Structure and biological function of human IgD. IX. Anti-IgD activation of human lymphocytes.

Human peripheral lymphocytes were stimulated to incorporate tritiated thymidine when cultured with anti-sigma. The stimulation of lymphocytes by anti-sigma inversely correlates to PHA-induced lymphocyte transformation. In addition, lymphocytes from individuals with low serum IgD levels exhibited a significant response to anti-sigma, whereas, those with normal or slightly elevated levels of serum IgD showed minimal stimulation. This study is the first to provide evidence that cell surface IgD may regulate metabolic functions of lymphocytes and is consistent with the idea that IgD is a 'triggering' receptor.     

Structure and biological functions of human IgD. VI. Serum IgD in patients with allergic bronchopulmonary aspergillosis.

Serum IgD levels were quantified in a large population of individuals with a variety of atopic and/or pulmonary disorders. As a group, patients with allergic bronchopulmonary aspergillosis (ABPA) have markedly elevated (5.03 mg%) serum IgD when compared with serum IgD levels from either normal individuals (2.17 mg%) or patients with a variety of pulmonary and/or atopic disorders. Evidence is presented suggesting that the increased serum IgD levels in ABPA patients are not necessarily related to a hyper-IgE or panhyperimmunoglobulinemia common to most of all these patients, but is rather associated with an additional quantitative difference in the immunological responsiveness of some of these individuals. An understanding of why these patients produce supranormal levels of IgD may reveal the basis for inducing an IgD antibody response and subsequently help in elucidating the biological function(s) of IgD.     

Structure and biological functions of human IgD. XIV. The development of a solid-phase radioimmunoassay for the quantitation of IgD in human sera and secretions.

A simple but sensitive radioimmunoassay method for the quantitation of IgD in sera and body fluids has been developed. Specifically purified chicken anti-delta was covalently coupled to cyanogen bromide activated filter paper discs and the discs were incubated together with the samples to be analyzed. The washed discs were next incubated with 125-I-labeled specifically purified chicken or rabbit anti-human delta-antibodies. IgD on the disc was quantified by reference to a standard curve prepared with purified human IgD. The sensitivity of this assay was approximately 10 ng/ml IgD. IgD was demonstrable in cord sera (18 of 18), hypogammaglobulinemic sera (15 of 15), amniotic fluids (6 of 6), salivas (8 of 10), normal cerebrospinal fluids (12 of 13) and cerebrospinal fluids from patients with subacute sclerosing panencephalitis (5 of 5). The finding of IgD in human secretions raises the interesting possibility that IgD may be a secretory antibody.     

Small intestinal bacterial overgrowth in patients with chronic lymphocytic leukaemia.

As part of a study to assess the possible contribution of lymphoid infiltration of the gastrointestinal mucosa to occult blood loss or malabsorption 20 patients with chronic lymphocytic leukaemia (CLL) had a lactulose hydrogen breath test. In 10 cases (50%) a small intestinal peak was detected, suggesting small bowel bacterial overgrowth, and this was confirmed in seven patients by the positive culture of jejunal aspirate. Of the patients with a positive hydrogen breath test, radiological examination showed a duodenal diverticulum in two but no anatomical abnormalities in the other cases. There was no evidence of achlorhydria and transit times were normal. There was no difference in the incidence of hypogammaglobulinaemia among those patients with evidence of small intestinal bacterial overgrowth and those without. Seven patients with a positive hydrogen breath test, however, had undetectable secretory piece in their jejunal aspirates whereas this was present in all patients with a normal breath test who had local immunoglobulin concentrations measured (p less than 0.05), indicating that the small intestinal bacterial overgrowth may be due to impaired local immunity.     

Electrophoresis of lymphocytes from normal human subjects and patients with chronic lymphocytic leukaemia

The electrophoretic mobility (EM) of peripheral blood lymphocytes from twelve normal subjects and four patients with chronic lymphocytic leukaemia (CLL) was studied in relation to the thymus-derived (T) lymphocytes and bone marrow-derived (B) lymphocytic system. An attempt was made to correlate electrophoretic results with other methods of identifying T and B lymphocytes—T cells by the formation of sheep-cell rosettes and B cells by the formation of erythrocyte–antibody–complement (EAC') rosettes and by staining for surface immunoglobulin.

In the normal subjects the majority of cells migrated quickly with a small `tail' of slower cells. It is suggested that the faster populations are T cells and the slower, B. In CLL the majority of the cells were slow migrators. There was agreement between the percentage of fast cells as assessed by electrophoresis with that of T cells by sheep-cell rosetting; there was also some agreement between the percentage of electrophoretically slow cells to B cells by EAC' rosettes or surface immunoglobulin.

It was possible to remove some of the B cells by density centrifugation after forming EAC' rosettes. This further defined the T cell peak on electrophoresis.

It is concluded that T and B cells carry different surface charge densities which permit them to be separated by electrophoresis and that the malignant B lymphocytes of CLL migrate electrophoretically in a similar fashion to normal B cells.

     

Blood dendritic cells in patients with chronic lymphocytic leukaemia

Myeloid and plasmacytoid dendritic cells (MDC, PDC) play a key role in the initiation of immune responses. We found a reduction of both DC subsets in 42 patients with chronic lymphocytic leukaemia (CLL) at diagnosis (P<0.0001 and 0.0001 vs. controls, respectively), likely related to the high secretion of CCL22 and CXCL12 (P=0.04 and 0.008 vs. controls, respectively) by leukaemic cells. However, CD14+ monocytes from CLL patients could give rise to functional IL-12p70-secreting monocyte-derived DCs, capable of inducing a type 1 polarization immunostimulatory profile. These monocyte-derived DCs from CLL patients efficiently migrate in response to CCL19/MIP-3beta chemokine, suggesting that functional autologous DCs can be generated for immunotherapeutic purposes to circumvent DC defects in CLL.     

Autoimmunity in chronic lymphocytic leukaemia.

The prevalence of autoantibodies in B cell chronic lymphocytic leukaemia (B-CLL) was investigated. A lower prevalence of autoimmune haemolytic anaemia than that found in other series was found: large numbers of non-progressive stage A disease cases were included, in which the prevalence of autoimmune haemolytic anaemia is low. Non-haematological autoantibodies were no commoner than in age matched controls. Whatever explanation is offered for autoimmune phenomena in B-CLL it must take account of the fact that those phenomena are virtually confined to autoantibodies against the formed elements of the blood.     

Non-cytotoxic antibodies in chronic lymphocytic leukaemia.

Non-cytotoxic Fc receptor blocking antibodies against autologous B lymphocytes were sought in sera from patients with chronic lymphocytic leukaemia (CLL), using a rosette inhibition assay. They were found in 11 of 52 (21%) of patients with CLL, but were not associated with previous blood transfusion or pregnancy, suggesting that they were unlikely to have resulted from allogeneic stimulation. Fc receptor blockade was more commonly detected in sera from patients with stage B rather than stage A CLL (Binet classification), though this did not achieve significance beyond the 90% level, and it was noted in 62.5% of those with lymphocyte doubling times of less than one year, compared with 36.3% of those whose lymphocyte doubling time was more than one year. The results indicate that autologous Fc receptor blocking antibody activity occurs in sera from patients with CLL, is more likely to be generated by the tumour itself than by allogeneic stimulation, and is associated with increased tumour load. Such antibodies may permit tolerance of tumour by the host.     

Serum paraproteins in chronic lymphocytic leukaemia.

The presence of paraproteins in the sera of 10 patients with chronic lymphocytic leukaemia (CLL) was investigated using immunoisoelectric focusing. Monoclonal immunoglobulins were found in nine of these 10 sera. Five sera contained a single monoclonal IgM paraprotein, one serum contained a single monoclonal IgG paraprotein, while three sera contained more than one monoclonal paraprotein--namely, IgM + IgD, IgM + IgG, and IgM + IgD + IgG. The results indicate that the malignant B cells of CLL may be at a later stage of differentiation than previously assumed and serum monoclonal immunoglobulin could be of value as a tumour marker.     

Angiogenesis in the bone marrow of patients with chronic lymphocytic leukaemia

Angiogenesis is a process of formation of new vessels from the preexisting ones. It is involved in many physiological processes, at the same time, however, it is involved also in the progress of tumoral growth. Although a lot is known about angiogenesis in solid tumors where it plays a role in tumoral invasion and its metastatic potential, in hematological malignancies it has been appreciated only recently. However, the results of studies on abnormal angiogenesis in hematological malignancies are inconsistent. Angiogenesis can be studied at different levels; histologically, it is studied in the infiltrated tissues (lymph nodes, bone marrow) and quantified as microvessel density (MVD). The aims of our study were to introduce the method of MVD quantification in the bonemarrow using immunohistochemical detection of endothelial markers (fVIII) and then evaluate MVD in bone marrow samples in a group of patients with chronic lymphocytic leukaemia (CLL) and compare the results with a control group of patients (CON). CLL is a typical malignancy of the hematopoietic tissue but the course and the prognosis of patients with this disease vary considerably. For this reason there is urgent need for novel prognostic markers in order to assess individual patient prognosis and tailor treatment. Angiogenesis is one of the possible markers which may add more informations about the course of this disease. So far only few studies have been published about angiogenesis measured as MVD in CLL patients andthe results are inconsistent. In our study, both the number and the area of microvessels were increased in bone marrow of patients with CLL, but the number and area of sinuses were not. It can be concluded that there are signs of abnormal angiogenesis in bone marrow of patients with CLL but larger study with longer follow-up is needed to give more specific information about prognostic value of these findings.     

Prognostic factors in chronic lymphocytic leukaemia: contribution of recent biological markers

Chronic lymphocytic leukemia (CLL) is the most common lymphoid hemopathy in elderly. Diagnosis of CLL is easily made with a full blood count and immunophenotyping, but there is an heterogeneity in clinical evolution. Until now, scheduling of treatment is based on Rai or Binet staging systems. These staging systems can not distinguish patients with a rapid evolution and thus who will need an earlier treatment. In order to detect these patients, it is useful to have some relevant markers to predict disease evolution. This article reviews recent biologic markers that can be used to evaluate long term prognosis of CLL patients.     

Role of BAFF and APRIL in human B-cell chronic lymphocytic leukaemia

B-cell chronic lymphocytic leukaemia (B-CLL) is the most prevalent leukaemia in Western countries and is characterized by the gradual accumulation in patients of small mature B cells. Since the vast majority of tumoral cells are quiescent, the accumulation mostly results from deficient apoptosis rather than from acute proliferation. Although the phenomenon is relevant in vivo, B-CLL cells die rapidly in vitro as a consequence of apoptosis, suggesting a lack of essential growth factors in the culture medium. Indeed, the rate of B-CLL cell death in vitro is modulated by different cytokines, some favouring the apoptotic process, others counteracting it. Two related members of the tumour necrosis factor family, BAFF (B-cell activating factor of the TNF family) and APRIL (a proliferation-inducing ligand), already known for their crucial role in normal B-cell survival, differentiation and apoptosis, were recently shown to be expressed by B-CLL cells. These molecules are able to protect the leukaemic cells against spontaneous and drug-induced apoptosis via autocrine and/or paracrine pathways. This review will focus on the role of BAFF and APRIL in the survival of tumoral cells. It will discuss the expression of these molecules by B-CLL cells, their regulation, transduction pathways and their effects on leukaemic cells. The design of reagents able to counteract the effects of these molecules seems to be a new promising therapeutic approach for B-CLL and is already currently developed in the treatment of autoimmune diseases.     

Spontaneous lymphocyte transformation in chronic lymphocytic leukaemia.

In ten patients with CLL and in eight controls T lymphocytes were separated out by the rosetting method. The in vitro reactivity of these cells and of unseparated lymphocytes was studied in cultures without stimulants. A net increase of spontaneous lymphocyte transformation was found in the populations enriched with T cells in patients with CLL with respect to analogous populations in the controls. This increase is interpreted as a cell-mediated immunological reaction and due to T-cell sensitization towards the neoplastic B cells.     

Direct effect of interleukin 2 on chronic lymphocytic leukaemia B cell functions and morphology.

The functional and morphological changes induced by recombinant interleukin 2 (IL-2) were studied in purified B cells from patients with untreated B chronic lymphocytic leukaemia (B-CLL). In eight of nine patients, purified B-CLL cells increased their DNA synthesis in response to IL-2 without preactivation in vitro. This response, studied in detail in three patients, was dose dependent and reached a maximum on day 5 or 6. IL-2 induced or increased IgM secretion in cultures from five of the nine patients studied. Two of this responsive group were particularly interesting as IL-2 not only stimulated IgM secretion but also induced the secretion of IgG. Immunoglobulin production was invariably monoclonal. B CLL cells incubated with IL-2 showed distinct morphological changes including an increase in the size of cytoplasm and enlargement of nuclei together with the appearance of nucleoli. These changes were present in all IL-2 treated cultures but were more pronounced in those containing immunoglobulin secreting cells. None of the IL-2 induced changes appeared to correlate with the clinical stage of the disease or the level of Tac antigen expression on the freshly isolated CLL B cells.     

Expression of natural killer cell activating receptors in patients with chronic lymphocytic leukaemia

Recent advances in chronic lymphocytic leukaemia (CLL) treatment, more particularly through upfront use of anti-CD20 monoclonal antibodies, have prolonged patient progression-free survival. Nonetheless, apart from allogeneic stem cell transplantation, no curative treatment is available. One possible explanation for the lack of cure in CLL could be a defective immune anti-tumour response. As the result of abnormal HLA class I molecule expression, CLL cells escape from specific T-lymphocyte immunity but should be the target for the innate natural killer (NK) cell-mediated immune response. Defective NK cytotoxicity as the result of decreased expression of the natural cytotoxicity receptors (NCRs) NKp30/NCR3, NKp44/NCR2 and NKp46/NCR1 has been described in haematological malignancies such as acute myeloid leukaemia. This prompted us to focus our attention on NCR expression on NK cells from patients with CLL. Although we failed to detect any difference between CLL patients and healthy age-matched controls, a precise analysis of clinical data showed a correlation between decreased NCR expression and poor prognosis factors such as low haemoglobin level, high (>30×10(9) per litre) lymphocyte count or elevated C-reactive protein. Together, these observations support the rationale for restoration of normal NK cell functions in patients with CLL, putatively through the use of immune therapy protocols that already have demonstrated some benefit in acute myeloid leukaemia such as interleukin-2 plus histamine dihydrochloride.     

Acquired factor XI inhibitor in chronic lymphocytic leukaemia.

A 71 year old man with chronic lymphocytic leukaemia (CLL) experienced excessive bleeding following transurethral resection of the prostate. Investigations showed a prolonged kaolin cephalin clotting time (KCCT) with low concentrations of factor XI. The prolonged KCCT was largely corrected by mixing with normal plasma but this correction was lost on incubation, confirming the presence of an inhibitor. He was treated with pulsed methylprednisolone and chlorambucil which resulted in the resolution of the bleeding problem and the loss of detectable circulating inhibitor.