Lentiviruses, as exemplified by visna virus of sheep, are nononcogenic retroviruses that cause slowly progressive diseases after prolonged periods of incubation. Earlier studies on visna have shown that the long incubation period of the disease is associated with constant production of minimal quantities of virus in tissues, whereas virus could be obtained by culturing monocytes and macrophages from explants of lymphatic tissues and inflamed organs. In this study the role of macrophages in lentivirus infection was explored using two sheep that were intrabronchially inoculated with virus.

Visna lentiviruses have a natural tropism for cells of the macrophage lineage of sheep and goats, but virus replication in these cells in vivo is restricted so that only small quantities of virus are produced. One restricting factor suggested in previous studies is that virus replication is dependent on the maturity of the cells: the more mature the cell, the less restrictive the replication of the virus. Since monocytes in peripheral blood are precursors of macrophages, we investigated the effect of cell maturation on virus replication under limited control conditions in vitro by inoculating blood leukocytes with virus and retarding the maturation of monocytes to macrophages during cultivation in serum-free medium. Using enzyme markers that identified the cells in their resting monocytic stage (peroxidase) and mature macrophage stage (acid phosphatase) along with quantitative in situ hybridization and immunocytochemistry with viral reagents to trace the efficiency of virus replication, we correlated virus replication with cell maturation. Only a few monocytes were susceptible to infection, and virus replication did not extend beyond a low level of transcription of viral RNA. In the acid phosphatase-positive, maturing macrophage, susceptibility of the cells to infection was increased and virus replication was greatly amplified to the level of translation of viral polypeptides. However, virus maturation was delayed by 3 days until further cell maturation had occurred. Thus, the entire life cycle of the virus, from its attachment to the target cell to its maturation in the cell, was dependent on the level of maturation/differentiation of the monocytic cell.

The purpose of this study was to identify an optimum targeted particulate formulation based on mannan (MN)-decorated poly(D, L-lactide-co-glycolide) (PLGA) nanoparticles (NPs), for efficient delivery of incorporated cargo to dendritic cells (DCs). In brief, NPs were formulated from two different types of PLGA; ester-terminated (capped) or COOH-terminated (uncapped) polymer. Incorporation of MN in NPs was achieved either through addition of MN during the process of NP formation or by attachment of MN onto the surface of the freeze dried NPs by physical adsorption or chemical conjugation (to COOH terminated polymer). The formulated NPs were characterized in terms of particle size, Zeta potential and level of MN incorporation. The effect of polymer type and the incorporation method on the extent of fluorescently labelled NP uptake by murine bone marrow-derived DCs have been investigated using flowcytometry. The results of this study showed MN incorporation to enhance the uptake of PLGA NPs by DCs. Among different MN incorporation strategies, covalent attachment of MN to COOH-terminated PLGA-NPs provided the highest level of MN surface decoration on NPs. Maximum NP uptake by DCs was achieved by COOH terminated PLGA NPs containing covalent or adsorbed MN. Therefore, a better chance of success for these formulations for active targeted drug and/or vaccine delivery to DCs is anticipated.

Visna and maedi of sheep are classical "slow" virus diseases that have a long incubation period and progressive debilitating clinical courses (1). Histologically, lesions are characterized by infiltration and proliferation of mononuclear cells in an active-chronic inflammatory process in specific organ systems such as the brain, lung, joints, and mammary gland (2). The etiological agents of the disease complex belong to a newly recognized taxonomic group called lentiviruses, which are nononcogenic, replication-competent retroviruses (1). The viruses cause persistent infections in their natural hosts and replicate at a restrictive level indefinitely (3, 4). In nonneural tissues such as the lung, chronic progressive lesions develop, whereas the lesions in the central nervous system (CNS) l appear to represent an episodic pathogenesis with repeated episodes of acute lesions leading to repair (5).Studies on host-virus interactions in vivo (6) have shown that cells of the macrophage lineage are the main virus host cells but that replication is confined to macrophages only in those tissues which develop inflammatory lesions. Thus, the minimally productive virus replication in specific populations of tissue macrophages forms the basis of persistent infection of the animal, "slow" or restricted virus replication in tissue, and lymphoproliferative pathologic responses that occur at the sites of virus replication. The mechanisms of the lesions remain unknown. The recent discovery (preceding article [7]) that an interferon (IFN) is induced during interaction between lymphocytes and lentivirus-infected macrophages provides a potentially important bridge linking the virus-infected

Mannan (MN) is the natural ligand for mannose receptors, which are widely expressed on dendritic cells (DCs). The purpose of this study was to assess the effect of formulation parameters on the immunogenicity of MN-decorated poly (D, L-lactide-co-glycolide) (PLGA) nanoparticles (NPs) in terms of their ability to stimulate DC phenotypic as well as functional maturation. For this purpose, NPs were formulated from either ester-terminated or COOH-terminated PLGA. Incorporation of MN in NPs was achieved through encapsulation, physical adsorption or chemical conjugation. Murine bone marrow derived DCs (BMDCs) were treated with various NP formulations and assessed for their ability to up-regulate DC cell surface markers, secrete immunostimulatory cytokines and to activate allogenic T cell responses. DCs treated with COOH-terminated PLGA-NPs containing chemically conjugated MN (MN-Cov-COOH) have shown superior performance in improving DC biological functions, compared to the rest of the formulations tested. This may be attributed to the higher level of MN incorporation in the former formulation. Incorporation of MN in PLGA NPs through chemical conjugation can lead to enhanced DC maturation and stimulatory function. This strategy may be used to develop more effective PLGA-based vaccine formulations.