Myeloma cells destroy bone by producing an osteoclast-stimulating factor that has chemical and biological characteristics similar to the bone-resorbing activity present in the supernatants of activated leukocyte cultures. Recently, a number of bone-resorbing leukocyte cytokines have been identified, including interleukin-1, lymphotoxin, and tumor necrosis factor. We have examined the products of human myeloma cells for the presence of these bone-resorbing cytokines. In a tumor cell line derived from a patient who had myeloma with osteolytic bone lesions and hypercalcemia, we found that the myeloma cells induced bone-resorbing activity and cytotoxic activity in vitro. Most of the bone-resorbing activity and all cytotoxic activity were suppressed by neutralizing antibodies to lymphotoxin. The myeloma cells expressed both lymphotoxin and tumor necrosis factor mRNA, but no tumor necrosis factor could be detected in the cell-culture medium. Interleukin-1 mRNA was not detected in the myeloma cells, and biologic activity of interleukin-1 was not measurable in the medium harvested from the cultured cells. The bone-resorbing activity induced by recombinant tumor necrosis factor and recombinant interleukin-1 was not affected by treatment with the lymphotoxin antibodies. When lymphotoxin was infused subcutaneously into normal mice (10 micrograms per day for three days), their plasma calcium levels increased. We also evaluated four established cell lines derived from three other patients with myeloma, and found a similar pattern of lymphotoxin expression in each. It appears that production of the bone-resorbing cytokine lymphotoxin is related to osteoclastic bone destruction and hypercalcemia in patients with myeloma.

Bone marrow samples from 55 patients with multiple myeloma (MM) and 23 patients with monoclonal gammopathy of undertermined significance (MGUS) were evaluated with a broad panel of monoclonal antibodies. Plasma cells from 78% (43/55) of patients with MM strongly expressed the natural killer cell antigen CD56 (NKH-1, Leu-19). Of the 23 patients with MGUS, none showed strong CD56 reactivity, although three had weak reactivity in less than 20% of plasma cells. Myeloma cells expressing CD56 did not coexpress the CD57 or CD16 antigens. Patients with CD56-positive plasma cells had both indolent and aggressive disease. However, the 12 CD56-negative patients had predominantly aggressive disease with an unexpected preponderance of kappa Bence Jones only myeloma (5/10[50%] evaluable patients). Polyclonal plasma cells from non-neoplastic tissue sites (normal bone marrows, lymph nodes, tonsillar biopsies, and gut-mucosa biopsies) showed a near absence of CD56. We conclude that isolated, strong CD56 expression is common in MM, but not in MGUS or reactive plasma cells. The potential biologic importance of CD56 positivity in myeloma is reviewed.

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Generation time ( G T ) of normal human bone marrow cells cultured in the C FC-gm assay was measured by using bromodeoxyuridine (BrdU) incorporation and sister chromatid differential staining. Cells were cultured in methylcellulose for 72 hr prior to the addition of BrdU and then harvested at 6-12 hr intervals for up to 72 hr. The time interval between the appearance of second and third division metaphases at the 50% level gave mean GTs which ranged from 32 to 43 hr. These values are longer than those reported for myeloblasts and promyelocytes but shorter than those previously reported for myelocytes.Human bone marrow functions as a continuously renewing cell system in which pluripotent stem cells give rise by proliferation and differentiation to functionally mature haematopoietic elements. Studies of the generation time of normal human bone marrow precursor cells in uiuo and in uifro are few, due both to ethical considerations (i.e. injecting normal individuals with [3H]TdR) and to difficulties in obtaining adequate cell numbers in uitro to apply traditional cytokinetic techniques.Bromodeoxyuridine (BrdU) incorporation and sister chromatid differential staining has proved to be a rapid means for identifying successive cell divisions in a proliferating population (Tice, Schneider & Rary, 1976; Crossen & Morgan, 1977a,b;Crossen et al., 1985). In this method, cells were cultured in medium that contained the thymidine analogue 5-bromodeoxyuridine (5-BrdU) which is readily incorporated into chromosomal DNA. Metaphases which have completed two complete cell cycles in medium containing BrdU have chromosomes in which the DNA of one chromatid has both strands substituted with BrdU while the other chromatid has only one strand substituted with BrdU. When stained by the sister chromatid exchange technique, the doubly-substituted chromosome appears pale staining while the singly substituted is dark staining. Cells that have completed one cell cycle have both chromatids dark staining while metaphases that have completed three cell cycles have three quarters of their chromatin pale staining. By harvesting the cultures at various time intervals, a detailed analysis of the proliferative characteristics of the culture can be obtained. Recent studies (Trent et af., 1986) indicate that a modification of the BrdU technique

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