Supernatants of acquired immunodeficiency syndrome-related Kaposi's sarcoma cells induce endothelial cell chemotaxis and invasiveness

Kaposi's sarcoma (KS) in general, and acquired immunodeficiency syndrome-related KS (AIDS-KS) in particular, is a highly invasive and intensely angiogenic neoplasm of unknown cellular origin. We have recently established AIDS-KS cells in long term culture and reported the development of KS-like lesions in nude mice inoculated with these cells. Here, we have examined the in vitro invasiveness of basement membrane by AIDS-KS cells, as well as the effect(s) of their supernatants on the migration and invasiveness of human vascular endothelial cells. AIDS-KS cells were highly invasive in the Boyden chamber invasion assay and formed invasive, branching colonies in a 3-dimensional gel (Matrigel). Normal endothelial cells form tube-like structures on Matrigel. AIDS-KS cell-conditioned media induced endothelial cells to form invasive clusters in addition to tubes. KS-cell-conditioned media, when placed in the lower compartment of the Boyden chamber, stimulated the migration of human and bovine vascular endothelial cells across filters coated with either small amounts of collagen IV (chemotaxis) or a Matrigel barrier (invasion). Basic fibroblast growth factor could also induce endothelial cell chemotaxis and invasion in these assays. However, when antibodies to basic fibroblast growth factor were used the invasive activity induced by the AIDS-KS-cell-conditioned media was only marginally inhibited, suggesting that the large quantities of basic fibroblast growth factor-like material released by the AIDS-KS cells are not the main mediators of this effect. Specific inhibitors of laminin and collagenase IV action, which represent critical determinants of basement membrane invasion, blocked the invasiveness of the AIDS-KS cell-activated endothelial cells in these assays. These data indicate that KS cells appear to be of smooth muscle origin but secrete a potent inducer of endothelial cell chemotaxis and invasiveness which could be responsible for angiogenesis and the resulting highly vascularized lesions. These assays appear to be a model to study the invasive spread and angiogenic capacity of human AIDS-related KS and should prove useful in the identification of molecular mediators and potential inhibitors of neoplastic neovascularization.     

Inhibition of tumor invasion of smooth muscle cell layers by recombinant human metalloproteinase inhibitor

Matrix metalloproteinases secreted by tumor cells play an important role in the proteolytic degradation of the extracellular matrix during invasion. In a previous study, we showed that the degradation of extra-cellular matrices by human HT 1080 fibrosarcoma cells is suppressed by endothelial cells. The identification of inhibitors of metalloproteinases secreted by endothelial cells led us to postulate that these inhibitors were responsible for the suppressive effect (Cancer Res., 46: 3580-3586, 1986). In the present study, we have investigated the inhibitory activity of one of these inhibitors designated metalloproteinase inhibitor (MI)/tissue inhibitor of metalloproteinases (TIMP)-2 on the degradation and invasion of rat smooth muscle cell matrices by two invasive tumor cell lines, the c-Ha-ras-1 transfected rat embryo cell line 4R and the HT 1080 human fibrosarcoma cell line. The inhibitor was obtained in recombinant form from the culture medium of Chinese hamster ovary cells transfected with a human MI complementary DNA. Recombinant MI/TIMP-2 inhibited several matrix metalloproteinases identified in the culture medium of the tumor cell lines including interstitial collagenase. Mr 72,000 gelatinase (type IV collagenase), and Mr 92,000 gelatinase. Approximately 70% inhibition of the degradation of smooth muscle cell matrices was observed when the recombinant inhibitor was present along with cultured cells at a concentration of 10 micrograms/ml. Similarly, inhibition of the penetration of a multilayer of growing smooth muscle cells and their surrounding matrix was demonstrated. The inhibitor had no effect on cell growth or attachment. Thus, recombinant MI/TIMP-2, like TIMP, is a potent inhibitor of tumor invasion. Since both inhibitors are secreted by endothelial cells (J. Biol. Chem., 264: 17445-17453, 1989), they may play an important role in protecting large blood vessels from invasion.     

Characterization of kaposis sarcoma-derived cell-cultures from an epidemic and a classic case

Cells derived from skin biopsies from two Kaposi's sarcoma patients, an elderly female with a sporadic non-AIDS form, and an AIDS-affected homosexual male, were established in culture. The classic patient had a few small lesions, while the epidemic case presented-large, disseminated, cutaneous and oral mucosa lesions. The cells obtained from both patients, termed IST-KS2 and AIDS-IST-KS3 respectively, had the characteristic spindle shape reported for Kaposi's sarcoma-derived cells. By immunocytochemistry they were both found to express the smooth muscle specific isoform of alpha actin. The KS cells expressed the fibroblastic antigen TE-7, which is not expressed in endothelial cells. Furthermore both KS cultures were negative for the endothelium associated markers Factor VIII, EN4 and PAL-E. They were also negative for the leukocyte antigen CD45, but were positive for vimentin. Immunocytochemistry studies were therefore suggestive of a primitive mesenchymal cell. When the KS-derived cells were grown on a gel of reconstituted basement membrane, both cultures formed large branching colonies characteristic of malignant cells of mesenchymal origin. No differences were observed between HIV-related and the sporadic KS-derived cultures studied. Fibroblasts and smooth muscle cells did not form branching colonies, while endothelial cells on matrigel differentiated forming tube-like structures. Supernatants from both sporadic and AIDS-related KS cell cultures had similar effects on endothelial cell growth in vitro and were also found to stimulate chemotaxis and chemoinvasion of normal vascular endothelial cells in the Boyden chamber assay, showing angiogenic potential in vitro. Our results demonstrate that long term cultures of spindle shaped cells derived from either HIV-associated and classic KS show the same histocytochemical phenotype, have invasiveness in matrigel similar to that of malignant sarcomas, and share in vitro angiogenic properties. Therefore, factors from the host are likely to be responsible for the divergent clinical picture of the classic and epidemic Kaposi's patients studied here.     

Testicular hyaluronidase induces tubular structures of endothelial cells grown in three-dimensional collagen gel through a CD44-mediated mechanism.

Cultured brain capillary endothelial cells grown in a 3-dimensional collagen gel can form tubular structures after stimulation by angiogenic factors. We found that treatment of such cultures with testicular hyaluronidase led to formation of tubular structures and cell survival. Anion-exchange chromatography of the enzyme preparation on a MonoQ column revealed the presence of the angiogenic factor basic fibroblast growth factor (bFGF) in the flow-through fraction, as determined by immunoblotting; part of the effect on endothelial cell morphogenesis could thus be ascribed to bFGF. However, adsorbed fractions eluted with increasing concentrations of NaCl, which exhibited hyaluronan-degrading activity at neutral pH, did not contain bFGF but were still able to induce tube-like structures of the endothelial cells. Streptomyces hyaluronidase failed to evoke the same effect. Interestingly, blocking of hyaluronan binding to CD44 receptors by the monoclonal antibody KM114 inhibited the effect of hyaluronidase, but not of bFGF, on endothelial cell tube formation. Our data suggest a CD44-mediated specific role for certain populations of testicular hyaluronidase in the induction of angiogenesis.     

The relevance of cell proliferation, vascular endothelial growth factor, and basic fibroblast growth factor production to angiogenesis and tumorigenicity in human glioma cell lines.

Tumor growth is partially dependent on angiogenesis, a process that relies on angiogenic factors. Tumorigenicity of cancer cells is thought to be associated with the production of various angiogenic factors that stimulate or inhibit the rate of endothelial cell migration and proliferation. However, the relative importance of specific individual factors originally studied in cancer cell lines has yet to be determined in vivo. In this study, we examined seven human glioma cell lines for dynamic changes of two major angiogenic factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), and for doubling time and tumorigenicity in nude mice. Various correlation studies demonstrated that in these glioma cell lines, VEGF expression correlated well with RBC density in tumor sections (r2 = 0.804) and with average tumor weight (r2 = 0.987). In contrast, bFGF expression in the observed glioma cell lines did not correlate with tumorigenicity (r2 = 0.001) or with VEGF expression (r2 = 0.255). Furthermore, there was no correlation between doubling time and tumorigenicity in these cell lines (r2 = 0.160). Taken together, these results suggest that VEGF plays a major role in glioma formation and that down-regulation of VEGF, rather than bFGF, would be a more effective choice for glioma gene therapy.     

Extracellular matrix-resident growth factors and enzymes: possible involvement in tumor metastasis and angiogenesis

Summary Neoplastic cells require an appropriate pericellular environment and new formation of stroma and blood vessels in order to constitute a soilid tumor. Tumor progression also involves degradation of various extracellular matrix (ECM) constituents. In this review we have focused on the possible involvement of ECM-resident growth factors and enzymes in neovascularization and cell invasion. We demonstrate that the pluripotent angiogenic factor, basic fibroblast growth factor (bFGF) is an ECM component required for supporting cell proliferation and differentiation. Basic FGF has been identified in the subendothelial ECM producedin vitro and in basement membranes of the cornea and blood vesselsin vivo. Despite the ubiquitous presence of bFGF in normal tissues, endothelial cell (EC) proliferation in these tissues is usually very low, suggesting that bFGF is somehow sequestered from its site of action. Our results indicate that bFGF is bound to heparan sulfate (HS) in the ECM and is released in an active form when the ECM-HS is degraded by cellular heparanase. We propose that restriction of bFGF bioavailability by binding to ECM and local regulation of its release, provides a novel mechanism for regulation of capillary blood vessel growth in normal and pathological situations. Heparanase activity correlates with the metastatic potential of various tumor cells and heparanase inhibiting molecules markedly reduce the incidence of lung metastasis in experimental animals. Heparanase may therefore participate in both tumor cell invasion and angiogenesis through degradation of the ECM-HS and mobilization of ECM-resident EC growth factors. The subendothelial ECM contains also tissue type- and urokinase type- plasminogen activators (PA), as well as PA inhibitor which may regulate cell invasion and tissue remodeling. Heparanase and the ECM-resident PA participate synergistically in sequential degradation of HS-proteoglycans in the ECM. These results together with similar observations on the properties of other ECM-immobilized enzymes and growth factors, suggest that the ECM provides a storage depot for biologically active molecules which are thereby stabilized and protected. This may allow a more localized, regulated and persistent mode of action, as compared to the same molecules in a fluid phase.     

Human chondrosarcoma secretes vascular endothelial growth factor to induce tumor angiogenesis and stores basic fibroblast growth factor for regulation of its own growth.

Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are well-known factors that induce neovascularization in many tumors. The molecular mechanisms that regulate tumor angiogenesis in human chondrosarcoma are not clear. We assessed in this work the angiogenic activities of a human chondrosarcoma cell line (OUMS-27) in vivo and determined the efficacies of angiogenic factors derived from OUMS-27 cells on human umbilical vein endothelial cells (HUVECs) in vitro. Tumor xenografts induced an increase in the formation of neovessels, but the distributions of Ki-67 antigen, VEGF and bFGF were unaffected. We also demonstrated that OUMS-27 cells secreted VEGF(165) into the culture medium and that it was the maximal angiogenic factor to stimulate endothelial proliferation and migration in chondrosarcoma. Anti-VEGF antibodies induced an approximately 70% inhibition of these responses of HUVECs, but did not have any effect on OUMS-27 cells. Anti-bFGF antibodies suppressed not only the activities of HUVECs but also the growth of tumor cells in vitro. We indicate that angiogenesis is principally elicited by VEGF(165) and that tumorigenesis is mainly regulated by bFGF stored in the extracellular matrix of OUMS-27 cells. The present study may offer the availability of combination therapies for inhibition of VEGF and bFGF action on vascular endothelial cells and chondrosarcoma cells, respectively.     

TGF-alpha as well as VEGF, PD-ECGF and bFGF contribute to angiogenesis of esophageal squamous cell carcinoma

It has been demonstrated that vascular endothelial growth factor (VEGF) is associated with tumor progression as an angiogenic factor in esophageal squamous cell carcinoma (SCC)s. However, the role of other angiogenic factors such as transforming growth factor-alpha (TGF-alpha), platelet-derived endothelial cell growth factor (PD-ECGF), and basic fibroblast growth factor (bFGF) are still unknown in esophageal SCCs. In this study, we detected the expression of VEGF, TGF-alpha, PD-ECGF and bFGF in tissue specimens from 96 patients with SCC of the esophagus by immunohistochemical staining. To evaluate angiogenesis, endothelial cells were stained immunohistochemically and microvessel density (MVD) was counted in 24 cases. The positive rates for VEGF, TGF-alpha, PD-ECGF and bFGF were 65% (62/96), 67% (64/96), 66% (63/96), and 49% (47/96), respectively. Only TGF-alpha expression had a strong correlation with the average MVD (p=0.0059). However, the MVD increased as the number of positive factors for these 4 factors increased (p=0.0023). The expression of all of these factors significantly correlated to the depth of tumor invasion, and lymph node metastasis. Finally, survival analysis of the patients revealed that VEGF, TGF-alpha, and PD-ECGF were significant prognostic factors. However, multivariate analysis revealed that these factors were not prognostic. Thus, we suggest that TGF-alpha as well as VEGF, PD-ECGF and bFGF may be associated with angiogenesis, and the progression and metastasis of esophageal squamous cell carcinoma.     

The role of human immunodeficiency virus-I in the pathogenesis of acquired immunodeficiency syndrome-related Kaposi's sarcoma: the importance of an inflammatory and angiogenic milieu

People infected with human immunodeficiency virus type-I (HIV-I) have an increased incidence of Kaposi's sarcoma (KS) and HIV-I infection alters the natural history of KS. The potent trans-activator HIV-I protein Tat plays a major role in the pathogenesis of acquired immunodeficiency syndrome (AIDS)-related KS. Among many of its KS-promoting activities, the Tat protein augments the angiogenic activities of basic fibroblast growth factor (bFGF), interferon gamma, and vascular endothelial growth factor (VEGF); it also mimics the effects of the extramedullary matrix proteins fibronectin and vitronectin, and increases the expression of matrix metalloproteinases. Inflammatory cytokines induce endothelial cells to acquire the phenotype and functional features of AIDS-related KS spindle cells. These cytokines act by both autocrine and paracrine mechanisms. The synergy between cytokines and the HIV-I Tat protein provides possible insight into why AIDS-related KS is more aggressive than the classic Mediterranean form, in which the HIV-I Tat protein does not play a role.     

Evidence for a proangiogenic activity of TNF-related apoptosis-inducing ligand

Starting from the observation that tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/Apo-2L protein is expressed in both malignant and inflammatory cells in some highly vascularized soft tissue sarcomas, the angiogenic potential of TRAIL was investigated in a series of in vitro assays. Recombinant soluble TRAIL induced endothelial cell migration and vessel tube formation to a degree comparable to vascular endothelial growth factor (VEGF), one of the best-characterized angiogenic factors. However, the proangiogenic activity of TRAIL was not mediated by endogenous expression of VEGF. Although TRAIL potentiated VEGF-induced extracellular signal-regulated kinase (ERK) phosphorylation and endothelial cell proliferation, the combination of TRAIL + VEGF did not show additive effects with respect to VEGF alone in inducing vessel tube formation. Thus, although TRAIL has gained attention as a potential anticancer therapeutic for its ability to induce apoptosis in a variety of cancer cells, our present data suggest that TRAIL might also play an unexpected role in promoting angiogenesis, which might have therapeutic implications.     

Thalidomide downregulates angiogenic genes in bone marrow endothelial cells of patients with active multiple myeloma.

PURPOSE: To study the antiangiogenic effect of thalidomide. PATIENTS AND METHODS: The expression of key angiogenic genes was studied in bone marrow endothelial cells (ECs) of patients with active and nonactive multiple myeloma (MM), monoclonal gammopathies unattributed/unassociated (MG[u]), diffuse large B-cell non-Hodgkin's lymphoma, in a Kaposi's sarcoma (KS) cell line, and in healthy human umbilical vein ECs (HUVECs) following exposure to therapeutic doses of thalidomide. RESULTS: Thalidomide markedly downregulates the genes in a dose-dependent fashion in active MMECs and KS cell line, but upregulates them or is ineffective in nonactive MMECs, MG(u)ECs, NHL-ECs, and in HUVECs. Secretion of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and hepatocyte growth factor also diminishes according to the dose in culture conditioned media (CM) of active MMECs and KS, whereas it does not change in the other CM. CONCLUSION: Inhibition by thalidomide is probably confined to the genes of active MMECs and KS. This would account for its higher efficacy in these diseases.     

Heparin oligosaccharides: inhibitors of the biological activity of bFGF on Caco-2 cells.

A number of growth factors, including members of the fibroblast growth factor (FGF) family - hepatocyte growth factor, vascular endothelial growth factor and heparin-binding epidermal growth factor - are dependent on heparan sulphate (HS) for biological activity mediated through their high-affinity signal-transducing receptors. This obligate requirement for HS prompted the search for antagonists of HS function that could be used as anti-growth factor drugs for the treatment of cancer. Basic FGF (bFGF) was the focus of this study. Caco-2, a human colon carcinoma cell line, was adapted to growth in serum-free medium so that investigation of its growth factor requirements for growth and migration could be performed in defined conditions (Jayson GC, Evans GS, Pemberton PW, Lobley RW, Allen T 1994, Cancer Res, 54, 5718-5723). This cell line multiplied and moved in a dose-dependent manner in response to bFGF. Here, we show that the mitogenic response to bFGF is dependent on the presence of heparan sulphate. A library of heparin oligosaccharides with uniform composition but variable length was generated [general formula [IdoA(2S)-GlcNS(6S)n], and oligosaccharides of defined lengths were tested for their ability to inhibit the biological activity of bFGF. While intact heparin and heparin-derived fragments of 12 monosaccharide units did not affect bFGF-induced cell division or bFGF-induced cell migration, octasaccharides and decasaccharides potently inhibited the bFGF-induced growth and migration responses. In particular, octasaccharides completely inhibited these biological activities at 10 microg ml-, a clinically achievable and tolerable concentration. This study shows that the length of an oligosaccharide determines its ability to block the biological activity of bFGF. The observation that the biological activity of cell-surface heparan sulphate can be antagonized in this way in a human carcinoma cell line suggests that oligosaccharides should be investigated further as anti-growth factor agents for the treatment of cancer. In addition, the results suggest that the clinical evaluation of low-molecular weight heparin (LMWH) as an anti-cancer agent might benefit from subfractionation of the LMWH, to remove oligosaccharides of 12 or more residues.     

Suramin inhibits bFGF-induced endothelial cell proliferation and angiogenesis in the chick chorioallantoic membrane.

The effects of suramin, an inhibitor of growth factor mitogenic activity, were evaluated on basic fibroblast growth factor (bFGF)-induced proliferation of bovine aortic endothelial cells and on angiogenesis in the chorioallantoic membrane (CAM) of chick embryos. The role of bFGF gene expression in endothelial cell growth was also investigated by using an antisense oligodeoxynucleotide to bFGF. The 4-fold increase in [3H]-thymidine uptake in endothelial cells in vitro upon stimulation with 10 ng ml-1 of bFGF was inhibited by suramin 300 micrograms ml-1. bFGF antisense oligomer (10 microM) reduced [3H]-thymidine incorporation in exponentially growing cells by 76%; this effect was reversed by bFGF 10 ng ml-1. In the CAM of chick embryos suramin 50 micrograms was a more potent inhibitor of angiogenesis than the combination of heparin 60 micrograms/hydrocortisone 50 micrograms; the mean value of the area with reduced vascularity was significantly larger in suramin-treated CAMs (2.4 cm2) than in heparin/hydrocortisone (0.6 cm2), while the reduction of vascular density was similar (- 35 and - 29% compared to controls, respectively), In conclusion, the effects of treatments with bFGF and bFGF antisense oligomer demonstrate that bFGF plays a relevant role in endothelial cell proliferation and may be the target of suramin since the drug is able to suppress basal and bFGF-induced endothelial cell growth; in addition to this, suramin is a more potent angiogenesis inhibitor in the CAM than the combination of heparin/hydrocortisone.     

Angiogenic switch occurs during the precancerous stage of human esophageal squamous cell carcinoma

We previously reported that vascular endothelial growth factor (VEGF) expression correlates with vessel density in human esophageal squamous cell carcinomas. However, tumor angiogenesis is not controlled simply by the presence of VEGF, and is likely regulated by several angiogenic factors produced by tumor and host cells. The goal of the present study was to determine the angiogenic profile of precancerous and cancerous lesions of the esophagus. Expression of mRNAs for VEGF, platelet derived endothelial cell growth factor (PD-ECGF), basic fibroblast growth factor (bFGF), and interleukin (IL)-8 was examined in six esophageal carcinoma cell lines and fresh biopsy specimens from 16 patients with invasive esophageal carcinoma by RT-PCR. Immunohistochemical analyses with antibodies against VEGF, PD-ECGF, bFGF, and IL-8 were performed on archival specimens of 60 normal esophageal mucosa, 11 dysplasias and 49 carcinomas of the esophagus. Microvessels were stained with anti-CD34 antibody and quantified by counting the number of vessels in a x200 field in the most vascularized areas of the tumor. Esophageal carcinoma cell lines and tumor tissues expressed mRNAs for one or more these angiogenic factors at various levels. An initial increase in vessel density and enhanced expression of PD-ECGF and VEGF were observed in dysplastic epithelium. Vessel density was significantly higher in more advanced lesions. bFGF and IL-8 were not expressed in dysplasias and mucosal carcinomas, but expression was increased in late stage squamous cell carcinoma. These findings suggest that the angiogenic switch is a very early event in the development of invasive carcinoma. Several different angiogenic factors produced by tumor cells and host cells may regulate angiogenesis during different steps of esophageal carcinogenesis.     

Activated mesothelial cells produce heparin-binding growth factors: implications for tumour metastases

Curative surgery for gastrointestinal malignancy is commonly thwarted by local tumour recurrence. The heparin-binding growth factors, basic fibroblast growth factor (bFGF), heparin-binding epidermal growth factor-like growth factor (HB-EGF) and vascular epidermal growth factor (VEGF) are all implicated in the metastatic process, but whether or not these essential growth factors are produced by the activated peritoneum is unknown. This study reveals that peritoneal mesothelial cells constitutively express mRNA for bFGF, HB-EGF and two VEGF spliced variants, VEGF121 and VEGF165. Mesothelial activation with interleukin (IL)-1b or tumour necrosis factor (TNF)-a produced an up-regulation of mRNA for HB-EGF and VEGF, but not bFGF expression. IL-6 failed to stimulate growth factor expression, whereas IL-2 produced a marked suppression in HB-EGF and bFGF, but not VEGF expression. Mesothelial cells were shown to predominantly express mRNA for the intermediate affinity (bg(c)) IL-2 receptor. Cytokine-induced growth factor up-regulation was confirmed at the protein level using Western blotting of mesothelial cell lysates for HB-EGF and culture supernatant enzyme-linked immunosorbent assay for VEGF. The production of these growth factors by human mesothelial cells may play a significant role in post-operative peritoneal tumour recurrence. Their common heparin-binding property offers a potential therapeutic target for manipulating the growth factor environment of the human peritoneum.     

Angiogenic phenotype induced by basic fibroblast growth factor transfection in brain microvascular endothelial cells

Basic fibroblast growth factor (bFGF) is expressed in the vascular endothelium of human brain tumors. To investigate the biological consequences of a possible autocrine modality of microvascular endothelial cell activation by endogenous bFGF in these tumors, mouse brain microvascular endothelial cells were stably transfected with a retroviral expression vector harboring a human bFGF cDNA. When grown on tissue culture plastic, bFGF-transfected clones show a transformed morphology and increased saturation density. bFGF-transfectants have an invasive behavior when seeded on three-dimensional fibrin gel and originate endothelial cell sprouts when embedded within fibrin. Also, bFGF-transfected cells undergo morphogenetic organization and produce a complex network of branching cord-like structures connecting foci of infiltrating cells when seeded on Matrigel, a laminin-rich extracellular matrix material. In contrast, parental and mock-transfected cells do not invade fibrin gels nor organize on Matrigel. These findings demonstrate that bFGF overexpression induces an angiogenic phenotype in brain microvascular endothelial cells characterized by an invasive behavior and morphogenic potential. They support the notion that neovascularization of brain tumors can be triggered by stimuli that induce vascular endothelium to produce its own autocrine factor(s).     

Angiogenic activity of mesenchymal stem cells in multiple myeloma

The pathophysiology of multiple myeloma-induced angiogenesis is complex and involves both direct production of angiogenic cytokines by plasma cells and their induction within the microenvironment. In this research, we investigated whether mesenchymal stem cells participated in inducing the angiogenic response in multiple myeloma, and explored the mechanism by which MSCs influence myeloma angiogenesis. We detected the concentration of angiogenic factors (bFGF, HGF, and VEGF) in the conditioned medium of mesenchymal stem cells and the capillary formation ability of mesenchymal stem cells in vitro. We found that conditioned medium of MSCs derived from MM significantly promoted the proliferation, chemotaxis, and capillary formation of human umbilical vein endothelial cells compared with that from normal donors. ELISA and RT-PCR were used to detect the mRNA and protein levels of angiogenic factors (bFGF, HGF, and VEGF) in the conditioned medium. We found that mRNA and protein levels of angiogenic factors were elevated in MSCs from multiple myeloma compared with normal donors.