TREM-1 regulates macrophage polarization in ureteral obstruction.

Chronic kidney disease (CKD) is an emerging worldwide public health problem. Inflammatory cell infiltration and activation during the early stages in injured kidneys is a common pathologic feature of CKD. Here, we determined whether an important inflammatory regulator, triggering receptor expressed on myeloid cells (TREM)-1, is upregulated in renal tissues collected from mouse ureteral obstruction-induced nephritis. TREM-1 is crucial for modulating macrophage polarization, and has a pivotal role in mediating tubular injury and interstitial collagen deposition in obstructive nephritis. Lysates from nephritic kidneys triggered a TREM-1-dependent M1 polarization ex vivo, consistent with the observation that granulocyte-macrophage colony-stimulating factor (GM-CSF)-derived M1 macrophages express higher levels of TREM-1 in comparison with M-CSF-derived cells. Moreover, agonistic TREM-1 cross-link significantly strengthens the inductions of iNOS and GM-CSF in M1 cells. These observations are validated by a strong clinical correlation between infiltrating TREM-1-expressing/iNOS-positive macrophages and renal injury in human obstructive nephropathy. Thus, TREM-1 may be a potential diagnostic and therapeutic target in human kidney disease.

CLEC4F is an inducible C-type lectin in F4/80-positive cells and is involved in alpha-galactosylceramide presentation in liver.

CLEC4F, a member of C-type lectin, was first purified from rat liver extract with high binding affinity to fucose, galactose (Gal), N-acetylgalactosamine (GalNAc), and un-sialylated glucosphingolipids with GalNAc or Gal terminus. However, the biological functions of CLEC4F have not been elucidated. To address this question, we examined the expression and distribution of murine CLEC4F, determined its binding specificity by glycan array, and investigated its function using CLEC4F knockout (Clec4f-/-) mice. We found that CLEC4F is a heavily glycosylated membrane protein co-expressed with F4/80 on Kupffer cells. In contrast to F4/80, CLEC4F is detectable in fetal livers at embryonic day 11.5 (E11.5) but not in yolk sac, suggesting the expression of CLEC4F is induced as cells migrate from yolk cells to the liver. Even though CLEC4F is not detectable in tissues outside liver, both residential Kupffer cells and infiltrating mononuclear cells surrounding liver abscesses are CLEC4F-positive upon Listeria monocytogenes (L. monocytogenes) infection. While CLEC4F has strong binding to Gal and GalNAc, terminal fucosylation inhibits CLEC4F recognition to several glycans such as Fucosyl GM1, Globo H, Bb3∼4 and other fucosyl-glycans. Moreover, CLEC4F interacts with alpha-galactosylceramide (α-GalCer) in a calcium-dependent manner and participates in the presentation of α-GalCer to natural killer T (NKT) cells. This suggests that CLEC4F is a C-type lectin with diverse binding specificity expressed on residential Kupffer cells and infiltrating monocytes in the liver, and may play an important role to modulate glycolipids presentation on Kupffer cells.

Decoy receptor 3 enhances tumor progression via induction of tumor-associated macrophages.

Tumor-associated macrophages (TAMs) are the major component of tumor-infiltrating leukocytes. TAMs are heterogeneous, with distinct phenotypes influenced by the microenvironment surrounding tumor tissues. Decoy receptor 3 (DcR3), a member of the TNFR superfamily, is overexpressed in tumor cells and is capable of modulating host immunity as either a neutralizing decoy receptor or an effector molecule. Upregulation of DcR3 has been observed to correlate with a poor prognosis in various cancers. However, the mechanisms underlying the DcR3-mediated tumor-promoting effect remain unclear. We previously demonstrated that DcR3 modulates macrophage activation toward an M2-like phenotype in vitro and that DcR3 downregulates MHC class II expression in TAMs via epigenetic control. To investigate whether DcR3 promotes tumor growth, CT26-DcR3 stable transfectants were established. Compared with the vector control clone, DcR3-transfectants grew faster and resulted in TAM infiltration. We further generated CD68 promoter-driven DcR3 transgenic (Tg) mice to investigate tumor growth in vivo. Compared with wild-type mice, macrophages isolated from DcR3-Tg mice displayed higher levels of IL-10, IL-1ra, Ym1, and arginase activity, whereas the expression of IL-12, TNF-α, IL-6, NO, and MHC class II was downregulated. Significantly enhanced tumor growth and spreading were observed in DcR3-Tg mice, and the enhanced tumor growth was abolished by arginase inhibitor N-ω-hydroxy-l-norarginine and histone deacetylase inhibitor sodium valproate. These results indicated that induction of TAMs is an important mechanism for DcR3-mediated tumor progression. Our findings also suggest that targeting DcR3 might help in the development of novel treatment strategies for tumors with high DcR3 expression.

Humoral immunity against capsule polysaccharide protects the host from magA+ Klebsiella pneumoniae-induced lethal disease by evading Toll-like receptor 4 signaling.

Klebsiella pneumoniae magA (for mucoviscosity-associated gene A) is linked to the pathogenesis of primary pyogenic liver abscess, but the underlying mechanism by which magA increases pathogenicity is not well elucidated. In this study, we investigated the role of the capsular polysaccharides (CPS) in the pathogenesis of magA(+) K. pneumoniae by comparing host immunity to magA(+) K. pneumoniae and a DeltamagA mutant. We found that Toll-like receptor 4 recognition by magA(+) K. pneumoniae was hampered by the mucoviscosity of the magA(+) K. pneumoniae CPS. Interestingly, monoclonal antibodies (MAbs) against magA(+) K. pneumoniae CPS recognized all of the K1 strains tested but not the DeltamagA and non-K1 strains. Moreover, the anti-CPS MAbs protected mice from magA(+) K. pneumoniae-induced liver abscess formation and lethality. This indicates that the K1 epitope is a promising target for vaccine development, and anti-CPS MAbs has great potential to protect host from K1 strain-induced mortality and morbidity in diabetic and other immunocompromised patients in the future.

Epigenetic control of MHC class II expression in tumor-associated macrophages by decoy receptor 3.

Decoy receptor 3 (DcR3) is a member of the TNF receptor superfamily and is up-regulated in tumors originating from a diversity of lineages. DcR3 is capable of promoting angiogenesis, inducing dendritic cell apoptosis, and modulating macrophage differentiation. Since tumor-associated macrophages (TAMs) are the major infiltrating leukocytes in most malignant tumors, we used microarray technology to investigate whether DcR3 contributes to the development of TAMs. Among the DcR3-modulated genes expressed by TAMs, those that encode proteins involved in MHC class II (MHC-II)-dependent antigen presentation were down-regulated substantially, together with the master regulator of MHC-II expression (the class II transactivator, CIITA). The ERK- and JNK-induced deacetylation of histones associated with the CIITA promoters was responsible for DcR3-mediated down-regulation of MHC-II expression. Furthermore, the expression level of DcR3 in cancer cells correlated inversely with HLA-DR levels on TAMs and with the overall survival time of pancreatic cancer patients. The role of DcR3 in the development of TAMs was further confirmed using transgenic mice overexpressing DcR3. This elucidates the molecular mechanism of impaired MHC-II-mediated antigen presentation by TAMs, and raises the possibility that subversion of TAM-induced immunosuppression via inhibition of DcR3 expression might represent a target for the design of new therapeutics.

Apoptosis of dendritic cells induced by decoy receptor 3 (DcR3).

Decoy receptor 3 (DcR3) is a soluble decoy receptor belonging to the tumor necrosis factor receptor (TNFR) superfamily, and its expression is not only up-regulated in cancer cells derived from various cell lineages, but also correlates with overall survival of patients with cancer. It has been shown that DcR3 sensitize cells of hematopoietic origin to TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis; therefore, we asked whether DcR3 down-regulated host immunity by inducing immune cell apoptosis. We demonstrate that DcR3 induces dendritic cell (DC) apoptosis by activating PKC-delta and JNK subsequently to up-regulate DR5 to recruit Fas-associated death domain (FADD) to propagate the apoptotic signals. The association of FADD with DR5 results in the formation of death-inducing signaling complex (DISC) to trigger the downstream apoptotic signaling cascade. PKC-delta is activated via cross-linking of heparan sulfate proteoglycan (HSPG) on DCs, because recombinant protein containing the heparin-binding domain (HBD) of DcR3 and the Fc portion of IgG(1), the HBD.Fc fusion protein, is also able to trigger DC apoptosis. This provides the first evidence that cross-linking of HSPG on DCs can activate PKC-delta to induce DC apoptosis via the formation of DR5 DISC, and elucidates a novel mechanism of DcR3-mediated immunosuppression.

Attenuation of Th1 response in decoy receptor 3 transgenic mice.

The soluble decoy receptor 3 (DcR3) is a member of the TNFR superfamily. Because DcR3 is up-regulated in tumor tissues and is detectable in the sera of cancer patients, it is regarded as an immunosuppressor to down-regulate immune responses. To understand the function of DcR3 in vivo, we generated transgenic mice overexpressing DcR3 systemically. In comparison with HNT-TCR (HNT) transgenic mice, up-regulation of IL-4 and IL-10 and down-regulation of IFN-gamma, IL-12, and TNF-alpha were observed in the influenza hemagglutinin(126-138) peptide-stimulated splenocytes of HNT-DcR3 double-transgenic mice. When infected with Listeria monocytogenes, DcR3 transgenic mice show attenuated expression of IFN-gamma as well as increased susceptibility to infection. The Th2 cell-biased phenotype in DcR3 transgenic mice is attributed to decreased IL-2 secretion by T cells, resulting in the suppression of IL-2 dependent CD4(+) T cell proliferation. This suggests that DcR3 might help tumor growth by attenuating the Th1 response and suppressing cell-mediated immunity.