Study on construction, expression, and biological activity of recombinant adenovirus of neurturin.
Neurturin (NTN) can improve the function and delay the rate of degeneration of dopaminergic neurons in Parkinson's disease (PD). However, its method of delivery to the central nervous system has not been established. Adenoviral vectors have been widely applied in gene therapy because of their high-efficiency gene transfer, easy manipulation, and safety. We used replication-defective adenovirus type 5 (Ad5) to construct a recombinant viral vector encoding full-length human NTN (Ad-NTN) and amplified Ad-NTN and the control (Ad-lacZ) in HEK 293 cells. NTN-specific expression in the Ad-NTN-infected HEK 293 cells was detected by RT-PCR and the immunofluorescent assay. However, no NTN expression was detected in the Ad-lacZ-infected HEK 293 cells. After incubation with the Ad-NTN-infected conditioned medium (CM), the dorsal root ganglia of chicken embryos examined in vitro exhibited radial neurite outgrowth around the ganglia. However, incubation with the Ad-lacZ-infected or blank CM resulted in a short or absent nerve process and the growth of only a few fibroblasts. Our findings indicated that recombinant Ad-NTN was specifically expressed in the host cells, and the expressed NTN possessed biological activity.
Li
Hongjun
H
Institute of Medical Biology, Chinese Academy of Medical Sciences, Peking Union Medical College, Kunming, Yunnan, P.R. China.
Yang
Fang
F
Shi
Rui
R
Wang
Wenju
W
Yan
Min
M
Xie
Tianhong
T
Ma
Kaili
K
Sun
Maosheng
M
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D002642:Chick Embryo; D003001:Cloning, Molecular; D005727:Ganglia, Spinal / Q000378:metabolism*; D005822:Genetic Vectors / Q000235:genetics; D007668:Kidney / Q000378:metabolism*; D051101:Neurturin / Q000235:genetics* / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000378:metabolism*; D014162:Transfection / Q000379:methods*
C495053:NRTN protein, human; D051101:Neurturin; D011994:Recombinant Proteins