The neurotrophic effects of PCa extend into the peritumoral "normal" pancreatic areas without neuro-cancer interactions. The neurotrophic characteristics of PCa can be mimicked by in vitro analyses and reveal NGF and Artemin as potential key players in the generation of pancreatic neuropathy in PCa.

2B4 (CD244) is an important activating receptor for the regulation of natural killer (NK) cell responses.Here we show that 2B4 is heavily and differentially glycosylated in primary human NK cells and NK cell lines. The differential glycosylation could be attributed to sialic acid residues on N-and O-linked carbohydrates. Using a recombinant fusion protein of the extracellular domain of 2B4, we demonstrate that N-linked glycosylation of 2B4 is essential for the binding to its ligand CD48. In contrast, sialylation of 2B4 has a negative impact on ligand binding, as the interaction between 2B4 and CD48 is increased after the removal of sialic acids. This was confirmed in a functional assay system, where the desialylation of NK cells or the inhibition of O-linked glycosylation resulted in increased 2B4-mediated lysis of CD48-expressing tumor target cells. These data demonstrate that glycosylation has an important impact on 2B4-mediated NK cell function and suggest that regulated changes in glycosylation during NK cell development and activation might be involved in the regulation of NK cell responses.Natural killer cells are the first lymphoid subpopulation in the defense against tumors and viral infection. Their activity is regulated by the interplay between inhibitory receptors, most of which recognize MHC class I expression on target cells, and activating receptors, which are engaged by various ligands (1). Important inhibitory receptors are members of the killer cell immunoglobulin-like receptor family as well as CD94/NKG2 heterodimers (2). Examples for activating receptors are NKG2D (CD314), DNAM-1 (CD226), and the well characterized natural cytotoxicity receptors (NCRs) 2 NKp30 (CD337), NKp44 (CD336), and NKp46 (CD335) (2, 3). These receptors transduce their activation signal via small adaptor proteins like CD3zeta or DAP10 (4). The human activating NK cell receptor 2B4 (CD244) belongs to the SLAM-related receptor (5) and is a 365-amino acid type 1 transmembrane protein with a calculated molecular weight of 39 kDa and a pI of 9.16 (6). It has a large intracellular domain containing four immunoreceptor tyrosine-based switch motifs that recruit adaptor molecules like SAP (SH2D1A or DSHP) or EAT2 after phosphorylation (7-9). The extracellular part of 2B4 consists of an N-terminal V-set Ig domain, a membrane-proximal C2-set Ig domain, and several N-glycosylation sites (6). The ligand of 2B4 is CD48, a glycosyl-phosphatidylinositol-anchored molecule with broad expression in the hematopoietic system (10, 11). Engagement of 2B4 by its ligand results in the recruitment and clustering of the receptor into lipid rafts and phosphorylation of its immunoreceptor tyrosine-based switch motif domains (12, 13). This initiates a signaling cascade, leading to polarization and the release of cytolytic granules into the contact zone between the NK and target cell (14, 15). Although 2B4 stimulation alone is sufficient to activate IL-2 stimulated NK cells, it also serves as an important coactivatory signal for the stimulation of rest...

Marked reduction in apoptosis is a hallmark of early colon tumour growth and the vast majority of these tumours exhibit a loss of expression of the glycoprotein carcinoembryonic-antigen-related cell adhesion molecule 1 (CEACAM1). We recently reported that the CEACAM1 functions as a mediator of apoptosis implicating this cell surface protein in early tumour development. However, the mechanistic involvement of CEACAM1 in cell death pathways is unclear. Here, we show that apoptosis triggers cleavage of the long form of CEACAM1 (CEACAM1-4L) at intracellular and extracellular sites in Jurkat cells and HEK293 cells. Signalling through CEACAM1 leads to caspase activation including caspase-1 and -3 and also involves non-caspase proteases. Moreover, we provide evidence that the naturally occurring CEACAM family member CEA is an inducer of CEACAM1-mediated apoptosis in HT29 colon cancer cells, an effect that depends on the abundance of CEACAM1 on the cell surface. Together, our results demonstrate that the CEACAM1-dependent cell death pathway involves dual cleavage of CEACAM1 and caspase activation and can be activated by CEA.

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Activation of natural killer cells can be mediated by different receptors. Stimulation of the receptors 2B4, NTB-A and CRACC, members of the SLAM-related receptor family, induces cytotoxicity and cytokine production. The surface expression of 2B4 and other activating natural killer cell receptors is down-modulated after receptor engagement, which results in a weaker response to consecutive stimulation. We tested whether this regulatory mechanism applies to all SLAM-related receptors expressed by primary human natural killer cells. After co-culture with target cells expressing the respective ligands different effects on receptor surface expression were observed. While 2B4 ex-pression was strongly reduced, NTB-A showed less prominent down-modulation and the expression level of CRACC remained unchanged. The expression levels of the receptor-proximal signaling molecules SAP, EAT-2 and FynT did not change after receptor engagement. Co-culture with target cells expressing the ligands for NTB-A or CRACC had no impact on subsequent NTB-A or CRACC-mediated NK cell activation.

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