Immunoglobulin Isotype Switching in a Plasma Cell Myeloma Patient Treated with High-dose Chemotherapy and Hematopoietic Stem Cell Transplantation

A malignant plasma cell clone usually produces a single abnormal monoclonal protein with a constant isotype. However, switching of paraprotein isotype has been reported to be a transient phenomenon associated with the recovery of B-cell function, and, in some cases, the switching might be misinterpreted as relapse. In August 2008, we encountered a case of a 59-year-old man with proteinuria and high IgG level (5.6 g/dL), kappa free light chain level of 2,660 mg/L, reversed A/G ratio (0.51), and multiple osteolytic lesions. Plasma cells, which accounted for 57% of all the nucleated cells, in bone marrow aspirates were positive for kappa immunostaining. Serum protein electrophoresis showed one M-spike, concentration of 4.87 g/dL in the beta region. Immunofixation electrophoresis revealed the peak as an IgG-kappa monoclonal protein; therefore, a diagnosis of plasma cell myeloma was made. Complete remission was achieved after chemotherapy, and autologous peripheral stem cell collection was performed. In March 2009, the patient underwent high-dose chemotherapy and autologous peripheral stem cell transplantation support. After 2 months, serum protein electrophoresis showed 2 M-spikes in the gamma region with positive IgM-lambda, IgG-lambda, and IgG-kappa, and these bands persisted. The electrophoretic mobility of the IgG-kappa protein was different from that of the original disease protein, and bone marrow results were the same as the previous ones. Although immunoglobulin isotype switch is known to have a benign course, it always requires careful monitoring because, in rare cases, true clonal switching may occur.

Vitamin C acts indirectly to modulate isotype switching in mouse B cells / 대한해부학회지

Vitamin C, one of essential micronutrients, has been reported to modulate the humoral immune responses in some mammals. We investigated whether vitamin C might modulate this response in mice by directly affecting B cells. Splenic B cells were isolated and activated by CD40- and B cell receptor-ligation in vitro. The cells were cultured with a pretreatment of vitamin C from 0 to 1 mM of concentrations. Vitamin C slightly increased apoptosis of B cells dose-dependently and behaved as an antioxidant. We found that in vivo administration of vitamin C by intraperitoneal injection affected isotype switching as previously reported: the titer of antigen-specific IgG1 antibody was decreased, while that of IgG2a was unaffected. Somewhat different from those observed in vivo, in vitro exposure to vitamin C slightly decreased isotype switching to IgG1 and increased isotype switching to IgG2a. Pretreatment with vitamin C in the safe range did not affect either proliferation of cultured B cells or the expression of CD80 and CD86 in those cells. Taken together, in vivo results suggest that vitamin C acts to modulate isotype switching in the mouse. However, because of our in vitro results, we suggest that the modulation exerted by vitamin C in vivo is by indirectly affecting B cells, perhaps by directly influencing other immune cells such as dendritic cells.

Characterization of Mouse B Lymphoma Cells (CH12F3-2A) for the Study of IgA Isotype Switching

BACKGROUND: It is well known that IgA isotype switching is induced by TGF-beta1. LPS-activated mouse normal B cells well differentiate into IgA secreting plasma cells under the influence of TGF-beta1. Nevertheless, there are lots of difficulties in studying normal B cells in detail because it is not simple to obtain highly purified B cells, showing low reproducibility and transfection efficacy, moreover impossible to keep continuous culture. To overcome these obstacles, it is desperately needed to develop B cell line which acts like normal B cells. In the present study, we investigated whether CH12F3-2A lymphoma cells are appropriate for studying IgA isotype switching event. METHODS: CH12F3-2A B cell line was treated with LPS and TGF-beta1, then levels of germ-line (GL) transcripts were measured by RT-PCR, and GLalpha promoter activity was measured by luciferase assay. In addition, membrane IgA (mIgA) expression and IgA secretion were determined by FACS and ELISA, respectively. RESULTS: TGF-beta1, regardless of the presence of LPS, increased level of GLalpha transcripts but not GLgamma2b transcripts. However, IgA secretion was increased dramatically by co-stimulation of LPS and TGF-beta1. Both mIgA and IgA secretion in the presence of TGF-beta1 were further increased by over-expression of Smad3/4. Finally, GLalpha promoter activity was increased by TGF-beta1. CONCLUSION: CH12F3-2A cell line acts quite similarly to the normal B cells which have been previously reported regarding IgA expression. Thus, CH12F3-2A lymphoma cell line appears to be adequate for the investigation of the mechanism(s) of IgA isotype switching at the cellular and molecular levels.

T Cell Dependent Antigen-Induced Immunoglobulin Isotype Swiching and Diifferentiation of Lymph Node / 대한면역학회지

Lymph nodes, one of peripheral lymphoid organs, are the sites, where the lymphocytes receive their initial instructions for producing effector functioning resulting in humoral or cell-mediated immunity. Each lymph node consists of an outer cortex in which there are aggregates of cells constituting the follicles, B-cell areas. Some follicles have central areas called germinal centers, which stain lightly. Germinal centers are B lymphoblast cell areas arising eccentrically in primary lymphoid follicles in response to T-cell dependent antigenic stimulation and are the generally accepted sites of generation of memory B cells and undergoing isotype switching and somatic mutation. We observed the morphologic, cellular, protein and molecular events arising in mouse popliteal lymph nodes in response to T-cell dependent antigenic stimulation. In this study mice were immunized into footpads with TNP-chicken ovalbumin. The germinal center formation in primary follicles of popliteal lymph nodes was first observed 6 days after immunization and germinal centers persisted until 24 days of immunization. Lymph node cells were stained with PE-labeled anti-B220 antibody and/or FITC labeled PNA and analyzed by using FACScan. B cells (B220(+) cell) in lymph node increased after 3 days and peaked between 6 and 18 days after immunization. The proportion of germinal center B cells (B220, PNA(high) cells) among lymph node B cells was low (2%) before immunization but increased at day 6 (9%) and reached the peak (30%) at day 18. The expression of IgG1 productive mRNAs and germline transcripts were observed by using RT-PCR. The expression of IgG1 productive mRNA was detected at day 10 and continued until 24 days after immunization. The expression of IgG1 germline transcripts was observed 10 days after immunization and rapidly declined over the next one week. IgG1 anti-TNP antibody, main isotype of anti-TNP antibodies, was first detected at day 14 and reached the peak level 24 days after immunization. Taken these data together, we can conclude that the first immunological event observed from mouse popliteal lymph node in response to T-cell dependent antigenic stimulation is the increase in the number of B cells, and this event is followed by appearance of germinal center B cells and at the same time by the formation of germinal center in primary lymphoid follicles. Once the germinal center is formed, the process of isotype switching to IgG1 occurs in lymph node and antigen-specific IgG1 antibody is produced.