A Quaroni1, J F Beaulieu. 1. Section of Physiology, Cornell University, Ithaca, New York, USA.
Abstract
BACKGROUND & AIMS: Intestinal epithelial cell lines capable of differentiating in monolayer culture have been difficult to obtain, prompting the use of novel approaches including immortalization with oncogenes or tumor viruses. The aim of this study was to obtain conditionally immortalized human intestinal epithelial cells (temperature-sensitive fetal human intestinal [tsFHI] cells) and study their growth and differentiation capabilities. METHODS: Intestinal cells from human fetuses were transformed with a temperature-sensitive simian virus 40 large tumor antigen, cloned, and screened for expression of intestinal markers. RESULTS: Establishment of tsFHI cells was crucially dependent on a growth medium based on OptiMEM. At 32 degrees C, the tsFHI cells proliferate and display crypt cell markers. Only a few scattered differentiated cells are observed together with cells displaying the morphological signs of apoptosis. A shift to 39 degrees C results in irreversible growth arrest and acquisition of an enterocyte-like phenotype. Expression of the cyclin-dependent kinase inhibitor p21/WAF1/Cip1 is induced within hours of a shift to 39 degrees C 1-2 days before corresponding increases in brush border enzymes, suggesting a role in the induction rather than in the maintenance of a differentiated phenotype. CONCLUSIONS: The tsFHI cells recapitulate key aspects of intestinal cell dynamics in vitro and seem particularly suited for the study of early events in cell differentiation.
BACKGROUND & AIMS: Intestinal epithelial cell lines capable of differentiating in monolayer culture have been difficult to obtain, prompting the use of novel approaches including immortalization with oncogenes or tumor viruses. The aim of this study was to obtain conditionally immortalized human intestinal epithelial cells (temperature-sensitive fetal human intestinal [tsFHI] cells) and study their growth and differentiation capabilities. METHODS: Intestinal cells from human fetuses were transformed with a temperature-sensitive simian virus 40 large tumor antigen, cloned, and screened for expression of intestinal markers. RESULTS: Establishment of tsFHI cells was crucially dependent on a growth medium based on OptiMEM. At 32 degrees C, the tsFHI cells proliferate and display crypt cell markers. Only a few scattered differentiated cells are observed together with cells displaying the morphological signs of apoptosis. A shift to 39 degrees C results in irreversible growth arrest and acquisition of an enterocyte-like phenotype. Expression of the cyclin-dependent kinase inhibitor p21/WAF1/Cip1 is induced within hours of a shift to 39 degrees C 1-2 days before corresponding increases in brush border enzymes, suggesting a role in the induction rather than in the maintenance of a differentiated phenotype. CONCLUSIONS: The tsFHI cells recapitulate key aspects of intestinal cell dynamics in vitro and seem particularly suited for the study of early events in cell differentiation.
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