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7. Molecular investigation of the cytokines produced by normal and malignant B lymphocytes

Author

Schena, M., Gianluca GAIDANO, Gottardi, D., Malavasi, F., Larsson, L. -G, Nilsson, K., and Caligaris-Cappio, F.

Subjects
B-Lymphocytes, Interleukin-6, Tumor Necrosis Factor-alpha, Blotting, Western, Palatine Tonsil, Proteins, Blotting, Northern, CD5 Antigens, Lymphocyte Activation, Antigens, CD, Transforming Growth Factor beta, Leukemia, B-Cell, Cytokines, Humans, Interleukin-2, Interleukin-4, Interleukin-1
Abstract

Different normal and malignant human B-cell populations were studied with a twofold aim: to define which cytokines are produced in vivo, and to assess the relationship between cytokine production and kinetic state. To analyse normal B-cells representative of different stages of activation and proliferation in vivo, we purified germinal centre (GC)-B blasts and mantle B (M-B) cells from tonsils. To compare malignant B lymphocytes with their closest normal equivalent cells, we separated malignant CD5+B lymphocytes from the peripheral blood of patients with B-chronic lymphocytic leukemia (B-CLL) and normal CD5+B lymphocytes from cord blood. The expression of interleukins (IL) IL-1 alpha, IL-1 beta, tumour necrosis factor alpha (TNF-alpha), transforming growth factor beta (TGF-beta), IL-2, IL-4, and IL-6 genes was analysed using Northern and Western blotting techniques. TNF-alpha mRNA is produced by resting (M-B) and actively proliferating (GC-B) normal B lymphocytes. TGF-beta mRNA is present at high levels in resting normal M-B cells, while the transcript levels are lower in proliferating GC-B and in activated CD5+B lymphocytes. IL-2 production is limited to the actively proliferating GC-B blasts, IL-1 beta and IL-6 to resting M-B cells. The cytokine production profile of CD5+ malignant B-CLL cells differs from that of their putative normal counterparts and is more like the profile of M-B cells, since B-CLL cells produce IL-1 beta, TNF-alpha, TGF-beta, and IL-6. These observations lead to the following conclusions: among normal B lymphocyte populations, resting M-B lymphocytes are the most active cytokine producers, and B-CLL malignant B cells reflect the production pattern of normal resting B lymphocytes.

Published
1992

8. A novel role of the CX3CR1/CX3CL1 system in the cross-talk between chronic lymphocytic leukemia cells and tumor microenvironment

Author

S. Matis, F. Blengio, Maria Bertolotto, Fabio Malavasi, Domenico Ribatti, Vito Pistoia, Luciano Ottonello, Silvia Deaglio, Davide Rossi, Gianluca Gaidano, Claudio Tripodo, Simonetta Zupo, Anna Corcione, Valentina Audrito, Elisa Ferretti, Ferretti, E, Bertolotto, M, Deaglio, S, Tripodo, C, Ribatti, D, Audrito, V, Blengio, F, Matis, S, Zupo, S, Rossi, D, Ottonello, L, Gaidano, G, Malavasi, F, Pistoia, V, and Corcione, A.

Subjects
Adult, Male, chemokines, chronic lymphocytic leukemia (CLL), nurselike cells (NLCs), tumor microenvironment, Cancer Research, Chemokine, Stromal cell, Chronic lymphocytic leukemia, CX3C Chemokine Receptor 1, Antigens, Differentiation, Myelomonocytic, C-C chemokine receptor type 7, Cell Communication, Chemokine receptor, Antigens, CD, immune system diseases, hemic and lymphatic diseases, medicine, Humans, Phosphorylation, Aged, Aged, 80 and over, Tumor microenvironment, biology, Chemokine CX3CL1, Chemistry, Chemotaxis, Hematology, Middle Aged, medicine.disease, Leukemia, Lymphocytic, Chronic, B-Cell, CX(3)CR1/CX(3)CL1 system, chronic lymphocytic leukemia, Leukemia, Haematopoiesis, Oncology, Cancer research, biology.protein, Female, Receptors, Chemokine, Lymph Nodes, Proto-Oncogene Proteins c-akt, Signal Transduction
Abstract

Several chemokines/chemokine receptors such as CCR7, CXCR4 and CXCR5 attract chronic lymphocytic leukemia (CLL) cells to specific microenvironments. Here we have investigated whether the CX(3)CR1/CX(3)CL1 axis is involved in the interaction of CLL with their microenvironment. CLL cells from 52 patients expressed surface CX(3)CR1 and CX(3)CL1 and released constitutively soluble CX(3)CL1. One third of these were attracted in vitro by soluble CX(3)CL1. CX(3)CL1-induced phosphorylation of PI3K, Erk1/2, p38, Akt and Src was involved in induction of CLL chemotaxis. Leukemic B cells upregulated CXCR4 upon incubation with CX(3)CL1 and this was paralleled by increased chemotaxis to CXCL12. Akt phosphorylation was involved in CX(3)CL1-induced upregulation of CXCR4 on CLL. In proliferation centers from CLL lymph node and bone marrow, CX(3)CL1 was expressed by CLL cells whereas CX(3)CR1 was detected in CLL and stromal cells. Nurselike cells (NLCs) generated from CLL patient blood co-expressed surface CX(3)CR1 and CX(3)CL1, but did not secrete soluble CX(3)CL1. Only half of NLC cell fractions were attracted in vitro by CX(3)CL1. In conclusion, the CX(3)CR1/CX(3)CL1 system may contribute to interactions between CLL cells and tumor microenvironment by increasing CXCL12-mediated attraction of leukemic cells to NLC and promoting directly adhesion of CLL cells to NLC.

Published
2011

9. C3b receptors mediate the growth factor-induced proliferation of malignant B-chronic lymphocytic leukemia lymphocytes.

Author

Caligaris-Cappio F, Schena M, Bergui L, Tesio L, Riva M, Rege-Cambrin G, Funaro A, and Malavasi F

Subjects
Aged, Cell Division drug effects, Cells, Cultured, Female, Humans, Interleukin-4, Lymphocytes pathology, Male, Middle Aged, Receptors, Complement 3b, Receptors, Complement 3d, Interleukins pharmacology, Leukemia, Lymphoid pathology, Lymphocytes drug effects, Receptors, Complement metabolism
Abstract

We have investigated the function of C3b receptor (CR1) in the malignant lymphocytes of B-chronic lymphocytic leukemia (B-CLL) mimicking the physiological ligand C3b with the anti-CR1 monoclonal antibody CB04 covalently linked to Sepharose CL-4B (CB04-S). The binding of insolubilized CB04-S to CR1 gave a progression signal to B-CLL cells which became B cell growth factor (BCGF)-responsive. The cells of 13 of 14 cases treated with CB04-S showed an active time-dependent proliferation when BCGF was added to the culture. After 72 hr of exposure to BCGF, the growth fraction evaluated with the Ki67 monoclonal antibody was 23.4 +/- 8.9 and the proportion of cells in S phase assessed by the bromodeoxyuridine incorporation technique was 18.6 +/- 8.5%. The proper sequence of CB04-S followed by BCGF was also important since the proliferation was halved when the sequence was reversed or the two signals were delivered concomitantly. CB04-S and BCGF alone failed to induce any significant proliferation; the percentage of cycling cells was less than 1% overlapping that of control culture cells. On the contrary, the proliferation of normal tonsil B cells was triggered both by CB04-S and by BCGF used as single agents (bromodeoxyuridine+ cells 12.7 +/- 5.1% and 20.0 +/- 7.3, respectively). Together these data indicate that malignant B-CLL cells need a sequential two-step signal based upon CR1 binding in order to be activated in vitro. This is a major difference with normal tonsil B lymphocytes whose proliferation is triggered both by CB04-S and by BCGF used as single agents.

Published
1987

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