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Controlling Chlamydia Infection
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but had no effect in D2 control cells. Genome microarray analysis on mRNA from peritoneal exudates of infected D2 mice indicated up-regulation of transcripts for proinflammatory cytokine and chemokine genes. Infected B6 mice had increased expression of regulators of inflammatory cytokine release, genes involved in differentiation and proliferation of macrophages, and NK cell activating genes. Histology and cytology confirmed a greater influx of neutrophils to the infection site in D2 mice but more macrophages in B6 mice. BALB/c mice lacking a chemokine gene did not recruit neutrophils to the C. psittaci infection site and survived. The authors suggest that differences in neutrophil recruitment regulated by Irgb10 and Iigp2 are responsible for the differential susceptibility to C. psittaci infection in B6 and D2 mice. CD8+ DC to Exosome: Gotcha!
Exosomes, the small membranous vesicles secreted by dendritic cells (DCs), carry MHC class I- and class II-peptide complexes. Although exosomes can activate Ag-specific T cells, their interactions with recipient DCs are unknown. On p. 1489 , Segura et al. used paraformaldehyde to fix DCs incubated in vitro either with the HY peptide or with MHC class II-HY peptide exosomes purified from the supernatant of LPS- and HY peptide-treated DCs. Only DCs incubated with exosomes activated HY-specific naive CD4+ T cells. T cell activation was abrogated by the addition of control exosomes or by the use of exosomes from Icam-1–/– DCs. Similarly, Lfa-1–/– DCs exposed to wild-type exosomes were less able to activate HY-specific T cells than were wild-type DCs. In vivo, there was a significant decrease in the proliferation of adoptively transferred HY-specific T cells in Lfa-1–/– mice compared with wild-type mice after footpad injection with HY-bearing exosomes. Flow cytometry demonstrated higher levels of LFA-1 on CD8+ DCs vs CD8– DCs from the spleens of wild-type mice, whereas LFA-1 was expressed only on CD8+ DCs from cutaneous lymph nodes. Only CD8+ DCs had I-Ab-HY complexes that activated HY-specific T cells in vitro 3 h after footpad injection of wild-type mice with HY-bearing exosomes; CD8+ and CD8– DCs both presented HY peptide in animals injected with HY peptide. CD8+ DCs were unable to capture HY-bearing exosomes in injected Lfa-1–/– mice. The authors conclude that MHC class II-HY peptide exosomes secreted by mature DCs are captured by LFA-1 molecules expressed on CD8+ DCs and are presented to and activate T cells without further processing.
Cell Density and MSC Plasticity
The clinical use of mesenchymal stromal cells (MSCs) in modulating immune responses requires a thorough understanding of the conditions under which they either become immunosuppressive or acquire APC functions. Romieu-Mourez et al. (p. 1549
) measured higher up-regulation of MHC class II molecules and OVA peptide presentation in IFN--treated primary mouse MSC cultures at high vs low cell density. Serum-starvation of high- or low-density MSC cultures increased their IFN--induced MHC class II expression and OVA peptide processing. Whereas the activations of STAT1 and IFN regulatory factor-1 were equivalent in the two IFN--treated cultures, the induction of type IV CIITA mRNA expression was greatest in cells of the high-density cultures. Expression of one TGF-
type I receptor was increased by low-density culture conditions. Growth of MSCs was inhibited by TGF- pretreatment; the cells had reduced expression of IFN--induced CIITA and MHC class II molecules, reduced OVA-peptide presentation, and increased phosphorylation of several proteins downstream of the TGF- type I receptor. High-density cultures of human MSCs from normal donors lost the rapidly self-renewing cells needed for multilineage differentiation and in vivo engraftment compared with low-density cultures. IFN- induced greater up-regulation of MHC class II molecules in low-density human MSC cultures, and prior treatment with TGF- blocked the IFN- effect. The authors point out that up-regulation of MHC class II expression and Ag presentation occurs in high-density mouse MSC cultures but in low-density human MSC cultures and that MSCs in both species are inhibited by pretreatment with TGF-.
Targeting Inflammation in Arthritis
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Prosurvival Role of MAPK Kinase 4
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B were induced by TNF in wild-type cells but were not expressed in TNF-treated mutant fibroblasts. NF-B activation was detected by EMSA only in wild-type cells treated with TNF, LPS, PMA, or cigarette smoke condensate and in TNF-treated mutant cells transfected with an MKK4 expression vector. The mutant cells also did not activate IB
kinase, did not phosphorylate or degrade IB protein, and did not induce phosphorylation or nuclear translocation of p65 in response to TNF treatment. Further, a transiently transfected NF-B-regulated reporter gene was activated only in TNF-stimulated wild-type, but not mutant, cells. Among several molecules in the TNF-signaling pathway individually cotransfected with the NF-B-regulated reporter gene into MKK4–/– cells, only p65 induced expression of the reporter. The data suggest that MKK4 protects mouse cells from TNF-induced apoptosis by modulating the NF-B signaling pathway at a step upstream of p65.
Function of IBNS in TCR Activation
Induction of IBNS, a new member of the IB family of NF-B inhibitors identified by the Clayton laboratory, is known to correlate with TCR signal strength. However, thefunction of IBNS is not known. In a continuation of their studies, Touma et al. (p. 1681
) found that mutant thymocytes, CD4+ T cells and CD8+ T cells from IBNS–/– mice, had reduced proliferation in response to plate-bound anti-CD3 mAbs, anti-CD3/anti-CD28 mAbs, and Con A compared with wild-type cells. Mutant cells carrying a TCR specific for a viral peptide had lower proliferation in response to viral peptide-loaded, irradiated splenic APCs. Microarray analysis identified eight genes that differed in expression between mutant and wild-type cells at rest and 30 genes that differed after activation. Cytokine multiplex analysis on activated mutant T cells detected reduced production of IL-2 and IFN-. Cotransfection of wild-type T cells with a vector expressing either IBNS or c-Rel increased the expression of a reporter plasmid controlled by the IL-2 gene promoter. Mutation of a NF-B site within the IL-2 promoter disrupted the IBNS but not the c-Rel effect, whereas mutation of a CD28 response element had the opposite effect. DNA pulldown experiments on nuclear lysates from virus-injected mice carrying the viral peptide-specific TCR showed that more p50 and c-Rel were made and bound to the IL-2 gene promoter up to 0.5 h after infection but binding then decreased; IBNS binding to the IL-2 gene promoter increased for at least 2 h as did IL-2 mRNA levels. The authors propose that IBNS interacts with unidentified DNA-binding proteins to form a complex that positively regulates the IL-2 promoter in TCR-activated T cells.
Stressed but Not Suppressed
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by cells activated by CpG, LPS, poly(I:C), or killed bacteria; a nonselective P1 adenosine receptor (P1R) blocking agent reversed the inhibition. Several adenosine 1 receptor (A1R) agonists had the same inhibitory effects as adenosine, and P1R or A1R antagonists reversed A1R blockages. HPLC UV scanning showed that bone marrow DCs removed adenosine from media faster than control fibroblasts; an adenosine transport inhibitor slowed the conversion of adenosine to inosine. Two anti-ADA Abs stained human splenic DCs and B cells, but not resting primary CD4+ or CD8+ T cells. Cytokine responses of activated DCs incubated in ADA-free serum also were inhibited by EHNA plus adenosine, and a coinjected ADA inhibitor prevented the movement of FITC-labeled beads mixed with killed mouse cells from the hind flanks of injected mice to draining lymph nodes. EHNA plus adenosine prevented in vitro presentation of MHC class I-restricted Ags by mouse DCs to T cells expressing peptide-specific TCRs. The data indicate that the high activity of ADA in activated DCs enables them to overcome suppression from adenosine acting on A1R. Summaries written by Dorothy L. Buchhagen, Ph.D.
Related articles in The JI:
, TGF-, and Cell Density
BNS in T Cell Cytokine Regulation As Revealed by Targeted Gene Disruption
B Activation and NF-B-Regulated Antiapoptotic Gene Products
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