Mechanism of antigen presentation after hypertonic loading of soluble antigens.

Hypertonic loading of proteins into cells has been used to introduce soluble proteins into the major histocompatibility complex class I pathway of antigen presentation followed by cytotoxic T-lymphocyte (CTL) induction. The precise mechanism for this pathway is not completely understood. The antigen is either processed and presented by/on the same cell or by professional antigen-presenting cells (APC) after taking up the antigen from damaged or apoptotic cells. After loading labelled ovalbumin (OVA), it could be co-precipitated with the proteasome complex, supporting the role of this pathway for antigen processing. The processing speed however, appeared to be slow since intact OVA could be detected inside the cells even after 18 hr. This corresponded well with the processing of OVA by isolated proteasomes. On the other hand, enough peptides for recognition of target cells by CTLs were generated in this reaction. One reason for the low level of processing might be that hypertonic loading may damage the cells and inhibit direct processing. In fact, at least 50% of the cells became positive for Annexin V binding after hypertonic loading which indicates severe membrane alterations usually associated with the progress of apoptosis. Annexin V binds to phosphatidylserine residues which also serve as ligand for CD36 expressed on monocytes and some immature dendritic cells. This may direct the phagocytic pathway to hypertonically loaded cells and thus enable professional APCs to present OVA-peptides. Therefore, in addition to the direct processing of OVA, CTLs can be primed by professional APC after uptake of apoptotic, OVA-loaded cells.