Splenic trapping of nanoparticles: Complementary approaches for in situ studies

Purpose. To identify more accurately in the spleen, the areas and the cells where nanoparticulate carriers were taken up from the blood flow, a series of complementary approaches were used. Methods. First, in and ex vivo examination of the whole spleen led to a global view of all the trapping areas. Then, histological studies on frozen sections of the same organ allowed for a more precise localization of these areas and image analysis gave an evaluation of tissue distribution of the nanoparticles. Finally, immunological and enzymological characteristics of the capturing cells were determined in situ, using monoclonal antibodies (F4/80 and anti-sialoadhesin) and cytochemichal reactions (esterases and acid phosphatase). Furthermore incubation of spleen slices with different nanoparticles was used so as to know if the capture was due to a high capturing capacity of these cells or to a high blood flow in their vicinity. Results. It was shown that more than 90% of the splenic capture was localized in the marginal zone of the follicles. The capturing cells form a special population of macrophages inserted in a reticular meshwork, showing low esterase and acid phosphatase activities, giving faint or no reaction with F4/80 or anti-sialoadhesin antibodies. The circulating nanoparticles were quickly trapped with rather low specificity by these cells. Conclusions. Combination of coherent approaches allowed for the tracking of capturing cells from in vivo observations to their in situ identification on immunological and enzymological criteria.