[Successful closure of a postoperative esophagobronchial fistula following esophageal resection using fibrin glue]. / Erfolgreicher Fibrinkleber-basierter Verschluss einer postoperativen ösophagobronchialen Fistel nach Ösophagusresektion.

UNLABELLED: HISTORY AND INTERVENTION: A 52-year-old female patient underwent open abdominothoracic cardia and esophageal resection with gastric transposition because of histologically diagnosed Barrett metaplasia with "high-grade" intraepithelial neoplasia (HGIEN) and parts of an invasive adenocarcinoma. The anastomotic insufficiency on the 10th postoperative day including an esophagobronchial fistula prompted to a subsequent surgical re-intervention with suture of the fistula, lavage and additional drainage, an endoscopic stenting of the fistula from esophageal site, as well as repeated (n = 22) bronchoscopic applications of fibrin glue (1-3 ml each) into the lumen of the fistula after each bronchoscopic lavage of the fistula until the complete closure was achieved. The changeful clinical course of 77 days on the surgical ICU was characterized by secondary complications such as pneumonia, mediastinitis and respiratory insufficiency with long-term artificial respiration and creation of a percutaneous dilatation tracheotomy. CONCLUSION: The application of fibrin glue can be considered a promising, minimally invasive therapeutic option in the management of postoperative fistula after esophageal resection, which requires expertise in decision-making and the finding-specific approach, in particular, if indicated inital steps of the sequential complication management such as surgical re-intervention and conventional endoscopic measures (stenting, Endo-VAC[-sponge]) do not provide great therapeutic potential any more due to the prolonged postoperative time course and the unfavorable local findings. In the presented case, modes of an assisted artificial respiration with low pressure and short phases of apnoe after fibrin glue application were the crucial predictions for an initial and favorable adhesion of this glue and finally for a successful sealing resulting in a sufficient closure of the fistula.

Human NK cells require caspases for activation-induced proliferation and cytokine release but not for cytotoxicity.

Natural killer (NK) cells are innate immune cells involved in antiviral defence and tumour surveillance. To fulfil these tasks, NK cells make use of two major effector functions, cytokine and chemokine release and cytotoxicity. In addition, NK cells proliferate in response to cytokines such as IL-2. NK cells possess a large array of activating and inhibitory receptors and their activation demands a complex crosstalk between those receptors. The signalling pathways leading to NK-cell activation are a field of intensive research. The first clue for signal specificity was provided by studies showing that a pathway leading to NF-κB activation selectively induces cytokine release, but is dispensable for cytotoxicity. Here, we demonstrate that in human NK cells caspase activity is required for the upregulation of select activation markers and IFN-γ and TNF production, but not for cytotoxicity. Interestingly, caspases have previously been linked in T cells to the same mechanism of NF-κB induction that is active in NK cells. Moreover, we provide evidence that caspases are involved in IL-2-induced proliferation. Thus, our data provide the basis for a novel approach using caspase inhibitors to generate cytotoxic NK cells, while simultaneously suppressing cytokine release.

Activation of ERK1/2 and cPLA(2) by the p55 TNF receptor occurs independently of FAN.

The generation of proinflammatory eicosanoids in response to tumor necrosis factor (TNF) involves the activation of cytosolic phospholipase A(2) (cPLA(2)), presumably by phosphorylation through extracellular signal-regulated kinases (ERK). Earlier results had suggested that a pathway involving the p55 TNF receptor (TNF-R55), neutral sphingomyelinase (N-SMase), and c-Raf-1 activates ERK and cPLA(2). We have previously shown that a cytoplasmic region of TNF-R55 distinct from the death domain regulates the activation of N-SMase through binding of the adapter protein FAN. Analysis of embryonal fibroblasts from FAN knockout mice revealed that TNF-induced activation of both ERK and cPLA(2) occurs without involvement of FAN. Furthermore, we provide evidence that the TNF-dependent activation of ERK and cPLA(2) requires the intact death domain of TNF-R55. Finally, we demonstrate that in murine fibroblasts cPLA(2) is phosphorylated in response to TNF solely by ERK, but not by p38 mitogen-activated protein kinase, suggesting a signaling pathway from TNF-R55 via the death domain to ERK and cPLA(2).

Sensitization to death receptor cytotoxicity by inhibition of fas-associated death domain protein (FADD)/caspase signaling. Requirement of cell cycle progression.

Upon binding of their ligands, death receptors belonging to the tumor necrosis factor (TNF) receptor family initiate a signaling pathway leading to the activation of caspases and ultimately apoptosis. TNF, however, in parallel elicits survival signals, protecting many cell types from cell death that can only be induced by combined treatment with TNF and inhibitors of protein synthesis. Here, we report that in NIH3T3 cells, apoptosis in response TNF and cycloheximide is not inhibited by the broad spectrum caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (zVAD. fmk). Moreover, treatment with zVAD.fmk sensitizes the cells to the cytotoxic action of TNF. Sensitization was also achieved by overexpression of a dominant-negative mutant of Fas-associated death domain protein and, to a lesser extent, by specific inhibition of caspase-8. A similar, but weaker sensitization of zVAD.fmk to treatment with the TNF-related apoptosis-inducing ligand (TRAIL) or anti-CD95 antibody was demonstrated. The unexpected cell death in response to TNF and caspase inhibition occurs despite the activation of nuclear factor kappaB and c-Jun N-terminal kinases. The mode of cell death shows several signs of apoptosis including DNA fragmentation, although activation of caspase-3 was excluded. TNF/zVAD.fmk-induced cell death is preceded by an accumulation of cells in the G(2)/M phase of the cell cycle, indicating an important role of cell cycle progression. This hypothesis is further strengthened by the observation that arresting the cells in the G(1) phase of the cell cycle inhibited TNF/zVAD.fmk-induced cell death, whereas blocking them in the G(2)/M phase augmented it.

Caspase-mediated inhibition of human cytosolic phospholipase A2 during apoptosis.

Activation of cytosolic phospholipase A2 (cPLA2) is an essential step in the initiation of the cascade of enzymatic reactions leading to the generation of proinflammatory lipid mediators. Hence, the regulation of cPLA2 is a key event in the induction of inflammatory responses. cPLA2 is activated, in part, by apoptotic stimuli such as TNF or Fas ligand. Apoptosis, however, does not provoke an inflammatory response. Here, we demonstrate that cPLA2 is cleaved by caspase-3 and/or a related caspase in HeLa cells undergoing apoptosis. Mutation of a predicted caspase-3 cleavage site abolishes cPLA2 processing both in vitro and in intact cells. The 70-kDa cleavage product of cPLA2 itself has no catalytic function, while inhibition of cleavage results in an increased enzymatic activity. Additionally, overexpression of the 70-kDa fragment appears to produce a dominant negative effect on endogenous cPLA2 activity. In HeLa cells, cPLA2 activity was dispensable for the course of apoptosis. We cannot rule out, however, that cPLA2 activity is involved in the induction of apoptosis in other cell types. Taken together, our results suggest that the enzymatic activity of cPLA2 is specifically inhibited by caspase-mediated cleavage during apoptosis. The inactivation of cPLA2 represents a previously unrecognized mechanism for avoiding an inflammatory reaction against apoptotic cells.

Cleavage of human cytosolic phospholipase A2 by caspase-1 (ICE) and caspase-8 (FLICE).

The activation of caspases appears to play a key role in programmed cell death. An increasing number of substrates have been identified that are cleaved by caspases. In a previous study, we have reported that human cPLA2 is proteolytically inactivated during apoptosis through cleavage by a caspase-3-like activity. Here, we show that in cotransfection experiments the previously identified cleavage site at Asp522 can be used by a wide variety of caspases belonging to different subfamilies. The formation of additional fragments implied differences in cleavage site usage between the closely related caspases-3 and -7. A different cleavage pattern of cPLA2 was observed with caspase-1. Mutational analysis identified the caspase-1 cleavage site at Asp459 within the sequence YQSD/N. Most interestingly, we found that even caspase-8, an upstream component of the proposed caspase cascade, cleaves cPLA2 in vitro. The presence of multiple cleavage sites warrants proteolysis and inactivation of the proinflammatory cPLA2 during apoptosis.

Molecular forms of soluble HLA in body fluids: potential determinants of body odor cues.

The major histocompatibility complex (MHC) has been linked to encoding for individual olfactory identity. Experiments in mice and rats proved that behavior and mating were, at least in part, determined by genes within the MHC. This study was aimed at investigating whether sHLA are excreted in human urine, saliva and sweat. In particular examination of the molecular forms in these fluids would give clues to whether break down forms of soluble MHC molecules might participate in shaping behavior. Major bands of 45, 40, and 23 kD were detectable. Increased levels of sHLA were measured using a quantitative ELISA in urine shortly before ovulation decreasing to normal levels thereafter. In animal models strain specific MHC-linked odor cues have been detected in urine. Thus, excretion of sHLA in urine might indicate a similar role for these molecules in humans.