Disturbances in lipid metabolism related to excessive food intake and sedentary lifestyle are among major risk of various metabolic disorders. Stearoyl-CoA desaturase-1 (SCD1) has an essential role in these diseases, as it catalyzes the synthesis of unsaturated fatty acids, both supplying for fat storage and contributing to cellular defense against saturated fatty acid toxicity. Recent studies show that increased activity or over-expression of SCD1 is one of the contributing factors for type 2 diabetes mellitus (T2DM). We aimed to investigate the impact of the common missense rs2234970 (M224L) polymorphism on SCD1 function in transfected cells. We found a higher expression of the minor Leu224 variant, which can be attributed to a combination of mRNA and protein stabilization. The latter was further enhanced by various fatty acids. The increased level of Leu224 variant resulted in an elevated unsaturated: saturated fatty acid ratio, due to higher oleate and palmitoleate contents. Accumulation of Leu224 variant was found in a T2DM patient group, however, the difference was statistically not significant. In conclusion, the minor variant of rs2234970 polymorphism might contribute to the development of obesity-related metabolic disorders, including T2DM, through an increased intracellular level of SCD1.

Dietary trans fatty acids (TFAs) have been implicated in serious health risks, yet little is known about their cellular effects and metabolism. We aim to undertake an in vitro comparison of two representative TFAs (elaidate and vaccenate) to the best-characterized endogenous cis-unsaturated FA (oleate). The present study addresses the possible protective action of TFAs on palmitate-treated RINm5F insulinoma cells with special regards to apoptosis, endoplasmic reticulum stress and the underlying ceramide and diglyceride (DG) accumulation. Both TFAs significantly improved cell viability and reduced apoptosis in palmitate-treated cells. They mildly attenuated palmitate-induced XBP-1 mRNA cleavage and phosphorylation of eukaryotic initiation factor 2α (eIF2α) and stress-activated protein kinase (SAPK)/c-Jun N-terminal kinase (JNK), but they were markedly less potent than oleate. Accordingly, all the three unsaturated FAs markedly reduced cellular palmitate incorporation and prevented harmful ceramide and DG accumulation. However, more elaidate or vaccenate than oleate was inserted into ceramides and DGs. Our results revealed a protective effect of TFAs in short-term palmitate toxicity, yet they also provide important in vitro evidence and even a potential mechanism for unfavorable long-term health effects of TFAs compared to oleate.

Phosphatidylinositol (PI) 3,5-bisphosphate (PI(3,5)P2) is a minor inositol-containing phospholipid that serves as an important regulator of endolysosomal functions. However, the precise sites of subcellular enrichment and molecular targets of this regulatory lipid are poorly understood. Here, we describe the generation and detailed characterization of a short engineered catalytic fragment of the human PIKfyve enzyme, which potently converts PI 3-phosphate to PI(3,5)P2. This novel tool allowed for the evaluation of reported PI(3,5)P2-sensitive biosensors and showed that the recently identified phox homology (PX) domain of the Dictyostelium discoideum (Dd) protein, SNXA, can be used to monitor the production of PI(3,5)P2 in live cells. Modification and adaptation of the DdSNXAPX-based probes into compartment-specific bioluminescence resonance energy transfer–based biosensors allows for the real-time monitoring of PI(3,5)P2 generation within the endocytic compartments of entire cell populations. Collectively, these molecular tools should allow for exciting new studies to better understand the cellular processes controlled by localized PI(3,5)P2 metabolism.

Trans fatty acids (TFAs) are not synthesized in the human body but are generally ingested in substantial amounts. The widespread view that TFAs, particularly those of industrial origin, are unhealthy and contribute to obesity, cardiovascular diseases and diabetes is based mostly on in vivo studies, and the underlying molecular mechanisms remain to be elucidated. Here, we used a hepatoma model of palmitate-induced lipotoxicity to compare the metabolism and effects of the representative industrial and ruminant TFAs, elaidate and vaccenate, respectively, with those of cis-oleate. Cellular FAs, triacylglycerols, diacylglycerols and ceramides were quantitated using chromatography, markers of stress and apoptosis were assessed at mRNA and protein levels, ultrastructural changes were examined by electron microscopy and viability was evaluated by MTT assay. While TFAs were just slightly more damaging than oleate when applied alone, they were remarkably less protective against palmitate toxicity in cotreatments. These differences correlated with their diverse incorporation into the accumulating diacylglycerols and ceramides. Our results provide in vitro evidence for the unfavorable metabolic features and potent stress-inducing character of TFAs in comparison with oleate. These findings strengthen the reasoning against dietary trans fat intake, and they can also help us better understand the molecular mechanisms of lipotoxicity.