Figure 6.

In vitro and in vivo competition assays demonstrated that oligo-fucoidan binds to C-type lectin-like receptor 2 (CLEC-2)–asialoglycoprotein receptors (ASGPR)1/2 in hepatocytes. (A) Flow cytometry profiles of Clone 9 cells. The black line histogram represents isotype control, the blue indicates cells stained by asialofetuin–fluorescein isothiocyanate (ASF-FITC), and the red line denotes cells treated with OF and stained by asialofetuin–FITC (ASF-FITC). (B) The mean fluorescence intensity (MFI) profiles of Clone 9 cells with or without OF treatment in the flow cytometry analysis. The black color represents isotype control, blue indicates cells with asialofetuin–FITC (ASF-FITC), and the red line denotes cells treated with OF and asialofetuin–FITC (ASF-FITC). (C) ⁶⁸Ga-NOTA-hexavalent lactoside (HL) positron emission tomography (PET) imaging of the murine livers demonstrates the in vivo ASGPR binding image in vivo. Phosphate-buffered saline (PBS) and OF (100 mg) were administrated through an intravenous injection with ⁶⁸Ga-NOTA-HL (⁶⁸Ga-2,2’,2’’-(1,4,7-triazonane-1,4,7-triyl) triacetic acid-hexavalent lactoside) injection (N = 3/group) simultaneously. (D) The relative radioactive intensity of ⁶⁸Ga-NOTA-HL in vivo between the PBS and OF-injected groups. The bars indicate the mean ± SD, where the black bar represents the PBS group and the red bar indicates the OF treatment group. The radioactivity (%) was compared to PBS (no fucoidan treatment) group. Statistical significance was calculated by Student’s t-test (*p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001).