Downregulation of miR-26b has been found in various cancers, but it has never been investigated in osteosarcoma. In this study, we demonstrated downregulation of miR-26b in osteosarcoma tissues, negatively correlated with the expression of connective tissue growth factor (CTGF) and Smad1. Luciferase reporter assay confirmed the interaction of miR-26b with the 3' untranslated regions (UTRs) of CTGF and Smad1. Transfection of miR-26b in osteosarcoma cells suppressed the expression of CTGF and Smad1, suggesting CTGF and Smad1 as direct targets of miR-26b. Overexpression of miR-26b inhibited the migration of osteosarcoma cells, which was reversed by overexpression of CTGF or Smad1. Knockdown of CTGF by small interfering RNA (siRNA) interference blocked the activation of Smad1, ERK1/2, and MMP2, which was opposite to the overexpression of CTGF. Differently, Smad1 did not significantly affect CTGF level, but mediated ERK1/2 phosphorylation and MMP2 activation. Furthermore, miR-26b inhibited lung metastasis of osteosarcoma in vivo. Our data indicated that downregulation of miR-26b in osteosarcoma elevated the levels of CTGF and Smad1, facilitating osteosarcoma metastasis.