Acute lymphoblastic leukaemia (ALL) is the commonest form of childhood malignancy, and most cases arise from B-cell clones arrested at the pre-B-cell stage of differentiation. The molecular events that arrest pre-B-cell differentiation in the leukaemic pre-B cells have not been well characterized. Here we show that the differentiation regulator SLP-65 (an adaptor protein also called BLNK or BASH) inhibits pre-B-cell leukaemia in mice. Reconstitution of SLP-65 expression in a SLP-65-/- pre-B-cell line led to enhanced differentiation in vitro and prevented the development of pre-B-cell leukaemia in immune-deficient mice. Tyrosine 96 of SLP-65 was required for this activity. The murine SLP-65-/- pre-B-cell leukaemia resembles human childhood pre-B ALL. Indeed, 16 of the 34 childhood pre-B ALL samples that were tested showed a complete loss or drastic reduction of SLP-65 expression. This loss is probably due to the incorporation of alternative exons into SLP-65 transcripts, leading to premature stop codons. Thus, the somatic loss of SLP-65 and the accompanying block in pre-B-cell differentiation might be one of the primary causes of childhood pre-B ALL.
In the periphery, B cells differentiate in germinal centres (GCs) of secondary lymphoid organs. Isolated GC cells die quickly in vitro by apoptosis. Therefore, cell lines originating from follicular lymphomas, which are the malignant counterparts of GC B cells, would provide a stable in vitro model to study the immunobiology of GC B cells. We have established three novel human follicular lymphoma cell lines that were characterized with special reference to immunophenotypic features, response to B-cell receptor (BCR) triggering, response to cytokines and cytokine mRNA expression. One of the cell lines, HF-1A3, has a phenotype of a centrocyte. It expresses surface immunoglobulin G (sIgG) and dies by apoptosis following BCR cross-linking. Co-stimulation with interleukin-6 (IL-6), IL-15 or interferon-g (IFN-g) rescues HF-1A3 cells from BCRinduced apoptosis. The second cell line, HF-28, also represents phenotypically an IgG þ centrocyte. Ligation of its BCR leads to the cell-cycle arrest at G1 instead of apoptosis. HF-28 cells express both CD45RA and RO isoforms, which is unusual in B lymphocytes apart from plasma cells, thus suggesting a transition to plasma cell phenotype. The third cell line, HF-4.9, which phenotypically represents an sIgM þ centroblast, responds by proliferation to BCR cross-linking. These cell lines offer a unique in vitro model to study antigenic selection and cytokine-mediated growth regulation of human GC B cells.
In the present study we used a human follicular lymphoma cell line, HF1A3, as an in vitro model for the antigen-driven selection process in germinal centers. Apoptosis can be induced in HF1A3 cells by B cell receptor (BCR) stimulation, but the molecular mechanisms and kinetics of this process are largely unknown. We demonstrate here that there is over 12 h delay between receptor activation and the execution phase of apoptosis, i.e. disruption of mitochondrial membrane potential, release of cytochrome c from mitochondria, caspase-3 activation and DNA fragmentation. New protein synthesis is required for mitochondrial alterations and subsequent apoptosis to occur, as these processes are completely blocked by the protein synthesis inhibitor cycloheximide. All the apoptotic events induced by BCR triggering are completely reversed by CD40 ligation with anti-CD40 antibody. CD40 ligation can reverse the apoptotic process in HF1A3 cells almost until the first mitochondrial events take place demonstrating that CD40-mediated protection operates very fast and at or before mitochondrial phase of apoptosis. Using specific inhibitors of cell signaling we could demonstrate that Raf-extracellular signal-regulated kinase, phosphatidylinositol 3-kinase, p38 or protein kinase C activation pathways are not involved in CD40-mediated protection from BCR-induced apoptosis in HF1A3 cells.
Aiolos is a chromatin remodeling transcription regulator that plays an antiproliferative role in B lymphocyte function. In contrast to the related Ikaros factors, mammalian Aiolos has not beenreported to generate splice variants. In addition, although human leukemic lymphoblasts express non‐DNA‐binding Ikaros isoforms with potential dominant negative effect on other interacting factors,the role of Aiolos in human lymphoid disorders has remained obscure. To address the question, why Aiolos should delineate from Ikaros in such a marked way, we have here analyzed whether also human Aiolos could generate alternate isoforms. According to the results obtained, both normal and neoplastic B lineage cells were found to express at least five novel Aiolos variants. Also structurally dominant negative variants with less than three DNA‐binding domains were identified. In conclusion, given the multiplicity of also human Aiolos isoforms and thereby the evidently more intricate contribution of Aiolos to the chromatin remodeling machinery, it is suggested, that not only Ikaros, but also Aiolos could participate in a more versatile manner in the regulation of B lymphocyte function.
Introduction Lambert–Eaton myasthenic syndrome (LEMS) is characterized by autoantibodies against voltage-gated calcium channels (VGCC) at the neuromuscular junction causing proximal muscle weakness, decreased tendon reflexes, and autonomic changes. The European LEMS registry aimed to collate observational safety data for 3,4-diaminopyridine phosphate (3,4-DAPP) and examine long-term outcomes for patients with LEMS. Methods Thirty centers across four countries participated in the non-interventional European LEMS registry. Any patients diagnosed with LEMS by means of clinical assessment and abnormal neurophysiological testing, or clinical assessment and positive for VGCC antibodies were eligible to participate. Patients were monitored using standard assessments for LEMS-related clinical manifestations. Results Among 96 evaluable participants, 50 (52.1%) were being treated with 3,4-DAPP, 21 (21.9%) with 3,4-diaminopyridine (3,4-DAP), and 25 (26.0%) with other treatments (e.g., pyridostigmine, corticosteroids, immunoglobulins, and azathioprine); 74 participants (77.1%) were exposed to 3,4-DAPP at any time. Quantitative myasthenia gravis scores were similar across treatment groups. Muscle strength was generally good and maintained during follow-up. Cerebellar ataxia, defined as a negative Romberg’s test and at least one other positive ataxia test, was observed in 30 (56.6%) patients. Most participants had reduced reflex tone and limited functioning. Sustained or improved functioning was observed in participants administered 3,4-DAPP. Inconsistent and sporadic functional improvement and regression was observed with 3,4-DAP and other treatments. Fifty-five treatment-related adverse events (AEs) were reported by 32 (33.3%) participants. Eight (8.3%) participants reported nine treatment-related serious AEs. No new safety signals were identified. Conclusion No new safety signals were observed following long-term management of LEMS with 3,4-DAPP. Supplementary Information The online version contains supplementary material available at 10.1007/s40120-022-00354-8.
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We have analysed separately the role of B‐cell receptor (BCR) stimulation and the soluble second signal in the T‐cell‐independent type 2 (TI‐2) B‐cell response. We were able to show that human B cells and macrophages (Mφ) could function together in TI‐type microbial response. Interestingly, BCR cross‐linking of peripheral blood (PB) B cells enhanced IgG production induced by Mφ‐derived growth factors whereas interleukin (IL)‐12 + IL‐18 had milder effect on IgG production. We demonstrated that B‐cell‐derived soluble mediators primed lipopolysaccharide (LPS)‐stimulated Mφ for tumour necrosis factor‐α (TNF‐α) and IL‐6 production significantly better than IFN‐γ, confirming the role of B cells in the activation of Mφ. We could show that human PB B cells were active cytokine producers and could be induced to produce interferon (IFN)‐γ mRNA in the presence of known Mφ cytokines, like IL‐12 and IL‐18. BCR stimulation also stabilized and enhanced the IFN‐γ mRNA production induced by IL‐12 and IL‐18. In addition, our novel finding was that a known Mφ cytokine, IL‐10, induced the expression of IFN‐γ mRNA from human B‐cell line (HF28R0) cells. In summary, we propose a model for the active role of B cells in the induction of the inflammatory response during TI antigen challenge in close collaboration with Mφ.
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