[Construction of recombinant adenovirus and mediated reported gene expression in the guinea pig cochlea].
OBJECTIVE
To purify P0 protein from guinea pig's inner ear by preparative SDS-PAGE and study the possible role it may play in the etiology of autoimmune inner ear disease.


METHODS
A mixture of membraneous proteins of inner ear was separated by preparative SDS-PAGE. The corresponding band at 30kd was cut and electrically eluted. The protein collected was identified by analytical SDS-PAGE and Western blot assay. A group of 20 guinea pigs were immunized with P0 protein emulsified in complete Freund's adjuvant, another 10 guinea pigs were immunized with complete Freund 's adjuvant only as control. The guinea pigs' hearing thresholds, serum IgG level and morphological changes in the inner ear were investigated. The distribution of P0 protein in the cochlear was detected by immunohistochemical technique.


RESULTS
The purity of the protein was demonstrated by a single band at the 30 kD site in SDS-PAGE, which was identified as P0 protein by western blot analysis assay. About 17.5% P0-immunized guinea pigs showed increased hearing thresholds, elevated IgG level (F =6.48, P <0. 01), as well as a decreased number of spiral ganglion cells and inflammatory cell infiltration in the cochlear nerve region. The P0 protein is distributed in the cochlear nerve and spiral ganglion only.


CONCLUSIONS
P0 protein from guinea pig's inner ear can be successfully purified by preparative SDS-PAGE and an animal model of experimental autoimmune inner ear disease induced by P0 protein is successfully established.
Department of Otorhinolaryngology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
Yingpeng
Y
Liu
Guopeng
G
Wang
Anmin
A
Shen
Jianting
J
Wang
Pei
P
Chen
Zeweng
Z
Li
Shusheng
S
Gong
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D003051:Cochlea / Q000378:metabolism; D004195:Disease Models, Animal; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005801:Genes, Homeobox; D017930:Genes, Reporter; D005822:Genetic Vectors; D006168:Guinea Pigs; D018993:Myelin P0 Protein / Q000302:isolation & purification*
D018993:Myelin P0 Protein