Madin-Darby canine kidney cells are increased in aerobic glycolysis when cultured on flat and stiff collagen-coated surfaces rather than in physiological 3-D cultures.
We investigate the influence of the dimensionality and the biochemistry of the culture system on the cellular functionality by analyzing the protein expression levels in Madin-Darby canine kidney (MDCK) cells grown in 3-D and 2-D substrates. We cultured MDCK cells on a hard and flat 2-D uncoated plastic surface, on a 2-D collagen-coated plastic surface and in 3-D collagen gel and employed 2-D gel electrophoresis, MALDI-TOF-MS, and LC-MS/MS analysis to identify the differentially regulated proteins. We found significant differences in the expression of antioxidant proteins, actin-binding proteins, glycolytic enzymes, and heat-shock proteins/chaperons among the three types of cultures. While MDCK cells cultured in 3-D collagen up-regulate antioxidant proteins and proteins involved in the dynamic remodeling of the actin cytoskeleton, 2-D collagen-coated plastic surfaces induce the up-regulation of glycolytic enzymes. Our data shows that the culture conditions have profound effects on the physiology of the cell. Culture in 3-D collagen induces a differentiated polarized phenotype. In contrast, collagen-coated 2-D substrates favor a tumor-like phenotype with increased glycolysis. Thus, the suitability of 2-D cultures to study the physiological behavior of cells, especially in drug discovery, bioprocessing, and toxicology, should be carefully reconsidered.
Pampaloni
Francesco
F
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany. francesco.pampaloni@embl.de
Stelzer
Ernst H K
EH
Leicht
Stefan
S
Marcello
Marco
M
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D003094:Collagen / Q000737:chemistry / Q000378:metabolism*; D004285:Dogs; D005786:Gene Expression Regulation; D006019:Glycolysis; D007668:Kidney / Q000166:cytology*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D054457:Tissue Scaffolds / Q000737:chemistry
D011506:Proteins; D003094:Collagen