Signal transduction triggered by lipid A-like molecules in 70Z/3 pre-B lymphocyte tumor cells.
The lipid A (endotoxin) moiety of lipopolysaccharide (LPS) elicits rapid cellular responses from many cell types, including macrophages, lymphocytes, and monocytes. In CD14 transfected 70Z/3 pre-B lymphocyte tumor cells, these responses include activation of the MAP kinase homolog, p38, activation of NF-kappaB, and transcription of kappa light chains, leading to the assembly of surface IgM. In this work, we explored the specificity of the response with regard to lipid structure, and the requirement for p38 kinase activity prior to NF-kappaB activation in control and CD14 transfected 70Z/3 (CD14-70Z/3) cells. A p38-specific inhibitor, SB203580, was used to block p38 kinase activity in cells. CD14-70Z/3 cells were incubated with 1-50 microM SB203580, and then stimulated with LPS. Nuclear extracts were prepared, and NF-kappaB activation was measured using an electrophoretic mobility shift assay. SB203580 did not inhibit LPS induced NF-kappaB activation. In addition, LPS failed to activate p38 tyrosine phosphorylation in 70Z/3 cells lacking CD14, in spite of rapid NF-kappaB activation and robust surface IgM production with appropriate higher doses of LPS. LPS stimulation of p38 phosphorylation, NF-kappaB activation, and surface IgM expression were all blocked completely by lipid A-like endotoxin antagonists whether or not CD14 was present. Acidic glycerophospholipids and ceramides did not mimic lipid A-like molecules either as agonists or antagonists in this system. Our data support the hypothesis that lipid A-mediated activation of cells requires stimulation of a putative lipid A sensor that is downstream of CD14, but upstream of p38 and NF-kappaB. (+info)
Rituximab therapy in hematologic malignancy patients with circulating blood tumor cells: association with increased infusion-related side effects and rapid blood tumor clearance.
PURPOSE: Rituximab was recently approved for use in relapsed, low-grade non-Hodgkin's lymphoma; however, few data exist regarding the safety of this agent in patients with a high number of tumor cells in the blood. METHODS AND RESULTS: After the observation at our institution of a rapid reduction of peripheral-blood tumor cells with associated severe pulmonary infusion-related toxicity in two patients with refractory hematologic malignancies, data on three additional cases were collected from physician-submitted reports of adverse events related to rituximab treatment. Five patients with hematologic malignancies possessing a high number of blood tumor cells were treated with rituximab and developed rapid tumor clearance. The median age was 68 years (range, 26 to 78 years). Patients were diagnosed with B-cell prolymphocytic leukemia (n = 2), chronic lymphocytic leukemia (n = 2), or transformed non-Hodgkin's lymphoma (n = 1). All of these patients had bulky adenopathy or organomegaly. All five patients developed a unique syndrome of severe infusion-related reactions, thrombocytopenia, rapid decrement in circulating tumor cell load, and mild electrolyte evidence of tumor lysis, and all required hospitalization. In addition, one patient developed ascites. These events resolved, and four patients were subsequently treated with rituximab without significant complications. CONCLUSION: Rituximab administration in patients who have a high number of tumor cells in the blood may have an increased likelihood of severe initial infusion-related reactions. These data also suggest that rituximab may have activity in a variety of other lymphoid neoplasms, such as chronic lymphocytic leukemia and B-cell prolymphocytic leukemia. (+info)
Inhibition of caspases inhibits the release of apoptotic bodies: Bcl-2 inhibits the initiation of formation of apoptotic bodies in chemotherapeutic agent-induced apoptosis.
During apoptosis, the cell actively dismantles itself and reduces cell size by the formation and pinching off of portions of cytoplasm and nucleus as "apoptotic bodies." We have combined our previously established quantitative assay relating the amount of release of [3H]-membrane lipid to the degree of apoptosis with electron microscopy (EM) at a series of timepoints to study apoptosis of lymphoid cells exposed to vincristine or etoposide. We find that the [3H]-membrane lipid release assay correlates well with EM studies showing the formation and release of apoptotic bodies and cell death, and both processes are regulated in parallel by inducers or inhibitors of apoptosis. Overexpression of Bcl-2 or inhibition of caspases by DEVD inhibited equally well the activation of caspases as indicated by PARP cleavage. They also inhibited [3H]-membrane lipid release and release of apoptotic bodies. EM showed that cells overexpressing Bcl-2 displayed near-normal morphology and viability in response to vincristine or etoposide. In contrast, DEVD did not prevent cell death. Although DEVD inhibited the chromatin condensation, PARP cleavage, release of apoptotic bodies, and release of labeled lipid, DEVD-treated cells showed accumulation of heterogeneous vesicles trapped in the condensed cytoplasm. These results suggest that inhibition of caspases arrested the maturation and release of apoptotic bodies. Our results also imply that Bcl-2 regulates processes in addition to caspase activation. (+info)
The human (PsiL+mu-) proB complex: cell surface expression and biochemical structure of a putative transducing receptor.
The surrogate light chain (PsiL) associates with mu and Igalpha-Igbeta chains to form the preB-cell receptor that plays a critical role in early B-cell differentiation. Discrepancies exist in human concerning the existence of PsiL+mu- proB cells and the biochemical structure of such a proB-cell complex remains elusive. Among new antihuman VpreB monoclonal antibodies (MoAbs), 5 of the gamma kappa isotype bound to recombinant and native VpreB protein with high affinity. They recognized 4 discrete epitopes, upon which 2 were in the extra-loop fragment. Such MoAbs detected the PsiL at the cell surface of either preB or on both proB and preB cells. The previously reported SLC1/SLC2 MoAbs recognize a conformational epitope specific for the mu/PsiL association in accordance with their preB-cell reactivity. Using the proB/preB 4G7 MoAb, PsiL cell surface expression was detected on normal bone marrow, not only on CD34(-)CD19(+) preB but also on CD34(+)CD19(+) proB cells. Futhermore, this MoAb identified PsiL+mu- fresh proB leukemic cells of the TEL/AML1 type. Biochemical studies showed that, at the proB stage, the PsiL is associated noncovalently with two proteins of 105 and 130 kD. Triggering of this complex induces intracellular Ca2+ flux, suggesting that the PsiL may be involved in a new receptor at this early step of the B-cell differentiation. (+info)
Bone marrow pre-B-1 (Bomb1): a quantitative trait locus inducing bone marrow pre-B-cell expansion in lymphoma-prone SL/Kh mice.
Abnormalities of regulatory genes in early B-cell development often lead to lymphomagenesis. Our previous study showed that there is an abnormal transient expansion of bone marrow (BM) pre-B cells in lymphoma-prone SL/Kh strain mice. Such expansion is a genetic property of SL/Kh stem cells rather than BM microenvironments. Using the percentage of BP1+ B220+ pre-B cells in total BM lymphoid cells as a quantitative parameter, we studied the genetic control of BM pre-B cells in 159 F2 offspring of crosses between SL/Kh and NFS/N mice and 334 back-crosses to SL/Kh mice. A highly significant quantitative trait locus was identified on the distal segment of chromosome 3, showing logarithm of odds scores of 22.7 in the F2 cohort and 10.7 in back-cross mice. This quantitative trait locus, named bone marrow pre-B-1, colocalized with lymphoid enhancer factor-1, which encodes a high mobility group DNA-binding protein that is expressed in T and pre-B cells. (+info)
Unravelling an HLA-DR association in childhood acute lymphoblastic leukemia.
Genetic and environmental factors play an interactive role in the development of childhood acute lymphoblastic leukemia (ALL). Since the demonstration of a major histocompatibility complex (MHC) influence on mouse leukemia in 1964, an HLA association has been considered as a possible genetic risk factor. Despite extensive efforts, however, no strong evidence comparable to the H-2(k) influence on mouse leukemia has been shown. The number of negative serological studies resulted in a loss of interest and consequently, no molecular HLA-DR association study has been published to date. We reconsidered the HLA-DR association in childhood ALL in 114 patients from a single center and 325 local newborn controls by polymerase chain reaction (PCR) analysis of the HLA-DRB1/3/4/5 loci. With conventional analysis, there was a moderate allelic association with the most common allele in the HLA-DR53 group, HLA-DRB1*04, in the whole group that was stronger in males (P =.0005, odds ratio = 2.9). When the other expressed HLA-DRB loci were examined, homozygosity for HLA-DRB4*01, encoding the HLA-DR53 specificity, was increased in patients (21.1% v 8.3%; odds ratio = 2.9, P =.0005). Consideration of gender showed that all of these associations were reflections of a male-specific increase in homozygosity for HLA-DRB4*01 (32.8% v 4. 0%; odds ratio = 11.7, 95% confidence interval [CI] = 4.9 to 28.0; P = 3 x 10(-8)). This highly significant result provided the long-suspected evidence for the HLA-DR influence on the development of childhood ALL while confirming the recessive nature of the MHC influence on human leukemogenesis as in experimental models. The cross-reactivity between HLA-DR53 and H-2Ek, extensive mimicry of the immunodominant epitope of HLA-DR53 by several carcinogenic viruses, and the extra amount of DNA in the vicinity of the HLA-DRB4 gene argue for the case that HLA-DRB4*01 may be one of the genetic risk factors for childhood ALL. (+info)
High frequency of adhesion defects in B-lineage acute lymphoblastic leukemia.
Aberrant proliferation, differentiation, and/or migration of progenitors observed in various hematological malignancies may be caused by defects in expression and/or function of integrins. In this study, we have developed a new fluorescent beads adhesion assay that facilitates flow cytometric investigation of lymphocyte function-associated antigen 1 (LFA-1)- and very late activation antigen-4 (VLA-4)-mediated functional adhesion in B-lineage acute lymphoblastic leukemia (ALL) of both the CD10(-) and CD10(+) (leukemic) cell population within one blood or bone marrow sample. Surprisingly, of the 20 B-lineage ALL patients investigated, 17 contained a leukemic cell population with LFA-1- and/or VLA-4-mediated adhesion defects. Five patients contained CD10(+) cells that did not exhibit any LFA-1-mediated adhesion due to the lack of LFA-1 surface expression. The CD10(+) cells from 10 ALL patients expressed LFA-1 that could not be activated by the phorbol ester phorbol 12-myristate 13-acetate (PMA), whereas the CD10(-) cells expressed a functional LFA-1. Seven patients contained CD10(+) cells that expressed a PMA-unresponsive form of VLA-4. The PMA unresponsiveness of the integrins LFA-1 and VLA-4 expressed by the CD10(+) cells may be due to mutations in the integrins itself, in protein kinases, or in other intracellular molecules involved in integrin adhesion. These data clearly demonstrate the importance of investigating integrin function in addition to integrin surface expression. The strikingly high frequency (85%) of adhesion defects in ALL could suggest a causal relationship between integrin-mediated adhesion and B-lineage ALL. (+info)
A DNA damage repair mechanism is involved in the origin of chromosomal translocations t(4;11) in primary leukemic cells.
Some chromosomal translocations involved in the origin of leukemias and lymphomas are due to malfunctions of the recombinatorial machinery of immunoglobulin and T-cell receptor-genes. This mechanism has also been proposed for translocations t(4;11)(q21;q23), which are regularly associated with acute pro-B cell leukemias in early childhood. Here, reciprocal chromosomal breakpoints in primary biopsy material of fourteen t(4;11)-leukemia patients were analysed. In all cases, duplications, deletions and inversions of less than a few hundred nucleotides indicative of malfunctioning DNA repair mechanisms were observed. We concluded that these translocation events were initiated by several DNA strand breaks on both participating chromosomes and subsequent DNA repair by 'error-prone-repair' mechanisms, but not by the action of recombinases of the immune system. (+info)