Recognition of CD133 and modulation of its expression interfere with Tf uptake and TfR accessibility in Caco-2 cells.

<p>A) AC133 antibody treatment inhibited Tf uptake. Constitutively CD133-expressing undifferentiated Caco-2 cells were exposed to Tf-Alexa 488 for 1 h at 37°C/5%CO₂ in the presence of 5 or 10 µg/ml of AC133 or IgG1κ immunoglobulin control. Tf-Alexa 488 that was effectively internalized within cells was then monitored by flow cytometry. Results are expressed as a % of Tf-Alexa 488 amounts that were internalized in the untreated control. Data represented mean ± s.e.m. from a triplicate obtained from one representative experiment. Dunnett's test: *p<0.05, ***p<0.001. B) Flow cytometric analysis of the expression of TfR (CD71) at the surface of CD133^high (Control siRNA) and CD133^low (CD133 siRNA) Caco-2 cells. Data represented mean ± s.e.m. obtained from three independent experiments. IgG2aκ immunoglobulins were used as negative control immunostaining. Dunnett's test: ***p<0.001. C) Analysis of the expression of AC133, TfR/CD71 and CHC within Caco-2 cells by immunocytochemistry combined with confocal laser scanning microscopy. Note the increase of CD71 expression while AC133 was depleted from the Control siRNA to the CD133 siRNA situation. Bar: 50 µm.