Plasticity of protein expression during culture of fetal skin cells.
In order to gain insight into the biology of fetal skin during culture, cellular proteins were studied during four culture passages (P00, P01, P04 as well as P10) using high-resolution two-dimensional (2-D) gel electrophoresis and mass spectrometry (MS). Bioinformatic analyses were focused on a region of each gel corresponding to pI between 4 and 8 and M(r) from 8000 to 35 000. In this area, 373 +/- 42 spots were detected (N = 18). Twenty-six spots presented an integrated intensity that increased in the higher passages, whereas five spots showed a progressively lower intensity in subsequent passaging. MS analysis was performed on spots that were unambiguously identified on preparative 2-D gels. Among the 26 spots showing an increased size between P00 and P10, 9 were identified, and corresponded to 3 proteins: (i) peptidyl-prolyl cis-trans isomerase A (P05092; cyclophilin A or cyclosporin A-binding protein), (ii) triosephosphate isomerase (P00938), and (iii) enoyl-CoA hydratase (P30084). Among these nine identified spots, three were absent at P00, but were present at P10. They corresponded to isoforms of peptidyl-prolyl cis-trans isomerase and triosephosphate isomerase, respectively. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses of the acidic isoforms of triosephosphate isomerase showed modifications of cysteine residues to cysteic acid. All these isoforms were clearly present in the skin cells of a 4-year-old child, as well as in skin cells from a 80-year-old man, at P00. These observations probably reflect either an oxidative stress related to cell culture, or, alternatively, maturation, differentiation and the aging of the cells.
Vuadens
Françoise
F
Service Régional Vaudois de Transfusion Sanguine, Lausanne, Switzerland.
Crettaz
David
D
Scelatta
Corinne
C
Servis
Catherine
C
Quadroni
Manfredo
M
Bienvenut
Willy-Vincent
WV
Schneider
Philippe
P
Hohlfeld
Patrick
P
Applegate
Lee A
LA
Tissot
Jean-Daniel
JD
D000368:Aged; D000369:Aged, 80 and over; D002478:Cells, Cultured; D002675:Child, Preschool; D021984:Cyclophilin A / Q000032:analysis; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004746:Enoyl-CoA Hydratase / Q000032:analysis; D005333:Fetus; D015870:Gene Expression; D006801:Humans; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000032:analysis; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000032:analysis*; D040901:Proteomics / Q000295:instrumentation / Q000379:methods*; D012867:Skin / Q000166:cytology* / Q000196:embryology; D023822:Tissue Engineering; D014305:Triose-Phosphate Isomerase / Q000032:analysis
D020033:Protein Isoforms; D011506:Proteins; D004746:Enoyl-CoA Hydratase; D021984:Cyclophilin A; D014305:Triose-Phosphate Isomerase