CXCL10 Can Inhibit Endothelial Cell Proliferation Independently of CXCR3

CXCL10 (or Interferon-inducible protein of 10 kDa, IP-10) is an interferon-inducible chemokine with potent chemotactic activity on activated effector T cells and other leukocytes expressing its high affinity G protein-coupled receptor CXCR3. CXCL10 is also active on other cell types, including endothelial cells and fibroblasts. The mechanisms through which CXCL10 mediates its effects on non-leukocytes is not fully understood. In this study, we focus on the anti-proliferative effect of CXCL10 on endothelial cells, and demonstrate that CXCL10 can inhibit endothelial cell proliferation in vitro independently of CXCR3. Four main findings support this conclusion. First, primary mouse endothelial cells isolated from CXCR3-deficient mice were inhibited by CXCL10 as efficiently as wildtype endothelial cells. We also note that the proposed alternative splice form CXCR3-B, which is thought to mediate CXCL10's angiostatic activity, does not exist in mice based on published mouse CXCR3 genomic sequences as an in-frame stop codon would terminate the proposed CXCR3-B splice variant in mice. Second, we demonstrate that human umbilical vein endothelial cells and human lung microvascular endothelial cells that were inhibited by CXL10 did not express CXCR3 by FACS analysis. Third, two different neutralizing CXCR3 antibodies did not inhibit the anti-proliferative effect of CXCL10. Finally, fourth, utilizing a panel of CXCL10 mutants, we show that the ability to inhibit endothelial cell proliferation correlates with CXCL10's glycosaminoglycan binding affinity and not with its CXCR3 binding and signaling. Thus, using a very defined system, we show that CXCL10 can inhibit endothelial cell proliferation through a CXCR3-independent mechanism

Colonic Patch and colonic SILT development are independent and differentially-regulated events

Intestinal lymphoid tissues have to simultaneously ensure protection against pathogens and tolerance towards commensals. Despite such vital functions, their development in the colon is poorly understood. Here, we show that the two distinct lymphoid tissues of the colon–colonic patches and colonic SILTs–can easily be distinguished based on anatomical location, developmental timeframe and cellular organization. Furthermore, whereas colonic patch development depended on CXCL13-mediated LTi cell clustering followed by LTα-mediated consolidation, early LTi clustering at SILT anlagen did not require CXCL13, CCR6 or CXCR3. Subsequent dendritic cell recruitment to and gp38+VCAM-1+ lymphoid stromal cell differentiation within SILTs required LTα; B cell recruitment and follicular dendritic cell differentiation depended on MyD88-mediated signalling, but not the microflora. In conclusion, our data demonstrate that different mechanisms, mediated mainly by programmed stimuli, induce the formation of distinct colonic lymphoid tissues, therefore suggesting that these tissues may have different functions

Non-redundant Requirement for CXCR3 Signaling during Tumoricidal T Cell Trafficking across Tumor Vascular Checkpoints

T cell trafficking at vascular sites has emerged as a key step in antitumor immunity. Chemokines are credited with guiding the multistep recruitment of CD8+ T cells across tumor vessels. However, the multiplicity of chemokines within tumors has obscured the contributions of individual chemokine receptor/chemokine pairs to this process. Moreover, recent studies have challenged whether T cells require chemokine receptor signaling at effector sites. Here, we investigate the hierarchy of chemokine receptor requirements during T cell trafficking to murine and human melanoma. These studies reveal a non-redundant role for GαI-coupled CXCR3 in stabilizing intravascular adhesion and extravasation of adoptively transferred CD8+ effectors that is indispensable for therapeutic efficacy. In contrast, functional CCR2 and CCR5 on CD8+ effectors fail to support trafficking despite the presence of intratumoral cognate chemokines. Taken together, these studies identify CXCR3-mediated trafficking at the tumor vascular interface as a critical checkpoint to effective T cell-based cancer immunotherapy

Expression of chemokine receptors on peripheral blood lymphocytes in multiple sclerosis and neuromyelitis optica

<p>Abstract</p> <p>Background</p> <p>The role of different chemokine receptors in the pathogenesis of multiple sclerosis (MS) has been extensively investigated; however, little is known about the difference in the role of chemokine receptors between the pathogenesis of neuromyelitis optica (NMO) and MS. Therefore, we examined the expression of chemokine receptors on peripheral blood lymphocytes (PBL) in MS and NMO.</p> <p>Methods</p> <p>We used flow cytometry to analyse lymphocyte subsets in 12 patients with relapsing NMO, 24 with relapsing-remitting MS during relapse, 3 with NMO and 5 with MS during remission.</p> <p>Results</p> <p>Compared with healthy controls (HC), the percentage of lymphocytes in white blood cells was significantly lower in NMO and MS patients. The percentage of T cells expressing CD4⁺CD25⁺and CD4⁺CD45RO⁺was higher, while that of CD4⁺CC chemokine receptor (CCR)3⁺(T helper 2, Th2) was significantly lower in MS patients than in HC. The ratios of CD4⁺CXC chemokine receptors (CXCR)3⁺/CD4⁺CCR3⁺(Th1/Th2) and CD8⁺CXCR3⁺/CD8⁺CCR4⁺(T cytotoxic 1, Tc1/Tc2) were higher in MS patients than in HC. The percentage of CD8⁺CXCR3⁺T cell (Tc1) and CD4⁺CXCR3⁺T cell (Th1) decreased significantly during remission in MS patients (<it>P <</it>0.05). No significant differences were identified in the expression of the chemokine receptors on PBL in NMO patients compared with MS patients and HC.</p> <p>Conclusions</p> <p>Th1 dominance of chemokine receptors on blood T cells and the correlation between CXCR3⁺T cell (Th1 and Tc1) and disease activity in MS patients were confirmed by analysing chemokines receptors on PBL. In contrast, deviation in the Th1/Th2 balance was not observed in NMO patients.</p

An IP-10 (CXCL10)-derived peptide inhibits angiogenesis

Angiogenesis plays a critical role in processes such as organ development, wound healing, and tumor growth. It requires well-orchestrated integration of soluble and matrix factors and timely recognition of such signals to regulate this process. Previous work has shown that newly forming vessels express the chemokine receptor CXC receptor 3 (CXCR3) and, activation by its ligand IP-10 (CXCL10), both inhibits development of new vasculature and causes regression of newly formed vessels. To identify and develop new therapeutic agents to limit or reverse pathological angiogenesis, we identified a 21 amino acid fragment of IP-10, spanning the α-helical domain residues 77-98, that mimic the actions of the whole IP-10 molecule on endothelial cells. Treatment of the endothelial cells with the 22 amino acid fragment referred to as IP-10p significantly inhibited VEGF-induced endothelial motility and tube formation in vitro, properties critical for angiogenesis. Using a Matrigel plug assay in vivo, we demonstrate that IP-10p both prevented vessel formation and induced involution of nascent vessels. CXCR3 neutralizing antibody was able to block the inhibitory effects of the IP-10p, demonstrating specificity of the peptide. Inhibition of endothelial function by IP-10p was similar to that described for IP-10, secondary to CXCR3-mediated increase in cAMP production, activation of PKA inhibiting cell migration, and inhibition of VEGF-mediated m-calpain activation. IP-10p provides a novel therapeutic agent that inhibits endothelial cell function thus, allowing for the modulation of angiogenesis. © 2012 Yates-Binder et al

Cure of Chronic Viral Infection and Virus-Induced Type 1 Diabetes by Neutralizing Antibodies

The use of neutralizing antibodies is one of the most successful methods to interfere with receptor–ligand interactions in vivo. In particular blockade of soluble inflammatory mediators or their corresponding cellular receptors was proven an effective way to regulate inflammation and/or prevent its negative consequences. However, one problem that comes along with an effective neutralization of inflammatory mediators is the general systemic immunomodulatory effect. It is, therefore, important to design a treatment regimen in a way to strike at the right place and at the right time in order to achieve maximal effects with minimal duration of immunosuppression or hyperactivation. In this review, we reflect on two examples of how short time administration of such neutralizing antibodies can block two distinct inflammatory consequences of viral infection. First, we review recent findings that blockade of IL-10/IL-10R interaction can resolve chronic viral infection and second, we reflect on how neutralization of the chemokine CXCL10 can abrogate virus-induced type 1 diabetes

Analysis of single nucleotide polymorphism in the promoter and protein expression of the chemokine Eotaxin-1 in colorectal cancer patients

<p>Abstract</p> <p>Background</p> <p>Previous studies suggest that chemokines (chemotactic cytokines) promote and regulate neoplastic progression including metastasis and angiogenesis. The chemokine eotaxin-1 is a powerful eosinophil attractant but also exerts chemotaxis of other leukocytes. Eotaxin-1 has been implicated in gastrointestinal disorders and may play an important role in colorectal mucosal immunity.</p> <p>Patients and methods</p> <p>The objective of this study was to assess the role of eotaxin-1 in colorectal cancer (CRC). Levels of eotaxin-1 protein in CRC tissues (n = 86) and paired normal mucosa were compared after determination by ELISA. Plasma eotaxin-1 levels from CRC patients (n = 67) were also compared with controls (n = 103) using the same method. Moreover, a TaqMan system was used to evaluate the -384A>G eotaxin-1 gene variant in CRC patients (n = 241) and in a control group (n = 253).</p> <p>Results</p> <p>Eotaxin-1 protein levels in colorectal tumours were significantly (P < 0.0001) higher than in normal tissue. Immunohistochemistry revealed eotaxin-1 expression in stromal cells such as fibroblasts and leukocytes of the CRC tissue. The plasma eotaxin-1 level in CRC patients was lower compared with controls (P < 0.0001). Patients with tumours classified as Dukes' stage B and C had lower levels than patients with tumours in Dukes' stage A. We found no difference in genotype distribution but noted a difference regarding allele distribution (P = 0.036) and a dominance of allele G in rectal cancer patients.</p> <p>Conclusion</p> <p>The up-regulated eotaxin-1 protein expression in cancer tissue may reflect an eotaxin-1 mediated angiogenesis and/or a recruitment of leukocytes with potential antitumourigenic role. We noticed a dominance of the G allele in rectal cancer patients compared with colon cancer patients that was independent of eotaxin-1 expression.</p