Activity and properties of α- l-fucosidase are dependent on the state of enterocytic differentiation of HT-29 colon cancer cells

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Previously we have demonstrated an impairment in the activity of α- l-fucosidase in colon tumours. In order to establish an in vitro model to study this enzyme in colon cancer, we have determined the activity and properties of the enzyme during the differentiation of HT-29 colon cancer cells. Cultures were committed to differentiate into enterocyte-like cells by placing them in a culture medium without glucose for 18–21 days. The state of differentiation was evaluated by assaying the activity of enterocytic marker enzymes, and the acquisition of enterocyte morphology was assessed by electron microscopy. The α- l-fucosidase activity was determined using a fluorometric method. Intracellular levels of α- l-fucosidase activity are lower in non-differentiated cells (3.0±1.01 U/mg) than in differentiated ones (9.2±4.09 U/mg) ( P<0.001). This variation is not due to a greater secretion of the enzyme to the culture medium, and properties such as pH optimum or the affinity towards substrate are not dependent on differentiation. The enzyme however, is more stable at acidic pH and at high temperatures, and V max is higher in differentiated cells. Moreover, in undifferentiated cells the enzyme is mainly in a monomeric form whereas multimeric forms of the enzyme appear only upon differentiation. Most of these changes are very similar to those previously observed between normal colon tissue and colon tumours. Thus, we suggest that differentiation of HT-29 colon cancer cells could be used as a model to study the alterations of the enzyme α- l-fucosidase during the progression of the tumoural process.