Characterization of the vitamin D receptor from the Caco-2 human colon carcinoma cell line: Effect of cellular differentiation☆

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Abstract

The human colon carcinoma cell line, Caco-2, is the only intestinal cell line to spontaneously differentiate in culture to a population exhibiting structural and biochemical characteristics of mature enterocytes. We conducted studies to establish the presence of the vitamin D receptor (VDR), determine changes in VDR concentration and affinity with differentiation and determine whether 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) mediates a functional response in this cell line. We found that Caco-2 cells possess a specific 1,25(OH)2D3 binding protein similar to the mammalian VDR. It has an equilibrium dissociation constant (Kd) of 0.72 nm, binds vitamin D analogues in order of their biological activities in vivo (1,25(OH)2D3 > 25(OH)D3 > 24,25(OH)2D3), sediments as a single peak on sucrose density gradients at 3.7 S, and is eluted from a DNA-cellulose column by 0.16 m KCl. The maximum number of binding sites was 2.6-fold greater in the differentiated cell (Day 15) compared to the preconfluent, undifferentiated (Day 4) cell (23 fmol/ mg protein vs 56 fmol/mg protein). Cell growth was reduced 59% when exposed to 10−7m 1,25(OH)2D3 for 8 days. Alkaline phosphatase activity significantly increased in cultures incubated with 10−8m 1,25(OH)2D3 for up to 4 days when treatment was started in both undifferentiated cells (Day 5) and differentiated cells (Day 11). These findings suggest that the VDR present in undifferentiated and differentiated Caco-2 cells is functional. Caco-2 cells provide a unique in vitro model to study vitamin D-regulated functions in differentiated mammalian enterocytes.

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    ☆

    This project has been funded with Federal funds from the Department of Agriculture, Agricultural Research Service under Contract 53-3K06-5-10. The contents of this publication do not necessarily reflect the views of the U.S. Department of Agriculture, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government. Additional support was provided by NIH Grant DK 35317 (R.T.F.).

    2

    This work was performed by Anna R. Giuliano as partial fulfillment for the Ph.D. degree in Nutrition at Tufts University and presented in part at the Seventy-third Annual Meeting of the Federation of American Societies for Experimental Biology, New Orleans, LA, March, 1989 (1989, FASEB J. 3, A1245). Dr. Giuliano's present address is Department of Biochemistry, Biological Sciences West 436, University of Arizona, Tucson, AZ 85721.

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