Uptake of ricinB-quantum dot nanoparticles by a macropinocytosis-like mechanism

BACKGROUND: There is a huge effort in developing ligand-mediated targeting of nanoparticles to diseased cells and tissue. The plant toxin ricin has been shown to enter cells by utilizing both dynamin-dependent and -independent endocytic pathways. Thus, it is a representative ligand for addressing the important issue of whether even a relatively small ligand-nanoparticle conjugate can gain access to the same endocytic pathways as the free ligand. RESULTS: Here we present a systematic study concerning the internalization mechanism of ricinB:Quantum dot (QD) nanoparticle conjugates in HeLa cells. Contrary to uptake of ricin itself, we found that internalization of ricinB:QDs was inhibited in HeLa cells expressing dominant-negative dynamin. Both clathrin-, Rho-dependent uptake as well as a specific form of macropinocytosis involve dynamin. However, the ricinB:QD uptake was not affected by siRNA-mediated knockdown of clathrin or inhibition of Rho-dependent uptake caused by treating cells with the Clostridium C3 transferase. RicinB:QD uptake was significantly reduced by cholesterol depletion with methyl-β-cyclodextrin and by inhibitors of actin polymerization such as cytochalasin D. Finally, we found that uptake of ricinB:QDs was blocked by the amiloride analog EIPA, an inhibitor of macropinocytosis. Upon entry, the ricinB:QDs co-localized with dextran, a marker for fluid-phase uptake. Thus, internalization of ricinB:QDs in HeLa cells critically relies on a dynamin-dependent macropinocytosis-like mechanism. CONCLUSIONS: Our results demonstrate that internalization of a ligand-nanoparticle conjugate can be dependent on other endocytic mechanisms than those used by the free ligand, highlighting the challenges of using ligand-mediated targeting of nanoparticles-based drug delivery vehicles to cells of diseased tissues.