Bone morphogenetic protein (BMP)-9 has been associated with insulin resistance and type 2 diabetes mellitus. However, methods for delivering exogenous BMP-9 genes in vivo are lacking. In this study, we developed a gene delivery system using polyethyleneimine (PEI)-based core-shell nanoparticles (PCNs) as gene delivery carriers, and investigated the effectiveness and safety for delivery of the shBMP-9 gene. PCNs possessed a well-defined core-shell nanostructure with hydrophobic polymer cores and dense PEI shells of uniform particle size and highly positively charged surfaces. In vitro evaluation suggested that PCNs had high loading capacity for exogenous genes and low cytotoxicity toward hepatocytes. The transfection efficiency of PCNs/pENTR-shBMP9 complexes was higher than that of commercial lipofectamine 2000/shBMP9. In vivo studies showed that PCNs/pENTR-shBMP9 transfection led to a significant decrease in hepatic BMP9 expression compared with pENTR-shBMP9 transfection. Under high fat diet (HFD) feeding, PCNs/pENTR-shBMP9 mice exhibited aggravated glucose and insulin tolerance. At a molecular level, PCNs/pENTR-shBMP9 mice displayed elevated PEPCK protein levels and lower levels of InsR and Akt phosphorylation than pENTR-shBMP9 mice. These results suggest that the biological effects of PCNs/pENTR-shBMP9 in vivo are much more effective than those of pENTR-shBMP9. Therefore, the polyethyleneimine (PEI)-based core-shell nanoparticle can be applied as promising nanocarriers for effective and safe gene delivery.