A preliminary study on the interaction between Asn-Gly-Arg (NGR)-modified multifunctional nanoparticles and vascular epithelial cells

Liu, Chunxi; Liu, Tingxian; Yu, Xiaoyue; Gu, Yizhu

As shown in Fig. 3 , when HUVEC were incubated with an anti-CD13 antibody and fixed without warming (0 min), red fluorescence was observed evenly on the cell surfaces. In the same cells, labelling for CAV1 was observed in green. No co-localization occurred between CD13 and CAV1. After cells bound with the antibodies were incubated for 10 min at 37 1C, the labelling of CD13 and CAV1 on the HUVEC surface showed a uniform punctate distribution, and no co-localization between CD13 and CAV1 was observed. When the incubation time was 30 min at 37 1C, the labelling of CD13 and CAV1 showed in spots, and a small extent of co-localization between CD13 and CAV1 was observed (white arrows). With an extended incubation time at 37 1C, co-localization of CD13 with CAV1 became more frequent after 60 min (white arrows). When the incubation time extended to 2 and 3 h, CD13 and CAV1 gathered into clusters and co-localized extensively (white arrows). These results demonstrated that the antibody against CD13 bound to the cell surface evenly when incubated on ice, but the bound antibodies became sequestered to CAV1-positive patches when the temperature increased to 37 1C. Additionally, the results indicate that binding of anti-CD13 antibody and incubation at 37 1C could cause clustering of CD13 and its co-localization with CAV1. These results are exciting and imply that targeting CD13 could initiate a subsequent interaction with CAV1, although there was no colocalization between CD13 and CAV1 in the absence of antibody against CD13. The accumulation of cross-linked molecules in CAV1positive areas was similar to the previously reported behavior of glycosylphosphatidylinositol (GPI)-anchored proteins and glycosphingolipids 22, 39, 40 .