Abstract
Obesity increases the risk for metabolic and cardiovascular disease, and adipose tissue plays a central role in this process. Ceramide, the key intermediate of sphingolipid metabolism, also contributes to obesity-related disorders. We show that a high fat diet increased ceramide levels in the adipose tissues and plasma in C57BL/6J mice via a mechanism that involves an increase in gene expression of enzymes mediating ceramide generation through the de novo pathway (e.g. serine palmitoyltransferase) and via the hydrolysis of sphingomyelin (acid sphingomyelinase and neutral sphingomyelinase). Although the induction of total ceramide in response to the high fat diet was modest, dramatic increases were observed for C16, C18, and C18:1 ceramides. Next, we investigated the relationship of ceramide to plasminogen activator inhibitor-1 (PAI-1), the primary inhibitor of plasminogen activation and another key player in obesity. PAI-1 is consistently elevated in obesity and thought to contribute to increased artherothrombotic events and more recently to obesity-mediated insulin resistance. Interestingly, the changes in ceramide were attenuated in mice lacking PAI-1. Mechanistically, mice lacking PAI-1 were protected from diet-induced increase in serine palmitoyltransferase, acid sphingomyelinase, and neutral sphingomyelinase mRNA, providing a mechanistic link for decreased ceramide in PAI-1-/- mice. The decreases in plasma free fatty acids and adipose tumor necrosis factor-alpha in PAI-1-/- mice may have additionally contributed indirectly to improvements in ceramide profile in these mice. This study has identified a novel link between sphingolipid metabolism and PAI-1 and also suggests that ceramide may be an intermediary molecule linking elevated PAI-1 to insulin resistance.
Highlights
Sphingolipids, such as ceramide, sphingosine, and sphingosine 1-phosphate, have been implicated in the pathogenesis of obesity, insulin resistance [3, 4], and cardiovascular disease [5,6,7,8]
This study demonstrates that plasma and adipose ceramide levels are increased in C57BL/6J mice fed an high fat diet (HFD) via a mechanism that involves coordinated changes in the expression of ceramide-generating and -metabolizing enzymes
Our results suggest that the decrease in ceramide synthesis might be a possible contributing mechanism for both the improved insulin sensitivity observed in this study and previously [38, 39] and improved cardiovascular complications [42] observed in mice lacking plasminogen activator inhibitor-1 (PAI-1)
Summary
Mice—C57BL/6J (8-week-old) male wild type (WT) and agematched PAI-1-deficient (PAI-1Ϫ/Ϫ) mice (Jackson Laboratory) were placed for 16 weeks on either an HFD (D12492; Research Diets, New Brunswick, NJ) in which 60% of the total calories were derived from fat (soybean oil and lard) or a control low fat diet (LFD; D12450B) in which 10% of the total calories were derived from fat. The total kcal % in the high and low fat diets was identical This HFD induces obesity, hyperphagia, and insulin resistance in C57BL/6J mice [30]. RNA Analysis—Real time reverse transcription-PCR was performed in an iCycler (Bio-Rad) as previously described [25, 31, 32]. For real time PCR, cDNA was prepared from 1 g of total RNA by using random hexamers and Moloney murine leukemia virus reverse transcriptase (PerkinElmer Life Sciences) in a final reaction volume of 20 l. Measurement of Ceramide Levels—Ceramide levels were analyzed by high performance liquid chromatography-tandem mass spectroscopy as previously described [33] (Lipidomics Core, Medical University of South Carolina). Statistical Analysis—Statistical comparison of results was performed using the unpaired Student’s t test
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
