter. Employing Cre-LoxP system that can regulate expression of genes, we succeeded to obtain the recombinant adenovirus that express Bcl-2.(2) It was confirmed that genes introduced by adenovirus vecotrs were expressed in the chick DRG neurons and chick motoneurons in vitro. Introduced human Bcl-2 could rescue DRG neurons and motoneurons from cell death in vitro.(3) It is also confirmed that adenovirus vectors injected into leg bud were retrogradely transported to the motoneurons in the lateral motor column. Bcl-2 introduced with this retrograde transport could rescue some motoneurons after limb bud removal.(4) In the rat C6 glioma cell line system, human Bcl-2 introduced by adenovirus vector induced cell death.(5) We could not find rescuing effects of Crm A gene introduced by adenovirus vector in the DRG neuron system.