Expression Of stearoyl-CoA Desaturase Research Articles

Targeting stearoyl-CoA desaturase 1 to repress endometrial cancer progression.

Stearoyl-CoA desaturase 1 (SCD1) is an established molecular target in many primary tumors including breast, lung, pancreatic, colon and hepatocellular carcinomas. However, its potential role in supporting endometrial cancer growth and progression has not yet been determined. In this study, we evaluated the value of SCD1 as a candidate therapeutic target in human endometrial cancer. Compared with secretory and post-menopausal endometrium, SCD1 was highly expressed in normal endometrium of proliferative phase, endometrial hyperplasia and endometrial carcinoma, while was absent or low expression in non-malignant control stromal cells and adjacent normal endometrium. Knockdown of SCD1 significantly repressed endometrial cancer cell growth and induced cell apoptosis. Both short hairpin RNA targeted knockdown and chemical inhibitor of SCD1 suppressed the foci formation of AN3CA, a metastatic endometrial cell line. Xenograft model further demonstrated that reduced SCD1 expression impaired endometrial cancer growth in vivo. Taken together, these findings indicate that SCD1 is a potentially therapeutic target in human endometrial cancer. Inhibiting lipid metabolism in cancer cells would be a promising strategy for anti-cancer therapy.

Lung fibroblasts promote metastatic colonization through upregulation of stearoyl-CoA desaturase 1 in tumor cells.

As a rate-limiting step in metastasis, metastatic colonization requires survival signals from supportive stroma. However, the mechanisms driving this process are incompletely understood. Here, we showed that the proliferation of B16F10 cells was promoted when cocultured with lung fibroblasts. Meanwhile, co-injection of B16F10 tumor cells with mouse lung fibroblasts significantly increased lung metastasis. Based on GEO database, we identified stearoyl-CoA desaturase 1 (SCD1) as a novel factor promoting metastatic colonization. Importantly, we found that fibroblast-secreted cathepsin B (CTSB) induced the upregulation of SCD1 in B16F10 through Annexin A2 (ANXA2) and PI3K/Akt/mTOR pathway. The elevated SCD1 induced a higher ratio of monounsaturated fatty acids to saturated fatty acids in B16F10 cells. The changes in fatty acid composition contributed to tumor cell proliferation and metastatic colonization. Furthermore, targeting SCD1 effectively inhibited lung metastasis and prolonged the overall survival of mice. Meanwhile, the expression of SCD1 was negatively correlated with disease-free survival in five types of cancer patients. Collectively, our study identifies SCD1 as a critical modulator of tumor cell proliferation that is activated by cathepsin B, secreted by lung fibroblasts at the metastatic niche. Our novel findings provide potential therapeutic targets to prevent tumor metastasis.

Peroxisome proliferator-activated receptor gamma (PPARγ) agonist fails to overcome trans-10, cis-12 conjugated linoleic acid (CLA) inhibition of milk fat in dairy sheep

The trans-10, cis-12 conjugated linoleic acid (CLA) causes milk fat depression by downregulating expression of genes and transcription factors involved in lipogenesis and it has been proposed that peroxisome proliferator-activated receptor gamma (PPARγ) can be inhibited by trans-10, cis-12 CLA. The PPARγ is a nuclear receptor activated by natural or synthetic ligands and promotes expression of lipogenic genes and its effect on mammary lipogenesis and the interaction with trans-10, cis-12 CLA in lactating ewes was evaluated using thiazolidinedione (TZD), a chemical PPARγ agonist. A total of 24 lactating ewes were randomly assigned to one of the following treatments for 7 days: (1) Control (5 ml/day of saline solution); (2) TZD (4 mg/kg of BW/day in 5 ml of saline solution); (3) CLA (27 g/day with 29.9% of trans-10, cis-12); (4) TZD+CLA. Compared with Control, milk fat content was not changed by TZD, but was decreased 22.3% and 20.5% by CLA and TZD+CLA treatments. In the mammary gland, TZD increased PPARγ gene expression by 174.8% and 207.8% compared with Control and TZD+CLA treatments, respectively. Conjugated linoleic acid reduced sterol regulatory element-binding transcription protein 1 (SREBP1) gene expression 89.2% and 75.3% compared with Control and TZD+CLA, respectively, demonstrating that TZD fails to overcome CLA inhibition of SREBP1 signaling. In adipose tissue, the expression of SREBP1 and stearoyl CoA desaturase 1 (SCD1) genes were increased by the TZD+CLA treatment, compared with the other treatments. Conjugated linoleic acid decreased milk fat concentration and expression of lipogenic genes, while TZD had no effect on milk fat concentration, expression of lipogenic enzymes or regulators in the mammary gland and failed to overcome the inhibition of these by CLA. Therefore, CLA inhibition of milk fat synthesis was independent of the PPARγ pathway in lactating dairy ewes.

Epigenetic mechanisms contribute to decrease stearoyl-CoA desaturase 1 expression in the liver of dairy cows after prolonged feeding of high-concentrate diet

Subacute ruminal acidosis (SARA) of dairy cattle is a widely occurring but not very overt metabolic disorder thought to impair milk composition. The enzyme stearoyl-CoA desaturase 1 (SCD1) is rate-limiting for the formation of Δ-9 unsaturated fatty acids and thus crucially involved in controlling lipid metabolism in the liver. It is known that SCD1 expression is downregulated during SARA, but the underlying molecular mechanisms are unknown. To study these mechanisms, we enrolled 12 healthy multiparous mid-lactation Holstein cows into a diet-induced SARA experiment. Six cows were fed a high-concentrate diet for 18 weeks (60% content of high-concentrate to 40% forage; HC group), whereas the others received a low-concentrate diet ad libitum (40% high-concentrate content to 60% forage; LC group). Sustained low ruminal pH values (pH 5.6 maintained for 4 h/d) and reduced milk yield performance (2.07 kg/d less than LC cows) verified that SARA had been induced in the HC group. Results showed a significantly decreased concentrations of cis-9 monounsaturated long-chain fatty acids in plasma collected from hepatic but not portal veins. This was matched by reduced SCD1 mRNA and protein concentrations in HC livers. The expression levels of genes related to lipid formation (DGAT1 and PLIN2) were downregulated during SARA, whereas those of catabolic genes (CPT1A, CPT2, and ACOX1) and some inflammatory genes were upregulated. Expression of SCD1 was downregulated through reduced transcription and abundance of the transcription factor sterol regulatory element-binding protein 1 (SREBP1c).This effect was augmented by local chromatin tightening and DNA methylation at and around the SREBP1c binding site in the SCD1 promoter. Chromatin immunoprecipitation assays confirmed that SARA reduced SREBP1c binding at the SCD1 promoter; hence, epigenetic mechanisms are involved in regulating the expression of genes related to long-chain fatty acid modification, partially through downregulation of both SCD1 and SREBP1c in the liver. Our results suggest that in addition to inflammatory genes, SCD1 is also involved in SARA-induced epigenetic regulation and its associated metabolic changes. This knowledge might help to provide a target for intervening against the detrimental metabolic effects of SARA.

Carcass traits, fatty acid composition, gene expression, oxidative stability and quality attributes of different muscles in Dorper lambs fed Nigella sativa seeds, Rosmarinus officinalis leaves and their combination.

ObjectiveThis study examined the influence of dietary supplementation of Nigella sativa seeds, Rosmarinus officinalis leaves and their combination on carcass attributes, fatty acid (FA) composition, gene expression, lipid oxidation and physicochemical properties of longissimus dorsi (LD), semitendinosus (ST), and supraspinatus (SS) muscles in Dorper lambs.MethodsTwenty-four Dorper lambs (18.68±0.6 kg, 4 to 5 months old) were randomly assigned to a concentrate mixture containing either, no supplement (control, T1), 1% Rosmarinus officinalis leaves (T2), 1% Nigella sativa seeds (T3), or 1% Rosmarinus officinalis leaves+1% Nigella sativa seeds (T4) on a dry matter basis. The lambs were fed the treatments with urea-treated rice straw for 90 days, slaughtered and the muscles were subjected to a 7 d postmortem chill storage.ResultsThe T2 lambs had greater (p<0.05) slaughter and cold carcass weights than the control lambs. Dietary supplements did not affect (p>0.05) chill loss, dressing percentage, carcass composition, intramuscular fat and muscle pH in Dorper lambs. Meat from supplemented lambs had lower (p<0.05) cooking and drip losses, shear force, lightness, and lipid oxidation and greater (p<0.05) redness compared with the control meat. The impact of dietary supplements on muscle FA varied with muscle type. Diet had no effect (p>0.05) on the expression of stearoyl-CoA desaturase and lipoprotein lipase genes in LD and ST muscles in Dorper lambs. The T2 and T3 diets up regulated the expression of AMP-activated protein kinase alpha 2 gene in LD and ST muscles and up regulated the expression of sterol regulatory element-binding protein 1 in ST muscle in Dorper lambs.ConclusionDietary supplementation of Nigella sativa seeds and Rosmarinus officinalis leaves had beneficial effects on meat quality in Dorper lambs.

GLP-1 Elicits an Intrinsic Gut–Liver Metabolic Signal to Ameliorate Diet-Induced VLDL Overproduction and Insulin Resistance

Perturbations in hepatic lipid and very-low-density lipoprotein (VLDL) metabolism are involved in the pathogenesis of obesity and hepatic insulin resistance. The objective of this study is to delineate the mechanism of subdiaphragmatic vagotomy in preventing obesity, hyperlipidemia, and insulin resistance. By subjecting the complete subdiaphragmatic vagotomized mice to various nutritional conditions and investigating hepatic de novo lipogenesis pathway, we found that complete disruption of subdiaphragmatic vagal signaling resulted in a significant decrease of circulating VLDL-triglyceride compared with the mice obtained sham procedure. Vagotomy further prevented overproduction of VLDL-triglyceride induced by an acute fat load and a high-fat diet-induced obesity, hyperlipidemia, hepatic steatosis, and glucose intolerance. Mechanistic studies revealed that plasma glucagon-like peptide-1 was significantly raised in the vagotomized mice, which was associated with significant reductions in mRNA and protein expression of SREBP-1c (sterol regulatory element-binding protein 1c), SCD-1 (stearoyl-CoA desaturase-1), and FASN (fatty acid synthase), as well as enhanced hepatic insulin sensitivity. In vitro, treating mouse primary hepatocytes with a glucagon-like peptide-1 receptor agonist, exendin-4, for 48 hours inhibited free fatty acid, palmitic acid treatment induced de novo lipid synthesis, and VLDL secretion from hepatocytes. Elevation of glucagon-like peptide-1 in vagotomized mice may prevent VLDL overproduction and insulin resistance induced by high-fat diet. These novel findings, for the first time, delineate an intrinsic gut-liver regulatory circuit that is mediated by glucagon-like peptide-1 in regulating hepatic energy metabolism.

O23-5 - Cardiac Sirt1 Has a Protective Role in Lipotoxic Cardiomyopathy through Maintaining Membrane Fatty Acid Composition

Backgrounds: Lipotoxicity causes cardiac dysfunction in diabetes and obese. One potential mechanism is the effects of membrane fatty acid composition. Sirt1 is activated by exercise and calorie restriction, both of which have proved to be effective for lipotoxicity. We investigated a role of Sirt1 in the homeostasis of membrane fatty acid composition and cardiac function in lipotoxic cardiomyopathy. Methods and Results: (1) In neonatal rat cardiomyocytes, cell death induced by palmitate, a saturated fatty acid, was exacerbated in Sirt1-knockdown (Sirt1 KD) and was ameliorated by Sirt1 activator (nicotinamide mononucleotide). (2) Palmitate-overload changed membrane fatty acid composition; it increased palmitate and decreased oleate, a monounsaturated fatty acid. Such a change in membrane fatty acid composition was exaggerated in Sirt1 KD. (3) The expression of stearoyl-CoA desaturase 1 (SCD1), which converts saturated fatty acid to monounsaturated fatty acid, was increased by palmitate. This SCD1 induction was not observed in Sirt1 KD. (4) SCD1 overexpression could rescue both palmitate-induced membrane fatty acid composition change and cell death in Sirt1 KD. (5) Cardiomyocyte-specific Sirt1-knockout mice fed a high fat diet showed more severe diastolic dysfunction with an increase in cardiomyocyte death, fibrosis, and hypertrophy compared with control mice. Conclusions: Cardiac Sirt1 confers resistance against lipotoxic cardiomyopathy by preventing an increase in membrane saturated fatty acid content via the induction of SCD1.

Zinc mediates the SREBP-SCD axis to regulate lipid metabolism in Caenorhabditis elegans

Maintenance of lipid homeostasis is crucial for cells in response to lipid requirements or surplus. The SREBP transcription factors play essential roles in regulating lipid metabolism and are associated with many metabolic diseases. However, SREBP regulation of lipid metabolism is still not completely understood. Here, we showed that reduction of SBP-1, the only homolog of SREBPs in Caenorhabditis elegans, surprisingly led to a high level of zinc. On the contrary, zinc reduction by mutation of sur-7, encoding a member of the cation diffusion facilitator (CDF) family, restored the fat accumulation and fatty acid profile of the sbp-1(ep79) mutant. Zinc reduction resulted in iron overload, which thereby directly activated the conversion activity of stearoyl-CoA desaturase (SCD), a main target of SREBP, to promote lipid biosynthesis and accumulation. However, zinc reduction reversely repressed SBP-1 nuclear translocation and further downregulated the transcription expression of SCD for compensation. Collectively, we revealed zinc-mediated regulation of the SREBP-SCD axis in lipid metabolism, distinct from the negative regulation of SREBP-1 or SREBP-2 by phosphatidylcholine or cholesterol, respectively, thereby providing novel insights into the regulation of lipid homeostasis.

Blockade of Stearoyl-CoA-desaturase 1 activity reverts resistance to cisplatin in lung cancer stem cells

Poor prognosis in lung cancer has been attributed to the presence of lung cancer stem cells (CSCs) which resist chemotherapy and cause disease recurrence. Hence, the strong need to identify mechanisms of chemoresistance and to develop new combination therapies. We have previously shown that Stearoyl-CoA-desaturase 1 (SCD1), the enzyme responsible for the conversion of saturated to monounsaturated fatty acids is upregulated in 3D lung cancer spheroids and is an upstream activator of key proliferation pathways β-catenin and YAP/TAZ. Here we first show that SCD1 expression, either alone or in combination with a variety of CSCs markers, correlates with poor prognosis in adenocarcinoma (ADC) of the lung. Treatment of lung ADC cell cultures with cisplatin enhances the formation of larger 3D tumor spheroids and upregulates CSCs markers. In contrast, co-treatment with cisplatin and the SCD1 inhibitor MF-438 reverts upregulation of CSCs markers, strongly synergizes in the inhibition of 3D spheroids formation and induces CSCs apoptosis. Mechanistically, SCD1 inhibition activates endoplasmic reticulum stress response and enhances autophagy. These data all together support the use of combination therapy with SCD1 inhibitors to achieve better control of lung cancer.

Peripheral neuropeptide Y differentially influences adipogenesis and lipolysis in chicks from lines selected for low or high body weight

Neuropeptide Y (NPY) stimulates appetite and promotes lipid deposition. We demonstrated a differential sensitivity in the food intake response to central NPY in chicks from lines selected for low (LWS) or high (HWS) body weight, but have not reported whether such differences exist in the periphery. At 5days, LWS and HWS chicks were intraperitoneally injected with 0 (vehicle), 60, or 120μg/kg BW NPY and subcutaneous adipose tissue and plasma were collected at 1, 3, 6, 12, and 24h (n=12). NPY injection increased glycerol-3-phosphate dehydrogenase (G3PDH) activity at 1 and 3h and reduced plasma non-esterified fatty acids (NEFAs) at 1 and 12h. G3PDH activity was greater in HWS than LWS while NEFAs were greater in LWS. At 1h, peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer binding protein (C/EBP)α, and microsomal triglyceride transfer protein (MTTP) mRNAs were reduced in NPY-injected chicks whereas NPY receptor 1 (NPYR1) was increased. Expression of stearoyl-CoA desaturase (SCD1) was increased by NPY at 1h in HWS but not LWS. PPARγ (3 and 6h), C/EBPβ (3h), C/EBPα (6h) and NPYR1 and 2 (24h) mRNAs were greater in NPY- than vehicle-injected chicks. At several times, adipose triglyceride lipase, MTTP, perilipin 1, NPYR1, and NPYR2 mRNAs were greater in LWS than HWS, while expression of SCD1, glycerol-3-phosphate acyltransferase 3 and lipoprotein lipase was greater in HWS than LWS. Thus, NPY promotes fat deposition and inhibits lipolysis in chicks, with line differences indicative of greater rates of lipolysis in LWS and adipogenesis in HWS.

Dihydrosterculic acid from cottonseed oil suppresses desaturase activity and improves liver metabolomic profiles of high-fat–fed mice

Polyunsaturated fatty acid (PUFA)–rich diets are thought to provide beneficial effects toward metabolic health in part through their bioactive properties. We hypothesized that increasing PUFA intake in mice would increase peroxisome proliferator activated receptor delta (PPARδ) expression and activity, and we sought to examine the effect of different PUFA-enriched oils on muscle PPARδ expression. One of the oils we tested was cottonseed oil (CSO) which is primarily linoleic acid (53%) and palmitic acid (24%). Mice fed a CSO-enriched diet (50% energy from fat) displayed no change in muscle PPARδ expression; however, in the liver, it was consistently elevated along with its transcriptional coactivator Pgc-1. Male mice were fed chow or CSO-, saturated fat (SFA)–, or linoleic acid (18:2)–enriched diets that were matched for macronutrient content for 4 weeks. There were no differences in food intake, body weight, fasting glucose, glucose tolerance, or energy expenditure between chow- and CSO-fed mice, whereas SFA-fed mice had increased fat mass and 18:2-fed mice were less glucose tolerant. Metabolomic analyses revealed that the livers of CSO-fed mice closely matched those of chow-fed but significantly differed from SFA- and 18:2-enriched groups. Fatty acid composition of the diets and livers revealed an impairment in desaturase activity and the presence of dihydrosterculic acid (DHSA) in the CSO-fed mice. The effect of DHSA on PPARδ and stearoyl-CoA desaturase-1 expression mimicked that of the CSO-fed mice. Taken together, these data suggest that DHSA from CSO may be an effective means to increase PPARδ expression with concomitant suppression of liver stearoyl-CoA desaturase-1 activity.

Abstract 4772: Stearoyl-CoA Desaturase (SCD1) regulates liver tumor initiating cells through modulating ER stress

Abstract Sorafenib is the FDA approved drug for patients with advanced hepatocellular carcinoma (HCC), but the survival benefit is only modest, partly due to drug resistance. Increasing evidence shows that tumor-initiating cells (T-ICs) are intrinsically resistant to conventional treatments, and targeting signaling pathways in T-ICs provides potential therapeutic targets for HCC. In our established enriched T-IC population, we identified the activation of a lipogenesis pathway in which expression of stearoyl-CoA desaturase-1 (SCD1), an enzyme involved in the conversion of saturated into monounsaturated fatty acids, was most significant. SCD1 overexpression is frequently found in HCC samples and is significantly associated with poorer patient survival. Using overexpression and knockdown approaches, SCD1 was found to regulate the traits of T-ICs, including tumorigenicity, self-renewal, differentiation, drug resistance and the expression of liver T-IC markers. Interestingly, SCD1 was markedly upregulated in our established sorafenib-resistant cell lines as well as patient-derived xenografts (PDTXs), and its overexpression predicts the clinical response of HCC patients to sorafenib treatment. Consistently, pharmacological inhibition of SCD1 suppressed T-IC phenotypes and enhanced sensitivity to sorafenib treatment. Using a patient-derived xenograft model, we found that a novel SCD1 inhibitor (SSI-4) in combination with sorafenib demonstrated a maximal tumor suppressive effect. Induction of endoplasmic reticulum (ER) stress-mediated T-IC differentiation can account for the enhanced sensitivity towards sorafenib treatment upon SCD1 suppression. In conclusion, SCD1-induced ER stress may specifically increase the sensitivity of liver T-ICs to the effects of sorafenib treatment. Targeting SCD1 in combination with sorafenib therapy may be a novel therapeutic regimen against HCC. Citation Format: Kin Fai Ma, Eunice Yuen Ting Lau, Irene Oi Lin Ng, Kin Wah Lee. Stearoyl-CoA Desaturase (SCD1) regulates liver tumor initiating cells through modulating ER stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4772. doi:10.1158/1538-7445.AM2017-4772

Abstract 192: Targeting stearoyl CoA desaturase 1 (SCD1) in hepatobilliary carcinoma

Abstract Background: The paucity of effective therapeutic agents for hepatocellular cancer (HCC) underscores the critical need for more effective therapeutic strategies. Recent studies indicate lipid biosynthesis and desaturation is required for HCC survival. Targeting these may prove beneficial because such changes contribute to therapeutic resistance. Stearoyl CoA desaturase (SCD1), a key mediator of fatty acid (FA) biosynthesis and rate-limiting in conversion of saturated fatty acids (SFAs) to mono-unsaturated fatty acids (MUFAs), is upregulated in HCC and many other cancers. As such, we therapeutically targeted a novel lipogenic tumor survival mechanism mediated by SCD1 as a means to combat the chemoresistance associated with HCC. In so doing, we evaluated a novel lead SCD1 inhibitor in HCC. Methods: Paraffin embedded patient HCC tissues were examined for SCD1 expression. Using combined computational and synthetic chemistry approaches, we synthesized four novel specific SCD1 inhibitors with SSI-4 being the lead SCD1 inhibitor. HCC cell lines were examined using proliferation assays for response to SSI-4. IC50 concentrations for blocking SCD1 enzyme activity was determined. Blood half-life and bioavailability of single dose SSI-4 was determined. Mechanisms of action of SCD1 were examined that included Endoplasmic reticulum (ER) stress. In vivo, antitumor activity was determined using HCC patient derived xenograft (PDX) mouse models. Results: We identified elevated SCD1 mRNA and protein in HCCs tissues. SSI-4 dose-dependently inhibits cell proliferation in HCC cell lines with specificity demonstrated by oleic acid (MUFA) co-culture. Single dose oral gavage SSI-4 demonstrated a half-life of ~4 hours and excellent oral bioavailability. SSI-4 was well tolerated with long-term daily dosing. SSI-4 treatment of HCC cells and tumors led to endoplasmic reticulum (ER) stress followed by apoptotic cell death. Single agent SSI-4 demonstrated antitumor activity in HCC PDX mouse models with suppression of ER stress regulated proteins. Conclusions: Targeting a novel lipid metabolic pathway in HCC may provide effective therapy for aggressive HCC. Citation Format: John Alton Copland, Laura A. Marlow, Ilah Bok, James L. Miller, Matsuda Akiko, Yan W. Asmann, Vivekananda Sarangi, Steven R. Alberts, Kabir Mody, Lewis R. Roberts, Mark J. Truty, Tushar C. Patel. Targeting stearoyl CoA desaturase 1 (SCD1) in hepatobilliary carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 192. doi:10.1158/1538-7445.AM2017-192

Abstract 2048: Saturated fatty acids in cell membrane phospholipids play an important role in chemoresistance in colorectal cancer

Abstract [Background and Aim] Resistance to chemotherapy represents the major obstacle to survival in patients with chemoresistant colorectal cancer. Various factors, such as drug efflux, autocrine survival signaling, and alterations in DNA damage repair mechanisms, can contribute to chemoresistance; however, the actual underlying mechanism is yet to be elucidated. The aim of this study was to elucidate the mechanism behind anticancer drug resistance based on lipidomics. [Materials and Methods] 1) Spheroids were derived from three colorectal cancer patient-derived xenograft models (A, B, and C) that were incubated with chemotherapeutic agents, such as 5-fluorouracil (5-FU). We analyzed the response rate of spheroids under 5-FU treatment. Global lipid analysis was performed on 5-FU resistance spheroids (A) and sensitive spheroids (C) using a Q Exactive LC-MS/MS mass spectrometer, and lipid identification was performed using the Lipid Search software. 2) We tried to identify genes that were responsible for changes in phospholipid profiles in the cell membrane. 3) To determine phospholipid profile changes in the cell membrane that occur in colon cancer cell lines (HCT116, SW480) during anticancer drug resistance, the genes responsible for such resistance were knocked down using siRNA. [Results and discussion] 1) Response to 5-FU was different in individual spheroids. Global lipid analysis and chemosensitivity assay revealed that phospholipids of 5-FU resistant spheroids were rich in saturated fatty acids (SFAs), while those of the 5-FU sensitive spheroids were rich in monounsaturated fatty acids (MUFAs). 2) The enzyme stearoyl-CoA desaturase (SCD) converts SFAs to MUFAs in the phospholipids. Based on these findings we hypothesized that decreased SCD expression is possibly associated with chemoresistance. 3) Cell lines with SCD knock down demonstrated increased SFA containing phospholipids and chemoresistance towards 5-FU. Viable cells after the administration of the anti-cancer agent confirmed that the expression of CD44 was upregulated. Therefore, changes in the phospholipid composition of cell membranes regulate 5-FU resistance in colon cancer. [Conclusions] Identification and analysis of the role of membrane lipids has become possible using high-sensitivity and high-resolution mass spectrometers and the relevant software, such as Lipid Search. Changes of phospholipid composition regulates chemoresistance. In future, investigation of the pathophysiology of membrane lipids in cancer may help to reveal novel strategies for cancer therapy. Citation Format: Takanori Hiraide, Takanori Sakaguchi, Satoru Furuhashi, Tomohiro Matsumoto, Yusuke Ozaki, Ryota Kiuchi, Makoto Takeda, Yasushi Shibasaki, Yoshifumi Morita, Hirotoshi Kikuchi, Megumi Baba, Mitsutoshi Setou, Hiroyuki Konno. Saturated fatty acids in cell membrane phospholipids play an important role in chemoresistance in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2048. doi:10.1158/1538-7445.AM2017-2048

Abstract 4331: Novel Wnt-SCD-LRP5/6 pathway linking liver fibrosis to cancer

Abstract Activated hepatic stellate cells (HSCs) are primarily responsible for the genesis of liver fibrosis which promotes liver tumor development. Stearoyl-CoA desaturase (SCD) which synthesizes monounsaturated fatty acid (MUFA such as oleic acid or palmitoleic acid), is implicated in metabolic syndrome, tumorigenesis and stemness. Indeed, we demonstrate SCD upregulation in both HSCs and liver tumor cells in patients. Further, SCD correlates with the advancing grade of HCC and mortality in patients. However, its causality in tumorigenesis and mesenchyme-tumor crosstalk, and underlying mechanisms, are unknown. The present study aimed to determine whether and how SCD promotes and links liver fibrosis and cancer. In both HSCs and liver tumor-initiating stem cell-like cells (TIC), we reveal that the Wnt effector β-catenin is a potent co-activator for SREBP-1-dependent Scd transcription as demonstrated by promoter-reporter assay and re-ChIP analysis. We also discover β-catenin which drives SCD expression is in turn stabilized by SCD-derived MUFA. To identify molecular targets of MUFA mediating this SCD-β-catenin positive loop, we performed MUFA-nano-beads pull-down assay with TIC lysate, ESI-MS based identification of MUFA interacting proteins, computational docking analysis, co-IP and ribonucleoprotein-IP. These analyses reveal that MUFA is essential for LRP5/6 expression: MUFA interferes Transportin-1 and Ran-1 interaction, resulting in suppressed nuclear import of the mRNA binding protein HuR, HuR cytoplasmic accumulation, and increased HuR binding to and stabilization of Wnt co-receptor Lrp5/6 mRNA. As such, genetic or pharmacologic SCD inhibition reduces cytosolic HuR, LRP5/6 expression, β-catenin stabilization, HSC activation and TIC self-renewal and attenuates liver fibrosis and tumorigenesis in mice. Moreover, conditional genetic ablation of Scd2 in HSCs not only abrogates HSC-mediated promotion of TIC-derived tumorigenesis in a xenograft mouse model but also inhibits DEN-induced liver tumor development in Col1a1-Cre; Scd2flox/flox mice (tumor volume reduced to 21.5 + 7.9 vs. 360.9 + 193 mm3 in Scd2flox/flox mice; tumor multiplicity decreased to 18 + 5.4 vs. 45 + 8.4 in Scd2flox/flox mice, both p&amp;lt;0.05). Collectively, our findings support that a newly disclosed Wnt-SCD-LRP5/6 loop may serve as a novel therapeutic target for liver cancer and underlies a known crosstalk between liver mesenchyme and liver tumor development. Citation Format: Keane Lai, Soo-Mi Kweon, Feng Chi, Edward Hwang, Raymond Wu, Yasuaki Kabe, Ramachandran Murali, Lopa Mishra, James Ntambi, Hidekazu Tsukamoto. Novel Wnt-SCD-LRP5/6 pathway linking liver fibrosis to cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4331. doi:10.1158/1538-7445.AM2017-4331

MiR-212-5p suppresses lipid accumulation by targeting FAS and SCD1.

MicroRNAs, a class of small noncoding RNAs, are implicated in controlling a variety of biological processes. We have shown that leucine deprivation suppresses lipogenesis by inhibiting fatty acid synthase (FAS) expression in the liver previously; the aim of our current study is to investigate which kind of microRNA is involved in the regulation of FAS expression in response to leucine deprivation. Here, we indicated that microRNA-212-5p specifically binds to mouse FAS 3'UTR and inhibits its activity. Leucine deficiency significantly increased the mRNA levels of miR-212-5p in the livers of mice. Further studies proved that miR-212-5p also directly binds to the 3'UTR of stearoyl-CoA desaturase-1 (SCD1) to inhibit its activity. Overexpression of miR-212-5p decreases the protein levels of FAS and SCD1 in vitro and in vivo, and silencing of miR-212-5p has the opposite effects in mouse primary hepatocytes. Moreover, overexpression of miR-212-5p significantly decreases triglyceride (TG) accumulation in primary hepatocytes and in the livers of mice injected with adenovirus-mediated overexpressing of miR-212-5p (Ad-miR-212). Interestingly, inhibition of miR-212-5p reverses the suppressive effects of leucine deficiency on FAS and SCD1 expression, as well as TG accumulation in mouse primary hepatocytes. Finally, we demonstrate that leucine deficiency induces the expression of miR-212-5p in a GCN2/ATF4-dependent manner. Taken together, our results demonstrate a novel function of hepatic miR-212-5p in the regulation of lipid metabolism which represents a potential therapeutic target for the treatment of non-alcohol fatty liver diseases (NAFLD).

Altered expression of epidermal lipid bio-synthesis enzymes in atopic dermatitis skin is accompanied by changes in stratum corneum lipid composition

BackgroundThe barrier dysfunction in atopic dermatitis (AD) skin correlates with stratum corneum (SC) lipid abnormalities including reduction of global lipid content, shorter ceramide (CER) as well as free fatty acid (FFA) chain length and altered CER subclass levels. However, the underlying cause of these changes in lipid composition has not been fully investigated. AimWe investigated whether the expression of CER and FFA biosynthesis enzymes are altered in AD skin compared with control skin and determine whether changes in enzyme expression can be related with changes in lipid composition. MethodsIn AD patients and controls the expression of enzymes involved in the biosynthesis of FFAs and CERs was analyzed in relation to the SC lipid composition. These enzymes include stearoyl CoA desaturase (SCD), elongase 1 (ELOVL1) and ELOVL6 involved in FFA synthesis and β-glucocerebrosidase (GBA), acid-sphingomyelinase (aSmase), ceramide synthase 3 (CerS3) involved in CER synthesis. In TH2 treated human skin equivalents (AD HSEs) mimicking lesional AD skin, the mRNA expression of these enzymes was investigated. ResultsThe results reveal an altered expression of SCD and ELOVL1 in AD lesional skin. This was accompanied by functional changes displayed by increased unsaturated FFAs (SCD) and reduced FFA C22-C28 (ELOVL1) in AD lesional skin. The expression of GBA, aSmase and CerS3 were also altered in lesional skin. The CER composition in AD lesional skin showed corresponding changes such as increased CER AS and NS (aSmase) and decreased esterified ω-hydroxy CERs (CerS3). In support of the results from AD skin, the AD HSEs showed reduced mRNA ELOVL1, GBA and a Smase levels. ConclusionThis study shows that alterations in the expression of key enzymes involved in SC lipid synthesis contribute to changes in the lipid composition in AD skin and inflammation may influence expression of these enzymes.

Changes in LXR signaling influence early-pregnancy lipogenesis and protect against dysregulated fetoplacental lipid homeostasis.

Human pregnancy is associated with enhanced de novo lipogenesis in the early stages followed by hyperlipidemia during advanced gestation. Liver X receptors (LXRs) are oxysterol-activated nuclear receptors that stimulate de novo lipogenesis and also promote the efflux of cholesterol from extrahepatic tissues followed by its transport back to the liver for biliary excretion. Although LXR is recognized as a master regulator of triglyceride and cholesterol homeostasis, it is unknown whether it facilitates the gestational adaptations in lipid metabolism. To address this question, biochemical profiling, protein quantification, and gene expression studies were used, and gestational metabolic changes in T0901317-treated wild-type mice and Lxrab−/− mutants were investigated. Here, we show that altered LXR signaling contributes to the enhanced lipogenesis in early pregnancy by increasing the expression of hepatic Fas and stearoyl-CoA desaturase 1 (Scd1). Both the pharmacological activation of LXR with T0901317 and the genetic ablation of its two isoforms disrupted the increase in hepatic fatty acid biosynthesis and the development of hypertriglyceridemia during early gestation. We also demonstrate that absence of LXR enhances maternal white adipose tissue lipolysis, causing abnormal accumulation of triglycerides, cholesterol, and free fatty acids in the fetal liver. Together, these data identify LXR as an important factor in early-pregnancy lipogenesis that is also necessary to protect against abnormalities in fetoplacental lipid homeostasis.

Influence of temperature and growth selection on turkey pectoralis major muscle satellite cell adipogenic gene expression and lipid accumulation

Immature poults have an inefficient thermoregulatory system, and therefore extreme ambient temperatures can impact their internal body temperature. Satellite cells, the only posthatch myonuclei source, are multipotential stem cells and sensitive to temperature. Selection for faster-growing, high-yielding birds has altered satellite-cell properties. The objective of the current study was to determine how temperature affects adipogenic properties of satellite cells isolated from the pectoralis major (p. major) muscle of Randombred Control line (RBC2) and F line turkeys selected only for increased 16-wk body weight from the RBC2 line. Satellite cells were cultured at 2°C incremental temperatures between 33 and 43°C and compared to cells cultured at the control temperature of 38°C to ascertain temperature effects on lipid accumulation and expression of adipogenic genes: CCAAT/enhancer-binding protein-β (C/EBPβ), peroxisome proliferator-activated receptor-γ (PPARγ), and stearoyl-CoA desaturase (SCD). During proliferation, the amount of quantifiable lipid in both F and RBC2 satellite cells increased at temperatures above 38°C (P < 0.01) and decreased at temperatures below 38°C (P < 0.01). Above 38°C, RBC2 satellite cells had more lipid (P = 0.02) compared to the F line, whereas there were few differences between lines below 38°C. At 72 h of proliferation, expression of C/EBPβ, PPARγ, and SCD decreased (P ≤ 0.02) as temperatures increased from 33 to 43°C in both cell lines. During differentiation expression of C/EBPβ increased (P < 0.01) as temperatures increased from 33 to 43°C in both cell lines. In F line satellite cells, PPARγ expression decreased (P < 0.01) with increasing temperatures during differentiation, whereas there was no linear trend in RBC2 cells. During differentiation expression of SCD increased as temperatures increased (P < 0.01) in RBC2 cells, and there was no linear trend within the F line. Results from the current study suggest that environmental temperature can affect p. major satellite cellular fate; however, selection for increased body weight had little impact on these cellular responses.

Cysteine deprivation prevents induction of peroxisome proliferator-activated receptor gamma-2 and adipose differentiation of 3T3-L1 cells

Plasma cysteine is strongly associated with body fat mass in human cohorts and diets low in cysteine prevents fat accumulation in mice. It is unclear if plasma cysteine affects fat development or if fat accumulation raises plasma cysteine. To determine if cysteine affects adipogenesis, we differentiated 3T3-L1 preadipocytes in medium with reduced cysteine. Cells incubated in media with 10–20μM cysteine exhibited reduced capacity to differentiate into triacylglycerol-storing mature adipocytes compared with cells incubated with 50μM cysteine. Low cysteine severely reduced expression of peroxisome proliferator-activated receptor gamma2 (Pparγ2) and its target genes perlipin1 (Plin1) and fatty acid binding protein-4 (Fabp4). Expression of stearoyl-CoA desaturase-1 (Scd1), known to be repressed with cysteine depletion, was also reduced with low cysteine. Medium depletion of the essential amino acids leucine, valine, and isoleucine had only a modest effect on adipocyte specific gene expression and differentiation. Stimulation with the PPARγ agonist BRL-49653 or addition of a hydrogen sulfide donor enhanced differentiation of 3T3-L1 cells cultured in low cysteine. This demonstrates that the ability to induce PPARγ expression is preserved when cells are cultured in low cysteine. It therefore appears that cysteine depletion inhibits adipogenesis by specifically affecting molecular pathways required for induction of PPARγ expression, rather than through a general reduction of global protein synthesis. In conclusion, we show that low extracellular cysteine reduces adipocyte differentiation by interfering with PPARγ2 and PPARγ target gene expression. Our results provide further evidence for the hypothesis that plasma cysteine is a casual determinant for body fat mass.