The excessive intake of saturated fatty acids (SFA) causes obesity and liver steatosis, which are major risk factors for insulin resistance and type 2 diabetes. Although the expression of certain microRNAs (miRNAs) targeting the insulin signaling molecules are regulated aberrantly in SFA-induced obesity, their implications on hepatic insulin resistance are largely unknown. This study examined the associations of miR-424-5p, which is induced by SFA, with the development of insulin resistance. SFA palmitate (PA)-treated HepG2 cells and high fat diet (HFD)-induced obese mouse livers showed an impairment of insulin signaling due to a significant decrease in INSR and IRS-1 expression. Based on expression profiling and qRT-PCR analysis, miR-424-5p, which presumably targets the 3'UTR of INSR, was upregulated in both PA treated HepG2 cells and the liver of HFD-fed mice. miR-424-5p was found to target the 3'UTR of INSR directly and downregulated INSR expression at the post-transcriptional step. Furthermore, the over expression of miR-424-5p suppressed INSR expression significantly, leading to impaired insulin signaling and glycogen synthesis in hepatocytes. A novel mechanism for how SFA-induced miR-424-5p impairs insulin signaling through the targeting of 1NSR is reported. In addition, the crucial role and underlying mechanism of miR-424-5p in the obesity-induced hepatic insulin resistance is explained. (C) 2018 Elsevier Inc. All rights reserved.