Multiple myeloma (MM) pathogenesis and progression largely rely on the cells and extracellular factors in the bone marrow (BM) microenvironment. Compelling studies have identified tumour exosomes as key regulators in the maintenance and education of the BM microenvironment by targeting stromal cells, immune cells, and vascular cells. However, the role of MM exosomes in the modification of the BM microenvironment and MM progression remains unclear. Here, we explored the functions of MM exosomes in angiogenesis and immunosuppression in vitro and in vivo. Murine MM exosomes carrying multiple angiogenesis-related proteins enhanced angiogenesis and directly promoted endothelial cell growth. Several pathways such as signal transducer and activator of transcription 3 (STAT3), c-Jun N-terminal kinase, and p53 were modulated by the exosomes in endothelial and BM stromal cells. These exosomes promoted the growth of myeloid-derived suppressor cells (MDSCs) in naive mice through activation of the STAT3 pathway and changed their subsets to similar phenotypes to those seen in MM-bearing mice. Moreover, MM exosomes up-regulated inducible nitric oxide synthase and enhanced the immunosuppressive capacity of BM MDSCs in vivo. Our data show that MM exosomes modulate the BM microenvironment through enhancement of angiogenesis and immunosuppression, which will further facilitate MM progression. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Multiple myeloma (MM) drug resistance (DR) is a multistep transformation process based on a powerful interplay between bone marrow stromal cells and MM cells that allows the latter to escape anti-myeloma therapies. Here we present an overview of the role of the bone marrow microenvironment in both soluble factors-mediated drug resistance (SFM-DR) and cell adhesion-mediated drug resistance (CAM-DR), focusing on the role of new players, namely miRNAs, exosomes and cancer-associated fibroblasts.

Objective. To investigate whether immunologic abnormalities in patients with Behçet's disease (BD) are related to abnormalities of the Th1/Th2 ratio.Methods. Th1/Th2 cytokine production by peripheral blood lymphocytes (PBL) from 31 patients with BD, 11 patients with inflammatory arthritis, and 10 healthy blood donors was evaluated by intracellular immunofluorescence staining. Serum interleukin-12 (IL-12) levels were measured using an enzyme amplified-sensitivity immunoassay. The effect of recombinant IL-12 (rIL-12) on spontaneous and Fas-mediated apoptosis of phytohemagglutinin (PHA)-stimulated PBL was evaluated by flow cytometry using propidium iodide (PI) staining and a bromodeoxyuridine (BrdU)/PI procedure.Results. Intracellular immunofluorescence staining of IL-2, IL-4, and interferon-␥ (IFN␥) in CD3؉ lymphocytes from BD patients demonstrated a strong polarization of the immune response toward the Th1 pathway that correlated with the progression of BD. Peripheral Th1 cells were significantly increased in patients with active disease (n ‫؍‬ 14) as compared with those in patients in complete remission (n ‫؍‬ 17), patients with inflammatory arthritis, and normal donors. In addition, serum IL-12 levels were

In this study, flow cytometry was used to evaluate interleukin-6 (IL-6) production by bone marrow mononuclear cells from 47 patients with multiple myeloma (MM) in different clinical stages and 15 patients with monoclonal gammopathy of undetermined significance. In patients with MM, autocrine IL-6 production paralleled the clinical disease stage. The largest proportion of syndecan-1 ؉ /IL-6 ؉ cells was detected in patients with resistant relapse or primary refractory disease, suggesting that tumor progression involves expansion of myeloma cells producing IL-6. The authors assessed autocrine IL-6 production and in vitro proliferation and apoptosis of myeloma cells in 6 myeloma cell clones (MCCs) and in 2 myeloma cell lines, namely IM-9 and U-266-1970, which showed different sensitivities to the addition of exogenous IL-6. Autocrine IL-6 production was observed in IL-6-independent MCC-2, MCC-3, and MCC-5 cloned from patients with aggressive disease and in the IM-9 cell line. In contrast, IL-6-dependent MCC-1, MCC-4, and MCC-6 were syndecan-1 ؉ and IL-6 ؊ . Blocking experiments with anti-IL-6 monoclonal antibody from clone AH65, which binds IL-6-IL-6R␣ complexes, prevented cell proliferation of IL-6 ؉ MCCs. Flow cytometry evaluations after propidium iodide staining revealed different susceptibilities of MCCs to cell death. IL-6-producing MCCs showed minimal spontaneous and dexamethasone-induced apoptosis, whereas a regular amplitude of apoptosis occurred in the IL-6 ؊ MCCs. These data provide evidence that autocrine IL-6 reflects a highly malignant phenotype of myeloma cells. In fact, autocrine IL-6 production and deregulated apoptosis may induce expansion of selective IL-6 ؉ myeloma cells resistant to spontaneous and drug-induced cell death. IntroductionInterleukin 6 (IL-6), a pleiotropic cytokine produced by a variety of cells, is the most important growth factor for human multiple myeloma (MM). 1-3 Several findings support in vivo and in vitro roles for IL-6 in the disease: specifically, (1) serum IL-6 and IL-6R levels were found to correlate with disease activity 4-6 ; (2) therapy with anti-IL-6 monoclonal antibody (mAb) transiently reversed disease manifestations 7 ; (3) in vitro proliferation of myeloma cells was suppressed by neutralizing mAbs to either IL-6 or its cellular receptors [8][9] ; and (4) inactivation of IL-6 messenger RNA by antisense oligonucleotides inhibited proliferation of plasma cells. 10 Furthermore, other cytokines, such as IL-1, IL-3, and granulocytemacrophage colony-stimulating factor, regulate myeloma cell proliferation in synergy with IL-6 11 or by inducing IL-6 production in myeloma cells or the tumor environment. [12][13] The cellular origin of IL-6 is controversial. Several authors [13][14][15][16] showed that it is produced by the myeloma cells themselves (autocrine hypothesis). Other studies, [17][18][19] however, point to its paracrine production by cells in the bone marrow (BM) and suggest that proliferation of myeloma cells depends on close contact with stromal cells. [20][21...

The role of cancer-associated fibroblasts (CAFs) has not been previously studied in multiple myeloma (MM). Here, cytofluorimetric analysis revealed higher proportions of bone marrow (BM) CAFs in patients with active MM (both at diagnosis and relapse) compared with patients in remission or those with monoclonal gammopathy of undetermined significance or deficiency anemia (controls). CAFs from MM patients produced increased levels of transforming growth factor-β, interleukin-6, stromal cell-derived factor-1α, insulin-like growth factor-1, vascular endothelial growth factor and fibroblast growth factor-2 and displayed an activated and heterogeneous phenotype, which supported their origin from resident fibroblasts, endothelial cells and hematopoietic stem and progenitor cells via the endothelial-mesenchymal transition as well as mesenchymal stem cells via the mesenchymal transition, as both of these processes are induced by MM cells and CAFs. Active MM CAFs fostered chemotaxis, adhesion, proliferation and apoptosis resistance in MM cells through cytokine signals and cell-to-cell contact, which were inhibited by blocking CXCR4, several integrins and fibronectin. MM cells also induced the CAFs proliferation. In syngeneic 5T33MM and xenograft mouse models, MM cells induced the expansion of CAFs, which, in turn, promoted MM initiation and progression as well as angiogenesis. In BM biopsies from patients and mice, nests of CAFs were found in close contact with MM cells, suggesting a supportive niche. Therefore, the targeting of CAFs in MM patients may be envisaged as a novel therapeutic strategy.