RESUMO
OBJECTIVE: Multiple myeloma (MM) is caused by abnormal cloning of plasma cells. miR-184 is abnormally expressed in several types of tumors, but its expression and role in MM have not been reported. METHODS: The bone marrow samples of healthy controls and MM patients were collected, and plasma cells were sorted. The multiple myeloma cell line OPM-2 was cultured and assigned into miR-NC+siRNA-NC group, miR-184 inhibitor+siRNA-NC group, and miR-184 inhibitor+siRNA-Notch1 group. Cell proliferation was assessed by MTT assay. Clone formation was evaluated by colony formation assay. Cell apoptosis activity was tested with flow cytometry. Notch1 and cleaved caspase3 protein expressions were detected. RESULTS: MiR-184 expression was increased in myeloma plasma cells (P<0.05). Transfection of miR-184 inhibitor can downregulate miR-184 expression, increase the levels of Notch1 and cleaved caspase3, inhibit OPM-2 cell proliferation, restrain colony formation, enhance caspase3 activity, and suppress tumor cell invasion (P<0.05). However, administration of siRNA-Notch1 retarded the effect of miR-184 inhibitor by decreasing the expressions of Notch1 and cleaved caspase3, enhancing colony formation and tumor cell invasion, as well as inhibiting caspase3 activity and cell proliferation. CONCLUSION: Our data indicated that miR-184 expression is increased in myeloma plasma cells. Down-regulation of miR-184 promotes MM cell apoptosis and inhibits proliferation and colony formation by regulating Notch1 expression.
Assuntos
Apoptose , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , MicroRNAs , Mieloma Múltiplo , Receptor Notch1 , Humanos , Mieloma Múltiplo/genética , Mieloma Múltiplo/patologia , Mieloma Múltiplo/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Receptor Notch1/metabolismo , Receptor Notch1/genética , Proliferação de Células/genética , Apoptose/genética , Linhagem Celular Tumoral , Masculino , Feminino , Caspase 3/metabolismo , Caspase 3/genética , Pessoa de Meia-Idade , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismoRESUMO
INTRODUCTION: Osteosarcoma, the most prevalent primary malignancy of the bone, is often presented with high-grade subclinical metastatic disease that metastasizes at very early stages. Exosomes, as molecular information carriers, may play a potent role in the occurrence and development of tumors through oncogenic molecular reprogramming of tumor-associated macrophages (TAMs). In this study, we will investigate the effect of osteosarcoma-derived exosomes on the polarization of TAMs and decipher its underlying molecular mechanism. MATERIAL AND METHODS: Osteosarcoma-derived exosomes from MG63 cells were isolated and characterized by transmission electron microscopy, and nano-particle size analysis. Double fluorescence staining was performed to confirm the macrophages phagocytosis of exosomes. Western blot, qRT-PCR, and transwell assays were conducted to assess the effect of exosomes on migration, invasion, and macrophage differentiation. The mouse model of osteosarcoma was established to evaluate the effects of exosomes on lung metastasis in vivo. RESULTS: MG63 exosomes were successfully isolated and verified to be phagocytized by macrophages through fluorescence confocal microscopy. The results revealed that osteosarcoma cells could induce M2 type differentiation of macrophages largely through Tim-3 mediated by exosomes, which in turn could promote the migration, invasion, epithelial-mesenchymal transition (EMT), and lung metastasis of osteosarcoma cells through the secretion of cytokines including IL-10, TGF-ß, and VEGF. CONCLUSIONS: Our results demonstrated that osteosarcoma-derived exosomes induced M2 polarization of macrophages and promoted the invasion and metastasis of tumors through Tim-3; besides, the study also suggests a novel therapeutic target for future studies.
