Dendritic cells (DCs) in peripheral tissues can stimulate and bias naive CD4+ T cells or affect anergy or induction of T regulatory cells. However, it is not known how different populations of immature or semimature human skin DCs influence T cell responses. Morelli et al. (p. 7905 ) purified skin migratory DCs (smiDCs) after their spontaneous migration from human epidermal/dermal explants. Flow cytometric analyses of surface markers indicated several states of maturation/activation. Surface expression patterns of CD1a and CD14 identified three smiDC populations: CD1a+CD14 epidermal Langerhans’ cells (LCs), CD1aCD14 dermal DCs (DDCs), and CD1aCD14+ LC precursors. All smiDCs expressed CCR7 and secreted high levels of IL-10 and TGF-β1 and low levels of IL-12/23p40 but no IL-12p70. LCs and DDCs induced high proliferation and a potent Th1 response with high IFN-γ secretion in allogeneic naive CD4+ T cells in MLC; LC precursors induced only a weak response. CD4+ T cell proliferation and IFN-γ secretion in response to LC precursors were enhanced by higher LC:T cell ratios, longer interaction time, or addition of recombinant human IL-23. Treatment of LCs and DDCs with blocking mAbs to MHC class II, adhesion, or costimulatory molecules abrogated their ability to induce T cell proliferation and secretion of IFN-γ and IL-5. Anti-IL-23 mAb diminished IFN-γ, but not IL-5, secretion and inhibited the T cell response to LCs and DDCs. The authors conclude that the ability of smiDCs to stimulate CD4+ T cells correlates with their APC maturation stage and is dependent on secretion of IL-23.

Protein tyrosine phosphatase α (PTPα) positively regulates the src family tyrosine kinase, fyn, in several cell types. A similar function in immune cells has not been established. Maksumova et al. (p. 7947 ) found increased phosphotyrosine content and fyn phosphorylating activity in unstimulated thymocytes from PTPα−/− vs wild-type mice. Fyn activities were similar in cells from both strains after stimulation. Two critical fyn tyrosine residues, one in the activation loop and one in a regulatory loop, were found to have increased levels of phosphorylation in PTPα−/− thymocyte lysates by anti-phosphoamino acid-specific Ab probing of immunoblots. Both tyrosine residues were dephosphorylated on fyn immunoprecipitated from COS-1 cells cotransfected with fyn plus wild-type PTPα but not in cells transfected with fyn plus a catalytically inactive PTPα mutant. Fyn associated with lipid rafts was twice as active in unstimulated PTPα−/− vs wild-type cells. However, raft-associated fyn activity increased in wild-type, but decreased in PTPα−/−, cells after stimulation. A phosphorylated lipid raft-associated transmembrane protein in PTPα−/− thymocytes was dephosphorylated rapidly after cell stimulation. A src family kinase inhibitor prevented the increased phosphorylation of the transmembrane protein in PTPα−/− cells. Proliferation and IL-2 production were impaired for stimulated PTPα−/− thymocytes but were normal for stimulated lymph node T cells from wild-type and PTPα−/− mice. No alterations in fyn tyrosine phosphorylation or activity were seen in the lymph node T cells. The data indicate that PTPα negatively regulates fyn-mediated tyrosine phosphorylation in resting thymocytes before TCR activation.

Transcription of adjacent Il4 and Il13 genes is enhanced during development of Th2 cells in mice but silenced during Th1 cell development. No function has been determined for RNA transcripts mapping to noncoding intergenic regions of the Il4-Il13 locus in Th2 cells. Baguet et al. (p. 8146 ) used real-time RT-PCR to determine the distribution pattern of intergenic transcripts in a fully differentiated Th2 cell line. Transcripts from resting cells were mapped into several DNase I hypersensitive sites within and adjacent to the Il4-Il13 locus. PMA/ionomycin-mediated cell activation increased transcription levels within the genes but had no effect on intergenic transcription. Studies with a reversible inhibitor of RNA polymerase II showed that the enzyme was responsible for intergenic transcription but did not influence histone acetylation within the locus as determined by chromatin immunoprecipitation analysis. Intergenic transcript distribution in naive CD62LhighCD4+ T cells from spleen and lymph nodes of young BALB/c mice was similar to that of the Th2 cell line and of in vitro Th2-primed cells. In contrast, intergenic transcripts were no longer detectable in Th1-primed cells. However, nuclear run-on analysis of radiolabeled sense and antisense RNA showed that intergenic transcripts with the potential to form dsRNA were produced at the Th2 locus in both resting Th1 and Th2 cells. The authors propose that intergenic transcripts are degraded or processed in a Th1-specific manner to silence the Il4-Il13 locus and are not involved in maintenance of histone acetylation or cytokine transcription.

Although TNF-α is known to have a central role in synovitis and joint destruction in rheumatoid arthritis (RA), it is unclear whether TNF-α-mediated inflammation results in loss of tolerance to autoantigens. Hayer et al. (p. 8327 ) found pronounced IgG reactivities to heterogeneous nuclear ribonucleoprotein-A2 (hnRNP-A2) and some IgM reactivity with two stress proteins in sera from mice transgenic for the membrane-bound form of human TNF-α (hTNFtg). The IgG reactivity developed shortly after the onset of erosive arthritis in this mouse model of RA. Mouse sera detected four protein bands corresponding to hnRNP-A2, alternatively spliced variants, and a closely related hnRNP on immunoblots of HeLa cell nuclear proteins. The dominant epitope was mapped to an hnRNP-A2 peptide containing aa 50–70. Anti-hnRNP-A2 Ab levels were lower in sera from mice treated with anti-TNF-α mAb or osteoprotegerin, an inhibitor of osteoclast differentiation and proliferation, and in hTNFtg mice lacking a functional c-fos gene. T cells from spleen or peripheral lymph nodes of hTNFtg mice did not proliferate in vitro or produce IFN-γ or IL-4 in response to hnRNP-A2/B1 or purified protein derivative stimulation. Highly elevated expression of hnRNP-A2 in synovial macrophages and fibroblasts in inflamed joints of hTNFtg mice was demonstrated by immunohistochemistry. Immunoblotting on protein extracts of hindpaws detected hnRNP-A2, one splice variant, and a unique splice variant also seen in three nonlymphoid organs. Immunization of hTNFtg mice with hnRNP-A2/B1 or peptide 50–70 elicited greater inflammation and joint erosion compared with controls. The data demonstrate that TNF-α-induced inflammation in a murine RA model results in autoantibody production against overexpressed hnRNP-A2.

Cyclooxygenase (COX) inhibition during allergic sensitization and allergen challenge increases airway hyperresponsiveness (AHR) and inflammation and, as shown previously by Peebles et al., increases levels of leukotriene metabolites in mice. Factors controlling the heightened allergic response are unknown. Peebles et al. (p. 8253 ) detected more total cells, eosinophils, and lymphocytes in bronchoalveolar fluid from OVA-sensitized and OVA-challenged wild-type or 5-lipoxygenase-deficient mice treated with the COX inhibitor indomethacin (OVA-indo) than from either mouse strain treated with OVA only. AHR in response to methacholine challenge, concentrations of IL-5 and IL-13 in supernatants of ground lungs and OVA-specific IgE serum concentrations of OVA-sensitized mice were greatest in the wild-type OVA-indo animals. Increased numbers of total cells and eosinophils were found in bronchoalveolar fluid from IL-13−/− OVA-indo mice compared with controls following OVA challenge, whereas AHR was greatest in OVA-indo IL-5−/− mice. OVA-challenged wild-type OVA-indo mice had higher levels of IL-5 and IL-13 in ground lung supernatants compared with wild-type OVA mice, and IL-5 levels were elevated in lung supernatants of IL-13−/− OVA-indo mice compared with IL-13−/− OVA mice. The authors conclude that in a model of allergic inflammation under conditions of COX inhibition, IL-13 increases AHR but IL-5 increases airway eosinophilia. Neither response is dependent on leukotrienes.

Infection of immunocompromised individuals with the extracellular fungus Pneumocystis results in interstitial pneumonia with accumulation of CD8+ T cells. Yet, the mechanisms responsible for the ensuing lung damage are not known. Meissner et al. (p. 8271 ) infected CD4+ T cell-depleted perforin−/−, fas−/−, or IFN-γ−/− mice with Pneumocystis to show that their earlier observation of lung damage following the influx of CD8+ T cells in CD4+ T cell-depleted wild-type mice was independent of the deleted molecules. Pneumocystis-infected SCID mice reconstituted with polyclonal wild-type splenic CD8+ T cells had greater lung damage and influx of 10 times the number of lymphocytes than controls receiving CD8+ T cells from mice transgenic for a viral Ag-specific TCR or unreconstituted infected SCID mice. TNF-α levels in the bronchoalveolar fluid were most elevated in animals given wild-type cells. Greater lung damage also occurred in Pneumocystis-infected CD4+ T cell-depleted β2-microglobulin−/− or CD8α-chain−/− mice reconstituted with wild-type CD8+ T cells compared with wild-type controls; no significant differences were seen between CD1d−/− and wild-type mice. Lethally irradiated Rag-1−/− mice reconstituted with Rag-1−/− CD8+ T cells, infected with Pneumocystis, and adoptively transferred with wild-type CD8+ T cells had increased lung damage compared with similarly treated β2-microglobulin−/− mice. Antibiotic treatment of Pneumocystis-infected CD4+ T cell-depleted wild-type mice prevented lung damage. The data show that interaction of CD8+ T cells with Pneumocystis Ag-loaded MHC class I molecules on radiation-resistant lung stromal cells results in lung damage in this model of fungal-induced pneumonia.

Joint inflammation and destruction in animal models of rheumatoid arthritis (RA) are ameliorated by enhanced apoptosis. The Pope laboratory and others have shown that the Bcl-2 subfamily member Mcl-1 is essential for survival of hemopoietic cell types, but any role for Mcl-1 in survival of other cell types has not been established. Liu et al. (p. 8337 ) detected increased Mcl-1 expression in synovial lining, fibroblasts in the sublining and blood vessels of patients with RA compared with osteoarthritis patients or arthritis-free controls by immunohistochemistry. The intensity of Mcl-1 staining and the percentage of Mcl-1-positive fibroblasts correlated with the degree of inflammation in each group. Mcl-1 protein and mRNA expression was greatest in the RA synovial fibroblasts and increased after exposure to TNF-α or IL-1β. Introduction of an adenovirus vector expressing full-length antisense (AS) human Mcl-1 decreased Mcl-1 protein levels and led to loss of mitochondrial transmembrane potential and apoptotic cell death accompanied by cytochrome c release from mitochondria and activation of caspases 9 and 3. RA cells infected with a vector expressing Mcl-1 or Bcl-xL did not undergo apoptosis. Analysis of cell fractions and confocal microscopy detected Bax translocation from cytosol to mitochondria in RA cells infected with AS Mcl-1; silencing of Bax, Bak, or Bim by small interfering RNAs prevented apoptosis of those vector-infected cells. The authors conclude that Mcl-1 promotes survival of synovial fibroblasts in RA by preserving mitochondrial integrity and preventing apoptosis mediated by Bax, Bak, or Bim.

Summaries written by Dorothy L. Buchhagen, Ph.D.