Dendritic cells (DC) efficiently take up antigens by macropinocytosis and mannose receptor-mediated endocytosis. Here we show that endocytosis of mannose receptor-antigen complexes takes place via small coated vesicles, while non-mannosylated antigens were mainly present in larger vesicles. Shortly after internalization the mannose receptor and its ligand appeared in the larger vesicles. Within 10 min, the mannosylated and non-mannosylated antigens co-localized with typical markers for major histocompatibility complex class II-enriched compartments and lysosomes. In contrast, the mannose receptor appeared not to reach these compartments, suggesting that it releases its ligand in an earlier endosomal structure. Moreover, we demonstrate that mannosylation of protein antigen and peptides resulted in a 200-10,000-fold enhanced potency to stimulate HLA class II-restricted peptide-specific T cell clones compared to non-mannosylated peptides. Our results indicate that mannosylation of antigen leads to selective targeting and subsequent superior presentation by DC which may be applicable in vaccine design.

Embryonic stem cells (ESCs) can be induced to differentiate into nerve cells, endowing them with potential applications in the treatment of neurological diseases and neural repair. In this work, we report for the first time that sulfated chitosan can promote the neural differentiation of ESCs. As a type of sulfated glycosaminoglycan analog, sulfated chitosan with well-defined sulfation sites and a controlled degree of sulfation (DS) were prepared through simple procedures and the influence of sulfated glycosaminoglycan on neural differentiation of ESCs was investigated. Compared with other sulfation sites, 6-O-sulfated chitosan showed the most optimal effects. By monitoring the expression level of neural differentiation markers using immunofluorescence staining and PCR, it was found that neural differentiation was better enhanced by increasing the DS of 6-O-sulfated chitosan. However, increasing the DS by introducing another sulfation site in addition to the 6-O site to chitosan did not promote neural differentiation as much as 6-O-sulfated chitosan, indicating that compared with DS, the sulfation site is more important. Additionally, the optimal concentration and incubation time of 6-O-sulfated chitosan were investigated. Together, our results indicate that the sulfate site and the molecular structure in a sulfated polysaccharide are very important for inducing the differentiation of ESCs. Our findings may help to highlight the role of sulfated polysaccharide in inducing the neural differentiation of ESCs.

In this work, a novel gene delivery strategy was proposed based on silicon nanowire arrays modified with high-molecular-weight 25 kDa branched polyethylenimine (SN-PEI). Both the plasmid DNA (pDNA) binding capacity and the in vitro gene transfection efficiency of silicon nanowire arrays (SiNWAs) were significantly enhanced after modification with high-molecular-weight bPEI. Moreover, the transfection efficiency was substantially further increased by the introduction of free pDNA/PEI complexes formed by low-molecular-weight branched PEI (bPEI, 2 kDa). Additionally, factors affecting the in vitro transfection efficiency of the novel gene delivery system were investigated in detail, and the transfection efficiency was optimized on SN-PEI with a bPEI grafting time of 3 h, an incubation time of 10 min for tethered pDNA/PEI complexes consisting of high-molecular-weight bPEI grafted onto SiNWAs, and with an N/P ratio of 80 for free pDNA/PEI complexes made of low-molecular-weight bPEI. Together, our results indicate that high-molecular-weight bPEI modified SiNWAs can serve as an efficient platform for gene delivery.

Langerhans cells (LC) represent the dendritic cell (DC) lineage in the epidermis. They capture and process antigens in the skin and subsequently migrate to the draining lymph nodes to activate naive T cells. Efficient uptake and processing of protein antigens by LC would, therefore, seem a prerequisite. We have now compared the capacity of human epidermal LC, blood-derived DC and peripheral blood mononuclear cells to endocytose and present (mannosylated) antigens to antigen-specific T cells. Moreover, we have determined the expression of mannose receptors, and the composition of the intracellular endosomal/lysosomal MHC class II-positive compartment. The results indicate that LC have poor endocytic capacity and do not exploit mannose receptor-mediated endocytosis pathways. Furthermore, the composition of the class II compartment in LC is distinct from that in other antigen-presenting cells and is characterized by the presence of relatively low levels of lysosomal markers. These results underscore the unique properties of LC and indicate that LC are relatively inefficient in antigen uptake, processing and presentation. This may serve to avoid hyper-responsiveness to harmless protein antigens that are likely to be frequently encountered in the skin due to (mechanical) skin damage.

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The synthesis of a series of analogues of the a-glucosidase inhibitor 1-deoxynojirimycin (dNM, 1) and of the a-mannosidase inhibitor 1-deoxymannojirimycin (dMM, 3) is described. The ability of dNM, dMM and a series of N-alkylated dNM and dMM derivatives to interfere with biosynthesis, transport and maturation of the glycoprotein a ,-antitrypsin in human hepatoma HepG2 cells and with the syncytium-inducing capacity of HIV-infected cells was investigated. A strong correlation was observed between a-glucosidase inhibition found in HepG2 cells and antiviral activity in HIV-I-infected cells. N-Butyl-(35), N-pentyl-(38), N-benzyl-(41) and N-decyl-dNM (47) showed high activity in both assays. N-Decyl-dNM was active at 0.01 mM but showed drug-related cell toxicity at concentrations exceeding 0.1 mM. Branching of the N-alkyl side chain reduced the activity of dNM derivatives considerably. N-Benzyl-6-0-butyryl-dNM (42) and N-decyl-6-0-benzoyl-dNM (48) showed activity comparable to that of N-benzyl-dNM (41) and N-decyl-dNM (47), respectively. None of the dMM analogues prevented HIV-induced syncytia formation. The 3-hydroxyl group in dNM and in dMM plays a crucial role in the interaction of these drugs with the corresponding processing glycosidases: 3-0-methyl-dNM (49) and 3-0-methyl-dMM (56) were inactive in both assays. The plasma membrane constitutes a permeability barrier for castanospermine (CAS, 2), but not for dNM, since the activity of CAS in streptolysin-0-permeabilized HepG2 cells was significantly higher than that in intact HepG2 cells. Finally, an overview is given of structures, other than the many N-substituted derivatives that were described, related to dNM and dMM that have appeared in the literature in recent years.i' Present address: h Nomenclature and abbreviations: AIDS = Acquired immune&ficiency syndrome, Bn = benzyl, CAS = castanospermine (2), CI,. T M B = chlorine-3,3',S,S'-tetramethylbenzidine, d N M = 1deoxynojirimycin = 1,s-dideoxy-1 ,S-imino-D-glucitol (I), dMM = 1deoxymannojirimycin = 1,5-dideoxy-l,S-imino-D-mannitol (3), HIV = human immunodeficiency virus, HM = high mannose, 2.6-lutidine = 2,6.dimethylpyridine, SDS-PAGE = Sodium-dodecyl-sulphate poly.acrylamide-gel-electro~horesis, sw = swainsonine (4, triflate = trifluoromethane sulphonate.

1993 organic chemistry, review organic chemistry, review Z 0200 31 -303 Chemical Modification of Azasugars, Inhibitors of N-Glycoprotein-Processing Glycosidases and of HIV-I Infection. Review and Structure-Activity Relationships -(113 refs.) -(VAN DEN BROEK, L. A. G. M.; VERMAAS, D. J.; HESKAMP, B. M.; VAN BOECKEL, C. A. A.; TAN, M. C. A. A.; BOLSCHER, J. G. M.; PLOEGH, H. L.; VAN KEMENADE, F. J.; DE GOEDE, R. E. Y.; MIEDEMA, F.; Recl. Trav. Chim. Pays-Bas 112 (1993) 2, 82-94; Dep. Med. Chem. III, Organon Int. BV, 5340 BH Oss, Neth.; EN)