Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P10415 (Bcl-2)
 33,771 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The t(14;18) of human follicular B cell lymphoma translocates the Bcl-2 gene into the Ig H chain locus and markedly deregulates Bcl-2 expression. We sought to determine if Bcl-2 could be directly implicated in a growth-factor pathway. Consequently, we introduced a retrovirus containing the murine Bcl-2 gene (N2-M-Bcl-2) or the parental retrovirus (N2) into a series of factor-dependent hemopoietic cell lines. Overexpressed Bcl-2 resulted in no long term IL-2, IL-3, or IL-6 independent clones, indicating that Bcl-2 could not spare the need for a specific ligand-receptor interaction. However, Bcl-2 did extend the short term survival of IL-3-dependent cell lines after factor deprivation. Although viable, IL-3-deprived pro B lymphocytes (FL5.12) bearing N2-M-Bcl-2 were in Go, and deregulated Bcl-2 did not obviously influence cell-cycle progression. Bcl-2 predominant effects were to delay the onset of cell death and to modestly augment viable cell growth in the first 48 h after IL-3 deprivation. This death sparing was associated with increased levels of Bcl-2 RNA and protein in factor-deprived cells possessing N2-M-Bcl-2. This result was not restricted to prolymphocytes because an IL-3-dependent mast cell line (32D) as well as a promyeloid line (FDC-P1) demonstrated the same response to Bcl-2. Moreover, the effect was not limited to the IL-3/IL-3R signal transduction pathway in that promyeloid cells maintained in granulocyte-macrophage-CSF or IL-4 displayed a similar response. Yet, Bcl-2-enhanced cell survival was not universal as an IL-2-dependent T cell line, and an IL-6-dependent myeloma line demonstrated no consistent effect upon IL withdrawal. Thus, Bcl-2 appears to interfere with cell death but in a cell type and/or factor-restricted fashion.
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PMID:Deregulated Bcl-2 gene expression selectively prolongs survival of growth factor-deprived hemopoietic cell lines. 218 93

Flt3/flk-2 ligand (flt3-L) is a potent costimulator of normal bone marrow (BM) myeloid progenitors. Flt3-L is produced by BM stromal cells and its receptor is expressed in the majority of acute myeloid leukemia (AML) cases. Therefore, flt3-L may play a role in the paracrine and/or autocrine loops sustaining leukemic cell growth. We evaluated the effects of recombinant human flt3-L on proliferation, apoptosis, and Bcl-2 and Bax expression in primary AML cells and compared them with those of stem cell factor (SCF). Mononuclear BM cells from patients with newly diagnosed AML were cultured in serum-free conditions with flt3-L, SCF, granulocyte colony-stimulating factor (G-CSF) and granulocyte macrophage-colony-stimulating factor (GM-CSF) alone and in combination. In 9 of 10 samples, flt3-L significantly increased [3H]thymidine uptake (geometric mean stimulation index, 7.5; range, 2.4 to 41.5). Flt3-L also increased the number of AML blast colonies by 126% (range, 61% to 181%). In these 9 samples, flt3-L significantly enhanced the proliferative response triggered by G-CSF or GM-CSF. Flt3-L prevented apoptosis in AML blasts. It reduced the number of apoptotic cells by 36% +/- 3.9% compared with control cultures. Combining flt3-L with G-CSF or GM-CSF doubled the antiapoptotic effect. Cellular Bcl-2 and Bax levels were determined separately for apoptotic and nonapoptotic cells by flow cytometry. Cells undergoing spontaneous apoptosis had low Bcl-2 and high Bax levels, whereas nonapoptotic cells had high Bcl-2 and low Bax levels. Flt3-L alone or in combination with G-CSF or GM-CSF did not upregulate Bcl-2. However, Bax expression decreased in viable cells in the presence of these cytokines and the lowest level was achieved when a combination of flt3 and GM-CSF was used. Proliferative and viability effects of flt3-L were similar to those of SCF. Our results demonstrate that flt3-L acts as a stimulatory factor for primary AML cells. The antiapoptotic effects of flt3-L or its combinations with G-CSF or GM-CSF correlate with their ability to prevent upregulation of Bax.
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PMID:Flt3 ligand stimulates proliferation and inhibits apoptosis of acute myeloid leukemia cells: regulation of Bcl-2 and Bax. 891 65

The signal pathway for control of apoptosis in human neutrophils is currently unknown. In this study, we provide the first evidence that a Src family tyrosine kinase, Lyn, plays a key role in inhibition polymorphonuclear (PMN) cell death. Several nuclear proteins associated with apoptosis, i.e., p53, cdc2, and Rb, were absent from PMN. Bcl-2, known to inhibit apoptosis, was also not expressed. Programmed cell death that rapidly occurred in PMN could be arrested by granulocyte-macrophage CSF (GM-CSF), but this activation did not induce p53, cdc2, retinoblastoma, or Bcl-2 expression. Instead, GM-CSF produced a rapid activation of Lyn and Hck, but not Fgr, tyrosine phosphorylation within 1 min. Co-immunoprecipitation studies indicated that only Lyn, but not Hck, was physically coupled to GM-CSF receptor. By histologic assessment and evaluation of DNA fragmentation, only antisense Lyn, but not antisense Hck or antisense Fgr, could reverse the cell survival advantage provided by GM-CSF. Therefore, the physical coupling of Lyn to GM-CSF receptor and its early activation are required for inhibition or delay of apoptosis in PMN.
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PMID:Critical role of Lyn kinase in inhibition of neutrophil apoptosis by granulocyte-macrophage colony-stimulating factor. 894 27

The phosphatidylinositol 3-kinase (PI3K)-signaling pathway has emerged as an important component of cytokine-mediated survival of hemopoietic cells. Recently, the protein kinase PKB/akt (referred to here as PKB) has been identified as a downstream target of PI3K necessary for survival. PKB has also been implicated in the phosphorylation of Bad, potentially linking the survival effects of cytokines with the Bcl-2 family. We have shown that granulocyte/macrophage colony-stimulating factor (GM-CSF) maintains survival in the absence of PI3K activity, and we now show that when PKB activation is also completely blocked, GM-CSF is still able to stimulate phosphorylation of Bad. Interleukin 3 (IL-3), on the other hand, requires PI3K for survival, and blocking PI3K partially inhibited Bad phosphorylation. IL-4, unique among the cytokines in that it lacks the ability to activate the p21ras-mitogen-activated protein kinase (MAPK) cascade, was found to activate PKB and promote cell survival, but it did not stimulate Bad phosphorylation. Finally, although our data suggest that the MAPK pathway is not required for inhibition of apoptosis, we provide evidence that phosphorylation of Bad may be occurring via a MAPK/ERK kinase (MEK)-dependent pathway. Together, these results demonstrate that although PI3K may contribute to phosphorylation of Bad in some instances, there is at least one other PI3K-independent pathway involved, possibly via activation of MEK. Our data also suggest that although phosphorylation of Bad may be one means by which cytokines can inhibit apoptosis, it may be neither sufficient nor necessary for the survival effect.
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PMID:Dissociation of cytokine-induced phosphorylation of Bad and activation of PKB/akt: involvement of MEK upstream of Bad phosphorylation. 963 68

Stat3, a member of STAT, is activated by a variety of cytokines such as IL-6 family of cytokines, granulocyte CSF, epidermal growth factor, and leptin. A recent study with mice genetically deficient in the Stat3 gene has revealed its important role in the early embryogenesis. To assess the function of Stat3 in adult tissues, we disrupted the Stat3 gene specifically in T cells by conditional gene targeting using Cre-loxP system. In Stat3-deficient T cells, IL-6-induced proliferation was severely impaired. IL-6 did not enhance cell cycle progression, but prevented apoptosis of normal T cells. In contrast, IL-6 did not prevent apoptosis of Stat3-deficient T cells. Antiapoptotic protein, Bcl-2, was normally up-regulated in response to IL-6 even in Stat3-deficient T cells. These results demonstrate that Stat3 activation is involved in IL-6-dependent T cell proliferation through prevention of apoptosis independently of Bcl-2.
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PMID:Stat3 activation is responsible for IL-6-dependent T cell proliferation through preventing apoptosis: generation and characterization of T cell-specific Stat3-deficient mice. 2579 90

The ubiquitin-proteasome pathway is the principal mechanism for the degradation of short-lived proteins in eukaryotic cells. Here we examine the possibility that ubiquitin-proteasome is involved in regulating the levels of Bcl-2, which is abundantly expressed in M-07e cells, a granulocyte/macrophage colony-stimulating factor (GM-CSF)-dependent human leukaemic cell line. Apoptosis in M-07e cells, induced by GM-CSF withdrawal, was associated with a gradual cleavage of Bcl-2 into a 22 kDa fragment. Treatment of M-07e cells with benzyloxycarbonyl-Leu-Leu-l-leucinal (Z-LLL-CHO; MG-132), a reversible ubiquitin-proteasome inhibitor, markedly accelerated the cleavage of Bcl-2 and promoted cell death through the apoptotic pathway. The cleavage of Bcl-2 was inhibited by a caspase-3 (CPP32)-specific inhibitor [acetyl-Asp-Glu-Val-Asp-CHO (DEVD-CHO)] but not caspase 1 inhibitor (acetyl-Tyr-Val-Ala-Asp-CHO), suggesting that Bcl-2 is a proteolytic substrate of a caspase-3-like protease activated during apoptosis. The simultaneous addition of recombinant human GM-CSF (rhGM-CSF) to M-07e cultures delayed the activation of caspase 3 and Bcl-2 cleavage triggered by Z-LLL-CHO, suggesting that the activation of the GM-CSF signalling pathway can partly overcome the apoptotic effect induced by Z-LLL-CHO. Apoptosis induced by inhibition of the proteasome pathway was verified in studies with lactacystin, a highly specific and irreversible proteasome inhibitor. Lactacystin-induced apoptosis in M-07e cells was remarkably similar to that induced by Z-LLL-CHO, which included caspase 3 activation, cleavage of Bcl-2 into a 22 kDa fragment and, ultimately, cell death. These results showed that inhibition of the ubiquitin-proteasome pathways can lead to the activation of a DEVD-CHO-sensitive caspase and induces Bcl-2 cleavage, which might have a role in mediating apoptosis in M-07e cells.
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PMID:Inhibition of ubiquitin-proteasome pathway activates a caspase-3-like protease and induces Bcl-2 cleavage in human M-07e leukaemic cells. 1022 67

Sera from healthy subjects receiving recombinant human granulocyte colony-stimulating factor (rHuG-CSF) to mobilize CD34(+) peripheral blood progenitors (PBPC) have been recently shown to induce unresponsiveness of allogeneic lymphocytes to mitogenic challenge. In the present investigation, the effects of rHuG-CSF on the early stages of lymphocyte activation-induced apoptosis and on lymphocyte cell cycle entry were evaluated. Sera were obtained from HLA-identical donors receiving rHuG-CSF to mobilize CD34(+) PBPC for allogeneic transplantation. Normal peripheral blood mononuclear cells (PBMC) were challenged with phytohemagglutinin (PHA) in the presence of serum collected before (preG) or after rHuG-CSF administration (postG). Mitochondrial function, that is, incorporation of 3,3'-dihexyloxacarbocyanine iodide [DiOC(6)(3)] and generation of reactive oxygen species (ROS) as well as expression of c-Myc and Bcl-2 family members (Bcl-2, Bcl-X(L), Bax) were evaluated by multiparameter flow cytometry. The activation-induced fragmentation of genomic DNA was detected by highly sensitive LM-PCR assay.CD4(+)DiOC(6)(3)(low) and CD8(+)DiOC(6)(3)(low) T lymphocytes increased and reached 32% (range 27%-38%) and 20% (range 15%-23%) of circulating T cells, respectively, on day 4 of rHuG-CSF administration. Hypergeneration of ROS could be demonstrated in 65% (range 58%-82%) of CD4(+) T lymphocytes and in 0.4% (range 0.2%-0. 8%) of circulating CD8(+) T cells. rHuG-CSF determined no alteration of mitochondrial function if added to allogeneic PBMC in vitro, thus suggesting indirect effects mediated by soluble factors; on the contrary, when PBMC were challenged with PHA in the presence of postG serum, both perturbation of mitochondrial transmembrane potential (Deltapsi(m)) and hypergeneration of ROS were induced, and lymphocytes were predominantly arrested in a G(0) -like phase of the cell cycle and displayed genomic DNA fragmentation. Interestingly, the preincubation of PBMC with a blocking antibody directed against CD95 abrogated the perturbation of lymphocyte Deltapsi(m), suggesting that the CD95 signaling pathway might play a role in the induction of apoptosis after PHA stimulation in the presence of postG serum. Moreover, Bax protein was overexpressed in postG (median fluorescence intensity = 180, range 168-186) compared with preG cultures (median fluorescence intensity = 75, range 68-80; p < 0.01), while no differences in Bcl-2, Bcl-X(L), and c-Myc staining intensity were observed. Our findings demonstrate a humoral-mediated rHuG-CSF-induced dissipation of lymphocyte mitochondrial Deltapsi(m); these effects might be mediated by Bax overexpression, with imbalance between apoptosis-promoting and apoptosis-inhibiting Bcl-2 family members and with subsequent induction of mitochondrial permeability transition. Whether immune dysfunction will favorably impact on incidence and severity of acute graft vs host disease after allogeneic PBPC transplantation remains to be determined.
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PMID:Granulocyte colony-stimulating factor perturbs lymphocyte mitochondrial function and inhibits cell cycle progression. 1088 Jul 47

Bcl-2 is an integral intracellular membrane protein that can protect cells from apoptosis induced by multiple insults in a variety of cell types. During apoptosis, Bcl-2 was cleaved into a shortened fragment (Bcl-2/Delta34) by a caspase-3-like protease in human Mo7e megakaryocytic leukemia cells deprived of exogenous rhGM-CSF. Results from cell fractionation and immunoblot analyses indicated that both Bcl-2 and Bcl-2/Delta34 were located exclusively on the mitochondria of Mo7e cells. Treatment of isolated mitochondria with recombinant caspase-3 induced the same cleavage of Bcl-2 in vitro and caused the release of cytochrome c from the mitochondria into the supernatant. The antiapoptotic effect of Bcl-2/Delta34 was investigated using an in vitro protein translation approach. Both Bcl-2/Delta34 and Bax proteins generated in wheat germ extract were readily relocated to the mitochondria isolated from control Mo7e cells. Insertion of Bax, but not Bcl-2/Delta34, into mitochondria triggered a rapid release of cytochrome c from the mitochondria. Coimmunoprecipitation studies showed that, unlike Bcl-2, the cleaved Bcl-2 fragment was no longer functional for dimerization with either Bcl-2 or Bax. Taken together, these findings showed that the integrity of Bcl-2 is necessary for its function of heterodimerization with Bax, which appears to be one of the mechanisms of antiapoptotic effect of Bcl-2.
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PMID:Apoptosis of Mo7e leukemia cells is associated with the cleavage of Bcl-2 into a shortened fragment that is not functional for heterodimerization with Bcl-2 and Bax. 1108 88

The main objectives of this study were to determine the feasibility of administering high doses of cyclophosphamide plus recombinant human granulocyte-colony stimulating factor (rhG-CSF) every 14-21 days to patients with follicular small cleaved cell lymphoma. For each patient, the treatment was not considered feasible if fewer than four cycles of cyclophosphamide chemotherapy could be administered on schedule (i.e. at least every 29 days) or (1) hospitalization of the patient for longer than three days was necessary for neutropenic fever (38 degrees C) or bacteriologically documented infection in > 50% of the cycles, or (2) grade > or = 2 hemorrhage in association with thrombocytopenia of grade > or = 3 severity occurred in > 50% of the cycles or (3) non-hematologic toxicity (excluding nausea/vomiting and alopecia) of grade > or = 3 occurred in > 50% of cycles. The goal was to have a treatment program feasible in 75% or more of the treated patients. The secondary objectives were to determine the toxicities, the complete and partial response rates, and the time to treatment failure (TTF). The trial also attempted to assess the effectiveness of this treatment program in eradicating Bcl-2 rearrangements by PCR, and to assess complete remission duration in relationship to PCR results in patients who respond to this chemotherapy program. Patients were required to have histologically documented non-Hodgkin's lymphoma of the subtypes follicular, predominantly small cleaved cell (IWF-B) or follicular mixed, (IWF-C). Patients were required to have Stage IV disease including histologic evidence of bone marrow involvement. Measurable disease was required and patients were also required to have one of the following risk factors: > or = 2 extranodal sites, node or nodal group > or = 5 cm. Submission of fresh bone marrow for molecular genetic studies for the presence of Bcl-2-Ig fusion DNA was mandatory in previously untreated patients. Patients had to be between 18 and physiologic age 55 years (carefully selected patients over age 55 years were also eligible), expected survival > 2 years, performance status 0-1, and have adequate renal, hepatic and bone marrow function, and a cardiac ejection fraction > or = 50%. Cyclophosphamide 4.5 g/m2 i.v. was given with mesna every 14 days with rhG-CSF support. Twenty-nine patients were accrued to this trial. The median follow-up time is 5.0 years, with a range of 2.5-6.7 years. The overall response rate was 75% (9 CRs 37.5%, 9PRs 37.5%). The median duration of survival is 5.53 years. The 1-year estimated probability of freedom from treatment failure was 50% and of survival at 1 year was 92%. No strong association was observed between TTF and age, symptomatic stage, histology performance status, number of extranodal sites or baseline Bcl-2 status. At 3 years the survival of all patients was 78% and failure free survival was 17%. 15 (62%) of the 24 eligible previously untreated patients met the criteria for feasibility specified in the protocol. The 95% CI for the feasibility rate is (44 and 82%). Twenty-two of the 24 (92%) previously untreated patients had specimens submitted for testing for Bcl-2 rearrangements. Thirteen of the 22 (59%) were found to have rearrangements at baseline. Post-treatment specimens were submitted for seven of the 13 patients. Four of the seven converted to Bcl-2 negative following treatment. Eight of 13 Bcl-2 positive patients (62%) had a clinical response to treatment. The 95% exact binomial CI for the total response rate in this subgroup is (28 and 88%). This study demonstrates that repetitive doses of cyclophosphamide at 4.5 g/m2 every two weeks with rhG-CSF support can be administered to selected younger patients with advanced follicular lymphoma with morphologic involvement of the bone marrow with acceptable non-hematologic toxicity.
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PMID:High dose cyclophosphamide plus recombinant human granulocyte-colony stimulating factor (rhG-CSF) in the treatment of follicular, low grade non-Hodgkin's lymphoma: CALGB 9150. 1191 6

Phosphoinositide 3-kinase (PI3-kinase)-dependent phosphorylation of the proapoptotic Bcl-2 family member Bad has been proposed as an important regulator of apoptotic cell death. To understand the importance of this pathway in nontransformed hematopoietic cells, we have examined the effect of survival cytokines on PI3-kinase activity and Bad expression and phosphorylation status in human neutrophils. Granulocyte macrophage-colony-stimulating factor (GM-CSF) and tumor necrosis factor-alpha (TNF-alpha) both reduced the rate of apoptosis in neutrophils cultured in vitro for 20 hours. Coincubation with the PI3-kinase inhibitor LY294002, which in parallel experiments abolished GM-CSF-primed, fMLP-stimulated superoxide anion production and GM-CSF-stimulated PtdIns(3,4,5)P(3) accumulation, inhibited the GM-CSF and TNF-alpha survival effect. In contrast, the MAP kinase kinase (MEK1/2) inhibitor PD98059 and the protein kinase A inhibitor H-89 had only a marginal effect on GM-CSF-mediated neutrophil survival. GM-CSF substantially increased Bad phosphorylation at Ser112 and Ser136 and increased the cytosolic accumulation of Bad. GM-CSF also regulated Bad at a transcription level with a marked decrease in mRNA levels at 4 hours. TNF-alpha caused a biphasic effect on the rate of morphologic apoptosis, which corresponded to an early increase, and a late inhibition, of Bad mRNA levels. LY294002 inhibited GM-CSF- and TNF-alpha-mediated changes in Bad phosphorylation and mRNA levels. These data suggest that the survival effect of GM-CSF and TNF-alpha in neutrophils is caused by a PI3-kinase-dependent phosphorylation and cytosolic translocation of Bad, together with an inhibition of Bad mRNA levels. This has important implications for the regulation of neutrophil apoptosis in vivo.
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PMID:Role of PI3-kinase-dependent Bad phosphorylation and altered transcription in cytokine-mediated neutrophil survival. 1223 75


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