The Glucosylceramide Synthase InhibitorN-(5-Adamantane-1-yl-methoxy-pentyl)-deoxynojirimycin Induces Sterol Regulatory Element-Binding Protein-Regulated Gene Expression and Cholesterol Synthesis in HepG2 Cells

Abstract

Recent findings have implicated glycosphingolipids as modulators of insulin receptor activity. Studies with C57BL/6J ob/ob mice have shown that insulin sensitivity is enhanced by the synthetic hydrophobic iminosugar N-(5-adamantane-1-yl-methoxy-pentyl)-deoxynojirimycin (AMP-DNM) that inhibits glucosylceramide synthase. Here, we treated the liver hepatoma cell line HepG2 with AMP-DNM, resulting in a 70% reduction of glycosphingolipids, and we analyzed the effect on gene expression. Using whole human genome 44K oligonucleotide arrays, we identified 89 genes that were significantly (p Ͻ 0.01) up-or down-regulated by AMP-DNM treatment. Of the 56 up-regulated genes, 17 were direct target genes for transcription factors sterol regulatory element-binding protein (SREBP) 1 or SREBP2, which activate genes in the sterol biosynthesis pathway. An increase in cholesterol production rate confirmed that the induction of SREBP target genes seen at the mRNA level resulted in activation of the cholesterol biosynthesis pathway. It is interesting to note that the cholesterol content of the cells did not increase. It is noteworthy that no effects were found on expression of genes related to cell receptor signaling pathways, neither on toxicity nor cell growth. Our findings indicate that inhibition of glucosylceramide synthase with AMP-DNM leads to activation of SREBP target genes and synthesis of cholesterol in HepG2 cells.

Phosphatidylinositol 3-Kinase/AKT Pathway Regulates the Endoplasmic Reticulum to Golgi Traffic of Ceramide in Glioma Cells

“…In this context it is interesting to note that the PI3K/Akt pathway can positively regulate cholesterol biosynthesis by favoring the activating processing of SCAP/SREBP on its ER to Golgi movements (53). Furthermore, both Cer biosynthesis in the ER (57) and the biosynthesis of complex sphingolipids have been positively correlated with the activating processing of SREBP (58,59), independently of cellular cholesterol concentration. Thus we can hypothesize a role for the PI3K/Akt pathway to coordinate the metabolism of SM and cholesterol, membrane lipids known to co-localize in cell membrane lipid rafts.…”

Compounds of the sphingomyelin-ceramide-glycosphingolipid pathways as secondary messenger molecules: new targets for novel therapies for fatty liver disease and insulin resistance

“…The metabolism of SL is dysregulated in obesity, in which increased adipose tissue mass affects whole body insulin resistance and cardiovascular disease risk via adipose-tissue derived inflammatory cytokines that induce chronic inflammation, as well as increasing cardiovascular risk while the antagonizing insulin signaling and mitochondrial function, thereby impairing glucose homeostasis (23,149,153). These data suggest that SLs are effective targets for the treatment of diseases associated with chronic inflammation (23,20,24).…”

“…In ob/ob mice, HFD mice, and ZDF rats, AMP-DNM normalized the elevated tissue of GC and GM3 levels, subsequently lowering the circulating glucose level, hence improving oral glucose tolerance and insulin sensitivity in muscle and liver (5,6,161). AMP-DNM resulted in a 70% reduction of GSLs in the liver hepatoma cell line HepG2 (20,21,165). The effect was associated with a significant up-or downregulation of target genes for the transcription factors SREBP1 or SREBP2, which activate genes in the sterol biosynthesis pathway.…”

“…The effect was associated with a significant up-or downregulation of target genes for the transcription factors SREBP1 or SREBP2, which activate genes in the sterol biosynthesis pathway. The associated increase in cholesterol production was as a result of the activation of the cholesterol biosynthesis pathway (20,21,165). AMP-DNM treatment decreased plasma level of triglycerides and cholesterol, while neutral sterol, biliary lipids, and bile flow excretion is increased twofold (22,23,166).…”

“…SLs are also relevant in the pathogenesis of these diseases. Obesity-related inflammatory pathways modulate SL metabolism (20,23,24). Metabolomics profiling, liquid chromatography mass spectrometry, and gene expression analyses showed increased hepatic Cer and a higher levels of SLs in NASH (21,22,28,30,36,38).…”