MicroRNAs (miRNAs) are essential to the progression of osteosarcoma. Previous research using osteosarcoma samples confirmed that miR-377 expression is less than that observed in normal human osteoblast expression. These data suggest a role for miR-377 in osteosarcoma that warrants investigation. To address this concept, we measured miR-377 expression in two cell models, and we also observed that miR-377 was less expressed in osteosarcoma MG-63 cells compared to the hFOB1.19 human fetal osteoblastic cell line. Moreover, miR-377 overexpression reduced cell proliferation and suppressed invasion of MG-63 cells but had no effect on MG-63 cell apoptosis. Because cyclin-dependent kinase 6 (CDK6) may be a potential target of miR-377 in osteosarcoma cells, we overexpressed CDK6 and observed that overexpression attenuated tumor suppressive effects of miR-377 on cell proliferation. Our data suggest that miR-377 can suppress proliferation in MG-63 cells in part by targeting CDK6.