Drug-induced apoptosis in lung cnacer cells is not mediated by the Fas/FasL (CD95/APO1) signaling pathway.

Anticancer drugs exert at least part of their cytotoxic effect by triggering apoptosis. We previously identified chemotherapy-induced apoptosis in lung cancer cells and suggested a role for p53 alternative or complementary pathways in this process. Recently, a role for the Fas/FasL (CD95/Apo1) signaling system in chemotherapy-induced apoptosis was proposed in some cell types. In the present work, the involvement of the Fas/FasL system in drug-induced apoptosis in lung cancer cells was investigated upon exposure to four cytotoxic drugs (cisplatin, gemcitabine, topotecan, and paclitaxel). We assessed the expression of Fas and FasL and the function of the Fas pathway in six lung cancer cell lines (H460, H322, GLC4, GLC4/ADR, H187, and N417). All lung cancer cell lines expressed Fas and FasL at RNA and protein levels, and apoptosis could be induced in four of six cell lines upon exposure to the Fas agonistic monoclonal antibody (mAb) CLB-CD95/15. Nevertheless, after drug exposure, no significant FasL up-regulation was observed, whereas the Fas expression was increased in the wild-type p53 cell line H460, but not in the other lines, proved to be mutant p53 by direct gene sequencing. Moreover, no correlation was observed in lung cancer cell lines between sensitivity to drugs and to a Fas agonistic mAb, and preincubation of cells with either the Fas-antagonistic mAb CLB-CD95/2 or a FasL-neutralizing mAb did not protect from drug-induced apoptosis. Taken together, these observations strongly argue against a role of the Fas/FasL signaling pathway in drug-induced apoptosis in lung cancer cells. Interestingly, caspase-8 activation was observed upon drug exposure, independently from Fas/FasL signaling.

Extracellular granzymes A and B in humans: detection of native species during CTL responses in vitro and in vivo.

Activated CTLs and NK cells induce apoptosis via multiple mechanisms, including that termed granule exocytosis. The latter pathway consists of vectorial secretion of perforin and a family of granule-associated serine proteases (granzymes) to the target cell. To establish whether granzymes are released extracellularly during cytolytic reactions in vivo, ELISAs that measure the native enzymes were developed and were found to specifically detect granzyme A (GrA) and granzyme B (GrB) at picogram concentrations. Low levels of GrA and GrB were present in plasma of healthy individuals (GrA, 33.5 pg/ml (median); GrB, 11.5 pg/ml (median)), whereas significantly higher levels were present in patients with ongoing CTL response, i.e., patients suffering from infections by EBV or HIV type 1. Markedly elevated levels were also noted in synovial fluid of patients with active rheumatoid arthritis. The measurement of soluble granzymes should be useful to assess clinical disorders associated with activated CTL and NK cells. Furthermore, these results suggest that granzymes mediate biologic effects beyond their described role in apoptotic cell death.

Nonfunctioning malignant paraganglioma of the posterior mediastinum with spinal cord compression. A case report.

STUDY DESIGN: Case report and review of the literature. OBJECTIVE: To present a case with an uncommon nonfunctioning malignant paraganglioma of the posterior mediastinum with compression of the spinal cord. An update survey is given with respect to diagnostic, histopathologic, and therapeutic aspects of paragangliomas. SUMMARY OF BACKGROUND DATA: Paragangliomas in the posterior mediastinum are uncommon and rarely may produce spinal cord compression. To illustrate the low incidence, only two cases of paraganglioma of the posterior mediastinum were reported by the Mayo Clinic, Rochester, Minnesota, during a 40-year period. METHOD: Case report and review of the literature. RESULT: The diagnosis was made by modern imaging techniques (computed tomography, magnetic resonance imaging, and metaiodobenzylguanidine scan) and verified by computed tomography guided per thoracic puncture. A wide local resection was performed; the patient is disease-free 1 year after surgery. CONCLUSION: The uncertainty of prognosis and possibility of local recurrence of paragangliomas even after a long period emphasizes the importance of wide local surgical resection with or without adjuvant therapy and makes long-term follow-up and continued surveillance of the patient mandatory.

Evidence that intestinal intraepithelial lymphocytes are activated cytotoxic T cells in celiac disease but not in giardiasis.

To further define intraepithelial lymphocytes (IELs) in celiac disease (CD) and giardiasis, IELs were probed for the presence of cytolytic granules containing granzyme B (GrB) and T-cell-restricted intracellular antigen (TIA)-1. The expression of TIA-1, GrB, CD3 (T-cell-receptor-associated complex), and Leu-7 (subset of natural killer cells) was studied by a sensitive three-step immunoperoxidase technique. Stained IELs were determined quantitatively, and results were expressed as number of stained IELs per 100 epithelial cells (ECs). The relative content in labeled lamina propria lymphocytes was determined and expressed as the percentage of all lamina propria cells counted. When compared with controls, CD3+ and GrB+ IELs were significantly increased (P < 0.0004) in CD paralleled by an increase in TIA-1+ IELs (P < 0.0004). In CD, the highest numbers of IELs containing GrB were found in subjects with a flat mucosa (median, 38 IELs/100 ECs, P < 0.0004), followed by cases with shortened and blunted villi (median, 8 IELs/100 ECs, P < 0.0004) and, finally, CD patients with an intact villous architecture (median, 0.5 IELs/100 ECs, P < 0.02). Except for cases with giardiasis, Leu-7+ IELs were virtually absent in all groups as were GrB+ IELs in the controls and in subjects with giardiasis. In the lamina propria of CD subjects, GrB+ lymphocytes were also significantly increased (P < 0.001), whereas controls and cases with giardiasis were essentially free of GrB+ cytotoxic T lymphocytes. The percentage of CD3+ lamina propria lymphocytes was nearly equal in all groups. In humans and mice, extensive studies revealed a GrB expression to be absolutely restricted to activated cytotoxic T lymphocytes and natural killer cells. TIA-1, on the other hand, is considered a marker of resting T lymphocytes possessing cytolytic potential. We therefore conclude that IELs are cytotoxic T cells that are in a resting state in the normal small bowel and in giardiasis. In CD, they become activated as suggested by the GrB positivity of their granules.