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To elucidate the transcriptomic changes of long noncoding RNAs (lncRNAs) in high-fat diet (HFD)-fed mice, we defined their hepatic transcriptome by RNA sequencing. Aberrant expression of 37 representative lncRNAs and 254 protein-coding RNAs was observed in the livers of HFD-fed mice with insulin resistance compared with the livers from control mice. Of these, 24 lncRNAs and 179 protein-coding RNAs were upregulated, whereas 13 lncRNAs and 75 protein-coding RNAs were downregulated. Functional analyses showed that the aberrantly expressed protein-coding RNAs were enriched in various lipid metabolic processes and in the insulin signaling pathway. Genomic juxtaposition and coexpression patterns identified six pairs of aberrantly expressed lncRNAs and protein-coding genes, consisting of five lncRNAs and five protein-coding genes. Four of these protein-coding genes are targeted genes upregulated by PPARα. As expected, the corresponding lncRNAs were significantly elevated in AML12 cells treated with palmitic acid or the PPARα agonist, WY14643. In Hepa1-6 cells, knockdown of NONMMUG027912 increased the cellular cholesterol level, the expression of cholesterol biosynthesis genes and proteins, and the HMG-CoA reductase activity. This genome-wide profiling of lncRNAs in HFD-fed mice reveals one lncRNA, NONMMUG027912, which is potentially regulated by PPARα and is implicated in the process of cholesterol biosynthesis.

We describe the development of a method for the extraction and analysis of 62 sterols, oxysterols, and secosteroids from human plasma using a combination of HPLC-MS and GC-MS. Deuterated standards are added to 200 μl of human plasma. Bulk lipids are extracted with methanol:dichloromethane, the sample is hydrolyzed using a novel procedure, and sterols and secosteroids are isolated using solid-phase extraction (SPE). Compounds are resolved on C18 core-shell HPLC columns and by GC. Sterols and oxysterols are measured using triple quadrupole mass spectrometers, and lathosterol is measured using GC-MS. Detection for each compound measured by HPLC-MS was ∪ 1 ng/ml of plasma. Extraction efficiency was between 85 and 110%; day-to-day variability showed a relative standard error of