Characterization of myeloid and plasmacytoid dendritic cells in human lung.

Dendritic cells (DCs) are bone marrow-derived mononuclear cells that play a central role in the initiation of immune responses. Because human lung DCs have been incompletely characterized, we enumerated and phenotyped mononuclear cell populations from excess lung tissue obtained at surgery. Myeloid DCs (MDCs) were identified as CD1c(+)CD11c(+)CD14(-)HLA-DR(+) cells and comprised approximately 2% of low autofluorescent (LAF) mononuclear cells. Plasmacytoid DCs (PDCs) were characterized as CD123(+)CD11c(-)CD14(-)HLA-DR(+) cells and comprised approximately 1.0% of the LAF mononuclear cells. Cells enriched in MDCs expressed CD86, moderate CD80, and little CD40, but cells enriched in PDCs had little to no expression of these three costimulatory molecules. CD11c(+)CD14(-) lineage-negative (MDC-enriched) LAF cells were isolated and shown to be much more potent in stimulating an alloreaction than CD11c(+)CD14(+) lineage-negative (monocyte-enriched) LAF cells. PDC-enriched cells were more capable of responding to a TLR-7 agonist by secreting IFN-alpha than MDC-enriched cells. MDC-enriched cells were either CD123(+) or CD123(-), but both subsets secreted cytokines and chemokines typical of MDC upon stimulation with a TLR-4 agonist and both subsets failed to secrete IFN-alpha upon stimulation with a TLR-7 agonist. By immunohistochemistry, we identified MDCs throughout different anatomical locations of the lung. However, our method did not allow the localization of PDCs with certainty. In conclusion, in the human lung MDCs were twice as numerous and expressed higher levels of costimulatory molecules than PDCs. Our data suggest that both lung DC subsets exert distinct immune modulatory functions.

Flt3 ligand preferentially increases the number of functionally active myeloid dendritic cells in the lungs of mice.

In the present study, we investigated the effects of in vivo Flt3L administration on the generation, phenotype, and function of lung dendritic cells (DCs) to evaluate whether Flt3L favors the expansion and maturation of a particular DC subset. Injection of Flt3L into mice resulted in an increased number of CD11c-expressing lung DCs, preferentially in the alveolar septa. FACS analysis allowed us to quantify a 19-fold increase in the absolute numbers of CD11c-positive, CD45R/B220 negative DCs in the lungs of Flt3L-treated mice over vehicle-treated mice. Further analysis revealed a 90-fold increase in the absolute number of myeloid DCs (CD11c positive, CD45R/B220 negative, and CD11b positive) and only a 3-fold increase of lymphoid DCs (CD11c positive, CD45R/B220 negative, and CD11b negative) from the lungs of Flt3L-treated mice over vehicle-treated mice. Flt3L-treated lung DCs were more mature than vehicle-treated lung DCs as demonstrated by a significantly higher percentage of cells expressing MHC class II, CD86, and CD40. Freshly isolated Flt3L lung DCs were not fully mature, because after an overnight culture they continued to increase accessory molecule expression. Functionally, Flt3L-treated lung DCs were more efficient than vehicle-treated DCs at stimulating naive T cell proliferation. Our data show that administration of Flt3L favors the expansion of myeloid lung DCs over lymphoid DCs and enhanced their ability to stimulate naive lymphocytes.

Initiation of lung immunity: the afferent limb and the role of dendritic cells.

The lung is the largest epithelial surface in the body and constitutes a major portal of entry for microorganisms. Dendritic cells (DCs) are bone marrow-derived cells that populate the respiratory tract from the nasal mucosa to the lung pleura. DCs are dexterous cells playing a role in both the innate and the adaptive immune responses. DCs can respond to dangers in the lung by immediately generating protective cytokines that stimulate natural killer (NK) cells to produce cytokines and kill targets. Direct (by pathogen recognition receptors) and indirect (by inflammatory cytokines) recognition of the infectious agent can also trigger signaling pathways in the DCs that activate an integrated developmental program called maturation, which transforms DCs into efficient antigen-presenting cells (APCs) for naive T cell stimulation. Furthermore, DCs play a role in generating effector B cells in the lymph nodes draining the lung. This review focuses on how DCs sense the health of the lung and translate messages regarding the lung microenvironment to cells of the innate and adaptive immune response to initiate effector functions to clear an invading microorganism [T helper 1 (Th1) or Th2 response] or generate tolerance [T regulatory (Tr), anergy, or deletion] to inhaled antigen when no danger is sensed.

Dendritic cells: immune regulators in health and disease.

Dendritic cells (DCs) are bone marrow-derived cells of both lymphoid and myeloid stem cell origin that populate all lymphoid organs including the thymus, spleen, and lymph nodes, as well as nearly all nonlymphoid tissues and organs. Although DCs are a moderately diverse set of cells, they all have potent antigen-presenting capacity for stimulating naive, memory, and effector T cells. DCs are members of the innate immune system in that they can respond to dangers in the host environment by immediately generating protective cytokines. Most important, immature DCs respond to danger signals in the microenvironment by maturing, i.e., differentiating, and acquiring the capacity to direct the development of primary immune responses appropriate to the type of danger perceived. The powerful adjuvant activity that DCs possess in stimulating specific CD4 and CD8 T cell responses has made them targets in vaccine development strategies for the prevention and treatment of infections, allograft reactions, allergic and autoimmune diseases, and cancer. This review addresses the origins and migration of DCs to their sites of activity, their basic biology as antigen-presenting cells, their roles in important human diseases and, finally, selected strategies being pursued to harness their potent antigen-stimulating activity.

3-phosphoglycerate kinase: a glycolytic enzyme protein present in the cell wall of Candida albicans.

We have used a polyclonal antiserum to cell wall proteins of Candida albicans to isolate several clones from a cDNA lambda gt11 expression library. Affinity-purified antibody prepared to the fusion protein of one clone identified a 40 kDa moiety present in cell wall extracts from both morphologies of the organism. Indirect immunofluorescence demonstrated expression of this moiety at the C. albicans cell surface. Sequencing of a pBluescript II genomic clone identified with the cDNA clone revealed an open reading frame for a 417 amino acid protein. The nucleotide sequence showed significant homology with 3-phosphoglycerate kinase (PGK) genes, with 88%, 77% and 76% nucleotide homology with the PGK genes from Candida maltosa, Saccharomyces cerevisiae and Kluyveromyces lactis, respectively. The deduced amino acid sequence was consistent with this identification of the sequence as PGK1 of C. albicans. This finding was confirmed by a positive immunological response of a commercially available purified PGK from S. cerevisiae with the affinity-purified antibody against the fusion protein of the cDNA clone. The presence of PGK in the cell wall was confirmed by two additional methods. Cell wall protein were biotinylated with a derivative that does not permeate the cell membrane to distinguish extracellular from cytosolic proteins. Biotinylated PGK was detected among the biotinylated proteins obtained following streptavidin affinity chromatography. Immunoelectron microscopy revealed that the protein was present at the outer surface of the cell membrane and cell wall as well as expected in the cytoplasm. Northern blot analysis revealed that the gene transcript was present in C. albicans cells growing under different conditions, including different media, temperatures and morphologies. Most of the enzyme activity was found in the cytosol. Low enzymic activity was detected in intact cells but not in culture filtrates. These observations confirmed that PGK is a bona fide cell wall protein of C. albicans.