Key Points• Exosomal NKp30-ligand BAG6 is crucial for detection of tumor cells by NK cells in vitro and in vivo.• Soluble plasma factors including BAG6 suppress NK cell cytotoxicity and promote evasion of CLL cells from NK cell anti-tumor activity.Natural killer (NK) cells are a major component of the anti-tumor immune response. NK cell dysfunctions have been reported in various hematologic malignancies, including chronic lymphocytic leukemia (CLL). Here we investigated the role of tumor cellreleased soluble and exosomal ligands for NK cell receptors that modulate NK cell activity. Soluble CLL plasma factors suppressed NK cell cytotoxicity and downregulated the surface receptors CD16 and CD56 on NK cells of healthy donors. The inhibition of NK cell cytotoxicity was attributed to the soluble ligand BAG6/BAT3 that engages the activating receptor NKp30 expressed on NK cells. Soluble BAG6 was detectable in the plasma of CLL patients, with the highest levels at the advanced disease stages. In contrast, NK cells were activated when BAG6 was presented on the surface of exosomes. The latter form was induced in non-CLL cells by cellular stress via an nSmase2-dependent pathway. Such cells were eliminated by lymphocytes in a xenograft tumor model in vivo. Here, exosomal BAG6 was essential for tumor cell killing because BAG6-deficient cells evaded immune detection. Taken together, the findings show that the dysregulated balance of exosomal vs soluble BAG6 expression may cause immune evasion of CLL cells. (Blood. 2013;121(18):3658-3665) IntroductionChronic lymphocytic leukemia (CLL) patients suffer from severe immune defects resulting in increased susceptibility to infections and failure to generate an anti-tumor immune response. 1 Natural killer (NK) cells, lymphocytes of the innate immune system, are considered to be a major component of the immunosurveillance in leukemia. [2][3][4] However, little is known about the functionality of NK cells and their role in tumor immune escape in CLL.NK cells are tightly regulated by inhibitory or activating "missing self" and "induced self" signals sensed via cell surface receptors. 5 The best examined activating receptors are the Fc receptor CD16, NKG2D, and the natural cytotoxicity receptors (NCRs) NKp30, NKp44, and NKp46. Known ligands for NKG2D are the major histocompatibility complex (MHC) class I-related molecules MICA/B and the UL16-binding proteins (ULBP1, ULBP2, ULBP3, ULBP4, ULBP5, and ULBP6) that are induced upon cellular stress on target cells. 6,7 Only a few ligands for the NCRs have been identified to date. [8][9][10][11][12][13][14] Surprisingly, among novel ligands for NKp30 (BAG6 [BAT3], 10 B7-H6 11 ), NKp44 (proliferating cell nuclear antigen 12 ) and NKp46 (vimentin 13,14 ), only B7-H6 is a surface membrane ligand. BAG6, proliferating cell nuclear antigen, and vimentin are proteins without any classical transmembrane domain and are known to exert divergent intracellular functions, including protein sorting and transport, proliferation, and apoptosis. It is still ...

Active myofibroblast (MF) contraction contributes significantly to the increased intrahepatic vascular resistance that is the primary cause of portal hypertension (PHT) in cirrhosis. We sought proof of concept for direct therapeutic targeting of the dynamic component of PHT and markers of MF activation using short-term administration of the peptide hormone relaxin (RLN). We defined the portal hypotensive effect in rat models of sinusoidal PHT and the expression, activity, and function of the RLN-receptor signaling axis in human liver MFs. The effects of RLN were studied after 8 and 16 weeks carbon tetrachloride intoxication, following bile duct ligation, and in tissue culture models. Hemodynamic changes were analyzed by direct cannulation, perivascular flowprobe, indocyanine green imaging, and functional magnetic resonance imaging. Serum and hepatic nitric oxide (NO) levels were determined by immunoassay. Hepatic inflammation was assessed by histology and serum markers and fibrosis by collagen proportionate area. Gene expression was analyzed by quantitative reversetranscription polymerase chain reaction (qRT-PCR) and western blotting and hepatic stellate cell (HSC)-MF contractility by gel contraction assay. Increased expression of RLN receptor (RXFP1) was shown in HSC-MFs and fibrotic liver diseases in both rats and humans. RLN induced a selective and significant reduction in portal pressure in pathologically distinct PHT models, through augmentation of intrahepatic NO signaling and a dramatic reduction in contractile filament expression in HSC-MFs. Critical for translation, RLN did not induce systemic hypotension even in advanced cirrhosis models. Portal blood flow and hepatic oxygenation were increased by RLN in early cirrhosis. Treatment of human HSC-MFs with RLN inhibited contractility and induced an antifibrogenic phenotype in an RXFP1-dependent manner. Conclusion: We identified RXFP1 as a potential new therapeutic target for PHT and MF activation status. (HEPATOLOGY 2014;59:1492-1504 See Editorial on Page 1223 T he dynamic component of portal hypertension (PHT), mediated by active contraction of myofibroblasts (MFs) predominantly derived from activated hepatic stellate cells (HSCs), 1,2 accounts for about 30% of the increased intrahepatic vascular resistance (IHVR) in cirrhosis and is potentially reversible. 3,4 The increase in sinusoidal tone reflects a functional disturbance of the liver circulation in response to excessive production of contractile agonists (e.g., endothelin-1Abbreviations: a-SMA, alpha-smooth muscle actin; ALT, alanine aminotransferase; BDL, bile duct ligation; BOLD-MRI, blood oxygen dependent-magnetic resonance imaging; CCl4, carbon tetrachloride; CPA, collagen proportionate area; (e)NOS, (endothelial) nitric oxide synthase; ET-1, endothelin-1; (H2)-RLN, (human2)-relaxin; (h)HSC, (human) hepatic stellate cell; ICG, indocyanine green; IHVR, intrahepatic vascular resistance; L-NAME, L-N G -Nitroarginine methyl ester; MAP, mean arterial pressure; MF, myofibroblast; NO, nitric oxide; PHT...

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