Synthesis Of Monounsaturated Fatty Acids Research Articles

656 Psoriasis and obesity: Stearoyl-CoA desaturase 1 deficient mice are resistant to imiquimod-induced psoriasiform dermatitis

Psoriasis is a chronic inflammatory skin condition whose severity has been linked with obesity. In patients with psoriasis, the likelihood of developing new-onset obesity is higher than those without psoriasis. Stearoyl-CoA desaturase (SCD), is an endoplasmic reticulum resident desaturase, that acts as a catalyst in the synthesis of monounsaturated fatty acids. Mice lacking SCD1 are insulin sensitive, resistant to development of obesity and exhibit suppressed fatty acid synthesis. We have earlier shown that skin specific deletion of SCD1 in mice (SKO) results in epidermal proliferation, sebaceous gland hypoplasia and elevated interleukin 6 levels. We hypothesized that SCD1 deficiency will render mice susceptible to imiquimod (IMQ)-induced psoriasiform dermatitis. A topical dose of 62.5 mg IMQ cream was applied daily to the shaved back region of SKO and SCD1 proficient FLOX counterparts for 14 days after which the mice were euthanized. Cumulative average skin thickness in SKO animals following IMQ application, measured prior to excision was significantly (36%) less than that in FLOX animals. Quantitation of epidermal thickness in H&E stained skin sections validated this observation. Further, histological analysis revealed that in spite of skin inflammation and hair follicle dysplasia associated with their genetic background, the degree of IMQ associated inflammatory changes in the SKO mice were remarkably less than that in FLOX counterparts. Keratin 14, the prototypic marker of dividing basal keratinocytes was elevated in IMQ-treated FLOX skin. Moreover, fatty acid binding protein 5, upregulated in psoriasis tissue was reduced in SKO skin. Our data suggests a complex relationship between fatty acid metabolism and cytoskeletal proteins driving aberrant cell proliferation in psoriatic skin. We propose that SCD1 inhibition could be a novel approach for the management of psoriasis, especially in obese individuals.

Increasing jojoba-like wax ester production in Saccharomyces cerevisiae by enhancing very long-chain, monounsaturated fatty acid synthesis

BackgroundFatty acids (FAs) with a chain length of more than 18 carbon atoms (> C18) are interesting for the production of specialty compounds derived from these FAs. These compounds include free FAs, like erucic acid (C22:1-Δ13), primary fatty alcohols (FOHs), like docosanol (C22:0-FOH), as well as jojoba-like wax esters (WEs) (C38-WE to C44-WE), which are esters of (very) long-chain FAs and (very) long-chain FOHs. In particular, FAs, FOHs and WEs are used in the production of chemicals, pharmaceuticals and cosmetic products. Jojoba seed oil is highly enriched in diunsaturated WEs with over 70 mol% being composed of C18:1–C24:1 monounsaturated FOH and monounsaturated FA moieties. In this study, we aim for the production of jojoba-like WEs in the yeast Saccharomyces cerevisiae by increasing the amount of very long-chain, monounsaturated FAs and simultaneously expressing enzymes required for WE synthesis.ResultsWe show that the combined expression of a plant-derived fatty acid elongase (FAE/KCS) from Crambe abyssinica (CaKCS) together with the yeast intrinsic fatty acid desaturase (FAD) Ole1p leads to an increase in C20:1 and C22:1 FAs in S. cerevisiae. We also demonstrate that the best enzyme candidate for C24:1 FA production in S. cerevisiae is a FAE derived from Lunaria annua (LaKCS). The combined overexpression of CaKCS and Ole1p together with a fatty acyl reductase (FAR/FAldhR) from Marinobacter aquaeolei VT8 (MaFAldhR) and a wax synthase (WS) from Simmondsia chinensis (SciWS) in a S. cerevisiae strain, overexpressing a range of other enzymes involved in FA synthesis and elongation, leads to a yeast strain capable of producing high amounts of monounsaturated FOHs (up to C22:1-FOH) as well as diunsaturated WEs (up to C46:2-WE).ConclusionsChanging the FA profile of the yeast S. cerevisiae towards very long-chain monounsaturated FAs is possible by combined overexpression of endogenous and heterologous enzymes derived from various sources (e.g. a marine copepod or plants). This strategy was used to produce jojoba-like WEs in S. cerevisiae and can potentially be extended towards other commercially interesting products derived from very long-chain FAs.

Structural and physiological adaptations of soil microorganisms to freezing revealed by position-specific labeling and compound-specific 13C analysis

Psychrotolerant microbes are crucial for carbon cycling and biotechnological applications. Nonetheless, the mechanisms enabling their survival and functioning in frozen environments remain unclear. To elucidate adaptations of microbial cell membranes to freezing, we incubated soils with position-specific 13C labeled glucose at + 5 (control), − 5 and − 20 °C and quantified 13C in CO2 and phospholipid fatty acids. High oxidation of glucose C-1 at + 5 °C revealed a transformation via the pentose phosphate pathway. At subzero temperatures, however, the preferential oxidation of C-4 position suggested a switch to glycolysis. The threefold increase of Gram-negative phospholipid fatty acids in soil incubated at − 5 °C was accompanied by a twofold increase in 13C incorporation. This unequal increase of phospholipid fatty acids and incorporated 13C can be explained by simultaneous desaturation of existing fatty acid chains and the de novo synthesis of monounsaturated fatty acids, which indicates microbial growth. In contrast, Gram-positive bacteria incorporated 2 times higher 13C into their phospholipid fatty acids at − 20 °C than at − 5 and + 5 °C without a significant increase in their fatty acid contents. This reflects intensive repair of membranes damaged at − 20 °C without microbial growth. The fungal/bacterial ratio was 1.5 times lower at subzero temperatures than at + 5 °C, reflecting a shift in microbial community structure towards bacteria. Accordingly, soil microorganisms adapted to freezing by (1) switching their metabolic pathway from the pentose phosphate pathway to glycolysis, (2) modifying phospholipid fatty acids by desaturation and, (3) shifting microbial community structure towards Gram-negative bacteria by reducing the fungal population.

Vitamin A deficiency increases the oleic acid (C18:1) levels in the kidney of high fructose diet-fed rats.

Background & objectives:Stearoyl-CoA desaturase 1 (SCD1) is a key lipogenic enzyme responsible for endogenous synthesis of monounsaturated fatty acids (MUFA) and plays a key role in various pathophysiology, including fatty liver diseases. In this experimental study the impact of vitamin A deficiency was assessed on SCD1 regulation in relation to kidney biology, under high fructose (HFr) diet-fed condition in rats.Methods:Forty male weanling (21 day old) Wistar rats were divided into four groups control, vitamin A-deficient (VAD), HFr, VAD with HFr consisting of eight rats each, except 16 for the VAD group. The groups received one of the following diets: control, VAD, HFr and VAD with HFr for 16 wk, except half of the VAD diet-fed rats were shifted to HFr diet, after eight week period.Results:Feeding of VAD diet (alone or with HFr) significantly reduced the kidney retinol (0.51, 0.44 μg/g vs. 2.1 μg/g; P<0.05), while increased oleic (C18:1) and total MUFA levels (23.3, 22.2% and 27.3, 25.4% respectively vs. 14.7 and 16.6%; P<0.05) without affecting the SCD1, both at protein and mRNA levels, when compared with HFr. Comparable, immunohistological staining for SCD1 was observed in the distal convoluted tubules. Despite an increase in MUFA, morphology, triglyceride content and markers of kidney function were not affected by VAD diet feeding.Interpretation & conclusions:Feeding of VAD diet either alone or under HFr condition increased the kidney oleic acid (C18:1) levels and thus total MUFA, which corroborated with elevated SCD1 activity index, without affecting its expression status. However, these changes did not alter the kidney morphology and function. Thus, nutrient-gene regulation in kidney biology seems to be divergent.

Surplus dietary isoleucine intake enhanced monounsaturated fatty acid synthesis and fat accumulation in skeletal muscle of finishing pigs

BackgroundIsoleucine (Ile) has been implicated in the regulation of energy homeostasis and adipogenesis. However, the impact of surplus dietary Ile intake on muscle lipogenesis remains unknown. The present study aimed to investigate the impact of dietary supplementation of extra-Ile on lipogenesis, fatty acid profile and lipid accumulation in skeletal muscle in finishing pigs.MethodsForty-eight barrows with initial body weight of 77.0 ± 0.1 kg were allotted to one of two groups and fed diets containing 0.39%, 0.53% standardized ileal digestible (SID) Ile with six replicates per treatment and four pigs per replicate for 30 d.ResultsDietary Ile intake significantly improved the intramuscular fat (IMF) content and monounsaturated fatty acid (MUFA) concentration in the skeletal muscle (P < 0.05), and decreased the drip loss and shear force (P < 0.05) without influencing the growth performance of pigs (P > 0.05). Moreover, the phosphorylation of adenosine monophosphate activated protein kinase α (AMPKα) and acetyl coenzyme A carboxylase (ACC) proteins that monitor lipid metabolism were decreased in skeletal muscle of pigs offered extra-Ile diet (P < 0.05). The mRNA expression of adipose-specific genes adipocyte determination and differentiation factor 1 (ADD1), fatty acid synthase (FAS), and stearoyl-CoA desaturase (SCD) were upregulated and the activity of SCD was increased as well (P < 0.05).ConclusionsSurplus dietary Ile intake could increase IMF accumulation and MUFA synthesis in skeletal muscle through depressing the phosphorylation of AMPKα-ACC and stimulating the expression and activity of SCD, and increasing the capability of lipogenesis in skeletal muscle.

A method for the gross analysis of global protein acylation by gas-liquid chromatography.

Protein acylation is a posttranslational modification in which an amino acid residue of a protein is acylated by a fatty acid. This process plays a key role in regulating proteomic function. Studies of protein acylation have relied on the development and application of extremely complicated molecular methods. However, global protein acylation can be profiled following hydrolysis of fatty acyl groups from cellular proteins. The present study aimed to develop a method for analysis of global protein acylation using gas-liquid chromatography (GLC). The total protein was extracted from the human hepatocellular carcinoma (HepG2) cell line. Protein sedimentation and extensive wash were combined with differential O-, S-, or N-acyl hydrolysis using sodium hydroxide (NaOH), hydroxylamine (NH2 OH), or hydrochloric acid (HCl), respectively. GLC with a flame ionization detector system was used to analyze changes in the fatty acid composition of the released lipids. The effect of selective inhibition of monounsaturated fatty acid (MUFA) synthesis on global protein acylation and the expression of reprogramming markers were determined to further validate the proposed profiling approach. In all hydrolysis conditions, the amount of myristate released was significantly higher than of other fatty acids. Notable differences were observed in the release of individual fatty acids among the hydrolyzing agents. Only NH2 OH could release significant amounts of palmitoleate (>2.5-fold vs. NaOH and HCl). The acylation assay indicates that treatment with a chemical inhibitor of monounsaturated fatty acid synthesis led to an overall increase in saturated fatty acid O- and N-acylation, and a decrease in palmitoleate O- and S-acylation of cellular proteins (<-15%). This was accompanied by significant reductions in the gene expression of the reprogramming markers Oct4 (-26%, P < 0.01) and Sox2 (-40%, P < 0.01). GLC-based analysis of global protein acylation affords a semi-quantitative method that can be used to assess the gross changes in the protein acylation profile during cell differentiation and reprogramming. © 2018 IUBMB Life, 71(3):340-346, 2019.

Potential contribution of monounsaturated fatty acids to cardiovascular risk in chronic kidney disease.

INTRODUCTION Patients with chronic kidney disease (CKD) are particularly susceptible to cardiovascular disease (CVD). Increased synthesis of endogenous monounsaturated fatty acids (MUFAs) by stearoyl-CoA desaturase-1 (SCD1) might predispose to cardiovascular complications. OBJECTIVES The aim of this study was to examine the serum MUFA content in patients at subsequent stages of CKD, and to evaluate associations between MUFA content and SCD1 activity, patients' diet, and cardiovascular risk. PATIENTS AND METHODS Serum fatty acid composition was evaluated in 177 patients with subsequent stages of CKD (1-2, 3a, 3b, 4-5, hemodialysis, peritoneal dialysis, and after kidney transplantation), and in 30 healthy controls. Gas chromatography-mass spectrometry was used for the measurement. RESULTS Serum MUFA content was shown to increase with subsequent stages of CKD and to be correlated with various risk factors of CVD, including serum triacylglycerols, HDL cholesterol (P <0.01), and high-sensitivity C-reactive protein (P <0.05). Moreover, also the prevalence of CVD was shown to increase with CKD progression. Estimated SCD1 activity was associated with serum MUFA content (P <0.01), but no association was found between dietary MUFA intake and serum MUFA levels. CONCLUSIONS Our results indicate that the elevation of serum MUFA levels in CKD patients may contribute to an increased risk of CVD during CKD progression, mainly due to increased endogenous MUFA synthesis by SCD1.

Increased hydrophilic plasma bile acids are correlated with protection from adiposity in skin-specific stearoyl-CoA desaturase-1 deficient mice.

Stearoyl-CoA desaturase 1 (SCD1) catalyzes the rate limiting step in monounsaturated fatty acid synthesis by inserting a double bond at the delta-9 position of long-chain fatty acids. SCD1 converts stearate (18:0) to oleate (18:1n9) and palmitate (16:0) to palmitoleate (16:1n7), respectively. Mice with global and skin-specific deletion (SKO) of SCD1 exhibit increased whole body energy expenditure and protection against diet-induced adiposity, hepatic steatosis, insulin sensitivity and glucose intolerance. The mechanisms that link cutaneous lipid homeostasis with whole body energy balance are presently unknown. In this study, we reveal that SKO mice demonstrate increased skin surface free cholesterol, decreased circulating total cholesterol and increased taurine-conjugated and hydrophilic bile acids. Tauro-β-muricholic acid, which is a marker of extrahepatic bile acid synthesis, is significantly elevated in SKO plasma. Bile acid signaling through the bile acid-specific receptor TGR5 is known to be protective against obesity and metabolic disease; a phenotype that is similar to SKO mice. We therefore examined TGR5 expression and its downstream mediator, DIO2, in various tissues and found that both TGR5 and DIO2 expression were significantly increased in brown adipose tissue. In sum, we suggest that skin-derived bile acids are involved in the lean and metabolically healthy phenotype of SKO mice.

Downregulation of CPT2 promotes tumorigenesis and chemoresistance to cisplatin in hepatocellular carcinoma

BackgroundCancer cells often have characteristic changes in metabolism. Besides Warburg effect, abnormal lipid metabolism is also considered as one of the most typical metabolic symbols of cancer. Thus, understanding the mechanisms of cell metabolic reprogramming may provide a potential avenue for cancer treatment.Materials and methodsIn total, 41 pairs of matched samples of primary hepatocellular carcinoma (HCC) and adjacent non-cancerous liver tissues were collected. Afterward, we performed quantitative reverse transcriptase polymerase chain reaction to investigate carnitine palmitoyltransferase-2 (CPT2) expression and then systematically analyzed its relationship with clinicopathologic features. We further performed proliferation, colony formation, migration and invasion, drug resistance, and lipogenesis assays to determine the function of CPT2 in HCC.ResultsIn this study, we have identified CPT2 which is the rate-limiting enzyme of fatty acid oxidation, downregulated in HCC and was significantly associated with tumor histological differentiation and venous invasion. In vitro studies demonstrated that knockdown of CPT2 remarkably enhanced the tumorigenic activity and metastatic potential of hepatoma cells. In addition, CPT2 silencing induced chemoresistance to cisplatin. Mechanistically, low expression of CPT2 promoted cancer cell lipogenesis via upregulation of stearoyl-CoA desaturase-1, the key enzyme involved in the synthesis of monounsaturated fatty acids, at both mRNA and protein levels in hepatoma cell line.ConclusionAltogether, our findings demonstrate that CPT2 has a critical role in HCC progression and chemoresistance and may potentially serve as a novel prognostic marker and therapeutic target for HCC treatment.

IL‐6 and Bile Acids are Skin‐Derived Factors that Regulate Whole‐Body Metabolism in SCD1 Deficient Mice

Stearoyl‐CoA desaturase 1 (SCD1) is a lipogenic enzyme that catalyzes the rate limiting step in monounsaturated fatty acid synthesis by inserting a double bond at the delta‐9 position of long‐chain fatty acids. Specifically, SCD1 converts stearate (18:0) to oleate (18:1n9) and palmitate (16:0) to palmitoleate (16:1n7). Global deletion of SCD1 in mice (GKO) provides complete resistance to both high‐carbohydrate and high‐fat diet‐induced obesity. Furthermore, GKO mice also demonstrate improved insulin sensitivity and glucose clearance. Surprisingly, skin‐specific deletion of SCD1 also provides protection from high‐fat diet‐induced obesity and similar metabolic improvements to GKO mice. Histological analysis of SKO skin demonstrates dermal deviations that include 1) an increased number of hair follicles despite progressive hair‐loss, 2) increased macrophage infiltration and 3) differentially expressed IL‐6 that is localized around hair follicles. Co‐localization of macrophage and IL‐6 markers reveal that IL‐6 is not produced by macrophages but instead is produced by cells of the hair follicles. In plasma, IL‐6 protein levels are 25‐fold increased relative to controls, and downstream IL‐6‐mediated metabolic effects are increased at least 5‐fold in distal tissues such as white adipose tissue and liver. Another skin‐derived factor in SKO mice is bile acids, in that, genetic analysis of bile acid synthesis genes in the skin and liver of SKO mice show unexpected and divergent expression. Cyp7a1 the rate‐limiting bile acid synthesis gene is unchanged in SKO liver, but the extrahepatic equivalent Cyp7b1 is significantly increased in skin. In plasma, bile acid composition is not only increased but is also significantly skewed towards increased levels of tauro‐β‐muricholic acid which is indicative of increased extrahepatic bile acid synthesis in mice. Furthermore, the bile acid pool is also more hydrophilic which is a phenotype that is protective against the development of diseases of the metabolic syndrome. Together, changes in both IL‐6 and bile acid composition in SKO mice strongly suggest that skin metabolism is an important component of whole‐body metabolism, and this opens another avenue for the treatment of metabolic diseases.Support or Funding InformationSabrina Dumas was supported by USDA Hatch W2005This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Compensatory increases in tear volume and mucin levels associated with meibomian gland dysfunction caused by stearoyl-CoA desaturase-1 deficiency

The stearoyl-CoA desaturase (SCD) family of enzymes catalyzes monounsaturated fatty acid synthesis by inserting a cis double bond at the Δ9 position of saturated fatty acids. Disruption of these enzymes has been reported to induce a severe dry skin phenotype. Since lipid abnormalities in the meibomian glands have been associated with dry eye, we analyzed selected eye tissues contributing to tear volume and composition in genetically SCD-1-deficient mice (SCD-1 KO), including the lacrimal glands and conjunctiva. Previous histopathological analysis had revealed atrophy and loss of meibomian glands; taken together with the increased goblet cell and MUC5AC expression in the conjunctiva reported here, these findings suggest that the tear volume and mucin levels secreted are enhanced in the absence of lipid secretion as a compensatory mechanism. The expression of lipid metabolism genes in lacrimal glands was decreased in SCD1 KO mice. Thus, these results provide new pathophysiological mechanisms to pursue with regard to meibomian gland dysfunction. In addition, lack of SCD-1 causes a compensatory increase in the tear volume and mucin levels associated with changes in expression of lipid metabolism genes. These results may be useful as a new concept for dry eye treatment strategies.

Skeletal muscle expression of p43, a truncated thyroid hormone receptor α, affects lipid composition and metabolism.

Thyroid hormone is a major regulator of metabolism and mitochondrial function. Thyroid hormone also affects reactions in almost all pathways of lipids metabolism and as such is considered as the main hormonal regulator of lipid biogenesis. The aim of this study was to explore the possible involvement of p43, a 43 Kda truncated form of the nuclear thyroid hormone receptor TRα1 which stimulates mitochondrial activity. Therefore, using mouse models overexpressing p43 in skeletal muscle (p43-Tg) or lacking p43 (p43-/-), we have investigated the lipid composition in quadriceps muscle and in mitochondria. Here, we reported in the quadriceps muscle of p43-/- mice, a fall in triglycerides, an inhibition of monounsaturated fatty acids (MUFA) synthesis, an increase in elongase index and an decrease in desaturase index. However, in mitochondria from p43-/- mice, fatty acid profile was barely modified. In the quadriceps muscle of p43-Tg mice, MUFA content was decreased whereas the unsaturation index was increased. In addition, in quadriceps mitochondria of p43-Tg mice, we found an increase of linoleic acid level and unsaturation index. Last, we showed that cardiolipin content, a key phospholipid for mitochondrial function, remained unchanged both in quadriceps muscle and in its mitochondria whatever the mice genotype. In conclusion, this study shows that muscle lipid content and fatty acid profile are strongly affected in skeletal muscle by p43 levels. We also demonstrate that regulation of cardiolipin biosynthesis by the thyroid hormone does not imply p43.

SCD1 Confers Temozolomide Resistance to Human Glioma Cells via the Akt/GSK3β/β-Catenin Signaling Axis.

Resistance to temozolomide (TMZ), the standard chemotherapy agent for glioblastoma (GBM), poses a major clinical challenge to GBM prognosis. Understanding the mechanisms of TMZ resistance can help to identify novel drug targets and more effective therapies. Recent studies suggest that bioenergetic alterations of cancer cells play important roles in drug resistance. In our study, the altered metabolism of cancer cells was observed using a metabolic PCR array. We found that stearoyl-coenzyme A desaturase 1 (SCD1), a key rate-limiting enzyme for synthesis of monounsaturated fatty acids, was significantly upregulated in TMZ-resistant GBM cells compared to their parental counterparts. Overexpression of SCD1 promoted resistance to TMZ in parental GBM cells, whereas SCD1 downregulation by siRNA could re-sensitize TMZ-resistant cells in vitro. Combinational treatment of TMZ and an SCD1-specific inhibitor showed a combined inhibitory effect on TMZ-resistant glioma cells. We also observed that overexpression of SCD1 promoted Akt/GSK3β/β-catenin signaling, while silencing of SCD1 inhibited the signaling. The combination of an Akt activator with exogenous SCD1 or the combined inhibition of Akt and enforced expression of SCD1 resulted in the most significant changes of Akt signaling. Functionally, significantly lower viability and mobility rates were observed in TMZ-resistant cells when treated with Akt inhibitors and an SCD1 inhibitor simultaneously compared to when treated individually. In conclusion, our study identified SCD1 along with its functional pathway as a novel target in the development of TMZ resistance. SCD1 inhibition used alone or in combination with Akt inhibition could effectively overcome TMZ resistance in gliomas.

Plasma fatty acids and risk of colon and rectal cancers in the Singapore Chinese Health Study

Fatty acid composition in plasma captures both dietary intake and endogenous synthesis. Prospective analyses of plasma fatty acid composition are needed to establish the role of monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs) on risk of developing colorectal cancer. To evaluate associations between plasma fatty acid composition and colon or rectal cancer risk separately, a nested case-control study of 350 colorectal (211 colon and 139 rectal) cancer cases and an equal number of individually matched control subjects was conducted within the Singapore Chinese Health Study, a cohort of 63,257 men and women recruited between 1993 and 1998. Fatty acids in pre-diagnostic plasma were quantified using gas chromatography–tandem mass spectrometry. Conditional odds ratios (ORs) and 95% confidence intervals (CIs) comparing highest to lowest quartiles are presented. For colon cancer, inverse associations were reported with higher essential PUFAs, α-linolenic acid (OR = 0.41; 95% CI: 0.23, 0.73; Ptrend = 0.005) and linoleic acid (OR = 0.43; 95% CI: 0.23, 0.82; Ptrend = 0.008). Higher desaturase activity in the n-6 PUFA synthesis pathway estimated by the arachidonic:linoleic acid ratio was associated with increased colon cancer risk (OR = 3.53; 95% CI: 1.82, 6.85; Ptrend = 0.006), whereas higher desaturase activity in the MUFA synthesis pathway estimated by the oleic:stearic acid ratio was associated with decreased colon cancer risk (OR = 0.42; 95% CI: 0.19, 0.92; Ptrend = 0.024). There was no significant association between the essential fatty acids or the desaturase indices and rectal cancer risk. Endogenous synthesis of arachidonic and oleic acids has an impact on colon cancer development.

Oleate activates SREBP-1 signaling activity in SCD1-deficient hepatocytes.

Stearoyl-CoA desaturase-1 (SCD1) is a key player in lipid metabolism. SCD1 catalyzes the synthesis of monounsaturated fatty acids (MUFA). MUFA are then incorporated into triacylglycerols and phospholipids. Previous studies have shown that Scd1 deficiency in mice induces metabolic changes in the liver characterized by a decrease in de novo lipogenesis and an increase in β-oxidation. Interestingly, Scd1-deficient mice show a decrease in the expression and maturation of the principal lipogenic transcription factor sterol receptor element binding protein-1 (SREBP-1). The mechanisms mediating this effect on de novo lipogenesis and β-oxidation have not been fully elucidated. We evaluated the role of SCD1 on de novo lipogenesis and β-oxidation in HepG2 cells. We also used Scd1-deficient mice and two strains of transgenic mice that produce either oleate (GLS5) or palmitoleate (GLS3) in a liver-specific manner. We demonstrate that the expression of β-oxidation markers increases in SCD1-deficient hepatocytes and suggest that this is due to an increase in cellular polyunsaturated fatty acid content. We also show that the changes in the level of SREBP-1 expression, for both the precursor and the mature forms, are mainly due to the lack of oleate in SCD1-deficient hepatocytes. Indeed, oleate treatment of cultured HepG2 cells or hepatic oleate production in chow-fed GLS5 mice can restore SREBP-1 expression and increase hepatic de novo lipogenesis. Finally, we show that oleate specifically increases SREBP-1 nuclear accumulation, suggesting a central role for oleate in SREBP-1 signaling activity.

EGFR modulates monounsaturated fatty acid synthesis through phosphorylation of SCD1 in lung cancer

BackgroundEpidermal growth factor receptor (EGFR), a well-known oncogenic driver, contributes to the initiation and progression of a wide range of cancer types. Aberrant lipid metabolism including highly produced monounsaturated fatty acids (MUFA) is recognized as a hallmark of cancer. However, how EGFR regulates MUFA synthesis in cancer remains elusive. This is the focus of our study.MethodsThe interaction between EGFR and stearoyl-CoA desaturase-1 (SCD1) was detected byco-immunoprecipitation. SCD1 protein expression, stability and phosphorylation were tested by western blot. The synthesis of MUFA was determined by liquid chromatography-mass spectrometry. The growth of lung cancer was detected by CCK-8 assay, Annexin V/PI staining, colony formation assay and subcutaneous xenograft assay. The expression of activated EGFR, phosphorylated and total SCD1 was tested by immunohistochemistry in 90 non-small cell lung cancersamples. The clinical correlations were analyzed by Chi-square test, Kaplan-Meier survival curve analysis and Cox regression.ResultsEGFR binds to and phosphorylates SCD1 at Y55. Phosphorylation of Y55 is required for maintaining SCD1 protein stability and thus increases MUFA level to facilitate lung cancer growth. Moreover, EGFR-stimulated cancer growth depends on SCD1 activity. Evaluation of non-small cell lung cancersamples reveals a positive correlation among EGFR activation, SCD1 Y55 phosphorylation and SCD1 protein expression. Furthermore, phospho-SCD1 Y55 can serve as an independent prognostic factor for poor patient survival.ConclusionsOurstudy demonstrates that EGFR stabilizes SCD1 through Y55 phosphorylation, thereby up-regulating MUFA synthesis to promote lung cancer growth. Thus, we provide the first evidence that SCD1 can be subtly controlled by tyrosine phosphorylation and uncover a previously unknown direct linkage between oncogenic receptor tyrosine kinase and lipid metabolism in lung cancer. We also propose SCD1 Y55 phosphorylation as a potential diagnostic marker for lung cancer.

Stearoyl-CoA-desaturase 1 regulates lung cancer stemness via stabilization and nuclear localization of YAP/TAZ.

Recent evidences suggest that stearoyl-CoA-desaturase 1 (SCD1), the enzyme involved in monounsaturated fatty acids synthesis, has a role in several cancers. We previously demonstrated that SCD1 is important in lung cancer stem cells survival and propagation. In this article, we first show, using primary cell cultures from human lung adenocarcinoma, that the effectors of the Hippo pathway, Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ), are required for the generation of lung cancer three-dimensional cultures and that SCD1 knock down and pharmacological inhibition both decrease expression, nuclear localization and transcriptional activity of YAP and TAZ. Regulation of YAP/TAZ by SCD1 is at least in part dependent upon β-catenin pathway activity, as YAP/TAZ downregulation induced by SCD1 blockade can be rescued by the addition of exogenous wnt3a ligand. In addition, SCD1 activation of nuclear YAP/TAZ requires inactivation of the β-catenin destruction complex. In line with the in vitro findings, immunohistochemistry analysis of lung adenocarcinoma samples showed that expression levels of SCD1 co-vary with those of β-catenin and YAP/TAZ. Mining available gene expression data sets allowed to observe that high co-expression levels of SCD1, β-catenin, YAP/TAZ and downstream targets have a strong negative prognostic value in lung adenocarcinoma. Finally, bioinformatics analyses directed to identify which gene combinations had synergistic effects on clinical outcome in lung cancer showed that poor survival is associated with high co-expression of SCD1, β-catenin and the YAP/TAZ downstream target birc5. In summary, our data demonstrate for the first time the involvement of SCD1 in the regulation of the Hippo pathway in lung cancer, and point to fatty acids metabolism as a key regulator of lung cancer stem cells.

34 PRODUCTION OF TRANSGENIC CLONED BUFFALO EMBRYOS CONTAINING OVEREXPRESSED STEAROYL Co-A DESATURASE GENE FOLLOWING EFFICIENT TRANSFECTION

Stearoyl-CoA desaturase (SCD) is a rate-limiting enzyme that catalyses the synthesis of monounsaturated fatty acids and polyunsaturated fatty acids from saturated fatty acids, which are components of triglycerides, wax esters, cholesteryl esters, and membrane phospholipids. Alterations in phospholipid composition have been implicated in a variety of diseases including obesity and the associated metabolic syndrome. SCD also magnifies the conjugated linoleic acid (CLA) content in milk; CLA is a natural fat element, having reputed therapeutic health values including anti-carcinogenic properties. In light of this fact, this study was designed to amplify the levels of SCD at the gene level. In order to achieve the enhanced expression of SCD gene, combination of techniques were used. The somatic cells (fetal fibroblast) culture were established from ear pinnae obtained from bovine fetus procured from the abattoir and were cultured in basal medium, comprising DMEM with 10% fetal bovine serum, 1X NEAA and 1X PS antibiotics. These isolated cultured cells were transfected with a gene construct carrying buSCD gene (pAcGFPN1-buSCD) as BLGP-buSCD-BLG3′UTR-CMVP-EGFP-SV40. The buffalo fetal fibroblast cells were transfected using 3 methods: Nucleofection, Fugene and Lipofection. The successful transfection, as confirmed by PCR and Southern hybridisation, proved Nucleofection to be more efficient in transfecting cells among the techniques used, which were further maintained and selected by Geneticin (G418). These selected transfected cells were then used for nuclear transfer. Somatic cell nuclear transfer (SCNT) has provided an efficient pathway for the production of transgenic animals. Buffalo cumulus–oocyte complexes (COCs) were collected from ovaries collected from abattoir and matured in TCM-199 supplemented with 10% FBS, 5 µg mL−1 FSH, and 1 µg mL−1 β-oestradiol for 21 h in CO2 incubator at 5% CO2 in air and 38.5°C temperature with &gt;95% relative humidity. After 21 h, these COCs were denuded and subjected to zona removal. These zona-free oocytes were manually enucleated using microsurgical blades and 2 enucleated oocytes were fused with a transgenic cell using electro cell manipulator. Further, these reconstructed embryos were activated using calcium ionophore and cultured in IVC media thereafter for 8 days. The developmental competence rate as recorded on Day 8 was 53.26 ± 1.73%, 69.87 ± 6.24%, 62.99 ± 7.15%, 42.71 ± 5.05% and 28.00 ± 3.33% for 2-cell, 4-cell, 8–16 cell, morula, and blastocyst, respectively. When observed under fluorescence microscope, the embryos showed successful expression of GFP, which can be further used for animal production or further research analysis. In conclusion, amplified SCD at gene level will result in a boost to the dairy sector as well ameliorating human health due to its crucial role in anti-cancer, anti-diabetic, reduced cardio-vascular disease, and improved immune responses.

Long-chain n-3 PUFA supplied by the usual diet decrease plasma stearoyl-CoA desaturase index in non-hypertriglyceridemic older adults at high vascular risk

The activity of stearoyl-CoA desaturase-1 (SCD1), the central enzyme in the synthesis of monounsaturated fatty acids (MUFA), has been associated with de novo lipogenesis. In experimental models SCD1 is down-regulated by polyunsaturated fatty acids (PUFA), but clinical studies are scarce. The effect of long-chain n-3 PUFA (LCn-3PUFA) supplied by the regular diet, in the absence of fatty fish or fish oil supplementation, remains to be explored. We related 1-y changes in plasma SCD1 index, as assessed by the C16:1n-7/C16:0 ratio, to both adiposity traits and nutrient intake changes in a sub-cohort (n=243) of non-hypertriglyceridemic subjects of the PREDIMED (PREvención con DIeta MEDiterranea) trial. After adjustment for confounders, including changes in fasting triglycerides, plasma SCD1 index increased in parallel with body weight (0.221 [95% confidence interval, 0.021 to 0.422], P=0.031) and BMI (0.115 [0.027 to 0.202], P=0.011). Additionally, dietary LCn-3PUFA (but not MUFA or plant-derived PUFA) were associated with decreased plasma SCD1 index (-0.544 [-1.044 to-0.043], P=0.033, for each 1g/d-increase in LCn-3PUFA). No associations were found for other food groups, but there was a trend for fatty fish intake (-0.083 [-0.177 to 0.012], P=0.085, for each 10g/d-increase). Our data add clinical evidence on the down-regulation of plasma SCD1 index by LCn-3PUFA in the context of realistic changes in fish consumption in the customary, non-supplemented diet. http://www.Controlled-trials.com/ISRCTN35739639.

SCD1 deficiency protects mice against ethanol-induced liver injury

Stearoyl-CoA desaturase 1 (SCD1) is a delta-9 fatty acid desaturase that catalyzes the synthesis of mono-unsaturated fatty acids (MUFA). SCD1 is a critical control point regulating hepatic lipid synthesis and β-oxidation. Scd1 KO mice are resistant to the development of diet-induced non-alcoholic fatty liver disease (NAFLD). Using a chronic-binge protocol of ethanol-mediated liver injury, we aimed to determine if these KO mice are also resistant to the development of alcoholic fatty liver disease (AFLD).Mice fed a low-fat diet (especially low in MUFA) containing 5% ethanol for 10days, followed by a single ethanol (5g/kg) gavage, developed severe liver injury manifesting as hepatic steatosis. This was associated with an increase in de novo lipogenesis and inflammation. Using this model, we show that Scd1 KO mice are resistant to the development of AFLD. Scd1 KO mice do not show accumulation of hepatic triglycerides, activation of de novo lipogenesis nor elevation of cytokines or other pro-inflammatory markers. Incubating HepG2 cells with a SCD1 inhibitor induced a similar resistance to the effect of ethanol, confirming a role for SCD1 activity in mediating ethanol-induced hepatic injury.Taken together, our study shows that SCD1 is a key player in the development of AFLD and associated deleterious effects, and suggests SCD1 inhibition as a therapeutic option for the treatment of this hepatic disease.