Biomedical subjects

K Iwato

Publications and source records attributed to K Iwato.

At least 19 recordsLinked to original sources

PubMed1995

A specific chromosome abnormality of t(4;12)(q11-12;p13) in CD7+ acute leukaemia.

Three cases of acute leukaemia with t(4;12) (q11-12;p13) karyotypic abnormalities were analysed. They had the following common clinical and biological characteristics: (1) dysplasia of three haemopoietic lineages: (2) absent or low myeloperoxidase activity: and (3) retention of platelets in the peripheral blood and megakaryocytes in the bone marrow. There were increased numbers of basophils in the bone marrow and peripheral blood in two of the cases. In all, the blast cells displayed the unique immunophenotype CD7+CD13+CD34+HLA-DR+. The blasts analysed in one case expressed c-kit on the membrane surface. These findings suggest that the t(4;12) (q11-12;p13) abnormality is associated with a particular type of acute leukaemia, one in which the morphology and immunophenotype suggest that the translocation may have occurred at an early stage of haemopoiesis.

Acute Disease↗

PubMed1993

Identification of immature and mature myeloma cells in the bone marrow of human myelomas.

With regard to the expression of adhesion molecules, human myeloma cells freshly isolated from bone marrow were heterogeneous. By two-color analysis with anti-VLA-5 antibody (PE staining) and FITC-labeled anti-CD38 antibody, we found all myeloma cells located at CD38-strong positive (CD38++) fraction and identified two subpopulations among these myeloma cells: CD38++ VLA-5-(VLA-5-) myeloma cells and CD38++ VLA-5+ (VLA-5+) myeloma cells. To clarify the biologic character of these two subpopulations, the morphology, in vitro proliferative activity and in vitro M-protein secretion were examined in each fraction isolated by the purification procedure or a cell sorter. Morphologic examination showed that VLA-5- myeloma cells were mostly immature or plasmablastic and VLA-5+ cells were mature myeloma cells. Furthermore, VLA-5- myeloma cells proliferated markedly in vitro and responded to interleukin 6 (IL-6), a growth factor for myeloma cells, while VLA-5+ myeloma cells showed very low uptakes of 3H-thymidine and no responses to IL-6 but secreted higher amounts of M-protein (immunoglobulin) in vitro significantly. Therefore, we could clarify here heterogeneity of human myeloma cells in the bone marrow with regard to the expression of VLA-5, one of integrin adhesion molecules; VLA-5- myeloma cells were proliferative immature cells and VLA-5+ cells were mature myeloma cells.

ADP-ribosyl Cyclase↗

PubMed1993

Expressions of DNA topoisomerase I and II gene and the genes possibly related to drug resistance in human myeloma cells.

In order to clarify the mechanism of drug resistance in human myeloma cells, we investigated the expressions of DNA topoisomerase I and topoisomerase II gene and the genes possibly related to drug resistance; multi-drug resistant gene 1 (MDR-1), glutathione S-transferase class pi gene (GST-pi), by Northern blotting. Myeloma cells in eight of 15 cases prior to chemotherapy expressed topoisomerase I mRNA considerably, while the expression of topoisomerase II mRNA was detected weakly in only one of 16 myeloma patients. There was not any correlation between expression of topoisomerase I mRNA and clinical drug resistance. Significant expression of MDR-1 mRNA and P-glycoprotein was not detected in 25 cases of multiple myeloma prior to chemotherapy and even after several courses of VAD (vincristine, adriamycin and dexamethasone) therapy by Northern blotting and immunostaining using monoclonal anti-P-glycoprotein antibody (MRK-16), respectively. On the other hand, 16 of 21 myeloma cases showed significant expression of GST-pi protein and GST-pi mRNA with the various strengths, but there was no apparent correlation between GST-pi mRNA expression and clinical response. Therefore these data suggest that expression of the genes we tested may not determine the level of drug resistance in multiple myeloma, but lower or no significant expression of topoisomerase II mRNA in most myeloma cells indicates the possibility that topoisomerase II inhibitors such as VP-16 and topoisomerase II-mediated cytotoxic drugs such as adriamycin, are not so effective for the treatment of multiple myeloma.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1993

[Myeloma precursor cells].

Heterogenous biological character of myeloma cells was associated with different expression of adhesion molecules. Myeloma cells could be phenotypically divided into two subpopulations: CD38++/VLA5+/MPC-1+(VLA-5+) cells and CD38++/VLA5-/MPC-1-(VLA-5-) cells. VLA-5- myeloma cells were morphologically immature and proliferated markedly with response to IL-6 in vitro, while VLA-5+ cells showed very low uptakes of 3H-TdR but secreted higher amounts of M-protein in vitro. These results suggest VLA-5- cells are proliferative precursor in myeloma. With respect to VLA-5 and MPC-1 expression, myeloma precursor cells (CD38++/VLA-5-/MPC-1-/CD10-/CD24-) showed similar phenotype to germinal center B cells (CD38+/VLA-5-/MPC-1-/CD10+/CD24-), rather than that of pre-B cells in the bone marrow (CD38+/VLA-5+/MPC-1-/CD10+/CD24+). Identification of precursor cells and characterization of their growth is important for the understanding of pathophysiology of myeloma and the therapeutic strategy.

ADP-ribosyl Cyclase↗

PubMed1992

Long-term engraftment of fresh human myeloma cells in SCID mice.

Using highly purified myeloma cells from patient bone marrow, we established human-murine myeloma chimeras in severe combined immunodeficiency (SCID) mice and documented secretion of monoclonal human immunoglobulins (Hulgs) in the mice for up to 299 days. Monoclonality of circulating Hulgs was found only when highly purified myeloma cells were injected intraperitoneally. In contrast, injection of unfractionated myeloma marrow led to the development of polyclonal Hulgs in the SCID mice. The criteria for myeloma engraftment included prolonged presence of monoclonal Hulgs in the sera of SCID mice and/or detection of human myeloma cells in their tissues by immunohistochemical examination. Ninety-one percent (10/11) of the fresh purified myeloma specimens engrafted in the SCID mice. Fifty-five percent (6/11) of the patient samples resulted in human B-cell grafts, and 45% (5/11) were identifiable as human myeloma chimeras. Pathologic studies showed that most human plasmacytes were located in the peritoneal cavity but metastatic infiltrates were also found in other organs in 69% of the SCID-human myeloma chimeras. This chimeric model should provide a useful tool for characterization of growth modulation and microenvironmental interactions as well as a means of testing new therapeutic approaches to multiple myeloma.

Adult↗

PubMed1992

[Study on the therapy of multiple myelomas--initial induction therapy (MP, IFN alpha, steroid pulse) and maintenance therapy (VMP, MP continuous, VEP, MCNU)].

Untreated twenty patients of multiple myeloma were treated with the chemotherapy protocol as follows: Initial induction therapy;MP continuous or MP intermittent----IFN alpha----steroid pulse. Maintenance therapy;alkylating agents which have no cross resistance were used ((V) MP----(MP)----(V) EP----MCNU). Remission rate (CR+PR) after the initial MP therapy was 45%, and that after including IFN alpha and steroid pulse therapy was 50%, Fifty percent survival rate was almost as same as those reported previously (34M). Our protocol presented here was based on the idea that, initially, myeloma cells with proliferative activity could be affected by MP therapy, and subsequent IFN alpha therapy would have effect even on the residual myeloma cells. Serial checks of 3H-TdR uptake of myeloma cells during the therapy supported this idea. During the maintenance therapy, clinical responses to the initial induction therapy were not aggravated in the responded cases when evaluated by the variation of serum M-protein level. We propose that considering from a point of proliferative activity of myeloma cells is important for designing therapeutic protocols for multiple myeloma.

Adult↗

PubMed1991

Increased expression of the c-myc gene may be related to the aggressive transformation of human myeloma cells.

Alteration and abnormal expression of the c-myc oncogene were investigated in human multiple myeloma. Human myeloma cells were highly purified (more than 95%) from bone marrow aspirates in 14 cases of advanced multiple myelomas and one case of plasma cell leukaemia. Southern blotting revealed that a rearranged configuration of c-myc gene was found in only one case of them, but this was a novel truncation of the gene in its coding exon II; a rearranged 3.4 kb band was detected by digestion with Xba I using c-myc exon II probe, but no rearranged band was found using exon III probe. In this case, the truncated c-myc allele was not transcribed; normal sized (2.4 kb) c-myc mRNA was markedly expressed, but no aberrant mRNA was detected. On the other hand, by Northern blotting, the nine cases, including the case with the rearranged c-myc gene, showed increased expression of normal sized (2.4 kb) c-myc mRNA. Elevated c-myc mRNA expressions were well related to the in vitro proliferation (3H-TdR uptake), but not to IL-6 response. Interestingly, extremely high expressions of c-myc mRNA were detected in two cases of aggressive myelomas, including the case with the rearranged c-myc gene, and in one of plasma cell leukaemia. These two cases of aggressive myelomas were the ones who showed the markedly high 3H-TdR uptakes, and had the common clinical features with the formation of an extramedullary mass and very short survival. These results suggest that the activation of c-myc gene could induce high proliferative activities and the subsequent aggressive transformation of myeloma cells.

Blotting, Northern↗

PubMed1990

Effect of glucocorticoids on the biologic activities of myeloma cells: inhibition of interleukin-1 beta osteoclast activating factor-induced bone resorption.

Regulatory effects of glucocorticoids (dexamethasone) on myeloma cells as well as bone resorption in multiple myeloma were investigated. Glucocorticoids significantly inhibited proliferation of myeloma cells, and decreased the messenger RNA (mRNA) expressions of interleukin-6 (IL-6) and secretory type immunoglobulin G (IgG). The inhibitory effects of glucocorticoids on myeloma cell proliferation could be due to the decreased expression of IL-6 mRNA, decreased IL-6 production, and thus suppression of autocrine growth by IL-6, which is an autocrine growth factor for myeloma cells as reported previously (Nature 332:83, 1988). Glucocorticoids also inhibited M-protein secretion by decreasing the levels of secretory type Ig mRNA. On the other hand, because IL-1 beta rather than lymphotoxin is considered to be a major osteoclast activating factor (OAF) produced by myeloma cells, and glucocorticoids decreased the expression of IL-1 beta mRNA and markedly suppressed the bone resorbing activity induced by IL-1 beta OAF in 45Ca-release bone resorption assay, it is suggestive that glucocorticoids could inhibit bone resorption induced by IL-1 beta OAF in multiple myeloma. Therefore, from these data it is concluded that glucocorticoids could be more effective chemotherapeutic agents in multiple myeloma than we expected, especially with regards to the inhibitory effects on proliferation and M-protein secretion from myeloma cells, as well as bone resorption by myeloma cells.

Blotting, Northern↗

PubMed1989

Production of interleukin 1 beta, a potent bone resorbing cytokine, by cultured human myeloma cells.

Supernatants of freshly isolated human myeloma cell cultures were examined both for bone-resorbing activity (BRA) in vitro using newborn mouse calvaria, and for identification of the causal substances of the BRA. Eight of 14 culture supernatants of myeloma cells had BRA. All of these BRA-positive supernatants were from patients with marked destructive bone lesions of multiple myeloma. The presence of interleukin 1 (IL-1), especially IL-1 beta, was demonstrated in seven of these BRA-positive supernatants but not in BRA-negative supernatants. The concentrations of IL-1 beta were high enough to induce bone resorption in the newborn mouse calvaria assay and the BRA was totally abolished by pretreatment of the supernatants with anti-IL-1 beta antibody but not with either anti-IL-1 alpha antibody or normal serum. Other bone resorbing cytokines such as tumor necrosis factor or lymphotoxin were not present in high enough concentrations to stimulate bone resorption and their levels did not correlate with the BRA. IL-1 beta mRNA was also identified in BRA-positive myeloma cells. These results demonstrate that IL-1 beta is the principal agent of BRA present in supernatants of myeloma cell cultures, and also identify a possible role of IL-1 beta in destructive bone lesions in patients with multiple myeloma.

Adult↗

PubMed1989

Interleukin-1 beta rather than lymphotoxin as the major bone resorbing activity in human multiple myeloma.

Human myeloma cells were purified from bone marrow aspirates from four patients having advanced myeloma, including one with common acute lymphoblastic leukemia antigen-positive myeloma. All of these myelomas had marked bone lytic lesions. From the culture supernatants of these purified myeloma cells, bone-resorbing activities were significantly revealed by 45Ca-release bone resorption assay, and IL-1 activities were also detected by IL-1 bioassay (mouse thymocyte comitogenic assay). Sandwich enzyme immunoassay for IL-1 alpha or IL-1 beta revealed that IL-1 beta was responsible for IL-1 activity of these culture supernatants. Furthermore, the bone resorbing activities of these culture supernatants were completely neutralized by pretreatment of anti-IL-1 beta, but not anti-IL-1 alpha antibody. By Northern blot analysis, IL-1 beta mRNA was identified from these myeloma cells. Therefore, it is concluded that myeloma cells produce IL-1 beta, which acts as bone-resorbing activity in multiple myeloma.

Antigen-Antibody Reactions↗

PubMed1989

Altered cytokine activities are related to the suppression of synthesis of normal immunoglobulin in multiple myeloma.

The content of peripheral blood B cells (B1+) was reduced in patients of multiple myeloma (MM) and not in those with benign monoclonal gammopathy (BMG) compared to normal donors (P less than 0.01). This observation correlated with the suppression of synthesis of normal immunoglobulin (Ig) in MM. Thus, cytokine activities regulating the proliferation of normal mature B cells, such as B cell stimulatory factor 1 (BSF-1)/interleukin 4 (IL-4), B cell growth inhibitory factor (BIF) and IL-2 in peripheral blood T cells, and IL-1 in peripheral blood adherent cells, were investigated in patients with BMG (n = 7) and MM (n = 28). All patients of MM having a marked suppression of synthesis of all other normal Ig, had significantly lower levels of BSF-1 activity and inversely higher levels of BIF activity than those of normal donors. However, patients with BMG having no suppression of synthesis of normal Ig had BSF-1 and BIF activities similar to normal donors. There was no significant difference in IL-1 and IL-2 activities between both normal donors and BMG versus MM patients. These data show that in MM altered cytokine activities correlate with suppression of synthesis of normal Ig.

B-Lymphocytes↗

PubMed1989

BSF-2/IL-6 does not augment Ig secretion but stimulates proliferation in myeloma cells.

Human myeloma cells were highly purified from bone marrow aspirates of 21 patients with advanced immunoglobulin G (IgG)-type multiple myeloma. B-cell stimulatory factor 2 (BSF-2)/interleukin-6 (IL-6) was originally characterized as a cytokine that can enhance immunoglobulin secretion from activated normal B cells and increase the expression of secretory-type Ig mRNA in these B cells, but that does not augment proliferation of activated B cells. However, recombinant IL-6 (rIL-6) could not enhance M-protein (IgG) secretion in freshly isolated myeloma cells in vitro but could augment proliferation of myeloma cells, although myeloma cells constitutively expressed IL-6 receptors. Furthermore, expression of secretory-type IgG (gamma-chain) mRNA in myeloma cells was not changed in the presence of IL-6. These results show that IL-6 is not an enhancing factor in Ig secretion from myeloma cells, and thus signal transduction through IL-6 in myeloma cells may be altered as opposed to activated B cells.

Adjuvants, Immunologic↗

PubMed1989

Heterogenous response of B cell chronic lymphocytic leukaemia (B-CLL) cells to anti-human IgM antibody (anti-mu) and B cell stimulatory factor 1 (BSF-1).

B cell chronic lymphocytic leukaemia (B-CLL) cells from seven patients were examined for 3H-TdR uptake with anti-human IgM antibody (anti-mu) and B cell stimulatory factor 1 (BSF-1)/IL-4. B-CLL cells from one patient could proliferate with anti-mu plus BSF-1, but not with BSF-1 alone. B-CLL cells from two patients responded to BSF-1 alone; one could proliferate more by adding anti-mu, but the proliferation of the other cells was inhibited by adding anti-mu. On the other hand, B-CLL cells from the other four patients responded neither to anti-mu nor to anti-mu plus BSF-1. Surface densities of membrane IgM of B-CLL cells were also analysed by fluorescence activated cell sorter (FACS), but there was no correlation between proliferative response (DNA synthesis) of B-CLL cells to anti-mu and densities of surface membrane IgM. These results show that there is a functional heterogeneity of B-CLL cells with regard to proliferative response to anti-mu and BSF-1.

Aged↗

PubMed1989

[A case of refractory multiple myeloma demonstrating a relationship between the progression of the disease and in vitro myeloma cells activity].

In vitro proliferation (3H-TdR-uptake) and M-protein secretion rate by highly purified myeloma cells from bone marrow aspirates were examined serially to evaluate the progression of multiple myeloma in a patient who was refractory to conventional alkylating agents. Following the administration of IFN-alpha, serum M-protein decreased significantly, with the reduced in vitro spontaneous M-protein secretion rate from the separated myeloma cells. Similarly, when IFN-alpha as low as 10 u/ml was added in vitro, it also suppressed M-protein secretion from myeloma cells of this patient, suggesting that, the observed decrease of serum M-protein was due to diminished M-protein secretion by the myeloma cells themselves, as well as the reduction of the tumor cell burden. On the other hand the in vitro 3H-TdR uptake by the myeloma cells increased markedly with the decrease in the M-protein secretion rate. Five months after the initiation of IFN-alpha treatment, tumor formation at the lumbar vertebrae occurred when serum M-protein level was still low, followed by a bone marrow relapse. These results suggest that serial assessments of proliferation and M-protein secretion potential of myeloma cells in vitro can be helpful in predicting the progression of multiple myeloma.

Cell Division↗

PubMed1989

Interleukin-1 accelerates autocrine growth of myeloma cells through interleukin-6 in human myeloma.

Recombinant interleukin 1 alpha (rIL-1 alpha) augmented proliferation of freshly isolated myeloma cells as well as B-cell stimulatory factor 2 (BSF-2)/interleukin-6 (IL-6). Recombinant IL-1 alpha-induced proliferation was partially inhibited by anti-IL-6 antibody. In the culture supernatants of rIL-1 alpha-stimulated myeloma cells, IL-6 activities, which were measured by using an IL-6-dependent murine hybridoma clone, MH60.BSF2, were increased, when compared with those in the culture supernatants of nonstimulated myeloma cells. Furthermore, IL-6 messenger RNA (mRNA) expression was also augmented in IL-1 alpha-stimulated myeloma cells. Therefore rIL-1 alpha stimulates myeloma cells to produce IL-6, which consequently augments proliferation of myeloma cells. Thus, IL-1 can accelerate autocrine growth of myeloma cells through IL-6.

Adjuvants, Immunologic↗

PubMed1989

Sensitive inhibitory effect of interferon-alpha on M-protein secretion of human myeloma cells.

The effects of interferon-alpha (IFN alpha) on in vitro proliferation and M-protein secretion in human myeloma cells were investigated. Human myeloma cells were purified from bone marrow aspirates in 12 multiple myeloma patients. Purified myeloma cells were cultured for 48 hours with IFN alpha at its lower concentrations (0.1 to 100 U/mL). The cells were then pulsed with 3H-TdR for the last 12 hours and 3H-TdR uptake was measured (in vitro proliferation). After 48-hour culture, supernatants were harvested and the amount of M-protein in these fluids were measured by enzyme-linked immunosorbent assay (ELISA) (in vitro M-protein secretion). In vitro M-protein secretions of myeloma cells were significantly suppressed even at 0.1 U/mL of IFN alpha, while 3H-TdR uptakes were not so suppressed until 10 or 100 U/mL of IFN alpha were added. The expressions of secretory immunoglobulin (Ig) mRNA of these myeloma cells were also selectively suppressed by IFN alpha. Furthermore, after IFN alpha had been administered intramuscularly, 3 to 6 x 10(6) U/d for at least 1 month, in vitro M-protein secretions of these myeloma cells were decreased compared with those before IFN alpha administration. Therefore, these results suggest that IFN alpha has more sensitive inhibitory effect on M-protein secretion of human myeloma cells rather than on myeloma cell proliferation.

Blood Proteins↗

PubMed1988

Autocrine generation and requirement of BSF-2/IL-6 for human multiple myelomas.

Human B cell stimulatory factor 2 (BSF-2) was originally characterized and isolated as a T cell-derived factor that caused the terminal maturation of activated B cells to immunoglobulin-producing cells. Molecular cloning of the complementary DNA predicts that BSF-2 is a protein of relative molecular mass (Mr) 26,000 similar or identical to interferon beta 2, hybridoma plasmacytoma growth factor and hepatocyte stimulating factor. IL-6 has been proposed as a name for this molecule. It is now known that BSF-2 has a wide variety of biological functions and that its target cells are not restricted to normal B cells. Responses are also seen in T cells, plasmacytomas, hepatocytes, haematopoietic stem cells, fibroblasts and rat phoeochromocytoma, PC12 (Satoh, T. et al., manuscript in preparation). Of particular interest to this report is that human BSF-2 is a potent growth factor for murine plasmacytomas and hybridomas. This observation suggested to us that constitutive expression of BSF-2 or its receptor could be responsible for the generation of human myelomas. In this study we report that myeloma cells freshly isolated from patients produce BSF-2 and express its receptors. Moreover, anti-BSF-2 antibody inhibits the in vitro growth of myeloma cells. This is direct evidence that an autocrine loop is operating in oncogenesis of human myelomas.

Antibodies↗

PubMed1988

Decrease in BSF-2/IL-6 response in advanced cases of multiple myeloma.

Human myeloma cells freshly isolated from 40 patients with IgG multiple myeloma (MM, 10 in stage I and 30 in stage III), were cultured for 48 hours with recombinant B cell stimulatory factor 2 (rBSF-2)/interleukin-6 (IL-6), which is considered a major growth factor for myeloma cells. Uptake of 3H-thymidine by these purified myeloma cells was measured, and BSF-2 response was evaluated by stimulation index and delta cpm induced by rBSF-2. Myeloma cells from cases in stage I responded to rBSF-2 better than those in stage III. Moreover rBSF-2 responders also showed better response to chemotherapy. Therefore, these results suggest that in vitro response of myeloma cells to BSF-2 correlates with disease progression and clinical response in patients of MM.

Humans↗