Biomedical subjects

J Carracedo

Publications and source records attributed to J Carracedo.

26 records · Page 2Linked to original sources

PubMed1994

Pertussis toxin inhibits activation-induced cell death of human thymocytes, pre-B leukemia cells and monocytes.

Activation of human thymocytes and pre-B cells via the CD3/T cell receptor (TCR) complex or the IgM/B cell receptor complex, respectively, results in apoptotic cell death. Similarly, cross-linking of the activation marker CD69, which belongs to the natural killer complex, causes apoptosis of lipopolysaccharide-preactivated monocytes. Here we show that pertussis toxin (PTX) inhibits the activation-induced apoptosis of these three cell types, though it fails to prevent the programmed cell death that follows exposure of cells to the synthetic glucocorticoid dexamethasone (thymocytes, pre-B cells) or to interleukin 4 (monocytes). The capacity of pertussis toxin to suppress activation-induced death is not due to quenching of the activation signal, because thymocytes exposed to PTX are still capable of mobilizing Ca2+ after TCR-alpha/beta cross-linking and proliferate in response to costimulation with PTX and CD3/TCR ligation. The apoptosis-inhibitory effect of PTX depends on the presence of an intact adenosine diphosphate (ADP)-ribosylating moiety, since a mutant pertussis toxin molecule that lacks enzymatic activity, but still possesses the membrane translocating activity, fails to interfere with activation-induced cell death. A toxin that induces a different spectrum of ADP ribosylation than PTX, cholera toxin, fails to inhibit apoptosis. To suppress apoptosis, the intact PTX holotoxin must be added to cells before the lethal activation step; its addition 30 min after initial activation remains without effect on apoptosis. These data unravel a PTX sensitive signal transduction event that intervenes during an early step of activation-induced cell death of immune cells.

Apoptosis↗

PubMed1993

HLA class II-mediated aggregation is associated with the proliferation of B lymphocytes.

We have studied the role of LFA-1 antigens in human B lymphocyte aggregation, proliferation, and Ig production induced by a short stimulation via class II antigens. Cell stimulation with either bacterial superantigens or anti-DR mAbs rapidly induced homotypic cell aggregation. In response to IL-4, an increase in cell proliferation and Ig production was observed only when aggregation preceded addition of IL-4. The involvement of LFA-1 molecules in class II-induced aggregation was supported as LFA-1-deficient cells or B cells incubated with anti-LFA-1/ICAM-1 mAbs failed to aggregate after stimulation. The association between aggregation and subsequent Ig production and proliferation was further supported as, after IL-4 stimulation, in both LFA-1-deficient cells and B cells incubated with anti-LFA-1 mAbs, class II-mediated signals failed to increase Ig production or cell proliferation. These data suggest that in class II-stimulated cells, LFA-1-dependent aggregation has a major role in IL-4-dependent Ig production and proliferation of B cells.

B-Lymphocytes↗

PubMed1992

Mechanisms involved in NK resistance induced by interferon-gamma.

Human tumor cell lines were treated with interferon-gamma (IFN-gamma) and then used as target cells in NK assays to measure their ability to form conjugates and stimulate the production of NK cytotoxic factors (NKCF) and to determine their susceptibility to NKCF lysis. K562 and cell lines RS1, RS3, RS7, CAC, and CAP2, obtained from solid brain tumors, were used as targets, and peripheral blood lymphocytes (PBL) from normal donors were used as effector cells. IFN-gamma-treated cell lines had a decreased susceptibility to NKCF lysis and a decreased ability to induce the release of these factors without affecting target-effector cell binding. These results were not due to changes in HLA class I antigen expression, given that the level of HLA class I antigens on the tumor cell lines was not affected, the only exception being K562. In an attempt to further clarify the possible influence of HLA class I expression on K562, IFN-gamma-pretreated K562 cells were separated into HLA class I positive and HLA class I negative subsets for the NK assays. The results showed that both populations behaved similarly upon target-effector conjugate formation, whereas the HLA class I positive population showed a reduced susceptibility to lysis by NK cells and NKCF. Thus, these results establish that NK resistance induced by IFN-gamma is mediated by blocking the target cell's ability to activate NK cell triggering and release of NKCF and by blocking its susceptibility to lysis by these factors. This analysis helps to clarify not only the NK process but also the controversial regulatory effect of IFN in NK lysis.

Cell Adhesion↗

PubMed1992

HLA class-II-mediated homotypic aggregation: involvement of a protein tyrosine kinase and protein kinase C.

Homotypic aggregation of B-lymphocytes, B-cell lines and class-II-positive T cells via HLA class II molecules was examined. Signaling via DR antigens induced rapid aggregation in a dose- and time-dependent manner, maximum and stable aggregation was induced within 20 minutes. On the contrary, rapid signaling via DP or DQ required prestimulation with either PMA or anti-sIg. Aggregation was temperature and energy dependent. [Ca2+] and [Mg2+] concentrations and an intact cytoskeleton were required while neither mRNA or protein synthesis were required. Furthermore, FACS analysis revealed that aggregation was not directly correlated with cell surface expression of HLA class II molecules. Our results demonstrate that aggregation was mediated through a protein tyrosine kinase (PTK)-dependent pathway that preceded activation of protein kinase C (PKC) and failure to generate either the PTK signal or the PKC signal prevented aggregation. The contribution of a tyrosine kinase was further demonstrated by the total inhibition of aggregation following treatment with an anti-CD45 mAb.

Antibodies, Monoclonal↗

PubMed1990

Analysis of the mechanisms involved in NK resistance induced by a new tumor factor NK-RIF.

The mechanisms involved in susceptibility or resistance of neoplasic cells to lysis by NK cells are not well known. We have recently described a 12-kDa factor (NK-RIF), produced and released by different tumor cell lines, making K562 resistant to NK lysis without affecting the cytotoxic function of NK effector cells. In this paper we further study the mechanism involved in NK resistance of K562 mediated by NK-RIF and its biological implications. The results show that NK-RIF does not affect the binding capacity of target and effector cells nor the levels of HLA class I antigen expression on the target cells, as a proof that resistance to NK-mediated lysis is not always associated with a defect in target effector binding or with an increased MHC class I antigen expression. However NK-RIF-treated K562 loses its capacity to induce NK cell activation and the subsequent capacity to release NKCF and makes K562 resistant to lysis by NKCF. Therefore our results show that induction of resistance to NK cytotoxicity can be the result of the modulation of target structures responsible for inducing effector cell activation without affecting target/effector binding molecules. This indicates that the structures involved in adherence and activation of NK cells have a different nature and that molecules other than HLA participate in NK resistance.

Biological Factors↗

PubMed1990

Natural killer susceptibility is independent of HLA class I antigen expression on cell lines obtained from human solid tumors.

The susceptibility to natural killer (NK) cell-mediated cytotoxicity of 20 cell lines obtained from human solid tumors and their class I histocompatibility antigen (HLA) levels were studied in an attempt to determine whether major histocompatibility complex (MHC) products expressed on cells derived from human solid tumors influence NK susceptibility. The effect of interferon-gamma (IFN-gamma) treatment on these elements was also analyzed. The MHC class I (HLA-ABC, HLA-A and HLA-B) antigen levels and degree of NK lysis were very heterogeneous and no correlation was found on comparison. After treatment with IFN-gamma a marked decrease in NK susceptibility was observed in all the cell lines, including the control line K-562. However, the level of HLA class I expression was not modified in any of the lines with the exception of the K-562, which increased. In some cell lines the expression of HLA class I-like antigens. CDla, b and c, was also measured before and after IFN treatment; however, no correlation was found between CD1 levels and NK susceptibility. Consequently, from our results it is possible to conclude that HLA class I antigens do not play a decisive role in NK susceptibility of cell lines derived from human solid tumors and to suggest that molecules which are not HLA class I antigens but IFN-gamma inducible may confer NK resistance to these lines.

Antigens, CD1↗

PubMedDate not supplied

MHC class I expression on human tumour cells and their susceptibility to NK lysis.

Although natural killer (NK) activity is not restricted by the major histocompatibility complex (MHC), it has been suggested that the level of expression of MHC antigens by target cells may influence their lysis by NK cells. We have studied the NK susceptibility of 20 cell lines obtained from primitive and metastatic human tumours and the K562 cell line treated with gamma-interferon, phorbol ester TPA and tumour factor NK-RIF. When the levels of MHC class I antigen expression on the human tumour cell lines and their NK susceptibility were compared, no relationship between these two parameters was observed. Furthermore the treatment of K562 with either gamma-interferon, TPA or NK-RIF decreased its NK susceptibility independently of MHC class I expression. These results indicate that the MHC class I antigen is not the only factor directly involved in NK susceptibility and suggest that other membrane structures modulated by gamma-interferon, TPA or NK-RIF may also influence NK susceptibility.

Cytotoxicity, Immunologic↗

PubMedDate not supplied

Antibody-mediated pure red-cell aplasia (PRCA): the Spanish experience.

BACKGROUND: The incidence of antibody (Ab)-mediated pure red-cell aplasia (PRCA) in patients with chronic kidney disease (CKD) has increased between 1998 and 2002. After initially responding to treatment with recombinant human erythropoietic agents for CKD-associated anemia, patients became treatment-refractory and severely anemic. Although most PRCA cases have occurred in Europe, the varying epidemiologies among individual countries have not been well characterized. METHODS: We investigated Ab-mediated PRCA in 12 Spanish patients treated with epoetin alfa alone or prior to treatment with epoetin beta (n=1) or darbepoetin alfa (n=1). Serum Abs against epoetin alfa were detected by radioimmunoprecipitation (RIP) assay or bioassay. Following diagnosis of PRCA, erythropoietic treatment was stopped and patients received immunosuppressive therapy alone (n=11) or in combination with renal transplant (n=1). RESULTS: Treatments were administered for 16 months (average) before diagnosis of PRCA in bone marrow aspirates (n=8) or biopsies (n=4). At diagnosis, patients had an average of 0.68% blood reticulocytes and blood hemoglobin (Hb) level of 7.13 g/dL. Eight patients had anti-epoetin Abs detected by RIP, and 5 had neutralizing Abs measured in the bioassay. As of December 2003, 4 patients had died, 3 had no recovery, and 5 had recovered from anemia (blood Hb level, 9.9 g/dL). All 5 recovering patients received corticosteroid therapy alone, and 1 received a renal transplant as well as corticosteroids. CONCLUSIONS: Sudden onset of treatment-refractory anemia in CKD patients suggests a course of treatment cessation followed by diagnostic procedures for Ab-mediated PRCA, and immunosuppressive therapy. This study may serve as a model for a centralized global PRCA registry.

Antibodies↗