Two-dimensional electrophoresis analyses of atopic dermatitis and the chances to detect new candidate proteins by the variations in immobilized pH gradient strips.
BACKGROUND
Proteomic approaches, one of the high-throughput technologies, have been used to search for the proteins that are abnormally expressed in human diseases. The atopic dermatitis (AD)-associated genes or proteins are gradually being reported.


OBJECTIVE
In accordance with recent reports, we conducted the serial proteomic studies to compare with the protein expression level and to find a critical protein associated with AD.


METHODS
We applied two-dimensional electrophoresis (2-DE) coupled with MADI-TOF as well as LC-MS/MS to detect dysregulated protein in the AD proteome obtained from the patient-derived primary cells. Real-time PCR was also conducted to compare with the proteomic results in the transcriptional level.


RESULTS
We successfully detected new AD-associated proteins in the AD-derived fibroblasts 2D-PAGE studies due to the IPG strip variations after conducting MALDI-TOF mass spectrometry and identification of the proteins. From the real-time PCR quantifications, we found that the altered expression of caldesmon 1 isoform 5, nucleophosmin 1, esterase D and chloride intracellular channel 4 were well matched both at the transcriptional and translational levels, and this suggested that these proteins may have important involvement with the pathogenesis of AD.


CONCLUSIONS
By simply repeating the trials with changing the commercial strips with different lot numbers in the 2D-PAGE analysis, this provided us with new finding in the AD-derived samples. This approach we used may increase the chances of finding new candidate proteins in the clinical samples and it may also be applicable for proteomic studies of other diseases.
Park
Yong-Doo
YD
Department of Dermatology, Sungkyunkwan University School of Medicine, Samsung Medical Center, Seoul 135-710, Republic of Korea.
Lyou
You-Jeong
YJ
Yang
Jun-Mo
JM
D000328:Adult; D016022:Case-Control Studies; D002478:Cells, Cultured; D018118:Chloride Channels / Q000378:metabolism; D003876:Dermatitis, Atopic / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015603:Keratinocytes / Q000378:metabolism*; D008297:Male; D016133:Polymerase Chain Reaction; D040901:Proteomics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
C464459:CLIC4 protein, human; D018118:Chloride Channels