MyD88-dependent TLR1/2 signals educate dendritic cells with gut-specific imprinting properties.

Gut-associated dendritic cells (DC) synthesize all-trans retinoic acid, which is required for inducing gut-tropic lymphocytes. Gut-associated DC from MyD88(-/-) mice, which lack most TLR signals, expressed low levels of retinal dehydrogenases (critical enzymes for all-trans retinoic acid biosynthesis) and were significantly impaired in their ability to induce gut-homing T cells. Pretreatment of extraintestinal DC with a TLR1/2 agonist was sufficient to induce retinal dehydrogenases and to confer these DC with the capacity to induce gut-homing lymphocytes via a mechanism dependent on MyD88 and JNK/MAPK. Moreover, gut-associated DC from TLR2(-/-) mice, or from mice in which JNK was pharmacologically blocked, were impaired in their education to imprint gut-homing T cells, which correlated with a decreased induction of gut-tropic T cells in TLR2(-/-) mice upon immunization. Thus, MyD88-dependent TLR2 signals are necessary and sufficient to educate DC with gut-specific imprinting properties and contribute in vivo to the generation of gut-tropic T cells.

CCR7 essentially contributes to the homing of plasmacytoid dendritic cells to lymph nodes under steady-state as well as inflammatory conditions.

The chemokine receptor CCR7 represents an important determinant for circulating lymphocytes to enter lymph nodes (LN) via high endothelial venules. High endothelial venules also represent the major site of entry for plasmacytoid dendritic cells (pDC). In the steady-state, murine pDC have been suggested to home to LN engaging the chemokine receptors CXCR3, CXCR4, and CCR5, whereas responsiveness to CCR7 ligands is thought to be acquired only upon activation. In this study, we show that already resting pDC express minute amounts of CCR7 that suffice to trigger migration to CCL19/CCL21 in vitro. Upon activation with TLR ligands, CCR7 levels on pDC are strongly increased. Notably, CCR7-deficient mice display substantially reduced pDC counts in LN but not in bone marrow and spleen. Adoptive cell transfer experiments revealed that under both steady-state as well as inflammatory conditions, the homing of CCR7-deficient pDC is severely impaired, indicating that the reduced cell counts of naive pDC observed in CCR7(-/-) mice reflect an intrinsic homing defect of pDC. Together, these observations provide strong evidence that similar to naive lymphocytes, nonstimulated pDC exploit CCR7 to gain entry into LN. This adds to the repertoire of chemokine receptors permitting them to enter diverse tissues.

Circulating, Mycobacterium tuberculosis-specific lymphocytes from PPD skin test-negative patients with tuberculosis do not secrete interferon-gamma (IFN-gamma) and lack the cutaneous lymphocyte antigen skin-selective homing receptor.

Individuals with a negative intradermal reaction to tuberculin PPD have long been described in the Mycobacterium tuberculosis exposed, immune-competent population. Here, we studied PPD-specific blood T lymphocytes from these subjects for phenotypic markers relevant to skin migration, including the expression of the skin-selective homing receptor, the cutaneous lymphocyte-associated antigen (CLA). Out of 82 patients with active tuberculosis we identified four subjects who were repeatedly PPD skin test-negative. CD4 T lymphocytes specific to mycobacterial antigens were derived from these individuals, which (i) proliferated in vitro to M. tuberculosis antigens comparably to those from PPD+ patients; (ii) secreted comparable amounts of IL-2 but lower amounts of IFN-gamma; (iii) were confined within the CLA-negative T cell subset. We conclude that the negative tuberculin reaction in a small subset of patients exposed to mycobacteria is associated with impaired production of IFN-gamma by circulating PPD-specific T cells that are lacking CLA expression. On this basis in vitro proliferation to PPD can discriminate bona fide non-responders from infected patients with a deficit in the margination of M. tuberculosis-specific T lymphocytes.

beta7 Integrin expression is not required for the localization of T cells to the intestine and colitis pathogenesis.

beta7 Integrins have been shown to have an important role in the localization of T cells to the intestine. Utilizing two different experimental mouse models of inflammatory bowel disease (IBD), this study was undertaken to determine if beta7 integrin expression is critical for T cell localization to the intestine and colitis pathogenesis. Transfer of CD4+ CD45RBhigh cells into immunodeficient mice results in colitis. To examine the role of beta7 integrins, donor cells were obtained from beta7 integrin gene-deficient animals and disease induction was examined following transfer into severe combined immunodeficiency (SCID) mice. Additionally, beta7 integrin gene-deficient animals were crossed to IL-2-deficient mice and the onset of spontaneous colitis that normally occurs in IL-2-deficient animals was examined. No differences in the onset or severity of spontaneous colitis was noted in animals that were deficient in both beta7 integrin and IL-2. In contrast, the onset of colitis in recipients of T cells from beta7 integrin-deficient donors was delayed significantly. In mice receiving beta7 integrin negative cells, the initial lack of colitis appeared to correlate with fewer numbers of CD3+beta7 integrin -/- donor lymphocytes present in the host colon. The eventual development of disease, however, was associated with increased numbers of donor beta7 integrin -/- lymphocytes. These results show that beta7 integrin expression is not absolutely required for T cell localization to the intestine and colitis pathogenesis.

Impaired expression of integrin alpha-4 subunit in cultured mast cells derived from mutant mice of mi/mi genotype.

The mi locus encodes a member of the basic-helix-loop-helix-leucine zipper protein family of transcription factors (hereafter called MITF). We have reported that expression of several genes was impaired in cultured mast cells (CMCs) of mi/mi mice due to a defective transactivation ability of mutant MITF (mi-MITF). Because attachment of mi/mi CMCs to fibroblasts is impaired, we examined the expression of integrin genes in mi/mi CMCs in the present study. Among the integrin genes examined, the expression of integrin alpha4 subunit was barely detectable in mi/mi CMCs, and the alpha4 protein was not detected by flow cytometry either. The specific adhesion to vascular cell adhesion molecule-1 (VCAM-1), the ligand for alpha4 subunit, was observed in +/+ CMCs but not in mi/mi CMCs, indicating that the expression of integrin alpha4 subunit at a functional level did not occur in mi/mi CMCs. In the promoter region of the alpha4 subunit gene, there was a CACTTG motif to which normal MITF (+- MITF) bound. The coexpression of +-MITF but not of mi-MITF transactivated the promoter of the alpha4 subunit gene. The deletion or mutation of the CACTTG motif abolished the transactivation by +-MITF, suggesting that +-MITF directly transactivated the gene encoding alpha4 subunit of integrin.

High frequency of adhesion defects in B-lineage acute lymphoblastic leukemia.

Aberrant proliferation, differentiation, and/or migration of progenitors observed in various hematological malignancies may be caused by defects in expression and/or function of integrins. In this study, we have developed a new fluorescent beads adhesion assay that facilitates flow cytometric investigation of lymphocyte function-associated antigen 1 (LFA-1)- and very late activation antigen-4 (VLA-4)-mediated functional adhesion in B-lineage acute lymphoblastic leukemia (ALL) of both the CD10(-) and CD10(+) (leukemic) cell population within one blood or bone marrow sample. Surprisingly, of the 20 B-lineage ALL patients investigated, 17 contained a leukemic cell population with LFA-1- and/or VLA-4-mediated adhesion defects. Five patients contained CD10(+) cells that did not exhibit any LFA-1-mediated adhesion due to the lack of LFA-1 surface expression. The CD10(+) cells from 10 ALL patients expressed LFA-1 that could not be activated by the phorbol ester phorbol 12-myristate 13-acetate (PMA), whereas the CD10(-) cells expressed a functional LFA-1. Seven patients contained CD10(+) cells that expressed a PMA-unresponsive form of VLA-4. The PMA unresponsiveness of the integrins LFA-1 and VLA-4 expressed by the CD10(+) cells may be due to mutations in the integrins itself, in protein kinases, or in other intracellular molecules involved in integrin adhesion. These data clearly demonstrate the importance of investigating integrin function in addition to integrin surface expression. The strikingly high frequency (85%) of adhesion defects in ALL could suggest a causal relationship between integrin-mediated adhesion and B-lineage ALL.