Apical-to-basolateral transepithelial transport of human lactoferrin in the intestinal cell line HT-29cl.19A

Abstract

The role of human lactoferrin (hLf) in alimentary iron absorption across intestinal cells was explored using a human differentiated colon carcinoma cell line, HT-29cl.19A. The apical surface of HT-29cl.19A cell monolayers exhibited 1.5 x 10(6) specific binding sites for hLf per cell, with a dissociation constant of 8.3 x 10(-7) M. The apical-to-basolateral transport of 125I-labeled hLf (125I-hLf) or 125I-59Fe-labeled hLf (125I-[59Fe]hLf) was investigated using filter-grown HT-29cl.19A cell monolayers mounted in Ussing chambers. Transport of total (intact plus degraded) hLf, measured by the 125I flux, was not saturable up to an apical hLf concentration of 12.5 microM, whereas immunoreactive hLf transport measured by enzyme-linked immunoabsorbent assay was saturable at 3.75 microM. Lowering the temperature to 4 degrees C reduced the total and immunoreactive hLf fluxes by 77% and 90%, and colchicine (500 microM) reduced these fluxes by 30% and > 97%, respectively. These results indicate that both types of transport are transcellular. Electrophoretic analysis revealed that 125I-hLf transported to the basal compartment consisted of largely degraded fragments (< 2 and 10-20 kDa) and a small portion of intact hLf. At an apical concentration of 3.75 microM diferric 125I-[59Fe]hLf, the immunoreactive hLf flux (64.6 +/- 14.3 ng.h-1.cm-2) constituted approximately 12% of the total hLf flux (552.0 +/- 61.6 ng.h-1.cm-2) and was similar to the [59Fe]hLf-equivalent flux (77.0 +/- 12.3 ng.h-1.cm-2).(ABSTRACT TRUNCATED AT 250 WORDS)