The Influence Of Borna Disease Virus Phosphoprotein On Proliferation Of PC-12 Cells And Expression Of Tyrosine Hydroxylase

Objectives: Borna Disease Virus is a neurotropic, nonsegmented negative-strand RNA virus. Originally described as a disease of horses, Borna disease has also been found in sheep, llamas, ostriches, cats and cattle. Considering an even larger variety of species has been infected experimentally, the host range is likely to include all warm-blooded animals. Previous study suggested that Borna Disease Virus is related to many diseases, such as human psychiatric disease, multiple sclerosis, Guillain-Barre syndrome, autism and obesity. But we have little knowledge about the definite relationship and pathogenic mechanism. Borna Disease Virus phosphoprotein is a main contributor to BDV-induced neuronal abnormalities, it is rich in the brain of infected animal and it act as a central protein in the replication and transcription of virus particle. This study was aimed to construct a cell model expressing Borna Disease Virus phosphoprotein persistently, to investigate the influence of Borna Disease Virus on neuron cell proliferation and tyrosine hydroxylase expression in PC-12 cells, to provide a new insight to explore the pathogenic mechanism of Borna Disease.Methods:1. pEGFP-N1-P24 plasmid with Borna Disease Virus P24 fragment was transfected into PC-12 cells by positive ion liposome. Selected by G418, expression of Borna Disease Virus phosphoprotein in PC-12 cells was identified by both fluorescent microscope and polymerase chain reaction.2. Cells transfected with pEGFP-N1-P24 plasmid were compared with cells transfected pEGFP-N1 plasmid and untransfected cells in cell morphology by microscope, in cell proliferation by MTT, in expression of tyrosine hydroxylase by immunofluorescence and fluorescent real-time PCR.Results:1. Cell module persistently expressing BDV P24 was constructed successfully. BDV P24 mainly located in the nuclear, while pEGFP-N1 located in the whole cytoplasm. We detected BDV P24 fragment only in cells transfected with pEGFP-N1-P24 plasmid.2. BDV P24 inhibit cell proliferation, promote TH expression. Cells transfected with pEGFP-N1-P24 plasmid grew slower than cells transfected with pEGFP-N1 plasmid and untransfected cells. The difference is significant in statistics. 3. BDV P24 promote tyrosine hydroxylase expression in PC-12 cells. The mRNA and protein of tyrosine hydroxylase was much abundant in cells transfected with pEGFP-N1-P24 plasmid than in cells transfected with pEGFP-N1 and cells untransfected.Conclusion:We have constructed a cell model expressing BDV P24 persistently. BDV P24 didn't change the cell morphous, but did inhibit cell proliferation and enhance tyrosine hydroxylase expression. This change may induce excessive movement, extremist response, society capability impairment in BDV infected animal.