Primary cell cultures arising from normal kidney and renal cell carcinoma retain the proteomic profile of corresponding tissues.
Renal cell carcinoma (RCC) tissue is composed of a mixture of neoplastic and normal cells, which complicate proteome analysis. The aim of our study was to investigate whether it is feasible to establish primary cell cultures of RCC and of renal cortex maintaining the tissue phenotype along with a more homogeneous and enriched cytological material. Fourteen (82.3%) primary cultures from 17 surgical cases were established and characterized by morphology, growth rate, immunocytochemistry, and molecular analysis performed by Real-time PCR, Western blotting, two-dimensional electrophoresis (2-DE), and mass spectrometry. Cultures showed >90% cytokeratine-positive epithelial cells. In primary tumor cultures, the molecular phenotype of manganese superoxide dismutase and heat shock protein 27 was the same as that found in tumor tissues with overexpression and increased number of isoforms. Moreover, 27 out 28 specific proteins and their isoforms, present in spots excised from 2-DE gel of cortex or RCC cultures, corresponded to those identified on the 2-DE tissue cortex reference map, suggesting that these primary cultures retain the proteomic profile of the corresponding tissues.
Perego
Roberto A
RA
Department of Experimental, Environmental Medicine and Medical Biotechnologies, Milano-Bicocca University, Monza, Italy. roberto.perego@inimib.it
Bianchi
Cristina
C
Corizzato
Matteo
M
Eroini
Barbara
B
Torsello
Barbara
B
Valsecchi
Cristina
C
Di Fonzo
Andrea
A
Cordani
Nicoletta
N
Favini
Paolo
P
Ferrero
Stefano
S
Pitto
Marina
M
Sarto
Cecilia
C
Magni
Fulvio
F
Rocco
Francesco
F
Mocarelli
Paolo
P
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D015153:Blotting, Western; D002292:Carcinoma, Renal Cell / Q000378:metabolism*; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D015180:Electrophoresis, Gel, Two-Dimensional; D004847:Epithelial Cells / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007633:Keratins / Q000378:metabolism; D007668:Kidney / Q000378:metabolism*; D007680:Kidney Neoplasms / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry; D008875:Middle Aged; D018832:Molecular Chaperones; D009363:Neoplasm Proteins / Q000378:metabolism; D010449:Peptide Mapping; D010641:Phenotype; D010766:Phosphorylation; D020033:Protein Isoforms; D040901:Proteomics / Q000379:methods*; D012313:RNA / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012694:Serine / Q000737:chemistry; D013482:Superoxide Dismutase / Q000378:metabolism; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D055551:HSP27 Heat-Shock Proteins; C057725:HSPB1 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D009363:Neoplasm Proteins; D020033:Protein Isoforms; D012694:Serine; D012313:RNA; D007633:Keratins; D013482:Superoxide Dismutase