Interleukin-6 (B-cell stimulatory factor 2)-dependent growth of a Lennert's lymphoma-derived T-cell line (KT-3)

A T lymphoma cell line (KT-3) established from a patient with Lennert's lymphoma showed macrophage-dependent growth. Macrophage-derived factors were able to replace the macrophage functions. Experiments using a variety of cytokines demonstrated that KT-3 proliferated in response to recombinant interleukin-2 (rIL-2), rIL-4, or rIL-6 but did not proliferate in response to rIL-1 alpha, rIL-1 beta, rIL-3, recombinant granulocyte colony-stimulating factor (rG-CSF), rGM-CSF, recombinant interferon-alpha (rIFN-alpha), rIFN-gamma, recombinant tumor necrosis factor (rTNF-alpha), or native IFN-beta. Polyclonal rabbit anti-IL-6 antibody almost completely neutralized the activities of macrophage- derived factors or IL-6 but not IL-2 or IL-4. Scatchard plot analysis demonstrated that KT-3 cells indeed express IL-6 receptors. The results indicate that the macrophage-derived factor that supports the growth of KT-3 is IL-6 and suggest that macrophage-derived IL-6 may play an important role in the histopathogenesis of Lennert's lymphoma.     

多发性骨髓瘤血管内皮生长因子和白细胞介素-6相互作用的研究

目的　研究人类多发性骨髓瘤(MM)细胞系和MM患者骨髓基质细胞(BMSCs)之间相互作用对血管内皮生长因子(VEGF)和IL6分泌的调控作用,分析VEGF和IL6的相互作用在MM发病机制中的意义。方法　建立MMBMSCs和正常人BMSCs(NBMSCs)的培养体系,用IL6、抗IL6抗体、VEGF、抗VEGF抗体作用于BMSCs和(或)MM细胞系U266后,ELISA方法检测其VEGF和IL6的分泌量。结果　U266分泌VEGF,但不分泌IL6,而MMBMSCs和NBMSCs既分泌VEGF又分泌IL6。重组人VEGF刺激BMSCs后,以时间和剂量依赖性的方式诱导IL6的分泌,此效应可被抗VEGF抗体抑制。外源性IL6促进BMSCs分泌VEGF。当U266与BMSCs黏附后,VEGF分泌增加25~50倍,IL6增加55~90倍,两者差异有统计学意义(P<005);分别加入抗VEGF或抗IL6抗体,IL6或VEGF的分泌受抑。重组人IL6作用于U266,可诱导剂量依赖性的VEGF分泌的增加,此反应可被抗IL6抗体抑制。结论　在MM中,MM细胞和BMSCs之间的相互作用调节VEGF和IL6的分泌,促进MM细胞的生长和血管新生,在MM的发病机制中发挥重要作用,为针对骨髓微环境的靶位治疗提供了理论依据。     

Regulation of interleukin-4 production and cytokine-induced growth potential in peripheral T-cell non-Hodgkin's lymphomas

The malignant cells in tumour tissues produce cytokines/growth factors that may influence tumour growth, tumour immunogenicity and host immune response. We demonstrate that lymph node cell (LNC) purified neoplastic T cells from CD4⁺ peripheral T-cell lymphoma (CD4⁺ PTCL) and CD8⁺ PTCL spontaneously, and after stimulation with anti-CD3, secreted high amounts of interleukin-4 (IL-4) as compared to LNC-purified CD4⁺ and CD8⁺ non-malignant T cells. Furthermore, IL-4 was observed to be the most potent cytokine that induced in vitro proliferation and growth of the malignant T cells. Moreover, malignant T-cell-derived IL-4 secretion was augmented by exogeneous recombinant human interferon-gamma (IFN-γ) and was profoundly inhibited by IL-2. Because IL-4 was shown to be a locally active cytokine with a wide range of immunoregulatory properties, regulation of IL-4 production by IFN-γ and IL-2 in malignant T cells may be one of the important parameters to be assessed in the design of anticancer-specific immunotherapy. In summary, we report that malignant T cells produce IL-4, a type 2 cytokine (Th2 cell response) that acts as a growth factor and which may play a critical role in PTCL disease mechanism.     

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.     

An established CD4+ T lymphoma cell line derived from a patient with so- called Lennert's lymphoma: possible roles of cytokines in histopathogenesis

A T lymphoma cell line, KT-3, was established from the peripheral blood of a patient with so-called Lennert's lymphoma when the patient developed leukemic lymphoma. The KT-3 cells stain positively for alpha- naphthyl acetate esterase, PAS, and acid phosphatase. The surface phenotype is E rosette-positive, CD1-, CD2+, CD3-, CD4+, CD5+, CD8-, CD11-, CD20-, CD25 (anti-Tac)+, OKla-1+, HNK-1-, J-5-, and My9-, and the surface membrane immunoglobulin is negative, which is almost the same phenotype as the leukemic cells studied when the culture was started. Southern blot analysis showed rearrangement in both beta and gamma T cell receptor genes. Although KT-3 cells proliferate spontaneously and form clusters, they cease proliferating when they are cultured at low cell densities. They proliferate vigorously in response to macrophages, macrophage-derived factor, or recombinant interleukin 2 (rIL-2) added to the culture. Furthermore, they secrete interferon- gamma (IFN-gamma) spontaneously, and the secretion is augmented when they are stimulated with macrophage-derived factor. The macrophage- derived factor enhancing KT-3 cell growth is different from IL-1 alpha, IL-1 beta, IL-2, or IFN-gamma. To our knowledge, this is the first tumor cell line established from a patient with Lennert's lymphoma. The results conclusively confirmed that, at the least, Lennert's lymphoma included CD4+ T lymphoma and also suggested that cytokines or factors secreted by T lymphoma cells and epithelioid histiocytes play an important role in the histopathogenesis of Lennert's lymphoma.     

High serum levels of CA125 and interleukin-6 in a patient with Ki-1 lymphoma

We report a 53-year-old-man with an aggressive Ki-1 lymphoma who had high serum CA125, a marker protein of the epithelial ovarian cancer, and interleukin-6 (IL-6) concentrations. Both CA125 and IL-6 levels decreased after chemotherapy and elevated with disease progression. The patient's lymphoma cells obtained before chemotherapy grew continuously in vitro , were IL-6 dependent and were found to secrete CA125 in culture medium. These results indicate that CA125 can be secreted by Ki-1 lymphoma cells and IL-6 may promote the growth of Ki-1 lymphoma cells.     

Adult T-cell leukemia-derived factor/thioredoxin, produced by both human T-lymphotropic virus type I- and Epstein-Barr virus-transformed lymphocytes, acts as an autocrine growth factor and synergizes with interleukin 1 and interleukin 2.

Interleukin 1 (IL-1) has been obtained from the Epstein-Barr virus-infected B-lymphoblastoid cell line 3B6 and shown to be involved in autocrine growth of 3B6 B cells. Independently, adult T-cell leukemia-derived factor (ADF) was purified from human T-lymphotropic virus I-infected leukemic T-cell line (ATL-2) and reported as an interleukin 2 (IL-2) receptor-inducing factor. We have previously reported the same molecular mass, pI, and NH2-terminal amino acid sequence for both 3B6-derived IL-1 and ADF. cDNA cloning of ADF demonstrated high homology with the prokaryotic disulfide reducing enzyme thioredoxin. We show here that ADF and 3B6-derived IL-1 are identical. By RNA blot, 3B6 and ATL-2 cells were shown to contain high levels of 0.6-kilobase mRNA corresponding to ADF. Such message was not detected in resting peripheral blood lymphocytes but could be weakly induced by lymphocyte activation. Antibodies have been raised against synthetic peptides corresponding to the NH2 terminus and the COOH terminus of ADF. Immunoblotting and sequential immunoprecipitation with these antibodies revealed the same 13-kDa protein in 3B6 and ATL-2 cells. Recombinant ADF could sustain growth of 3B6 and ATL-2 cells at low cellular concentration without fetal calf serum; ADF, thus, appears involved in their autocrine growth. Similarly, recombinant ADF could enhance growth of other B-cell lines, including the Epstein-Barr virus-negative Burkitt lymphoma line BL41 and the lymphoblastoid cell lines CRAG8, CRB95, and 1G8. Finally, recombinant ADF exhibits marked synergism with other cytokines, such as IL-1 and IL-2, allowing virally infected lymphocytes to respond to suboptimal amounts of a variety of growth factors.     

Involvement of interleukin-10 (IL-10) and viral IL-6 in the spontaneous growth of Kaposi's sarcoma herpesvirus-associated infected primary effusion lymphoma cells.

Primary effusion lymphoma (PEL) is a distinct type of lymphoma associated with Kaposi's sarcoma-associated herpesvirus (KSHV) infection. To determine the factors responsible for the unrestrained proliferation of PEL, we have studied the growth factor requirements of the PEL-derived BCBL-1 and BC-1 cell lines. Both cell lines were found to be autocrine growth factor dependent and to release human interleukin-6 (IL-6), viral IL-6 (vIL-6), and human IL-10 in the culture supernatant. To establish whether these cytokines contribute to autocrine growth, neutralizing antibodies against human IL-6, vIL-6, human IL-10, and soluble IL-10 receptor were used. These experiments showed that human IL-10 and, to a lesser degree, vIL-6 serve as autocrine growth factors for BCBL-1 and BC-1 cells. Thus, human IL-10 and vIL-6 are growth factors released and used by PEL cells for autonomous proliferation and may be critical to the development and progression of PEL.     

High-level expression of a bioengineered, cysteine-free hepatocyte-stimulating factor (interleukin 6)-like protein.

Hepatocyte-stimulating factor, interferon-beta 2, B-cell stimulation factor 2, and hybridoma/plasmacytoma growth factor are identical proteins presently referred to as interleukin 6 (IL-6). Through the use of synthetic oligonucleotide technology, we have constructed a biologically active recombinant IL-6 (rIL-6) gene based on the sequence of a human IL-6 cDNA. The synthetic gene encodes a cysteine-free, bioengineered rIL-6 protein that is expressed at high levels in Escherichia coli as a tripartite fusion protein. Cleavage of the fusion protein with collagenase releases a 23-kDa rIL-6 protein that can be easily purified to homogeneity. We show that the rIL-6 protein displays a range of biological activities similar to those of natural human IL-6, as demonstrated by its ability to (i) protect cells from viral infection, (ii) stimulate the synthesis of fibrinogen in rat FAZA 967 cells, and (iii) induce the terminal differentiation of B cells, resulting in elevated secretion of immunoglobulin.     

Production of growth factors by malignant lymphoma cell lines

Fourteen Epstein-Barr virus (EBV)-negative cell lines were raised from bone marrow (BM), peripheral blood (PB), or lymph node samples of patients with intermediate- or high-grade malignant lymphoma. The cell lines were propagated in liquid suspension culture. They contain clonogenic progenitors capable of forming lymphoma colonies in semi- solid culture medium. Cells of these lines were used to examine the growth factor requirements of their clonogenic progenitors and to assess their ability to produce their own growth factors. Two of the cell lines (OCI-Ly9 and OCI-Ly13.1) required addition of exogenous factors for colony growth. These factors were routinely provided by media conditioned by phytohemagglutinin-stimulated leukocytes (PHA- LCM). Three lines formed some and nine lines gave rise to optimal numbers of colonies without addition of growth factors. Eight of these factor-independent lines were able to function as feeder cells and promoted colony formation by both factor-dependent lines. Cell lines that displayed feeder cell function released activities into supernatants able to replace their cellular source. Some of these endogenously produced growth-promoting activities could be replaced by known hematopoietic growth factors. Both factor-dependent cell lines were cultured with recombinant IL-1 alpha, IL-2, IL-3, IL-6, and GM colony-stimulating factor (CSF) and semipurified B-cell growth factor (BCGF) interleukin-4 (IL-4). A heterogeneous response pattern was observed. Both lines formed colonies with IL-4. The colonies were comparable in frequency and size with colonies observed with (PHA-LCM). OCI-Ly9 responded to IL-6 but showed no growth with IL-2. In contrast, the TAC-positive line OCI-Ly13.1 gave rise to colonies with IL-2 while remaining unresponsive to IL-6. A moderate number of colonies was observed when cells of this line were cultured with GM-CSF. Colony formation of both lines was uninfluenced by IL1 alpha or IL-3.     

Regulation of insulin-like growth factor-I and of insulin-like growth factor binding protein-1, -3 and -4 in cocultures of rat hepatocytes and Kupffer cells by interleukin-6.

BACKGROUND/AIMS: Catabolism is associated with decreased serum concentrations of insulin-like growth factor (IGF)-I and insulin-like growth factor binding protein (IGFBP)-3 associated with elevated IGFBP-3 protease activity and increased concentrations of IGFBP-1 and -4. The effects of the acute phase mediators interleukin (IL)-6, IL-1beta and tumor necrosis factor alpha (TNFalpha) on the biosynthesis of IGF-I and IGFBPs were studied in primary rat liver cells. METHODS: mRNA levels of IGF-I and of IGFBPs were analyzed by Northern blotting, secretion of IGFBPs by [(125)I]IGF-I ligand blotting. Proteolytic activity was measured using iodinated recombinant IGFBP-3 as the substrate. RESULTS: In hepatocytes, Kupffer cells (KC) and cocultures of hepatocytes with KC, IL-6 reduced IGF-I biosynthesis dose-dependently. IL-6 stimulated mRNA expression and protein secretion of IGFBP-1 and -4 in hepatocytes and that of IGFBP-3 in KC, respectively. In cocultures, biosynthesis of IGFBP-1, -3 and -4 was increased dose-dependently by IL-6, while the effects of IL-1beta or TNFalpha were less prominent. At neutral pH, proteolytic activity against IGFBP-3 was not detected in media of cocultures treated with IL-6. CONCLUSIONS: The alterations of IGF-I, IGFBP-1 and -4 observed in catabolism correlate with the effects of IL-6 on the biosynthesis of these components in primary rat liver cells, while a neutral IGFBP-3 protease was not detectable.     

Vascular endothelial growth factor and interleukin-6 in paracrine tumor-stromal cell interactions in multiple myeloma

Vascular endothelial growth factor (VEGF), a multifunctional cytokine, potently stimulates angiogenesis including tumor neovascularization. Although well established in solid tumors, the role of VEGF in bone marrow neoangiogenesis and paracrine tumor-stromal cell interactions in lymphohematopoietic malignancies has not been fully elucidated. In multiple myeloma (MM), marrow neovascularization parallels disease progression. This parallel prompted us to investigate the expression and secretion of VEGF by myeloma cells and its potential effects in myeloma-marrow stroma interactions. The biologically active splice variants VEGF165 and VEGF121 were expressed and secreted by myeloma cell lines and plasma cells isolated from the marrow of patients with MM. As shown by immunocytochemistry or RT-PCR, myeloma cells did not express or weakly expressed the VEGF receptors FLT-1 and FLK-1/KDR, indicating that autocrine stimulation is unlikely. In contrast, FLK-1/KDR was abundantly expressed by marrow stromal cells. Therefore, we studied the effects of VEGF on marrow stroma, focusing on the secretion of interleukin-6 (IL-6), a potent growth factor for myeloma cells and an inhibitor of plasma cell apoptosis. Exposure of stromal and microvascular endothelial cells to recombinant human (rh) VEGF165 or VEGF121 induced a time- and dose-dependent increase in IL-6 secretion (14- to 27-fold at 50 ng/mL after 24 hours, P <.001). Conversely, rhIL-6 stimulated VEGF expression and secretion in myeloma cell lines (40%-60%; P <.05) and to a variable degree (up to 5.3-fold; P <.005) in plasma cells purified from the marrow of patients with MM. This mutual stimulation suggests paracrine interactions between myeloma and marrow stromal cells triggered by VEGF and IL-6. (Blood. 2000;95:2630-2636)     

Interleukin-6 inhibits the proliferation of B-chronic lymphocytic leukemia cells that is induced by tumor necrosis factor-alpha or -beta

Tumor necrosis factor (TNF-alpha) acts as a growth stimulatory factor on leukemic B lymphocytes from many patients with chronic lymphocytic leukemia (CLL). Because TNF induces production of interleukin-6 (IL-6), which has been shown to be a growth factor for myeloma and other transformed B cells, we examined the possibility that IL-6 mediates the growth-stimulatory effect of TNF on B-CLL cells. In fact, we found that IL-6 is an inhibitor of B-CLL growth. The addition of recombinant human IL-6 markedly decreased the TNF-induced B-CLL growth, and this decrease was even greater when soluble IL-6 receptor, known to act as IL-6 agonist, was added with recombinant IL-6. Conversely, neutralizing monoclonal antibodies to IL-6 and to the IL-6 receptor potentiated the growth stimulation of TNF on B-CLL cells, in line with the possibility that IL-6 functions as a negative feedback regulator of an autocrine TNF action on these B-leukemic cells. Evidence is presented that production of IL-6 by monocytes and B cells of CLL patients is low, suggesting that administration of IL-6 may be beneficial in CLL to reduce the eventual growth stimulation by TNF and, possibly, also the deficiency in platelets and Ig production in this disease.     

Interleukin-10 is an autocrine growth factor for acquired immunodeficiency syndrome-related B-cell lymphoma [see comments]

Interleukin-10 (IL-10) is an acid-sensitive protein of 35 kD that has pleiotropic effects including inhibition of cytotoxic T-cell response, induction of major histocompatibility complex type II in B lymphocytes, induction of B-cell growth and differentiation, and autocrine growth factor activity in monocytes. We and others have shown that IL-10 is produced spontaneously by blood mononuclear cells from human immunodeficiency virus-seropositive patients. In an attempt to ascertain the potential role of IL-10 in acquired immunodeficiency syndrome (AIDS)-related B-cell lymphoma, we evaluated the expression of human IL-10 in both tumor-derived B-cell lines and primary tumor cells. Expression of human IL-10 (hIL-10) mRNA and protein was detected in four of five cell lines examined. An IL-10 antisense oligonucleotide inhibited IL-10 mRNA expression and IL-10 protein production. The proliferation of all B-cell lines was inhibited by an antisense oligonucleotide in a dose-dependent manner that was abrogated by the addition of recombinant hIL-10 protein. No effect of antisense oligonucleotide was observed in the B-cell line not producing hIL-10. Evaluation of primary tumor cells from patients with AIDS-lymphoma cells showed similar production and response to IL-10. These data suggest an autocrine growth mechanism for IL-10 in AIDS-related lymphoma cells and that IL-10 may be important in its pathogenesis.     

Cytokines secreted in response to Toll-like receptor ligand stimulation modulate differentiation of human Th17 cells

OBJECTIVE: Th17 cells (interleukin-17 [IL-17]-secreting T helper cells) have been implicated in the pathogenesis of rheumatoid arthritis and other autoimmune diseases, but the soluble factors that influence human Th17 differentiation have yet to be fully elucidated. This study was undertaken to investigate the hypothesis that the cytokines secreted by human peripheral blood mononuclear cells (PBMCs) in response to a subset of Toll-like receptor (TLR) ligands would influence Th17 polarization. METHODS: Supernatants from human PBMCs treated with a panel of TLR agonists were tested for their ability to induce de novo IL-17 production in naive T helper cells. Multiplex cytokine analysis was used to identify candidate cytokines for subsequent blocking and sufficiency experiments. RESULTS: Conditioned media from PBMCs stimulated with TLR-4 or TLR-8/7 agonists, but not from those stimulated with TLR-2/1, -3, or -9 agonists, evoked robust secretion of IL-17 by T helper cells, independent of coculture with antigen-presenting cells. Multiplex analysis of 22 cytokines and chemokines identified a 6-factor cytokine signature that significantly correlated with IL-17-inducing activity. T cell activation in the presence of recombinant IL-1beta, IL-6, and IL-23 reconstituted robust IL-17 production, and this was enhanced by transforming growth factor beta (TGFbeta). IL-6 suppressed the expression of forkhead box P3 and reversed TGFbeta-mediated inhibition of T cell proliferation, but did not trigger IL-17 secretion. IL-17 production was completely abrogated by anti-IL-1 or IL-1 receptor antagonist and partially inhibited by anti-IL-6, anti-IL-2, or exogenous retinoic acid, but not by anti-tumor necrosis factor alpha. IL-1beta and IL-6 independently induced IL-21 secretion, but the presence of IL-21 alone was not sufficient for IL-17 production. CONCLUSION: These results indicate that ligation of a subset of TLRs generates proinflammatory cytokines that combine to potentiate human Th17 differentiation.     

Autocrine growth stimulation of a human T-cell lymphoma line by interleukin 2.

The ability of tumor cells to produce and to respond to their own growth factor (autocrine secretion) may be of importance for their growth. We describe a human tumor cell line regulated by an autocrine secretion of the growth factor interleukin 2 (IL-2). This T-lymphocyte cell line, IARC 301, was established from a patient with a T-cell lymphoma in the absence of any added specific growth factor. It constitutively expresses biologically functional high-affinity cell-surface receptors for IL-2 as shown by the binding of both radiolabeled purified IL-2 and monoclonal antibodies to IL-2 receptors. In addition, it synthesizes IL-2, which is bound to cell surface receptors. Monoclonal antibodies directed against either IL-2 or the IL-2 receptor block IARC 301 cell growth. These findings demonstrate that the proliferation of this tumor cell line is mediated by an autocrine pathway involving endogenous IL-2 production and its binding to cell surface receptors.     

Constitutive expression and role in growth regulation of interleukin-1 and multiple cytokine receptors in a biphenotypic leukemic cell line.

A cell line (B1) was established from the bone marrow of a patient with a relapse of acute leukemia characterized by a 4;11 chromosomal translocation and biphenotypic features of early B and myeloid lineages. Analysis of the growth requirements of this cell line showed density-dependent growth and secretion of an autostimulatory growth factor, suggesting an autocrine mechanism. Several lines of evidence implicate the participation of interleukin-1 (IL-1) in the autocrine growth regulation of B1 cells. These cells constitutively express the messenger RNA (mRNA) for IL-1 and IL-1 receptor and secrete IL-1; recombinant IL-1 stimulated the growth of colonies when cells were seeded at low density, and anti-IL-1 antibodies inhibited the growth of colonies with cells seeded at higher density. B1 cells do not express detectable levels of mRNA for any of the other cytokines tested, and other cytokines failed to support the growth of B1 cells at low density. In addition, B1 cells express multiple cytokine receptor genes, including the receptors for IL-6, IL-7, tumor necrosis factor and gamma-interferon. Addition of the respective cytokines to the B1 cells resulted in inhibition of the growth of leukemic cells in vitro. The multiplicity of growth-inhibitory cytokine receptors on this leukemic cell line might be due to its biphenotypic lineage and may suggest new therapeutic possibilities in controlling leukemic cell proliferation.     

Stimulation of growth of Nb2 lymphoma cells by interleukin-2 in serum-free and serum-containing media

The human recombinant alanine-125 analogue of interleukin-2 (IL-2) causes a dose-dependent mitogenic response in rat lymphoma Nb2-11C cloned cells when tested in serum-containing medium and serum-free medium. IL-2 and hGH elicit their growth stimulation through different receptors since IL-2 does not compete with 125I-hGH for binding to Nb2 cells and Met14hGH, an antagonist of hGH, inhibits the hGH-stimulated growth of Nb2 cells but not that caused by IL-2. The tumor promoter, 12-O-tetradecanoyl phorbol-13-acetate (TPA) potentiates the action of hGH on Nb2 cells grown in both serum-containing and in serum-free medium. TPA to a variable degree also potentiates IL-2-stimulated growth of Nb2 cells when cultured in medium containing serum but has no effect on cells grown in serum-free medium. In conclusion, IL-2 is a potent mitogen for Nb2-11C cells.     

Transforming growth factor-beta1: differential effects on multiple myeloma versus normal B cells

Interleukin-6 (IL-6), a product of bone marrow stromal cells (BMSCs), is a growth factor for multiple myeloma (MM) cells. Transforming growth factor-beta1 (TGF-beta1) is also produced by BMSCs and can regulate IL- 6 secretion by several tissues, including BMSCs. The present study was designed to characterize in vitro tumor growth regulation by TGF-beta1 in MM. Sorted CD38+CD45RA- MM cells secreted significantly more TGF- beta1 (8.2 +/- 2.0 ng/mL) than peripheral blood mononuclear cells (P < .001), splenic B cells (P < .001), and CD40 ligand (CD40L) pretreated B cells (P < .05). TGF-beta1 secretion by MM-BMMCs (3.8 +/- 0.9 ng/mL) was significantly greater than by N-BMMCs (1.2 +/- 0.1 ng/mL, P < .001). MM-BMSCs also secreted significantly more TGF-beta1 (6.6 +/- 2.5 ng/mL, n = 11) than N-BMSCs (4.4 +/- 0.6 ng/mL, P < .02, n = 10) and N- BMSC lines (3.9 +/- 0.2 ng/mL, P < .02, n = 6). TGF-beta1 secretion was correlated with IL-6 secretion in MM-BMSCs. Anti-TGF-beta1 monoclonal antibody both blocked IL-6 secretion by BMSCs and inhibited the increments in IL-6 secretion by BMSCs induced by MM cell adhesion. Moreover, exogenous TGF-beta1 upregulated IL-6 secretion by MM-BMSCs, normal BMSCs, and CD38+ CD45RA- MM cells, as well as tumor cell proliferation. This is in contrast to the inhibitory effect of TGF- beta1 on proliferation and Ig secretion of normal splenic B cells. Finally, retinoblastoma proteins (pRB) are constitutively phosphorylated in MM cells; TGF-beta1 either did not alter or increased pRB phosphorylation. pRB are dephosphorylated in splenic B cells and phosphorylated in CD40L triggered B cells in contrast to its effects on MM cells, TGF-beta1 decreased phosphorylation of pRB in CD40L treated B cells. These results suggest that TGF-beta1 is produced in MM by both tumor cells and BMSCs, with related tumore cell growth. Moreover, MM cell growth may be enhanced by resistance of tumor cells to the inhibitory effects of TGF-beta1 on normal B-cell proliferation and Ig secretion.     

Interleukin-6 regulation of matrix metalloproteinase (MMP-2 and MMP-9) and tissue inhibitor of metalloproteinase (TIMP-1) expression in malignant non-Hodgkin's lymphomas.

We showed previously that human malignant non-Hodgkin's lymphomas (NHL) degrade extracellular matrix (ECM) components through the action of metalloproteinases and that elevated expression of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) correlated with a poor clinical outcome in patients with NHL. In the present study we sought to investigate whether there is any correlation between the expression of gelatinases (MMP-2 and MMP-9), TIMP-1, and the expression of cytokines and growth factors such as interleukin-1beta (IL-1beta), IL-6, IL-10, tumor necrosis factor alpha (TNF-alpha), transforming growth factor beta (TGFbeta), and basic fibroblast growth factor (bFGF) in human NHL. In lymphoma tissues obtained from 32 patients, elevated expression of IL-6 correlated significantly with elevated messenger RNA (mRNA) levels of MMP-9, MMP-2, and TIMP-1. Moreover, in human lymphoid cell lines of B- and T-cell origin (Raji, Jurkat, and NC-37), IL-6 stimulated production of MMP-9 and MMP-2 but not TIMP-1. In the Matrigel invasion assay IL-6 significantly upregulated transmigration of Raji and Jurkat cells, which in turn was inhibited by recombinant human TIMP-1 and anti-MMP-9 and MMP-2 antibodies. We postulate that IL-6 may play a role in the clinical aggressiveness of human NHL by stimulating MMP production.