Extracellular release of granzymes is considered to reflect the involvement of cytotoxic T lymphocytes and NK cells in various disease states. To obtain insight into granzyme release during bacterial infection, granzyme levels were measured during experimental human endotoxemia and in patients with melioidosis, a severe infection due to gram-negative bacteria. Plasma concentrations of granzyme A (GrA) and GrB increased transiently after endotoxin administration, peaking after 2-6 h. In patients with bacteremic melioidosis, GrA and GrB levels were elevated on admission and remained high during the 72-h study period. In whole blood stimulated with heat-killed Burkholderia pseudomallei, neutralization of tumor necrosis factor, interleukin-12, or interleukin-18 inhibited granzyme secretion, which was independent of interferon-gamma. Stimulation with endotoxin and other gram-negative and gram-positive bacteria also strongly induced the secretion of granzymes, suggesting that granzyme release is a general immune response during bacterial infection. The interaction between the cytokine network and granzymes may play an important immunoregulatory role during bacterial infections.

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Granzymes (Grs), serine proteases present in granules of effector lymphocytes, are involved in several host immune responses, including the activation of cell death and inflammatory pathways. The main goal of this study was to determine whether the local cell-mediated Gr pathway is activated during severe respiratory syncytial virus (RSV) lower respiratory tract illness (LRTI) in children. Tracheal aspirates (TA) from 23 children with RSV-LRTI and 12 controls without pulmonary disease were analyzed for Gr A and B. Bronchoalveolar lavage fluid samples from seven children with RSV-LRTI were analyzed for cellular expression of GrB. Levels of GrA and GrB in TA were significantly increased in RSV patients compared with controls and both Grs showed preserved activity. Gr levels correlated with the total leukocyte counts and IL-8 levels in the airways at several time points. However, no correlation between Gr levels and release of caspase-cleaved cytokeratin-18 was found. There was evidence for marked expression of GrB by both CD8 ϩ and CD4 ϩ T cells and natural killer cells in the respiratory tract. These findings suggest activation of the cell-mediated Gr pathway during severe

Granzymes are serine proteases released from the granules of cytotoxic lymphocytes during the induction of apoptosis. To evaluate the physiologic role of human granzyme K (GzmK), we developed a sensitive ELISA which was shown to specifically detect human GzmK in its active as well as its inactive conformation. Analysis of the lysate of lymphokine‐activated killer (LAK) cells by gel filtration revealed that GzmK seems to be complexed to proteoglycans within these cells. While the expression of GzmA and B by cytotoxic lymphocytes was strongly up‐regulated in response to several activating stimuli, GzmK expression did not increase significantly above constitutive levels, indicating differential regulation of these granzymes. However, low levels of GzmK were detected in plasma samples of healthy volunteers, which were in the same range as levels of GzmA and B. Furthermore, circulating levels of GzmK as well as of GzmA and B were significantly elevated in patients suffering from viral infections. We conclude that GzmK protein is produced by cytotoxic cells, and just as GzmA and B it can be released in a soluble form into the extracellular space. Furthermore, our data suggest that despite a more restricted cellular expression pattern, GzmK seems to participate in immune responses against several viruses.

SERPINB6 (PI6) is a member of the intracellular serine protease inhibitors (serpins). Previous studies showed that SERPINB6 is localized mainly in the cytoplasm of endothelial cells, some epithelial cells, monocytes, and neutrophils. In these cells SERPINB6 is thought to prevent cellular damage by scavenging leaking lysosomal proteases. We show here, using novel, well-defined monoclonal antibodies, that SERPINB6 is abundantly expressed by mast cells in all organs and by the human mast cell line HMC-1. Gel filtration experiments revealed that the latter cells contain a high-molecularweight form of SERPINB6, which consists of sodium dodecyl sulfate (SDS)-stable complexes of this inhibitor with monomeric ␤-tryptase. Expression of SERPINB6 by mast cells was compared with those of tryptase and CD117 (c-kit) in biopsies from patients with different forms of mast cell disease. In all cases the lesional mast cells expressed SERPINB6, and, in diffuse cutaneous mastocytosis and mastocytoma, SERPINB6 was expressed by a substantially higher number of mast cells when compared with tryptase. In conclusion, SERPINB6 is abundantly expressed by normal mast cells and by mast cells in mastocytoma lesions. We suggest that in mast cells, SERPINB6 serves to regulate the activity of endogenous ␤-tryptase in the cyto-