Internalization properties of the anti-tumor α-lactalbumin-oleic acid complex

α-Lactalbumin (α-LA) can bind oleic acid (OA) to form the anti-tumor α-LA-OA complex. Previous studies suggested α-LA-OA induced apoptosis or autophagy in an independent way. Furthermore, as a large molecule, α-LA-OA could enter tumor cells and accumulated in the nucleus, which was speculated as the basis of its anti-tumor activity. In this study, we evaluated the internalization property of α-LA-OA with and without endocytosis inhibitors using flow cytometry and laser scanning confocal microscopy. It was shown α-LA-OA transported from the cell membrane to the cytoplasm, then accumulated around the nucleus, which consequently began to shrink and condense. The α-LA component only located in the membrane whereas the OA component entered cytoplasm. When pre-treated cells with these inhibitors, the internalization of OA would all decrease while the interaction of α-LA with membrane did not influence. As for the complex, the internalization of α-LA-OA was completely blocked at 4°C and significantly decreased in the presence of cytochalasin D, an inhibitor of phagocytosis (p<0.01). In conclusion, the anti-tumor activity of α-LA-OA was mainly dependent on the OA component whereas the internalization mechanism was related to the α-LA component to be temperature-dependent and have a close relationship with the phagocytosis pathway.