Objectives <p>The treatment of bone defects caused by trauma and pathological factors was a common problem in clinic. Extracorporeal fucosylation has been proved to promote osteogenic differentiation. Nevertheless, the biological processes by which they promote osteogenesis are currently poorly understood. Long non-coding RNAs (lncRNAs) were essential for controlling osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). This study aimed to investigate whether LncRNA HIF1A-AS2 could mediate the effects of alpha-(1,3)-fucosyltransferase VI (FTVI) on osteogenic differentiation of BMSCs.</p> Methods <p>Rat BMSCs with lncRNA HIF1A-AS2 interference plasmid or the FTVI overexpression plasmid were co-cultured in osteogenic differentiation medium. The effects of fucosylation modification of FTVI on osteogenic differentiation of BMSCs were examined, with a focus on LncRNA HIF1A-AS2.</p> Results <p>FTVI could upregulate HIF1A-AS2, and inhibition of lncRNA HIF1A-AS2 in FTVI- transfected BMSCs could decrease type I collagen (Col I), runt-related transcription factor 2 (RUNX2), osteocalcin (OCN) and bone morphogenetic protein 2 (BMP2), vascular endothelial growth factor 165 (VEGF165) proteins expressions which were increased by FTVI. Furthermore, it was discovered that inhibiting lncRNA HIF1A-AS2 decreased the the homing ability of BMSCs demonstrating by antigen contents of HECA452 and CD15s.</p> Conclusions <p>According to these results, the effects of FTVI on osteogenic differentiation of BMSCs depend on the existence of lncRNA HIF1A-AS2. A better understanding of the pathophysiological mechanism of bone defect helped to provide theoretical basis for the reconstruction of bone damage using stem cells in clinic.</p>