B lymphocytes transformation

Devergne, O., Hatzivassiliou, E., Izumi, K. M., Kaye, K. M., Kleijnen, M. F., Kieff, E., and Mosialos, G. (1996). Association of TRAFl, TRAF2, and TRAF3 with an Epstein-Barr virus LMPl domain important for B-lymphocyte transformation Role in NF-kappaB activation. Mol. Cell. Biol. 16, 7098-7108. 

Lizundia R, Chaussepied M, Huerre M, WerUng D, Di Santo JP, Langsley G (2006) c-Jun NH2-terminal kinase/c-Jun signaling promotes survival and metastasis of B lymphocytes transformed by Theileria. Cancer Res 66 6105-6110... 

Immunoglobulins, or antibodies, are proteins produced by the pixsma cells of the bone marrow as part of the immune response. The plasma cells are B lymphocytes transformed after exposure to a foreign (or occasionally an endogenous) antigen. 

Inhibition and stimulation of LOX activity occurs as a rule by a free radical mechanism. Riendeau et al. [8] showed that hydroperoxide activation of 5-LOX is product-specific and can be stimulated by 5-HPETE and hydrogen peroxide. NADPH, FAD, Fe2+ ions, and Fe3+(EDTA) complex markedly increased the formation of oxidized products while NADH and 5-HETE were inhibitory. Jones et al. [9] also demonstrated that another hydroperoxide 13(5)-hydroperoxy-9,ll( , Z)-octadecadienoic acid (13-HPOD) (formed by the oxidation of linoleic acid by soybean LOX) activated the inactive ferrous form of the enzyme. These authors suggested that 13-HPOD attached to LOX and affected its activation through the formation of a protein radical. Werz et al. [10] showed that reactive oxygen species produced by xanthine oxidase, granulocytes, or mitochondria activated 5-LOX in the Epstein Barr virus-transformed B-lymphocytes. 

Monoclonal antibody technology entails isolation of such B-lymphocytes, with subsequent fusion of these cells with transformed (myeloma) cells. Many of the resultant hybrid cells retain immortal characteristics, while producing large quantities of the monospecific antibody. These hybridoma cells can be cultured long term to effectively produce an inexhaustible supply of the monoclonal antibody of choice. 

Although Epstein-Barr virus is capable of inducing cellular transformation, few antibody-producing B-lymphocytes display the viral cell surface receptor. Most, therefore, are immune... 

GM-CSF is undetectable in the serum of normal humans, and no normal cells have been shown to express this protein constitutively. Some transformed cells may constitutively express GM-CSF, and it is actively synthesised and secreted by antigen- and lectin-stimulated T cells and by endothelial cells and fibroblasts exposed to TNF, IL-1 or endotoxin. Other sources of GM-CSF include stimulated B lymphocytes, macrophages, mast cells and osteoblasts, whilst TNF and IL-1 can stimulate its production by acute myeloid leukaemia cells. Some solid tumours and synovial cells from rheumatoid joints may also express GM-CSF and this may be important in disease pathology. 

Literature data on cytotoxic effects of photoexcited fullerene C60 are controversial. In the studies on transformed B-lymphocytes of Raji fine, phototoxic action of water-soluble carboxy-C60 was not revealed even upon its concentration of 5 x 10 5 M (Irie et al., 1996). In the study (Kamat et al., 2000) damaging effect of fullerenes C60 in dependence on intensity of irradiation toward CHO cells has been demonstrated. Using microsomal fraction of rat liver that was treated with C -cyclodextrin complex, it was shown that already in 5-30 min after UV-irradiation the accumulation of LPO products occurs that is suppressed by antioxidants like ascorbic acid and a-tocopherol. Similar effect of fullerenes C60 has been revealed in microsomal fraction of the cells of ascitic sarcoma 180 (Kamat et al., 2000). 

In addition to these activities, IL-6 has been shown, in vitro at least, to regulate the growth and differentiation of various other cells — both normal and transformed. It inhibits the in vitro growth of human fibroblasts and endothelial cells. As both these cell types also synthesize IL-6, this may suggest the presence in vivo of a negative autocrine feedback loop (Figure 5.8). Plasmacytoma cells (transformed antibody-producing B lymphocytes), on the other hand, also secrete IL-6 and express and IL-6 receptor. In this case, however, the feedback loop is positive. 

Although Epstein-Barr virus (EBV) is capable of inducing cellular transformation, few antibody-producing B lymphocytes display the viral cell surface receptor. Most, therefore, are immune to EBV infection. Even upon successful transformation, most produce low-affinity IgM antibodies, and the cells are often unstable. Having said that, one monoclonal antibody approved for medical use (Humaspect, Table 10.4) is produced by a human lymphoblastoid cell line originally transformed by EBV. 

The production yields of human monoclonals upon immortalization of the human B lymphocyte (by whatever means) are also low. Recoveries of 10-20 /ig per 10 transformed cells are not unusual. Such figures are approximately 10-fold lower than production levels of murine monoclonals. In addition, immortalized human B lymphocytes grow to a much lower cell density than that achievable with its murine counterpart. These latter factors render the production of human monoclonals an expensive process. 

Antibodies are expressed by hybridoma cells formed by cell fusion of sensitized animal or human B lymphocytes with myeloma cells, or they are generated by EBV (Epstein-Barr virus) transformation of sensitized B lymphocytes. Other heterologous expression systems such as bacteria, yeast, insect cells, and mammalian cells have also been used for expression of antibodies and their fragments. However, because of renaturation problems, glycosylation, and expression levels, mammalian cells are mostly used for the expression of monoclonal antibodies. More recently, technologies have been extensively developed for the expression of antibodies in transgenic animals and transgenic plants. 

Casali, P., Inghirami, G., Nakamura, M., Davies, T. F., and Notkins, A. L. (1986). Human monoclonals from antigen-specific selection of B lymphocytes and transformation by EBV. Science 234, 476-479. 

Despite the well-studied respiratory burst it is now clear that superoxide is also released by a variety of non-phagocytic cells. For example human B-lympho-cytes, which have been transformed by Epstein-Barr virus, express a super-oxide-generating system similar in many respects to the NADPH-oxidase of neutrophils [80]. This oxidase in lymphocytes can be stimulated by cytokines, suggesting that superoxide release may be a normal function of these B-lympho-cytes [81], NADPH-oxidase also occurs in normal peripheral B-lymphocytes, but disappears from the cell surface during final differentiation to plasma cells. 

The basic principle of hybridoma technology is shown in Fig. 2.23. B cells obtained from the spleens of mice immunized with the antigen of interest are fused with estabhshed mouse myeloma cells. Myeloma cells are transformed cancerous cells of the B lymphocyte lineage with infinite growth capacity. While many myeloma cell hnes continue to secrete their antibody, others lose the ability to produce antibodies. Spleen cells from immunized animals are fused with non-antibody-producing myeloma cells, resulting in a hybrid cell which inherits the properties of both the fusion partners. Thus the hybrids are capable of continued growth in culture like the myeloma cell and also maintain the antibody production of the spleen B cell. 

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