In vitro culture with gemcitabine augments death receptor and NKG2D ligand expression on tumour cells

Much effort has been made to try to understand the relationship between chemotherapeutic treatment of cancer and the immune system. Whereas much of that focus has been on the direct effect of chemotherapy drugs on immune cells and the release of antigens and danger signals by malignant cells killed by chemotherapy, the effect of chemotherapy on cells surviving treatment has often been overlooked. In the present study, tumour cell lines: A549 (lung), HCT116 (colon) and MCF-7 (breast), were treated with various concentrations of the chemotherapeutic drugs cyclophosphamide, gemcitabine (GEM) and oxaliplatin (OXP) for 24 hours in vitro. In line with other reports, GEM and OXP upregulated expression of the death receptor CD95 (fas) on live cells even at sub-cytotoxic concentrations. Further investigation revealed that the increase in CD95 in response to GEM sensitised the cells to fas ligand treatment, was associated with increased phosphorylation of stress activated protein kinase/c-Jun N-terminal kinase and that other death receptors and activatory immune receptors were co-ordinately upregulated with CD95 in certain cell lines. The upregulation of death receptors and NKG2D ligands together on cells after chemotherapy suggest that although the cells have survived preliminary treatment with chemotherapy they may now be more susceptible to immune cell-mediated challenge. This re-enforces the idea that chemotherapy-immunotherapy combinations may be useful clinically and has implications for the make-up and scheduling of such treatments

TREM-1 links dyslipidemia to inflammation and lipid deposition in atherosclerosis.

Triggering receptor expressed on myeloid cells-1 (TREM-1) is a potent amplifier of pro-inflammatory innate immune responses, but its significance in non-infectious diseases remains unclear. Here, we demonstrate that TREM-1 promotes cardiovascular disease by exacerbating atherosclerosis. TREM-1 is expressed in advanced human atheromas and is highly upregulated under dyslipidemic conditions on circulating and on lesion-infiltrating myeloid cells in the Apoe(-/-) mouse model. TREM-1 strongly contributes to high-fat, high-cholesterol diet (HFCD)-induced monocytosis and synergizes with HFCD serum-derived factors to promote pro-inflammatory cytokine responses and foam cell formation of human monocyte/macrophages. Trem1(-/-)Apoe(-/-) mice exhibit substantially attenuated diet-induced atherogenesis. In particular, our results identify skewed monocyte differentiation and enhanced lipid accumulation as novel mechanisms through which TREM-1 can promote atherosclerosis. Collectively, our findings illustrate that dyslipidemia induces TREM-1 surface expression on myeloid cells and subsequently synergizes with TREM-1 to enhance monopoiesis, pro-atherogenic cytokine production and foam cell formation

Cellular expression, trafficking, and function of two isoforms of human ULBP5/RAET1G

Background: The activating immunoreceptor NKG2D is expressed on Natural Killer (NK) cells and subsets of T cells. NKG2D contributes to anti-tumour and anti-viral immune responses in vitro and in vivo. The ligands for NKG2D in humans are diverse proteins of the MIC and ULBP/RAET families that are upregulated on the surface of virally infected cells and tumours. Two splicing variants of ULBP5/RAET1G have been cloned previously, but not extensively characterised. Methodology/Principal Findings: We pursue a number of approaches to characterise the expression, trafficking, and function of the two isoforms of ULBP5/RAET1G. We show that both transcripts are frequently expressed in cell lines derived from epithelial cancers, and in primary breast cancers. The full-length transcript, RAET1G1, is predicted to encode a molecule with transmembrane and cytoplasmic domains that are unique amongst NKG2D ligands. Using specific anti-RAET1G1 antiserum to stain tissue microarrays we show that RAET1G1 expression is highly restricted in normal tissues. RAET1G1 was expressed at a low level in normal gastrointestinal epithelial cells in a similar pattern to MICA. Both RAET1G1 and MICA showed increased expression in the gut of patients with celiac disease. In contrast to healthy tissues the RAET1G1 antiserum stained a wide variety or different primary tumour sections. Both endogenously expressed and transfected RAET1G1 was mainly found inside the cell, with a minority of the protein reaching the cell surface. Conversely the truncated splicing variant of RAET1G2 was shown to encode a soluble molecule that could be secreted from cells. Secreted RAET1G2 was shown to downregulate NKG2D receptor expression on NK cells and hence may represent a novel tumour immune evasion strategy. Conclusions/Significance: We demonstrate that the expression patterns of ULBP5RAET1G are very similar to the well-characterised NKG2D ligand, MICA. However the two isoforms of ULBP5/RAET1G have very different cellular localisations that are likely to reflect unique functionality

New insights about host response to smallpox using microarray data

<p>Abstract</p> <p>Background</p> <p>Smallpox is a lethal disease that was endemic in many parts of the world until eradicated by massive immunization. Due to its lethality, there are serious concerns about its use as a bioweapon. Here we analyze publicly available microarray data to further understand survival of smallpox infected macaques, using systems biology approaches. Our goal is to improve the knowledge about the progression of this disease.</p> <p>Results</p> <p>We used KEGG pathways annotations to define groups of genes (or modules), and subsequently compared them to macaque survival times. This technique provided additional insights about the host response to this disease, such as increased expression of the cytokines and ECM receptors in the individuals with higher survival times. These results could indicate that these gene groups could influence an effective response from the host to smallpox.</p> <p>Conclusion</p> <p>Macaques with higher survival times clearly express some specific pathways previously unidentified using regular gene-by-gene approaches. Our work also shows how third party analysis of public datasets can be important to support new hypotheses to relevant biological problems.</p

Thermal- and Oxidative Stress Causes Enhanced Release of NKG2D Ligand-Bearing Immunosuppressive Exosomes in Leukemia/Lymphoma T and B Cells

Immune evasion from NK surveillance related to inadequate NK-cell function has been suggested as an explanation of the high incidence of relapse and fatal outcome of many blood malignancies. In this report we have used Jurkat and Raji cell lines as a model for studies of the NKG2D receptor-ligand system in T-and B cell leukemia/lymphoma. Using real-time quantitative RT-PCR and immunoflow cytometry we show that Jurkat and Raji cells constitutively express mRNA and protein for the stress-inducible NKG2D ligands MICA/B and ULBP1 and 2, and up-regulate the expression in a cell-line specific and stress-specific manner. Furthermore, we revealed by electron microscopy, immunoflow cytometry and western blot that these ligands were expressed and secreted on exosomes, nanometer-sized microvesicles of endosomal origin. Acting as a decoy, the NKG2D ligand-bearing exosomes downregulate the in vitro NKG2D receptor-mediated cytotoxicity and thus impair NK-cell function. Interestingly, thermal and oxidative stress enhanced the exosome secretion generating more soluble NKG2D ligands that aggravated the impairment of the cytotoxic response. Taken together, our results might partly explain the clinically observed NK-cell dysfunction in patients suffering from leukemia/lymphoma. The adverse effect of thermal and oxidative stress, enhancing the release of immunosuppressive exosomes, should be considered when cytostatic and hyperthermal anti-cancer therapies are designed

Semaphorins and their receptors: Novel features of neural guidance molecules

Semaphorins were originally identified as axon guidance cues involved in the development of the nervous system. In recent years, it is emerging that they also participate in various biological systems, including physiological and pathological processes. In this review, we primarily focus on our cumulative findings for the role of semaphorins and their receptors in the regulation of the immune system, while also summarizing recent progress in the context of cardiovascular system

NKG2D triggers cytotoxicity in mouse NK cells lacking DAP12 or Syk family kinases

In activated mouse natural killer (NK) cells, the NKG2D receptor associates with two intracellular adaptors, DAP10 and DAP12, which trigger phosphatidyl inositol 3 kinase (PI3K) and Syk family protein tyrosine kinases, respectively. Here we show that cytotoxicity, but not cytokine production, is triggered by NKG2D in activated NK cells lacking either DAP12 or the Syk family members Syk and ZAP70. Inhibition of PI3K blocks this cytotoxicity, suggesting that the DAP10-PI3K pathway is sufficient to initiate NKG2D-mediated killing of target cells. Our results highlight signaling divergence in the effector functions of NKG2D and indicate that alternative associations between a receptor and its adaptors may provide a single receptor with a dual 'on-switch', giving mouse NK cells more choices through which to trigger cytotoxicity

The NKG2D Ligands RAE-1δ and RAE-1ε Differ with Respect to Their Receptor Affinity, Expression Profiles and Transcriptional Regulation

BACKGROUND: RAE-1 is a ligand of the activating receptor NKG2D expressed by NK cells, NKT, γδT and some CD8(+)T lymphocytes. RAE-1 is overexpressed in tumor cell lines and its expression is induced after viral infection and genotoxic stress. We have recently demonstrated that RAE-1 is expressed in the adult subventricular zone (SVZ) from C57BL/6 mice. RAE-1 is also expressed in vitro by neural stem/progenitor cells (NSPCs) and plays a non-immune role in cell proliferation. The C57BL/6 mouse genome contains two rae-1 genes, rae-1δ and rae-1ε encoding two different proteins. The goals of this study are first to characterize the in vivo and in vitro expression of each gene and secondly to elucidate the mechanisms underlying their respective expression, which are far from known. PRINCIPAL FINDINGS: We observed that Rae-1δ and Rae-1ε transcripts are differentially expressed according to tissues, pathological conditions and cell lines. Embryonic tissue and the adult SVZ mainly expressed Rae-1δ transcripts. The NSPCs derived from the SVZ also mainly expressed RAE-1δ. The interest of this result is especially related to the observation that RAE-1δ is a weak NKG2D ligand compared to RAE-1ε. On the contrary, cell lines expressed either similar levels of RAE-1δ and RAE-1ε proteins or only RAE-1ε. Since the protein expression correlated with the level of transcripts for each rae-1 gene, we postulated that transcriptional regulation is one of the main processes explaining the difference between RAE-1δ and RAE-1ε expression. We indeed identified two different promoter regions for each gene: one mainly involved in the control of rae-1δ gene expression and the other in the control of rae-1ε expression. CONCLUSIONS/SIGNIFICANCE: RAE-1δ and RAE-1ε differ with respect to their function and the control of their expression. Immune function would be mainly exerted by RAE-1ε and non-immune function by RAE-1δ