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Intestinal DC Migration and Activation
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, and IFN-; plasmacytoid DCs (pDCs) purified from MLNs were the source of the cytokines. Anti-TNF- mAb treatment of mice abrogated DC accumulation in MLNs in response to oral R-848; CD86 expression was up-regulated on their MLN-DCs and pDCs. MLN-DCs and pDCs of mice deficient in IFN-
had minimal increase in CD86 expression after R-848 administration. Serum levels of IL-12p70, TNF- and IFN-, MLN-DCs numbers, and CD86 expression were reduced in R-848-fed mice previously given a pDC-depleting Ab. The authors use rat and mouse models to demonstrate TNF--dependent MLN accumulation of iL-DCs and their IFN--dependent peripheral activation in response to TLR7/8 stimulation. Control of NGAL Expression
Bacteria-induced inflammation of colon or lung epithelium results in increased expression of bacteriostatic neutrophil gelatinase-associated lipocalin (NGAL) protein and in NF-
B activation. However, Cowland et al. showed that NGAL gene expression is up-regulated only by IL-1 plus LPS and not by TNF-. On p. 5559
, Cowland et al. detected peak induction of mRNA for IB-
, an IL-1-inducible cofactor of NF-B, in a human lung epithelial cell line at 1.5 h after IL-1 stimulation; TNF- only weakly induced IB- gene transcription. Cycloheximide or a transfected small interfering RNA against IB- mRNA abrogated induction of NGAL mRNA in IL-1-stimulated cells within 24 h. Activity of a reporter plasmid under control of the NGAL promoter was increased in cells cotransfected with a plasmid expressing IB- and stimulated with either IL-1 or TNF-. Use of mutated NGAL promoter constructs demonstrated requirement for an intact NF-B binding site. Stimulation of cells with IL-1 for 3 h followed by TNF- stimulation without IL-1 resulted in NGAL protein accumulation in the medium; no accumulation was seen for cells stimulated with TNF- alone or unstimulated. The experiments demonstrate a critical role of the NF-B binding cofactor IB- at an early stage in induction of the antimicrobial protein NGAL in response to IL-1 signaling after bacterial infection.
CD4+ T Cell Response to Malarial Parasite
Although CD4+ T cells from both malaria-exposed and -nonexposed individuals respond to Plasmodium falciparum Ags, it is not known if the response of the nonexposed individual is a memory response that could interfere with vaccination against malaria. Ndungu et al. (p. 5504
) stimulated PBMCs from nonexposed individuals in vitro with a recombinant conserved cysteine interdomain region-1 (CIDR-1) domain of P. falciparum erythrocyte membrane protein-1 that binds CD36. Among the stimulated PBMCs, 45 and 4% of CD4+ T cells expressed CD69 and IFN-
, respectively, and 60 and 12% of CD56+ NK cells expressed CD69 and produced IFN-, respectively. A control erythrocyte membrane protein-1 domain did not induce CD69 expression or cytokine production. Whereas PBMCs from 18 and 16 of 33 malaria nonexposed donors produced IFN- and IL-10, respectively, only 8 gave a positive proliferative response to CIDR-1. Anti-MHC class II Ab did not inhibit the IFN- response to CIDR-1 stimulation of PBMCs from most nonexposed individuals tested but did inhibit proliferation and IFN- production by stimulated PBMCs from all malaria-exposed individuals tested. Myeloid, but not plasmacytoid, dendritic cells isolated from malaria-nonexposed PBMCs were stimulated by CIDR-1 or P. falciparum schizont-infected RBCs to produce IL-10 for 24 h; IL-12 production by the cells peaked between 6 and 12 h and then dropped to background level. The experiments demonstrate a nonmemory CD4+ T cell response in PBMCs from malaria-nonexposed individuals but a memory CD4+ T cell response in malaria-exposed individuals due to direct CIDR-1 stimulation of dendritic cells.
Mismatch Repair in B Cells
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1 and Sµ-S3 DNA junctions of in vitro-stimulated mlh3–/– B cells demonstrated decreased microhomologies and increased blunt junctions and insertions compared with stimulated wild-type controls. S1 and S3 region breakpoints preferentially targeted a specific mutational hot spot in the mutant cells. S-S breakpoints outside the hot spot entailing microhomologies and nonmicrohomologies were significantly altered in mlh3–/– vs mlh3+/+ cells in that there was a decrease in Sµ, S1, and S3 breakpoints outside the hot spot. Germinal center B220+PNAhigh cells isolated from Peyer’s patches of mlh3–/– and mlh3+/+ mice had comparable frequencies of VHDJH rearrangements involving JH4 intronic DNA. Sequences upstream of JH4 intronic DNA in mlh3–/– mice had more dC/dG mutations with dG being the most frequently mutated residue; the number of transitions in mutations was decreased in the mutant mice. Mutations at dC/dG that targeted the hot spot were twice as high in mlh3–/– mice, indicating a significant alteration in somatic hypermutation (SHM) compared with mlh3+/+ mice. An increase in drug resistance of mutant mouse embryonic fibroblasts in vitro showed that the change in SHM was genome wide. The experiments show that mouse MMR protein Mlh3 contributes to CSR in stimulated B cells and to genome-wide SHM. Catalase Inactivation in Asthma
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Ag-Induced IL-10+ Treg Cells
Although several different types of regulatory T (Treg) cells have been described, there is no clear characterization of peptide induced vs naturally occurring Treg cells. Nicolson et al. (p. 5329
) compared naturally occurring CD25+ Treg cells from spleens of naive Tg4 mice, transgenic for TCR and chains that recognize a specific epitope of myelin basic protein (MBP), with Treg cells from Tg4 mice injected with the specific MBP peptide (PI-Treg cells). After in vitro stimulation with peptide and IL-2, both populations of Treg cells were anergic to peptide restimulation compared with unstimulated naive CD4+ Tg4 cells. Naive Tg4 cells secreted IL-2, Ag-stimulated PI-Treg cells secreted IL-10, and Ag-stimulated CD25+ Treg cells secreted neither cytokine. CD25+ Treg cells and CD25-depleted PI-Treg cells suppressed Ag-induced proliferation of naive CD4+ Tg4 cells in vitro. Only CD25+ Treg cells expressed Foxp3 after Ag stimulation. Tg4 mice on a RAG–/– background lacked CD4+CD25high splenocytes yet generated IL-10-secreting PI-Treg cells after peptide injection that were anergic to restimulation. In response to intranasal administration of peptide, CFSE-labeled naive CD4+ Tg4 splenocytes divided readily in naive RAG–/– recipients but underwent fewer divisions in peptide-immunized RAG–/– recipients. No Foxp3 expression was detected in in vitro peptide-stimulated CD4+ T cells from peptide-immunized wild-type or RAG–/– Tg4 animals. The authors conclude that CD25 is not required for suppressive activity of IL-10-secreting PI-Treg cells and that only naturally occurring Treg cells express Foxp3.
TRAFs in B Cell Signaling
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B, TRAF1–/– B cells exhibited similar kinetics, and TRAF2–/– B cells had delayed kinetics compared with wild-type cells. Stimulated TRAF1–/–TRAF2–/– cells had no IB degradation and, in contrast to TRAF1–/– or TRAF2–/– cells, did not exhibit nuclear translocation of NF-B pathway factors or activate JNK. No signaling pathway defects were noted in any mutant cells stimulated with latent membrane protein 1 (LMP1) of EBV. TRAF1–/–TRAF2–/– B cells were unable to produce IgM in response to CD40 stimulation but did produce IgM in response to LMP1 stimulation; decreased IgM production in TRAF1–/– B cells was restored by activation of a stably transfected TRAF1 protein expression vector. TRAF2 and TRAF3 proteins were increased in immunoprecipitates of CD40 from the insoluble membrane raft fraction of CD40-stimulated TRAF1–/– B cell lysates compared with wild-type cells, whereas the amount of TRAF1 protein was decreased in rafts of stimulated TRAF2–/– B cells. Induced synthesis of TRAF1 protein in CD40-stimulated TRAF1–/– B cells decreased the amount of TRAF2 protein in the rafts. TRAF1 deficiency increased degradation of TRAF2 and TRAF3 proteins in response to CD40 stimulation, and this increase was abolished by induced expression of TRAF1 protein. The data from this loss-of-function model system demonstrate that TRAF1 and TRAF2 proteins interact to regulate CD40-mediated signaling in B cells but are not required for LMP1 signaling. Summaries written by Dorothy L. Buchhagen, Ph.D.
Related articles in The JI:
and Type 1 IFNs after Feeding a TLR7/8 Ligand
-Specific Up-Regulation of Neutrophil Gelatinase-Associated Lipocalin Is Controlled by IB-
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