The heterogeneity of leukemic reticuloendotheliosis, "hairy cell leukemia". Evidence for its monocytic origin.

The presence of B, T, and monocyte markers were studied on the spleen and peripheral blood mononuclear cells from two patients with leukemic reticuloendotheliosis. A high proportion of cells from both patients bore a receptor for cytophilic antibody, both in suspension and frozen tissue section. Cells in suspension lacked surface immunoglobulins or a receptor for sheep red blood cells. These results favor the evidence that "hairy cells" are monocytic in origin.     

Reactivity of acute lymphoblastic leukemia and normal bone marrow cells with the monoclonal anti-B-lymphocyte antibody, anti-Y 29/55

Malignant lymphocyte populations in peripheral blood of patients with B-cell chronic lymphocytic leukemia, leukemic variant of B-cell non-Hodgkin's lymphoma, and hairy cell leukemia can be characterized by the use of a monoclonal murine antibody (anti-Y 29/55) which is directed against a cell membrane component normally confined to the sessile nonrecirculating cells of the B-lymphocyte population in lymphoid tissues. The present report describes the reactivity of the anti-Y 29/55 antibody with bone marrow cells obtained from children with acute lymphoblastic leukemia using an indirect immunofluorescence method in combination with morphological and cytokinetic studies. In 25 patients (acute lymphoblastic leukemia subtype: 14 common; 4 pre-B-cell; 4 null; and 3 T-cell), a maximum of 2% of cells (small lymphocytes) were stained. One patient presented with blasts exhibiting cytoplasmic and surface immunoglobulin M (IgM) (pre-B-B-cell acute lymphoblastic leukemia). About 11% of this patient's blast cells showed a positive reaction with anti-Y 29/55. They could not be differentiated by morphological criteria from the anti-Y 29/55-negative blast cell population. In another patient with pre-B-B-cell acute lymphoblastic leukemia, only 1% of anti-Y 29/55-positive cells was found. In bone marrow of children with relative lymphocytosis, 1.4 to 8.7% of mononuclear cells reacted with anti-Y 29/55. Morphologically, these cells were small lymphocytes and predominantly expressed surface IgM. In two of these children, a further subdivision of bone marrow cells could be achieved by combining anti-Y 29/55 and cytoplasmic IgM reactivity with [3H]thymidine pulse labeling. These studies revealed that the actively proliferating, normal pre-B-cell population was anti-Y 29/55-nonreactive, whereas a nonproliferating population of anti-Y 29/55-reactive, cytoplasmic IgM-positive cells probably represented B-cells with surface immunoglobulin M reacting when cytoplasmic IgM was assessed. We conclude that the reactivity of the monoclonal anti-B-cell antibody (anti-Y 29/55) is restricted to surface immunoglobulin-positive bone marrow cells and that neither leukemic or normal pre-B-cells nor common, null-cell, or T-cell acute lymphoblastic leukemia blasts react.     

Hairy cell leukemia: establishment of a cell line and its characteristics

A hairy cell leukemia (HCL) line, ZK-H, was established from peripheral blood of a 69-year-old male patient. The ZK-H cells and the patient's original hairy cells shared the same surface properties; both possessed membrane-bound IgG with kappa light chains and villous surface structures. The ZK-H line carried Epstein-Barr virus (EBV)-determined nuclear antigen, but the patient's fresh leukemic cells lacked this antigen. Morphologically, the ZK-H cells appeared lymphoblastoid and more primitive than the preculture cells. The ZK-H line had a hyperdiploid chromosome constitution of 47 and trisomy no. 2. The presence of membrane-bound immunoglobulin and of B-cell tropic EBV in this cell line provides further evidence for the B-cell nature of HCL in this patient     

New cell line from hairy-cell leukemia: confirmation of leukemic cell origin by karyotype and Ig gene analysis

A hairy-cell leukemia (HCL) line, BNBH-I, was established from the peripheral blood of a 40-year-old male patient with HCL in a relatively stable clinical phase after splenectomy. The cells have since been growing continuously for more than 2 years. Their cell surface immunoglobulin (sIg) was identical with that found on the surface of freshly isolated leukemic cells, consisting of IgG-kappa. The BNBH-I cells were more mature than the original hairy cells in their degree of B-cell differentiation, as reflected by a decrease in sIg expression together with the appearance of some cytoplasmic Ig (cIg)+ cells, loss of EA gamma-rosette formation and reactivity with monoclonal antibody (MAb) FMC7, and an increase in the proportion of MAb PCA-I+ cells. The BNBH-I cells possessed the antigen recognized by Leu-M5, a highly specific MAb for HCL. Epstein-Barr virus nuclear antigen (EBNA) was present. Both the freshly isolated leukemic cells and the cell line had the 14q+ involving q32 chromosomal abnormality, and their Ig gene rearrangements were also identical. Following exposure to 12-O-tetradecanoylphorbol-13-acetate (TPA), both the freshly isolated leukemic cells and the BNBH-I cells adhered to culture dishes and extended long, thin processes, a phenomenon characteristic of HCL. These results indicate that the BNBH-I line was derived from the leukemic hairy cells.     

Diagnosis and monitoring in patients with hairy cell leukemia using the monoclonal antibody anti-HC2

Immunofluorescent staining of peripheral blood mononuclear cells with the monoclonal antibody anti-HC2 combined with phase microscopic examination identified leukemic hairy cells in nine of 13 patients (69%) evaluated at Memorial Hospital prior to treatment with recombinant alpha-interferon (rIFN-alpha A). The remaining four patients required further studies of bone marrow or peripheral blood cytospin samples for diagnosis. In some cases a low percentage of cells staining with anti-HC2 could be significantly increased by depleting T cells from the sample using sheep red blood cell rosetting. These 13 patients represent a subgroup of patients treated on a phase II rIFN-alpha A study at Memorial Hospital. Ten patients (77%) achieved a partial response in a median of 128 days (range, 64-234). One patient achieved a minor response and two patients failed treatment. Nonhematologic toxicity, consisting of fever and malaise, was transient in all patients. Serial determinations of HC2-positive cells in the peripheral blood closely paralleled disease activity in the bone marrow in one patient treated for 4 years with various therapeutic modalities. The antibody anti-HC2 may play a significant role in the diagnosis and monitoring of hairy cell leukemia using peripheral blood sampling.     

Hairy cell leukemia. An immunologic and ultrastructural study.

A case of hairy cell leukemia in a 39-year-old man is reported. Hairy cells from the peripheral blood, spleen, and bone marrow had lambda-type immunoglobulin on their surfaces; those from the peripheral blood and bone marrow also had IgD on their cell membranes. Frozen sections of spleen reacted with IgGEA, but not IgMEA or IgMEAC markers. Transmission electron microscopy revealed ribosome-lamella complexes in cells from the spleen, but not the peripheral blood. Scanning electron microscopy demonstrated a spectrum of cell surface morphology with many cells characterized by ridges and ruffles. The significance of these findings is considered and it is suggested that the hairy cell is a B lymphocyte.     

Colony formation in the absence of added growth factors by peripheral blood T-cell colony-forming cells of patients with T-cell malignancies

Clonogenic cells from peripheral blood of 13/16 patients with T-cell malignancies generated colonies in methylcellulose in absence of added growth factors or mitogenic stimulation. Spontaneous colonies were also obtained from purified cell fractions (E-OKT3- and/or E+ cells) in 73% and 57% of the patients, respectively. No spontaneous colony growth was observed with mononuclear cells of patients with solid tumors, non-T-cell leukemias or normal subjects. Colonies consisted of acid-phosphatase-positive, myeloperoxidase-and PAS-negative lymphoblasts bearing T-cell surface markers. Although the phenotype of pooled colony cells from either unfractionated mononuclear cells or E-OKT3- -derived colonies varied from patient to patient, the colonies, like fresh leukemic cells, were mostly composed of relatively immature cells as assessed by the high proportion (greater than 40%) of OKT6+ and OKT10+ cells and the low proportion (less than 40%) of OKT3+ and/or E+ cells. Cytogenetic analysis of colony cells revealed either normal metaphases or chromosome anomalies similar to those observed in fresh leukemic cells. Moreover, cells from primary colonies exhibited a capacity for self-renewal in the absence of added growth factors.     

Comparative antiproliferative and apoptotic effects of resveratrol,epsilon-viniferin and vine-shots derived polyphenols (vineatrols) on chronic B lymphocytic leukemia cells and normal human lymphocytes

Trans-resveratrol, its dimer epsilon-viniferin and two preparations of vineatrol (a grape-derived polyphenol fraction isolated from vine-shots extracts) were compared for their effects on the proliferation and survival of normal and leukemic human lymphocytes. The two different batches of vineatrol (vineatrol 10 and 25%) was obtained by HPLC fractionation and contained 10 and 25% trans-resveratrol, respectively. The different polyphenols were added to cultures of leukemic cells from chronic B cell malignancies (B-cell chronic lymphocytic leukemia, B-CLL or hairy cell leukemia, HCL) or normal peripheral blood-derived mononuclear cells (PBMC) as a control. The different polyphenols displayed anti-proliferative effect on the leukemic cells, as estimated by the observed inhibition of tritiated thymidine uptake and the reduction of cell recovery. Vineatrol 10% was the most potent whereas vineatrol 25% and resveratrol displayed comparable activity, epsilon-viniferin only exhibiting slight effets. The same order of potency was observed for their capacity to induce apoptosis in leukemic B cells. In contrast, the survival of normal peripheral blood mononuclear cells (PBMC) was little affected in the presence of these polyphenolic compounds and higher concentrations were required in order to elicit cell death. Polyphenol-driven apoptosis in chronic leukemic B cells was shown to correlate with an activation of caspase 3, a drop in the mitochondrial transmembrane potential, a reduction in the expression of the anti-apoptotic protein bcl-2, as well as a reduction in the expression of the inducible nitric oxide synthase (iNOS). Our data therefore indicate that vine-shoots may be a convenient and natural source of material for the purification of resveratrol and other polyphenolic compounds of putative therapeutic interest.     

Sezary syndrome in a patient with hairy cell leukemia in remission.

A 65-year-old man was evaluated for pancytopenia in March 1979, and found to have hairy cell leukemia (HCL). Treatment with splenectomy and subsequently interferon produced temporary remissions. In July 1985, the patient began intravenous deoxycoformycin (DCF) therapy, and after 1 year complete peripheral blood and bone marrow remission was achieved. Fourteen months after cessation of therapy, the patient developed a skin rash and was found to have cutaneous T-cell lymphoma and Sezary syndrome. Morphologic study of the hairy cells (HC) in the peripheral blood at presentation and the Sezary cells was distinct by light and electron microscopic study. Immunophenotyping of peripheral blood mononuclear cells showed clearly that the HC were of B-cell origin (CD20+, sIg+), whereas the lymphoid population at second presentation was T-cell (CD3+, CD4+, HLA-DR-). Clonal rearrangement of T-cell antigen receptor beta-chain gene was detected by Southern analysis of the Sezary cell population, whereas immunoglobulin heavy and light chain genes remained in germ line configuration. This is the first case of Sezary syndrome developing in a patient previously treated for HCL where studies have confirmed distinct B-cell and T-cell origin of the two neoplasms. The authors suggest that treatment and disease-related immunosuppression are possible etiologic factors in the development of this second lymphoid neoplasm.     

Establishment of a hairy cell leukemia cell line carrying Tac antigen and phagocytic activity with B-cell characteristics

A hairy cell leukemia cell line designated "Hair-M" was established in a suspension culture derived from the peripheral blood of an 86-year-old Japanese male with a diagnosis of hairy cell leukemia. The Hair-M cells were identified as having prominent hair-like cytoplasmic projections by examination with phase-contrast and scanning electron microscopy. These cells displayed ruffled membranes and stublike microvilli similar to those observed on the surfaces of cells in the peripheral blood of the patient. Immunologic and cytochemical studies on the Hair-M cells confirmed derivation from the clone of the patient's leukemia cells. Although the cultured Hair-M cells had definite B-cell characteristics, such as IgG kappa-chains on the surface and in cytoplasm, they also demonstrated Tac antigen, which is usually expressed on activated T-cells, and myelomonocyte antigens determined by OKM-1 and MCS-1 monoclonal antibodies. Other cell surface markers, including E(-), IgGFc(-), IgMFc(-), C3R(+), Ia-like antigen(+), OKT9(+), OKT10(+), and terminal deoxynucleotidyl transferase(-), were detected; no Epstein-Barr virus-determined nuclear antigen was detected. The karyotype of the Hair-M cells was determined to be 46XY with -11, -14, and two marker chromosomes. The Hair-M cells also had phagocytic activity to rabbit anti-human IgG serum-coated polyacrylamide gel particles.     

Expression of p35 in hairy leukemic cells from Japanese patients

Spiro and coworkers found a 35000 dalton membrane protein (p35) expressed abundantly in hairy cells of a subset of white, American patients with hairy cell leukemia. They subsequently identified p35 to be the human homologue of murine Ii, an electrophoretically invariant protein associated with Ia molecules. Using related analytical techniques, we have studied leukemic cells of two blood and one splenic samples from three Japanese patients with hairy cell leukemia and have demonstrated abundance of p35 in the splenic and one of the blood samples. The second blood sample was negative for p35 by our method of analysis, but the same hairy cells became positive for p35 at 2 months in culture and negative again at 4 months in culture. All 3 samples of this patient (fresh hairy cells and cultured cells at 2 and 4 months) remained consistently positive for Ia and for light chain-restricted immunoglobulin. Control cells from one each of acute lymphoblastic leukemia and Burkitt's lymphoma were negative for abundant p35 expression after culture as well as in the fresh state. One can conclude that the Japanese form of hairy cell leukemia, while varying in several distinct clinical and laboratory features from hairy cell leukemia in caucasians, is characterized also by abundant expression of p35 as seen in a subset of white patients. In addition, this expression of p35 is developed by hairy leukemic cells in culture initially but is lost upon longer term culture, in parallel with the level of tartrate-resistant acid phosphatase reactivity.     

Differentiation between benign and malignant human lymph nodes by means of immunologic markers.

A surface-marker assay combining immunofluorescence with anti-human immunoglobulin or anti-human brain serum (AHBS) and the formation of rosettes with untreated (E), antibody-sensitized (EA) and complement-coated (EAC) sheep erythrocytes was used to study mononuclear cell suspensions of human lymph nodes. The frequency of cells expressing more than one marker was increased in lymphoma nodes as compared to normal and hyperplastic nodes. The cells which simultaneously expressed complement receptors, surface immunoglobulin and the marker identified by AHBS represented the most prominent and characteristic subpopulation identified in neoplastic nodes. Distributions of cells with double and triple markers were studied by combining immunofluorescence with rosetting on frozen tissue sections. The multiple-marker cells had distributions that were characteristic in different human lymphomas. Benign and malignant human nodes could be distinguished on the basis of frequency and distribution of mononuclear cell populations carrying distinctive combinations of T- and B-cell surface markers.     

Genotypic and phenotypic evidence of T-cell leukemia in a patient successfully treated by interferon-alpha for typical hairy cell leukemia.

A 53-year-old man was diagnosed to have typical hairy cell leukemia. Immunophenotyping of frozen splenic tissue showed clonality of hairy cells for mu lambda, confirmed by the corresponding immunoglobulin gene rearrangements. The patient was successfully treated with interferon-alpha (IF-alpha). In the fifth year of treatment with IF-alpha the morphology of peripheral blood mononuclear cells (PBMC) and of bone marrow infiltration changed with the appearance of numerous small to intermediate shaped lymphocytes of a T-helper phenotype. Frank leukemia, resistant to IF-alpha treatment and ultimately aggressive chemotherapy, developed. Emergence of this second clonal disease was confirmed by rearrangement studies performed on PBMC; rearrangements of both alleles of the TCR beta were identified, whereas the JH and lambda IVS genes were in germline configuration. The outgrowth of a second, malignant T-cell clone paralleled by the disappearance (down-regulation?) of the initial B-cell clone while under cytokine treatment is consistent with the possibility that IF-alpha favoured the emergence of this second clone.     

Profil immunophénotypique des leucémies à tricholeucocytes à Madagascar

Hairy-cell leukemia (HCL) is a rare hematologic malignancy. Few studies have been carried out in this area in sub-Saharan Africa. The diagnosis is based upon the recognition of the characteristic ??hairy?? nature of the leukemic lymphoid cells in the peripheral blood smears and bone marrow smears and immunophenotypic analysis. Between 2009 and 2010, we have conducted an exploratory study to case report hairy cell leukemia in hematology CHU/Joseph Ravohangy Andrianavalona laboratory. Immunophenotypic analysis was done in CHR Sud Reunion and immunophenotypic profile has been compared to literature. For two years, two mens patients, 58 and 68 years old, were diagnosed as hairy cell leukaemic. From a clinical standpoint, patients all presented large spleen associated with severe pancytopenia. Haemogram and bone marrow aspiration showed infiltration by hairy cell leukemia. HCL had a characteristic immunophenotypic profile, the tumor cells express the markers CD11c, CD103, CD25 and CD123. But the first patient was positive for CD5 and CD23.     

T-cell lineage of a Friend virus-induced lymphatic leukemia in athymic rats

Athymic (rnu/rnu) and euthymic rats inoculated with the Friend virus-associated lymphatic leukemia virus developed lymphocytic leukemia. Neoplastic cells from these animals were evaluated by means of indirect immunofluorescence and flow cytofluorometry with monoclonal antibodies Ox-1, Ox-7, and W3/25, which react with surface antigens present on normal rat lymphoid cell populations. Lymphoid cells from leukemic animals revealed characteristic alterations in cell surface fluorescence profiles when compared to normal, healthy controls. Athymic and euthymic leukemic rats were similar in that many cells from both the spleen and bone marrow had markers on the cell surface normally found on thymocytes but not on mature peripheral lymphocytes. These studies provided evidence supporting the presence of T-lineage lymphocytes in the athymic rat. Further, this population of early or "pre"-T-lymphocytes included the predominant leukemia cell type induced by the Friend virus-associated lymphatic leukemia virus.     

Establishment and characterization of a new permanent cell line (GDM-1) from a patient with myelomonoblastic leukemia

The GDM-1 permanent cell line was established from the peripheral blood of a patient with a Philadelphia chromosome negative myeloproliferative disorder, after transformation to acute myelomonoblastic leukemia. The GDM-1 cells exhibited the same characteristics as those isolated from the peripheral blood of the patient prior to death: cells contained non-specific esterase sensitive to fluoride, myeloperoxidase, lysozyme (muramidase), and exhibited both Fc and complement (C₃) receptors but lacked B- and T-cell surface markers including T-associated antigens. E-rosetting capacity, surface and intracytoplasmic immunoglobulins and EBV determined nuclear antigen (EBNA). The GDM-1 cells bore the Ia receptor and the myeloid leukemia antigen (M-1). The karyotype of the cultured leukemic cells showed the same specific chromosomal abnormalities present in the monoblasts obtained from the peripheral blood prior to death, indicating that the cell line was derived from the original leukemic cells.     

Cryocrystalglobulinemia in hairy cell leukemia.

A patient with hairy cell leukemia (leukemic reticuloendotheliosis) was noted to have a spurious leukocytosis caused by a spontaneously crystallizing cryoglobulin. The cryoprotein was identified as IgG lambda. An intracytoplasmic immunoglobulin demonstrable within the hairy cell was also IgG lambda. The cryoglobulin spontaneously disappeared over a four day period. A reliable automated count on the Coulter Model S could be obtained by prewarming the blood specimen to 37 C.     

Induction of plasmacytoid and hairy cell features by phorbol esters (TPA) in B-lymphoma cells: attempted correlation with disease activity

Mononuclear cells concentrated from the blood of 16 non-Hodgkin's lymphoma (NHL) patients in the leukemic phase, were exposed to 10 ng/ml of TPA in an attempt to induce differentiation. Immunoglobulin (Ig) secretion, surface markers (SmIg, GP-70), tartrate resistant acid phosphatase (TRAP) and surface features were followed for up to six days in vitro. TPA induced 'hairy cell' like features in NHL cells as defined by cell morphology, ultrastructure, cell surface markers and the presence of TRAP. Unlike the results obtained in patients with CLL, cells from different patients at the same stage of disease reacted in a similar way. Differences were evident between NHL mononuclear cells obtained from patients in partial remission and active disease when compared with those derived from patients in complete remission. In the former group, NHL cells were maximally induced by TPA to secrete Ig and higher proportions of TRAP positive cells. In addition hairy cell features as seen by light and scanning electron microscopy were also more pronounced. TPA also induced the maximal expression of SmIg and GP-70 in cells derived from patients in this group. Patients with NHL in leukemic phase in complete remission did not express surface membrane GP-70 before or after TPA treatment while SmIg was expressed to some degree before TPA treatment and further induced following treatment with TPA. GP-70 appears to be a more reliable marker for follow-up of NHL patients than any other marker studied here.     

Establishment of a new human B cell line carrying t(11;14) chromosome abnormality

A new human cell line, SP-49, was established from the peripheral blood of a patient with leukemic conversion of diffuse lymphoma, medium-sized cell type. SP-49 cells were shown to have intracytoplasmic and surface immunoglobulin and Leu-12 antigen, but were negative for Epstein-Barr virus nuclear antigen. Chromosome analysis demonstrated that SP-49 had t(11;14) chromosome translocation involving 11q13 and 14q32, where oncogene bcl-1 and immunoglobulin heavy chain gene, respectively, were reportedly located.