scholarly journals Heterogeneous expression of a novel MPC-1 antigen on myeloma cells: possible involvement of MPC-1 antigen in the adhesion of mature myeloma cells to bone marrow stromal cells

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3721-3729 ◽  
Author(s):  
N Huang ◽  
MM Kawano ◽  
H Harada ◽  
Y Harada ◽  
A Sakai ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Cell Line ◽  
Bone Marrow Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Human Myeloma Cell Line ◽  
Heterogeneous Expression

Abstract Recent immunophenotypic analysis has shown that the heterogeneous expression of the adhesion molecule VLA-5 classifies myeloma cells into VLA-5+ mature and VLA-5- immature subpopulations. To further clarify the two myeloma subpopulations, we generated a monoclonal antibody, MPC- 1, by immunizing mice with an adherent human myeloma cell line, KMS-5. The MPC-1 antibody recognized a 48-Kd surface antigen on KMS-5 but not on U-266, a nonadherent human myeloma cell line. Specificity characterization showed that MPC-1 antigen was expressed on mature myeloma cells, normal plasma cells, and mature B cells, whereas pre-B cells and germinal center B cells lacked its expression. Monocytes and a human bone marrow stromal cell line, KM102, also expressed this antigen. Two subclones of MPC-1+ VLA-5+ (KMS-5Ad) and MPC-1-VLA-5+ (KMS- 5NAd) were separated from the KMS-5 cell line. The KMS-5NAd adhered to KM102 more tightly than did the KMS-5NAd, and the U-266 (MPC-1-VLA-5-) displayed almost no adherence to the KM102. The adhesion of the KMS-5Ad was partially inhibited by the MPC-1 antibody. These results, taken together, suggest that the MPC-1 antigen serves as a differentiation marker for B-lineage cells, including plasma cells, and may function as an adhesion molecule involved in the interaction of mature myeloma cells with bone marrow stromal cells.

scholarly journals Heterogeneous expression of a novel MPC-1 antigen on myeloma cells: possible involvement of MPC-1 antigen in the adhesion of mature myeloma cells to bone marrow stromal cells

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3721-3729 ◽  
Author(s):  
N Huang ◽  
MM Kawano ◽  
H Harada ◽  
Y Harada ◽  
A Sakai ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Cell Line ◽  
Bone Marrow Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Human Myeloma Cell Line ◽  
Heterogeneous Expression

Recent immunophenotypic analysis has shown that the heterogeneous expression of the adhesion molecule VLA-5 classifies myeloma cells into VLA-5+ mature and VLA-5- immature subpopulations. To further clarify the two myeloma subpopulations, we generated a monoclonal antibody, MPC- 1, by immunizing mice with an adherent human myeloma cell line, KMS-5. The MPC-1 antibody recognized a 48-Kd surface antigen on KMS-5 but not on U-266, a nonadherent human myeloma cell line. Specificity characterization showed that MPC-1 antigen was expressed on mature myeloma cells, normal plasma cells, and mature B cells, whereas pre-B cells and germinal center B cells lacked its expression. Monocytes and a human bone marrow stromal cell line, KM102, also expressed this antigen. Two subclones of MPC-1+ VLA-5+ (KMS-5Ad) and MPC-1-VLA-5+ (KMS- 5NAd) were separated from the KMS-5 cell line. The KMS-5NAd adhered to KM102 more tightly than did the KMS-5NAd, and the U-266 (MPC-1-VLA-5-) displayed almost no adherence to the KM102. The adhesion of the KMS-5Ad was partially inhibited by the MPC-1 antibody. These results, taken together, suggest that the MPC-1 antigen serves as a differentiation marker for B-lineage cells, including plasma cells, and may function as an adhesion molecule involved in the interaction of mature myeloma cells with bone marrow stromal cells.

IL-6 and IGF-1 Act in Synergy with HGF in Myeloma Cells by Modulating the Ras-MAPK Pathway.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4793-4793
Author(s):  
Håkon Hov ◽  
Erming Tian ◽  
Anders Waage ◽  
Magne Børset ◽  
Anders Sundan
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Growth Factors ◽  
Cell Line ◽  
Plasma Cells ◽  
Kinase Inhibitor ◽  
Mapk Pathway ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells

Abstract Multiple myeloma is an incurable malignancy of plasma cells homing to the bone marrow. Myeloma cells are dependent on factors in their microenvironment for survival and expansion. HGF may be produced both by myeloma cells and the bone marrow microenvironment and serum levels of HGF are known to be a prognostic factor in multiple myeloma. Both IL-6 and IGF-1 are known growth factors for myeloma cells. In the human myeloma cell line (HMCL) INA-6, HGF alone had low effect, but together with IL-6 and IGF-1 it became a potent growth factor increasing thymidin incorporation two- to three-fold above the levels obtained with IL-6 or IGF-1 alone. Similar results were obtained for the myeloma cell line OH-2. The ANBL-6 cell line harbours an autocrine growth promoting HGF-loop. When inhibiting this autocrine HGF loop with a specific c-Met receptor tyrosine kinase inhibitor (PHA-665752), IL-6- and IGF-1-induced proliferation was reduced by 80% and 50% respectively. Thus, in the prescence of HGF, both IL-6 and IGF-1 are dependent on the HGF-receptor c-Met for full effect on cell proliferation. There seems to be two interconnected explanations for the synergy between HGF- and either IL-6- or IGF-1-signalling in myeloma cells. IL-6 and IGF-1 treatment increased the expression of c-Met in INA-6 cells Secondly, we found that HGF was unique among the three growth factors in activating both Ras and p44/42 MAPK in INA-6 cells. Also in ANBL-6 cells, IGF and IL-6 was dependent on functional c-Met signalling to fully activate this pathway. Thus, the reason for synergy between HGF and IL-6 or IGF-1 seemed to be that full activation of the Ras-Mapk pathway through Gab1 and SHP-2 by these cytokines was dependent on operating c-Met signalling. Taken together, HGF and c-Met signalling would be attractive targets for therapy of multiple myeloma.

Enforced CD19 Expression Leads to Growth Inhibition and Reduced Tumorigenicity

Blood ◽  
1999 ◽  
Vol 94 (10) ◽  
pp. 3551-3558 ◽  
Author(s):  
Maged S. Mahmoud ◽  
Ryuichi Fujii ◽  
Hideaki Ishikawa ◽  
Michio M. Kawano
Keyword(s):  
Growth Inhibition ◽  
Cell Line ◽  
Myeloma Cell ◽  
Inhibitory Effect ◽  
Myeloma Cell Line ◽  
Growth Inhibitory Effect ◽  
Myeloma Cells ◽  
Growth Inhibitory ◽  
Human Myeloma Cell Line

In multiple myeloma (MM), the cell surface protein, CD19, is specifically lost while it continues to be expressed on normal plasma cells. To examine the biological significance of loss of CD19 in human myeloma, we have generated CD19 transfectants of a tumorigenic human myeloma cell line (KMS-5). The CD19 transfectants showed slower growth rate in vitro than that of control transfectants. They also showed a lower capability for colony formation as evaluated by anchorage-independent growth in soft agar assay. The CD19 transfectants also had reduced tumorigenicity in vivo when subcutaneously implanted into severe combined immunodeficiency (SCID)-human interleukin-6 (hIL-6) transgenic mice. The growth-inhibitory effect was CD19-specific and probably due to CD19 signaling because this effect was not observed in cells transfected with a truncated form of CD19 that lacks the cytoplasmic signaling domain. The in vitro growth-inhibitory effect was confirmed in a nontumorigenic human myeloma cell line (U-266). However, introduction of the CD19 gene into a human erythroleukemia cell line (K-562) also induced growth inhibition, suggesting that this effect is CD19-specific, but not restricted to myeloma cells. These data suggest that the specific and generalized loss of CD19 in human myeloma cells could be an important factor contributing to the proliferation of the malignant plasma cell clones in this disease.

scholarly journals Plasma cells induce apoptosis of pre-B cells by interacting with bone marrow stromal cells

Blood ◽  
1996 ◽  
Vol 87 (8) ◽  
pp. 3375-3383 ◽  
Author(s):  
T Tsujimoto ◽  
IA Lisukov ◽  
N Huang ◽  
MS Mahmoud ◽  
MM Kawano
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Cell Survival ◽  
Cell Lines ◽  
Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
B Cell Survival

By using two-color phenotypic analysis with fluorescein isothiocyanate- anti-CD38 and phycoerythrin-anti-CD19 antibodies, we found that pre-B cells (CD38+CD19+) signifcantly decreased depending on the number of plasma cells (CD38++CD19+) in the bone marrow (BM) in the cases with BM plasmacytosis, such as myelomas and even polyclonal gammopathy. To clarify how plasma cells suppress survival of pre-B cells, we examined the effect of plasma cells on the survival of pre-B cells with or without BM-derived stromal cells in vitro. Pre-B cells alone rapidly entered apoptosis, but interleukin-7 (IL-7), a BM stromal cell line (KM- 102), or culture supernatants of KM-102 cells could support pre-B cell survival. On the other hand, inhibitory factors such as transforming growth factor-beta1 (TGF-beta1) and macrophage inflammatory protein- 1beta (MIP-1beta) could suppress survival of pre-B cells even in the presence of IL-7. Plasma cells alone could not suppress survival of pre- B cells in the presence of IL-7, but coculture of plasma cells with KM- 102 cells or primary BM stromal cells induced apoptosis of pre-B cells. Supernatants of coculture with KM-102 and myeloma cell lines (KMS-5) also could suppress survival of pre-B cells. Furthermore, we examined the expression of IL-7, TGF-beta1, and MIP-1beta mRNA in KM-102 cells and primary stromal cells cocultured with myeloma cell lines (KMS-5). In these cells, IL-7 mRNA was downregulated, but the expression of TGF- beta1 and MIP-1beta mRNA was augmented. Therefore, these results suggest that BM-derived stromal cells attached to plasma (myeloma) cells were modulated to secrete lesser levels of supporting factor (IL- 7) and higher levels of inhibitory factors (TGF-beta1 and MIP-1beta) for pre-B cell survival, which could explain why the increased number of plasma (myeloma) cells induced suppression of pre-B cells in the BM. This phenomenon may represent a feedback loop between pre-B cells and plasma cells via BM stromal cells in the BM.

Multiple Myeloma: Interleukin-15 Antagonizes Bone Morphogenetic Protein-Induced Apoptosis and Growth Inhibition.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3444-3444
Author(s):  
Magne Rekvig ◽  
Anne-Tove Brenne ◽  
Torstein Baade Ro ◽  
Anders Waage ◽  
Magne Borset ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Growth Inhibition ◽  
Cell Line ◽  
Caspase 3 ◽  
Myeloma Cell ◽  
Stimulus Figure ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Induced Apoptosis

Abstract Multiple myeloma has two distinct features: Expansion of malignant plasma cells within the bone marrow accompanied by skeletal destruction. Bone morphogenetic proteins (BMPs) have been shown to induce apoptosis and inhibit growth in myeloma cells. BMPs are members of the TGF-β superfamily of proteins capable of inducing bone formation, and regulate proliferation, differentiation and apoptosis. We have investigated myeloma cell apoptosis and proliferation with BMP-4 and −6 in concert with the myeloma cell growth factors interleukin (IL)-2, IL-6, IL-10, IL-15, IL-21, tumor necrosis factor (TNF)-α and insulin-like growth factor (IGF)-1. Eight samples of highly purified myeloma cells from patients and a human myeloma cell line, IH-1 (Brenne AT et al. Blood. 2002 May 15;99(10):3756–62.), were used in this study. Cytokine concentrations used in the referred experiments were for BMP-4 20ng/ml, BMP-6 250ng/ml, IL-15 20ng/ml and IL-6 0,1ng/ml, respectively. Growth inhibition was measured in a proliferation assay by methyl-[3H]-thymidine incorporation and apoptosis by annexin V- FITC-binding/PI-uptake on flow cytometry. IL-15 antagonized growth inhibition (Figure A) and prevented apoptosis induced by BMP-4 (Figure B) and BMP-6 in the myeloma cell line IH-1. IL-15 also antagonized the growth inhibition induced by BMP-4 and/or BMP-6 in three out of eight patient samples. Neither IL-6, nor any of the other investigated cytokines were able to rescue the myeloma cells from growth inhibition and apoptosis induced by BMP-4 and -6. Among the investigated cytokines, we found that IL-15 has a unique capability to antagonize BMP- induced apoptosis and growth inhibition in myeloma cells. We examined cleavage of the proapoptotic protein caspase-3 and found that BMP-4 activated caspase-3 in the IH-1 cell line. This activation of caspase-3 was blocked by IL-15 but not by IL-6. We have demonstrated a possible mechanism for myeloma cells to escape apoptosis and growth-inhibition within the bone marrow. Intramedullar levels of IL-15 and BMPs may play a role in the pathogenesis of multiple myeloma. Figure A. Proliferation in response to BMP-4 stimulus Figure A. Proliferation in response to BMP-4 stimulus Figure B. Apoptosis in response to BMP-4 stimulus Figure B. Apoptosis in response to BMP-4 stimulus

Gene profiling of a myeloma cell line reveals similarities and unique signatures among IL-6 response, N-ras-activating mutations, and coculture with bone marrow stromal cells

Blood ◽  
2003 ◽  
Vol 102 (7) ◽  
pp. 2581-2592 ◽  
Author(s):  
Paula A. Croonquist ◽  
Michael A. Linden ◽  
Fangyi Zhao ◽  
Brian G. Van Ness
Keyword(s):  
Bone Marrow ◽  
Cell Line ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Expression Patterns ◽  
Myeloma Cell ◽  
Marrow Stromal Cells ◽  
Gene Profiling ◽  
Myeloma Cell Line ◽  
Ras Gene

Abstract ANBL-6, a myeloma cell line, proliferates in response to interleukin 6 (IL-6) stimulation, coculture with bone marrow stromal cells, and when harboring a constitutively active mutant N-ras gene. Eighteen samples, including 4 IL-6-treated, 3 mutant N-ras-transfected, 3 normal stroma-stimulated, 2 multiple myeloma (MM) stroma-stimulated, and 6 untreated controls were profiled using microarrays interrogating 12 626 genes. Global hierarchical clustering analysis distinguished at least 6 unique expression signatures. Notably, the different stimuli altered distinct functional gene programs. Class comparison analysis (P = .001) revealed 138 genes (54% involved in cell cycle) that distinguished IL-6-stimulated versus nontreated samples. Eighty-seven genes distinguished stroma-stimulated versus IL-6-treated samples (22% encoded for extracellular matrix [ECM] proteins). A total of 130 genes distinguished N-ras transfectants versus IL-6-treated samples (26% involved in metabolism). A total of 157 genes, 20% of these involved in signaling, distinguished N-ras from stroma-interacting samples. All 3 stimuli shared 347 genes, mostly of metabolic function. Genes that distinguished MM1 from MM4 clinical groups were induced at least by one treatment. Notably, only 3 genes (ETV5, DUSP6, and KIAA0735) are uniquely induced in mutant ras-containing cells. We have demonstrated gene expression patterns in myeloma cells that distinguish an intrinsic genetic transformation event and patterns derived from both soluble factors and cell contacts in the bone marrow microenvironment. (Blood. 2003;102:2581-2592)

Carcinoembryonic-Antigen-Related Cell Adhesion Molecule (CEACAM) Expression on Multiple Myelomas Inhibits Their Recognition by Autologous Memory CD8 T Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 738-738
Author(s):  
Mathias Witzens-Harig ◽  
Dirk Hose ◽  
Michael Hundemer ◽  
Simone Jünger ◽  
Anthony D. Ho ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Cell Responses ◽  
Healthy Donors

Abstract Introduction. Previous studies clearly demonstrated the spontaneous induction and accumulation of functionally competent myeloma antigen–specific memory CD8 T cell responses in the bone marrow of a large proportion of myeloma patients. However, other studies convincingly demonstrated that CD8 T cells from myeloma patients were incabable of lysing autologous myeloma cells. An explanation for this apparent discrepancy is still lacking. CEACAMs are induced on T cells during TCR-specific activation and mediate a rapid blocking of T cell effector functions upon homophilic binding with CEACAMs expressed on target cells. We here addressed the question if myeloma cells might escape recognition by autologous, myeloma-specific CD8 T cells through CEACAM expression. Methods. Presence of myeloma-specific CD8 T cells was analyzed by IFN-y Elispot assays using separated, bone marrow derived CD8 T cells and myeloma-associated antigen-pulsed autologous DCs or autolgous myeloma cells as antigen presenting cells. Expression of CEACAMs 1–21 was analyzed by differential gene expression profiling of sorted CD138+ myeloma cells from bone marrow of 140 myeloma patients and of respective plasma cells from 14 healthy donors. In addition, the expression of CEACAMs 1, 5, 6 and 8 was analyzed by flowcytometry on the myeloma cell line MM8226 and on CD138+ myeloma cells from altogether 7 myeloma patients. A role of CEACAM on T cell recognition of autologous myeloma cells was analyzed by coculture of CD8 T cells and sorted, autologous myeloma cells in the presence or absence of blocking antibodies against CEACAMs by IFN-γ Elispot assay. Results. We identified the presence of myeloma-specifcic CD8 T cells in app. 40% of tested myeloma patients (N=20). However, in none of the tested cases T cells were able to directly recognize autologous myeloma cells. Over expression of CEACAM mRNA was found for CEACAM1, 6 and 8 in myeloma cells of up to 20% of patients, but not in plasma cells of healthy donors. Flowcytometric analysis revealed the protein expression of CEACAMs 1 and 6 on 25–50 % of the myeloma cells of all 7 tested patients and on the myeloma cell line MM8226. Blocking of CEACAMs on sorted myeloma cells before their coculture with autologous CD8 T cells resulted in significant T cell responses to myeloma cells in all tested patients (N=6), while in none of these cases the T cells were able to respond to unblocked myeloma cells. Conclusions. We here demonstrate for the first time the expression of CEACAMs 1 and 6 on freshly isolated myeloma cells. Blocking of these CEACAMs resulted in a spontaneous CD8 T cell response against cocultured autologous myeloma cells which was undetectable in case of unblocked CEACAM expression despite the presence of myeloma-reactive memory T cells. We suggest that CEACAMs on myeloma cells inhibit the re-activation of tumour antigen specific CD8 T cells upon interaction with myeloma cells and may contribute to the well characterized immune resistance of multiple myeloma.

BLyS Regulates Human Myeloma Cell IL-6 Expression.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1412-1412
Author(s):  
Sook Kyung Chang ◽  
Diane F. Jelinek
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Cell Line ◽  
Tumor Cells ◽  
Myeloma Cell ◽  
Conditioned Media ◽  
Transcriptional Level ◽  
Myeloma Cells ◽  
Growth And Survival ◽  
Cells Cultured

Abstract Multiple myeloma (MM) is a malignant disease of plasma cells that accumulate in the bone marrow. The survival mechanisms of myeloma cells are still not fully understood. There are several cytokines that are known to support the survival and growth of myeloma cells, such as IL-6, IGF-I, TNF-a, and IL-1β. Recent evidence has also shown that the TNF family member, B lymphocyte stimulator (BLyS), is critical for normal B cell development and homeostasis, as well as for survival of malignant B cells, including MM. However, it should be noted that the precise mechanisms by which BLyS promotes the survival of MM cells remain uncertain. Our group has already reported that MM cell lines and primary myeloma cells can express BLyS and its receptors. Thus, exogenous BLyS augmented the survival of MM cells, including the IL-6 dependent MM cell line, KAS-6/1. To better study the mechanism underlying BLyS-mediated survival of myeloma cells, we sought to establish a KAS-6/1 cell line variant which was solely dependent on exogenous BLyS for growth and survival. We have previously reported that KAS-6/1 cells do not appear to express autocrine IL-6, and therefore are highly dependent on exogenous IL-6 for survival. To establish a BLyS-dependent KAS-6/1 variant, cells were cultured long-term in RPMI containing 0.5% BSA or 1% FCS in the absence of exogenous IL-6, and in the presence or absence of exogenous BLyS. KAS-6/1 cells cultured in the absence of cytokines were non-viable after two weeks of culture, whereas KAS-6/1 cells cultured with BLyS alone remained viable and proliferated, albeit at a lower rate than parental KAS-6/1 cells maintained with IL-6. We next focused on identifying the mechanisms of BLyS-dependent survival of the KAS-6/1/BLyS (KAS-6/B) variant cell line. Of interest, KAS-6/B cells placed in short term culture exhibited cytokine independent survival and proliferation and this was completely inhibited using a neutralizing IL-6 antibody. These results therefore suggested that the mechanism underlying BLyS-mediated survival and proliferation of the KAS-6/B cells is induction of an autocrine IL-6 pathway. This notion is further supported by the ability of BLyS to induce IL-6 expression at the transcriptional level, and at the protein level as revealed by an IL-6 specific ELISA. Consistent with the concept that BLyS induced autocrine IL-6 expression in the KAS-6/B cells, we also observed that STAT-3 was constitutively activated in these cells. In addition, we tested the ability of BLyS-conditioned media obtained from KAS-6/B cells to stimulate the proliferation of a different IL-6 dependent MM cell line, JMW. Whereas BLyS alone failed to stimulate JMW cell proliferation, BLyS-conditioned media obtained from the KAS-6/B cells stimulated proliferation, and in a manner that was inhibited by a neutralizing IL-6 antibody. Finally, BLyS stimulation of primary patient MM cells similarly resulted in increased IL-6 expression. IL-6 is known to play a major role in the malignant progression of MM by regulating the growth and survival of tumor cells. IL-6 levels often increase during disease progression, and it has been demonstrated that IL-6 may derive from the tumor cells themselves or from other cell types in the bone marrow. Because of this, it is important to fully understand what signals are capable of inducing IL-6 expression in this disease. In summary, our results demonstrate for the first time the ability of BLyS to maintain myeloma cell survival via the induction of an autocrine IL-6 pathway. Our results underscore the importance of IL-6 in this disease, and further demonstrate the complexity of the deregulated cytokine network in this disease.

Establishment and Characterization of a New Human Myeloma Cell Line WuS1.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5114-5114
Author(s):  
Wanling Sun ◽  
Yongji Wu ◽  
Hui Li ◽  
Xuan Wang ◽  
Nong Zou ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Growth Factors ◽  
Cell Line ◽  
Single Cells ◽  
Myeloma Cell ◽  
Nuclear Antigen ◽  
Myeloma Cell Line ◽  
Human Myeloma Cell Line ◽  
Human Myeloma Cell

Abstract A new human myeloma cell line WuS1 was established from the bone marrow of a 45-year-old Chinese male patient with IgGλ type multiple myeloma (stage IIIB). The growth of WuS1 cells is constitutively independent of exogenous growth factors of feeder cells. The WuS1 cell line proliferated consistently as free-floating single cells in suspension, sometimes in small clusters or slightly adherent on the bottom of the plastic culture flask, without forming clumps. The cell line has been maintained without any external growth factors for over a year, and cells frozen in liquid nitrogen can be revived successfully. The doubling time of the cells was about 11 hours and the colony-forming rate was 55.56±6.33%. WuS1 displayed immature plasma cell features with an obvious heterogeneity in size and a high nuclear-cytoplasmic ratio in Wright-Giemsa staining. Figure Figure They were positive for ALP, CE, ACP(not inhibited by tartrate) and PAS stainings and negative for POX and NBE. By transmission electron microscopy, the cytoplasm of WuS1 contained abundant mitochondria, and parallel endoplasmic reticulum or Golgi apparatus in some cells. The monoclonal immunoglobulin G and λ light chain were positive in cell lysate and not in cell culture supernatants by immnuoelectrophoresis. The cell surface antigens were positive for CD3, CD59, CD106 and CD138, and negative for CD4, CD5, CD8, CD10, CD13, CD14, CD19, CD20, CD22, CD29, CD31, CD33, CD34, CD38, CD44, CD49d, CD45, CD54, CD56 and HLA-DR by flow cytometry. Chromosomal analysis revealed a hypodiploidy and complex karyotype. WuS1 cells were negative for Epstein-Barr virus by PCR using EBV nuclear antigen-1 specific primers. Twelve SCID mice were injected with WuS1 cells intravenously or subcutaneously, and obvious tumor infiltration in bone marrow, liver, spleen, lung, kidney and injection site (subcutaneously group) were observed by pathologic examination. Figure Figure The novel WuS1 cell line will be useful in the study of the biology, etiology and treatment of multiple myeloma.

BP1 Expression in a Myeloma Cell Line: Modulation by Arsenic Trioxide and Ascorbic Acid.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5113-5113
Author(s):  
Khanh Do ◽  
Patricia Berg ◽  
Milcho Mincheff ◽  
Joao L. Ascensao
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Ascorbic Acid ◽  
B Cell ◽  
Cell Line ◽  
Arsenic Trioxide ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Cell Lineages

Abstract BP1 belongs to a family of homeobox genes which are not expressed in normal bone marrow cells but are seen in multiple leukemic cell lineages including erythroid, myeloid, and T- and B-cell lineages, and whose modulation is important in differentiation and oncogenic transformation (Shen et al. EMBO J 1992). Multiple Myeloma (MM) is a clonal B-cell neoplasm marked by proliferation of plasma cells within bone marrow. BP1 expression has not previously been demonstrated in MM cell lineages. Recent unpublished data demonstrated elevated BP1 expression in the bone marrow of Acute Promyelocytic Leukemia (APL) patients and its down regulation with the differentiating agent ATRA. With recent data demonstrating arsenic trioxide (As2O3) induced remission in chemorefractory APL and MM (Grad et al. Blood 2001), we hypothesized BP1 may share a role in this response conversion. In the current study, we treated ARH77 cells - a multiple myeloma cell line, with 0.25 – 3 uM As2O3 and adjuvant ascorbic acid (AA) for 24–96hrs; AA had been previously documented to potentiate As2O3-mediated cell death in multiple myeloma cells (Grad et al. Blood 2001). Viability, cell numbers and mRNA expression were determined at each time point. The % of viable cells at 72 and 96 hrs decreased to less than 10% of the control in cultures flasks containing 2 and 3uM; 0.25 uM had minimal effect in this assay. We then used 0.5 and 1uM, which caused growth arrest compared to controls at each interval. 1 uM caused a decrease of 12% viability at 72 hrs. A significant reduction in the expression of BP1 was seen in these cultures when balanced with beta-actin levels. Myeloma cells are marked by a myriad of antiapoptotic signaling mechanisms, which account for their acquired resistance to current chemotherapy. A better understanding of signaling pathways and genetic events may provide potential therapeutic targets and aid in correlation between genetic abnormalities and clinical outcomes.

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