The delay of anoikis due to the inhibition of protein tyrosine dephosphorylation enables the maintenance of normal rat colonocyte primary culture.
Apoptosis induced by detachment of cells from the extracellular matrix (anoikis) appears to be one of the main obstacles in attempts to establish long-term primary culture of normal colonocytes. In the present study, the dynamics of molecular events related to apoptosis of isolated normal rat colonocytes was investigated. The whole colonic crypts were isolated using collagenase/dispase digestion technique. DNA fragmentation typical for the apoptosis and the apoptotic morphology of cells were observed already at the end of their isolation. Considerable increase in caspase-3 activity was noted during the first two hours of cell cultivation. Delaying of apoptosis by treatment of cells with sodium orthovanadate, the specific protein tyrosine phosphatase inhibitor, was found to be possible. It may facilitate long-term culture of intestinal epithelial cells.
Dzierzewicz
Z
Z
Department of Molecular Biology, Biochemistry and Biopharmacy, Medical University of Silesia, Sosnowiec, Poland. zofiadz@slam.katowice.pl
Orchel
A
A
Parfiniewicz
B
B
Weglarz
L
L
Stojko
J
J
Swierczek-Zieba
G
G
Wilczok
T
T
D000818:Animals; D023102:Anoikis / Q000187:drug effects* / Q000502:physiology; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D003106:Colon / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D053938:DNA Fragmentation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D011930:Reaction Time / Q000187:drug effects / Q000502:physiology; D014638:Vanadates / Q000494:pharmacology
D004791:Enzyme Inhibitors; D014638:Vanadates; D017027:Protein Tyrosine Phosphatases; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases