Molecular Cloning and Characterization of a Novel CXC Chemokine Macrophage Inflammatory Protein-2γ Chemoattractant for Human Neutrophils and Dendritic Cells

Abstract

Chemokines play important roles in leukocyte trafficking as well as function regulation. In this study, we described the identification and characterization of a novel CXC chemokine from a human dendritic cell (DC) cDNA library, the full-length cDNA of which contains an open reading frame encoding 111 aa with a putative signal peptide of 34 aa. This CXC chemokine shares greatest homology with macrophage inflammatory protein (MIP)-2αβ, hence is designated as MIP-2γ. Mouse MIP-2γ was identified by electrocloning and is highly homologous to human MIP-2γ. Northern blotting revealed that MIP-2γ was constitutively and widely expressed in most normal tissues with the greatest expression in kidney, but undetectable in most tumor cell lines except THP-1 cells. In situ hybridization analysis demonstrated that MIP-2γ was mainly expressed by the epithelium of tubules in the kidney and hepatocytes in the liver. Although no detectable expression was observed in freshly isolated or PMA-treated monocytes, RT-PCR analysis revealed MIP-2γ expression by monocyte-derived DC. Recombinant MIP-2γ from 293 cells is about 9.5 kDa in size and specifically detectable by its polyclonal Ab developed by the immunization with its 6His-tagged fusion protein. The eukaryotically expressed MIP-2γ is a potent chemoattractant for neutrophils, and weaker for DC, but inactive to monocytes, NK cells, and T and B lymphocytes. Receptor binding assays showed that MIP-2γ does not bind to CXCR2. This implies that DC might contribute to the innate immunity through the production of neutrophil-attracting chemokines and extends the knowledge about the regulation of DC migration.

Lipoteichoic Acid Increases TLR and Functional Chemokine Expression while Reducing Dentin Formation in In Vitro Differentiated Human Odontoblasts

“…Using gene arrays, we compared the expression of chemokines by resting vs TLR2-activated odontoblasts. We observed the expression by resting odontoblasts of CCL2, CXCL12, and CXCL14, three chemokines known to drive iDCs recruitment in vitro (11,31,32,33). However, the chemokine the most strongly expressed by resting odontoblasts was CCL26, a natural antagonist for CCR1, CCR2, and CCR5 (34).…”

Murine CXCL14 Is Dispensable for Dendritic Cell Function and Localization within Peripheral Tissues

“…Human CXCL14, alternatively called BRAK and MIP-2␥, is constitutively produced in healthy epithelial tissues such as skin and gut. CXCL14 is also present in various tumors of epithelial origin, but the level of expression is heterogeneous, with the majority showing a downregulation with respect to healthy tissue (4,9,15,19,43,44,46). Recently, we have shown that human CXCL14 is selective for blood monocytes and CD14 ϩ DC precursors, either derived from CD34 ϩ hematopoietic progenitor cells or isolated from blood (19,42).…”

“…CXCL14 is a chemokine with unknown function and receptor selectivity (4,9,15). Human CXCL14, alternatively called BRAK and MIP-2␥, is constitutively produced in healthy epithelial tissues such as skin and gut.…”

“…Cells were separated by passing them through a 40-m-pore-size cell strainer and washed in PBS-5 mM EDTA-2% fetal calf serum. Prior to staining, blood, splenocytes, and bone marrow cells were subjected to red blood cell lysis using 5 to 10 ml of lysis buffer (0.15 M NH 4 Cl, 10 mM KHCO 3 , 0.1 M EDTA). Cells were routinely incubated with 2.4G2 (BD Biosciences) or, if a secondary anti-rat antibody was used, with 600 g/ml mouse IgG (Sigma) to block Fc receptors; cells were subsequently labeled with antibodies diluted in PBS-1% bovine serum albumin-0.05% azide for 30 min on ice.…”

Human membrane protein Tim-3 facilitates hepatitis A virus entry into target cells

“…It has been shown that mouse Tim-3 is involved in the interactions between Th1 cells and macrophages, which results in the expansion and activation of macrophages. As a homologue of mouse Tim-3, (14)(15)(16). One of the full-length cDNAs encodes a 301-residue type I integral membrane glycoprotein.…”