Human NK cells lyse Ab-coated target cells through the process of Ab-dependent cellular cytotoxicity (ADCC). Improving ADCC responses is desirable because it is thought to be an important antitumor mechanism for some Abs. NK cell inhibitory receptors, such as killer cell Ig-like receptors, engage with MHC class I molecules on self-cells to block NK cell activation. Accordingly, we enhanced ADCC responses by blocking NK cell inhibitory receptors, thus perturbing induction of the self-recognition signal. In a cell line model of anti-lymphoma therapy, the combination of rituximab with an Ab that blocks inhibitory self-recognition yielded increased NK cell-mediated target cell lysis when compared with rituximab alone. To validate this proof-of-concept, we then used a more representative approach in which an individual’s fresh primary NK cells encountered autologous, EBV-transformed B cells. In this system, rituximab and a combination of Abs that block NK cell inhibitory receptors yielded improved NK cell-mediated lysis over rituximab alone. The results show, for the first time, that disruption of inhibitory self-recognition can efficiently promote ADCC in a human model, applying an autologous system in which physiologic checkpoints are in place. This method provides an alternative approach to potentiate the therapeutic benefit of antitumor Abs that mediate ADCC.

The aim of this study is to determine the incidence of radiological pulmonary edema in elective liver transplant recipients and its relationship to perioperative factors and postoperative course. We reviewed 102 chest radiographs from 34 patients who had undergone orthotopic liver transplantation (OLT). Films were assessed by 2 trained radiologists for evidence of pulmonary edema using a standardized system. Clinical and outcome data from the 34 patients were also recorded. There was a high incidence (47%) of postoperative radiological pulmonary edema that was associated with deterioration in gaseous exchange, elevated pulmonary artery pressure, and increased duration of ventilator dependence and intensive care stay. Eighteen percent of the patients developed edema immediately after surgery, which was associated with greater pulmonary artery pressure and transfusion requirements during surgery. An additional 29% developed edema during the next 16 to 20 hours, but there was no association with fluid replacement. We conclude that pulmonary edema is common after OLT and will influence postoperative recovery in a substantial proportion of transplant recipients. Excess perioperative fluid replacement is unlikely to be the sole mechanism of edema in these patients. A fter elective orthotopic liver transplantation (OLT), pulmonary complications are an important source of morbidity that frequently impede recovery. 1,2 Their prevalence is shown by the fact that pulmonary failure is responsible for 25% of readmissions to the intensive therapy unit (ITU). 3 There is no single process accounting for the pulmonary complications, but they can be classified into infective and noninfective causes. Infective pulmonary complications become an important cause of morbidity during the first few months after transplantation but are uncommon in the early transplantation period (Ͻ2 days). Noninfectious complications, including atelectasis, pleural effusion, pulmonary edema, pulmonary embolism, and diaphragmatic dysfunction, make up the majority of early problems. Of all pulmonary complications after elective OLT, 50% to 64% 3,4 are noninfectious. This proportion is greater than that with other organ transplantation procedures. 5 Many episodes of pulmonary edema occur in patients with fulminant hepatic failure, 6 and in such patients these may predict acute lung injury. Some episodes of pulmonary edema that occur after elective OLT may represent less severe acute lung injury. These complications should perhaps be treated differently. The aim of the present study is to determine the incidence of radiological pulmonary edema after elective OLT. The relationship of edema to relevant clinical cardiorespiratory indices, transfusion requirements, and early postoperative ITU recovery was also investigated. MethodsData from 34 patients with chronic liver disease undergoing consecutive OLTs during a 1-year period were included on the study. Patients with fulminant hepatic failure were specifically excluded from the study.Patients were anesthetiz...

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KIR2DL4 (2DL4) is a member of the killer cell Ig-like receptor (KIR) family in human NK cells. It can stimulate potent cytokine production and weak cytolytic activity in resting NK cells, but the mechanism for 2DL4-mediated signaling remains unclear. In this study we characterized the signaling pathways stimulated by 2DL4 engagement. In a human NK-like cell line, KHYG-1, cross-linking of 2DL4 activated MAPKs including JNK, ERK, and p38. Furthermore, 2DL4 cross-linking resulted in phosphorylation of IkappaB kinase beta (IKKbeta) and the phosphorylation and degradation of IkappaBalpha, which indicate activation of the classical NF-kappaB pathway. Engagement of 2DL4 was also shown to activate the transcription and translation of a variety of cytokine genes, including TNF-alpha, IFN-gamma, MIP1alpha, MIP1beta, and IL-8. Pharmacological inhibitors of JNK, MEK1/2 and p38, blocked IFN-gamma, IL-8, and MIP1alpha production, suggesting that MAPKs are regulating 2DL4-mediated cytokine production in a nonredundant manner. Activation of both p38 and ERK appear to be upstream of the stimulation of NF-kappaB. Mutation of a transmembrane arginine in 2DL4 to glycine (R/G mutant) abrogated FcepsilonRI-gamma association, as well as receptor-mediated cytolytic activity and calcium responses. Surprisingly, the R/G mutant still activated MAPKs and the NF-kappaB pathway and selectively stimulated the production of MIP1alpha, but not that of IFN-gamma or IL-8. In conclusion, we provide evidence that the activating functions of 2DL4 can be compartmentalized into two distinct structural modules: 1) through transmembrane association with FcepsilonRI-gamma; and 2) through another receptor domain independent of the transmembrane arginine.

KIR2DL4 is a member of the killer cell Ig-like receptor (KIR) family expressed on human natural killer (NK) cells. A transmembrane Arginine (R) in KIR2DL4 binds to the transmembrane adaptor FcεRI-γ and stimulates potent IFN-γ production and weak cytolytic activity. In this study, we have found that the mutation of transmembrane R to Glycine (G) (RG mutant) in KIR2DL4 abrogates FcεRI-γ association, IFN-γ production, cytolytic activity and calcium responses. We further found that crosslinking of KIR2DL4 activates the MAP kinases JNK and ERK, as well as the NF-κB pathway in KHYG-1, a human NK-like cell line. Engagement of KIR2DL4 also activates the transcription of a variety of cytokine and chemokine genes including TNF-α, IFN-γ, MIP1α, MIP1β, IL-8 and RANTES. Pharmacological inhibitors of JNK and MEK1/2 blocked IFN-γ and IL-8 production, suggesting that these MAP kinases are regulating KIR2DL4 mediated cytokine production. Surprisingly, we have found that the RG mutant of KIR2DL4 still activates MAP kinases and the NF-κB pathway and differentially regulates chemokine production. In conclusion, our data suggests that the activating functions of KIR2DL4 are mediated by two distinct pathways in NK cells:through transmembrane association with FcεRI-γ andthrough its cytoplasmic tail, which is independent of the transmembrane R. This work was supported by National Institutes of Health Grant CA-100226 to K.S.C.

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