Lipogenesis Enzymes Research Articles

Dodecanedioic acid prevents and reverses metabolic-associated liver disease and obesity and ameliorates liver fibrosis in a rodent model of diet-induced obesity.

Dodecanedioic acid (DC12) is a dicarboxylic acid present in protective polymers of fruit and leaves. We explored the effects of DC12 on metabolic dysfunction-associated steatohepatitis (MASH) and obesity. DC12 supplementation (100 mg/kg/day) was added to a high-fat diet (HFD) for 8 weeks in rodents to assess its impact on obesity and MASH prevention. Rats given DC12 experienced significant reductions of weight gain, liver and visceral fat weight, and improved glucose tolerance and insulin sensitivity. Liver histology showed protection against diet-induced MASH, with reduced steatosis, hepatocyte ballooning, and fibrosis. For weight-loss and MASH reversion, rats were fed HFD for 14 weeks, followed by 6 weeks with or without DC12. DC12 supplementation (100 mg/kg/day) led to a significant reduction of weight gain and liver weight. DC12 induced white adipose tissue beiging and reduced adiposity with a decrease of visceral fat. It also improved glucose tolerance, insulin sensitivity, and reduced hepatic gluconeogenic gene expression. Liver histology revealed a significant reduction in steatosis, hepatocyte ballooning, and inflammation as well as fibrosis, indicating MASH reversal. DC12 reduced hepatic lipogenesis enzymes as well as de novo lipogenesis measured by deuterated water and increased fatty acid β-oxidation. Plasma lipid profile showed lower triglycerides and phosphatidylcholines in the DC12 group. Notably, DC12 decreased mINDY expression, the cell membrane Na+-coupled citrate transporter, reducing citrate uptake and de-novo lipogenesis, linking its effects to improved lipid metabolism and reduced steatosis. We found that during the hepatic first pass, half of the DC12 ingested with water was taken up by the liver. The concentration of DC12 in the portal vein falls within the range identified invitro as sufficient to inhibit citrate transport in hepatocytes.

GDF15 is required for maintaining subcutaneous adipose tissue lipid metabolic signature

Recent research has identified growth differentiation factor 15 (GDF15) as a crucial factor in various physiological and pathological processes, particularly in energy balance regulation. While the role of GDF15 in modulating energy metabolism through hindbrain GDNF family receptor alpha-like (GFRAL) signaling has been extensively studied, emerging evidence suggests direct peripheral metabolic actions of GDF15. Using knockout mouse models, we investigated GDF15 and GFRAL’s roles in adipose tissue metabolism. Our findings indicate that C57BL/6/129/SvJ Gdf15-KO mice exhibit impaired expression of de novo lipogenesis enzymes in subcutaneous adipose tissue (sWAT). In contrast, C57BL/6J Gfral-KO mice showed no impairments compared to wild-type (WT) littermates. RNA-Seq analysis of sWAT in Gdf15-KO mice revealed a broad downregulation of genes involved in lipid metabolism. Importantly, our study uncovered sex-specific effects, with females being more affected by GDF15 loss than males. Additionally, we observed a fasting-induced upregulation of GDF15 gene expression in sWAT of both mice and humans, reinforcing this factor’s role in adipose tissue lipid metabolism. In conclusion, our research highlights an essential role for GDF15 in sWAT lipid metabolic homeostasis. These insights enhance our understanding of GDF15’s functions in adipose tissue physiology and underscore its potential as a therapeutic target for metabolic disorders.

The Inhibitory Effects of Maclurin on Fatty Acid Synthase and Adipocyte Differentiation.

Obesity is a complex health condition characterized by excessive adipose tissue accumulation, leading to significant metabolic disturbances such as insulin resistance and cardiovascular diseases. Fatty acid synthase (FAS), a key enzyme in lipogenesis, has been identified as a potential therapeutic target for obesity due to its role in adipocyte differentiation and lipid accumulation. This study employed a multidisciplinary approach involving in silico and in vitro analyses to investigate the anti-adipogenic properties of maclurin, a natural phenolic compound derived from Morus alba. Using SwissDock software (ChEMBL version 23), we predicted protein interactions and demonstrated a high probability (95.6%) of maclurin targeting FAS, surpassing the interaction rates of established inhibitors like cerulenin. Docking simulations revealed maclurin's superior binding affinity to FAS, with a binding score of -7.3 kcal/mol compared to -6.7 kcal/mol for cerulenin. Subsequent in vitro assays confirmed these findings, with maclurin effectively inhibiting FAS activity in a concentration-dependent manner in 3T3-L1 adipocytes, without compromising cell viability. Furthermore, maclurin treatment resulted in significant reductions in lipid accumulation and the downregulated expression of critical adipogenic genes such as PPARγ, C/EBPα, and FAS, indicating the suppression of adipocyte differentiation. Maclurin shows potential as a novel FAS inhibitor with significant anti-adipogenic effects, offering a promising therapeutic avenue for the treatment and prevention of obesity.

Nitrogen limitation-induced adaptive response and lipogenesis in the Antarctic yeast Rhodotorula mucilaginosa M94C9.

The extremotolerant red yeast Rhodotorula mucilaginosa displays resilience to diverse environmental stressors, including cold, osmolarity, salinity, and oligotrophic conditions. Particularly, this yeast exhibits a remarkable ability to accumulate lipids and carotenoids in response to stress conditions. However, research into lipid biosynthesis has been hampered by limited genetic tools and a scarcity of studies on adaptive responses to nutrient stressors stimulating lipogenesis. This study investigated the impact of nitrogen stress on the adaptive response in Antarctic yeast R. mucilaginosa M94C9. Varied nitrogen availability reveals a nitrogen-dependent modulation of biomass and lipid droplet production, accompanied by significant ultrastructural changes to withstand nitrogen starvation. In silico analysis identifies open reading frames of genes encoding key lipogenesis enzymes, including acetyl-CoA carboxylase (Acc1), fatty acid synthases 1 and 2 (Fas1/Fas2), and acyl-CoA diacylglycerol O-acyltransferase 1 (Dga1). Further investigation into the expression profiles of RmACC1, RmFAS1, RmFAS2, and RmDGA1 genes under nitrogen stress revealed that the prolonged up-regulation of the RmDGA1 gene is a molecular indicator of lipogenesis. Subsequent fatty acid profiling unveiled an accumulation of oleic and palmitic acids under nitrogen limitation during the stationary phase. This investigation enhances our understanding of nitrogen stress adaptation and lipid biosynthesis, offering valuable insights into R. mucilaginosa M94C9 for potential industrial applications in the future.

Hepatic stearoyl-CoA desaturase-1 deficiency induces fibrosis and hepatocellular carcinoma-related gene activation under a high carbohydrate low fat diet

Stearoyl-CoA desaturase-1 (SCD1) is a pivotal enzyme in lipogenesis, which catalyzes the synthesis of monounsaturated fatty acids (MUFA) from saturated fatty acids, whose ablation downregulates lipid synthesis, preventing steatosis and obesity. Yet deletion of SCD1 promotes hepatic inflammation and endoplasmic reticulum stress, raising the question of whether hepatic SCD1 deficiency promotes further liver damage, including fibrosis. To delineate whether SCD1 deficiency predisposes the liver to fibrosis, cirrhosis, and hepatocellular carcinoma (HCC), we employed in vivo SCD1 deficient global and liver-specific mouse models fed a high carbohydrate low-fat diet and in vitro established AML12 mouse cells. The absence of liver SCD1 remarkably increased the saturation of liver lipid species, as indicated by lipidomic analysis, and led to hepatic fibrosis. Consistently, SCD1 deficiency promoted hepatic gene expression related to fibrosis, cirrhosis, and HCC. Deletion of SCD1 increased the circulating levels of Osteopontin, known to be increased in fibrosis, and alpha-fetoprotein, often used as an early marker and a prognostic marker for patients with HCC. De novo lipogenesis or dietary supplementation of oleate, an SCD1-generated MUFA, restored the gene expression related to fibrosis, cirrhosis, and HCC. Although SCD1 deficient mice are protected against obesity and fatty liver, our results show that MUFA deprivation results in liver injury, including fibrosis, thus providing novel insights between MUFA insufficiency and pathways leading to fibrosis, cirrhosis, and HCC under lean non-steatotic conditions.

The integral role of de novo lipogenesis in the preparation for seasonal dormancy

Animals can alter their body compositions in anticipation of dormancy to endure seasons with limited food availability. Accumulation of lipid reserves, mostly in the form of triglycerides (TAGs), is observed during the preparation for dormancy in diverse animals, including insects (diapause) and mammals (hibernation). However, the mechanisms involved in the regulation of lipid accumulation and the ecological consequences of failure to accumulate adequate lipid stores in preparation for animal dormancy remain understudied. In the broadest sense, lipid reserves can be accumulated in two ways: the animal either receives lipids directly from the environment or converts the sugars and amino acids present in food to fatty acids through de novo lipogenesis and then to TAGs. Here, we show that preparation for diapause in the Colorado potato beetle (Leptinotarsa decemlineata) involves orchestrated upregulation of genes involved in lipid metabolism with a transcript peak in 8- and 10-d-old diapause-destined insects. Regulation at the transcript abundance level was associated with the accumulation of substantial fat stores. Furthermore, the knockdown of de novo lipogenesis enzymes (ACCase and FAS-1) prolonged the preparatory phase, while the knockdown of fatty acid transportation genes shortened the preparatory phase. Our findings suggest a model in which the insects dynamically decide when to transition from the preparation phase into diapause, depending on the progress in lipid accumulation through de novo lipogenesis.

Inositol and taurine ameliorate abnormal liver lipid metabolism induced by high sucrose intake

Previous attempts to limit fat intake failed to reduce metabolic syndrome incidence, spotlighting excessive sucrose consumption phenomenon. Recent studies challenge the traditional belief, actually fructose is predominantly metabolized in the small intestine, not just the liver. Evidence supports sucrose overconsumption leads to hepatic lipid accumulation via gut microbiome modulation, but the mechanisms and prevention strategies remain unclear. This study investigates the impact of osmolytes inositol and taurine, which are abundant in citrus fruits and seafood respectively, on lipid metabolism in rats consuming excessive sucrose. Five-week-old male Wistar rats fed with control diet, high sucrose diet, and high sucrose diet with inositol or taurine for 28 days. High sucrose diet-induced accumulation of hepatic triglyceride level was reduced only by inositol. Both inositol and taurine decreased hepatic SREBP1, ChREBP, lipogenesis enzymes mRNA levels increased by a high sucrose diet. Taurine also significantly increased the fatty acid uptake major player CD36 mRNA levels in liver. Expression of genes related to fatty acid degradation and transport remained unchanged. In cecal microbiome, high sucrose diet groups showed a decrease in alpha diversity and diverged in beta diversity from the starch diet group, while inositol and taurine did not significantly affect these aspects. Ten bacterial features including genera such as Bacteroides, Parabacteroides, Odoribacter, Streptococcus, etc., were filtered out related to the ameliorative effects of inositol and taurine on lipid metabolism. Overall, both inositol and taurine modulated hepatic lipogenesis and microbiome. However, due to taurine's enhancement of fatty acid uptake, only inositol improved steatosis in high sucrose diet rats.

Multi-Omics Reveals Disrupted Immunometabolic Homeostasis and Oxidative Stress in Adipose Tissue of Dairy Cows with Subclinical Ketosis: A Sphingolipid-Centric Perspective.

Ketosis, especially its subclinical form, is frequently observed in high-yielding dairy cows and is linked to various diseases during the transition period. Although adipose tissue plays a significant role in the development of metabolic disorders, its exact impact on the emergence of subclinical ketosis (SCK) is still poorly understood. The objectives of this study were to characterize and compare the profiling of transcriptome and lipidome of blood and adipose tissue between SCK and healthy cows and investigate the potential correlation between metabolic disorders and lipid metabolism. We obtained blood and adipose tissue samples from healthy cows (CON, n = 8, β-hydroxybutyric acid concentration < 1.2 mmol/L) and subclinical ketotic cows (SCK, n = 8, β-hydroxybutyric acid concentration = 1.2-3.0 mmol/L) for analyzing biochemical parameters, transcriptome, and lipidome. We found that serum levels of nonesterified fatty acids, malonaldehyde, serum amyloid A protein, IL-1β, and IL-6 were higher in SCK cows than in CON cows. Levels of adiponectin and total antioxidant capacity were higher in serum and adipose tissue from SCK cows than in CON cows. The top enriched pathways in whole blood and adipose tissue were associated with immune and inflammatory responses and sphingolipid metabolism, respectively. The accumulation of ceramide and sphingomyelin in adipose tissue was paralleled by an increase in genes related to ceramide biosynthesis, lipolysis, and inflammation and a decrease in genes related to ceramide catabolism, lipogenesis, adiponectin production, and antioxidant enzyme systems. Increased ceramide concentrations in blood and adipose tissue correlated with reduced insulin sensitivity. The current results indicate that the lipid profile of blood and adipose tissue is altered with SCK and that certain ceramide species correlate with metabolic health. Our research suggests that disruptions in ceramide metabolism could be crucial in the progression of SCK, exacerbating conditions such as insulin resistance, increased lipolysis, inflammation, and oxidative stress, providing a potential biomarker of SCK and a novel target for nutritional manipulation and pharmacological therapy.

Assessing the ability of dihydrosterculic acid to activate PPARα

Peroxisome proliferator-activated receptor α (PPARα) is a ligand-activated transcription factor associated with the expression of specific target genes involved in fatty acid β-oxidation. Activation of PPARα could potentially be utilized to treat diseases associated with the accumulation of hepatic lipids such as non-alcoholic fatty liver disease (NAFLD), which currently impacts approximately 25% of the world’s population. At present, a class of synthetic ligands called fibrates can be prescribed to help manage NAFLD but are weak agonists for PPARα and therefore have limited effcacy. Consequently, alternative ligands for PPARα must be established and no natural endogenous ligands have been conclusively identified. This study expands on our previous work and aims to identify the direct effects of dihydrosterculic acid (DHSA), a cyclopropyl fatty acid found in cottonseed oil (CSO), on PPARα activity. To identify downstream effects of DHSA, male mice were placed on a CSO- (DHSA-containing) or isocaloric oil-enriched diet for 6 weeks and tissues were sent for genome-wide RNA sequencing analysis. CSO-fed mice demonstrated increased PPARα gene expression in the liver and concomitant decreased triglyceride in plasma (-75.1±11.5 milligrams/deciliter (mg/dL); p&lt;0.01) and liver (-86.7±71.7 mg/gram (g) protein; p=0.1) and decreased free fatty acid in liver (-16.5±3.4 micromolar (μM)/g protein; p&lt;0.01) relative to the oil-enriched control diet. Confirmational tissue analysis of liver samples show an increase in PPARα target gene expression in the DHSA containing groups compared to control diet groups, indicating that DHSA could potentially serve as an endogenous ligand for PPARα to increase its transcriptional activity to increase fat oxidation. To further test this hypothesis, female PPARα knockout mice were fed a DHSA containing diet or control diet for 4 weeks (n=8). Following the 4-week period, the chow-fed wild type females gained more weight (1.3±0.6 g; p&lt;0.05) than the PPARα knockout mice. Otherwise, there were no significant differences in body weight or food intake between diets or genotypes. Additional in vitro studies show increased PPARα target gene expression following treatments with DHSA, specifically for CPT1a (2.7±0.2 fold increase; p&lt;0.01), ACOX1 (1.3±0.1 fold increase; p&lt;0.05), and ACADVL (1.6±0.1 fold increase, p&lt;0.01), which encode key enzymes in lipid oxidation pathways. SCD1, the rate limiting enzyme in lipogenesis, was also downregulated (0.5±0.03 fold decrease, p&lt;0.01) in response to a 24-hour treatment with DHSA indicating a reduction in endogenous lipogenesis. From these results, we believe that DHSA could function through a PPARα-dependent mechanism. Further analysis of the tissues collected from the PPARα knockout mouse model will aid in determining the mechanism of DHSA as well as identifying its potential for the treatment of excess lipid accumulation. This project is funded by Cotton Incorporated. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Oral supplementation of gut microbial metabolite indole-3-acetate alleviates diet-induced steatosis and inflammation in mice

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries. There is growing evidence that dysbiosis of the intestinal microbiota and disruption of microbiota-host interactions contribute to the pathology of NAFLD. We previously demonstrated that gut microbiota-derived tryptophan metabolite indole-3-acetate (I3A) was decreased in both cecum and liver of high-fat diet-fed mice and attenuated the expression of inflammatory cytokines in macrophages and Tnfa and fatty acid-induced inflammatory responses in an aryl-hydrocarbon receptor (AhR)-dependent manner in hepatocytes. In this study, we investigated the effect of orally administered I3A in a mouse model of diet-induced NAFLD. Western diet (WD)-fed mice given sugar water (SW) with I3A showed dramatically decreased serum ALT, hepatic triglycerides (TG), liver steatosis, hepatocyte ballooning, lobular inflammation, and hepatic production of inflammatory cytokines, compared to WD-fed mice given only SW. Metagenomic analysis show that I3A administration did not significantly modify the intestinal microbiome, suggesting that I3A’s beneficial effects likely reflect the metabolite’s direct actions on the liver. Administration of I3A partially reversed WD-induced alterations of liver metabolome and proteome, notably, decreasing expression of several enzymes in hepatic lipogenesis and β-oxidation. Mechanistically, we also show that AMP-activated protein kinase (AMPK) mediates the anti-inflammatory effects of I3A in macrophages. The potency of I3A in alleviating liver steatosis and inflammation clearly demonstrates its potential as a therapeutic modality for preventing the progression of steatosis to non-alcoholic steatohepatitis (NASH).

Oral supplementation of gut microbial metabolite indole-3-acetate alleviates diet-induced steatosis and inflammation in mice.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries. There is growing evidence that dysbiosis of the intestinal microbiota and disruption of microbiota-host interactions contribute to the pathology of NAFLD. We previously demonstrated that gut microbiota-derived tryptophan metabolite indole-3-acetate (I3A) was decreased in both cecum and liver of high-fat diet-fed mice and attenuated the expression of inflammatory cytokines in macrophages and Tnfa and fatty acid-induced inflammatory responses in an aryl-hydrocarbon receptor (AhR)-dependent manner in hepatocytes. In this study, we investigated the effect of orally administered I3A in a mouse model of diet-induced NAFLD. Western diet (WD)-fed mice given sugar water (SW) with I3A showed dramatically decreased serum ALT, hepatic triglycerides (TG), liver steatosis, hepatocyte ballooning, lobular inflammation, and hepatic production of inflammatory cytokines, compared to WD-fed mice given only SW. Metagenomic analysis show that I3A administration did not significantly modify the intestinal microbiome, suggesting that I3A's beneficial effects likely reflect the metabolite's direct actions on the liver. Administration of I3A partially reversed WD-induced alterations of liver metabolome and proteome, notably, decreasing expression of several enzymes in hepatic lipogenesis and β-oxidation. Mechanistically, we also show that AMP-activated protein kinase (AMPK) mediates the anti-inflammatory effects of I3A in macrophages. The potency of I3A in alleviating liver steatosis and inflammation clearly demonstrates its potential as a therapeutic modality for preventing the progression of steatosis to non-alcoholic steatohepatitis (NASH).

Androgen signaling inhibits de novo lipogenesis to alleviate lipid deposition in zebrafish.

Testosterone is closely associated with lipid metabolism and known to affect body fat composition and muscle mass in males. However, the mechanisms by which testosterone acts on lipid metabolism are not yet fully understood, especially in teleosts. In this study, cyp17a1-/- zebrafish ( Danio rerio) exhibited excessive visceral adipose tissue (VAT), lipid content, and up-regulated expression and activity of hepatic de novo lipogenesis (DNL) enzymes. The assay for transposase accessible chromatin with sequencing (ATAC-seq) results demonstrated that chromatin accessibility of DNL genes was increased in cyp17a1-/- fish compared to cyp17a1+/+ male fish, including stearoyl-CoA desaturase ( scd) and fatty acid synthase ( fasn). Androgen response element (ARE) motifs in the androgen signaling pathway were significantly enriched in cyp17a1+/+ male fish but not in cyp17a1-/- fish. Both androgen receptor ( ar)-/- and wild-type (WT) zebrafish administered with Ar antagonist flutamide displayed excessive visceral adipose tissue, lipid content, and up-regulated expression and activity of hepatic de novo lipogenesis enzymes. The Ar agonist BMS-564929 reduced the content of VAT and lipid content, and down-regulated acetyl-CoA carboxylase a ( acaca), fasn, and scd expression. Mechanistically, the rescue effect of testosterone on cyp17a1-/- fish in terms of phenotypes was abolished when ar was additionally depleted. Collectively, these findings reveal that testosterone inhibits lipid deposition by down-regulating DNL genes via Ar in zebrafish, thus expanding our understanding of the relationship between testosterone and lipid metabolism in teleosts.

Effect of lipoic acid on the expression of nonalcoholic fatty liver disease-associated genes in the liver of rats fed a hypercaloric choline-deficient diet

Nonalcoholic fatty liver disease (NAFLD) is one of the most common chronic liver pathologies. Molecular mechanisms of NAFLD pathogenesis are complex and still require further clarification. Thus, this study aimed to investigate the impact of lipoic acid (LA), widely used as an antioxidant, on the development of different links of NAFLD, including de novo lipogenesis, antioxidant and xenobiotic-metabolizing enzymes by gene expression analysis. The experiment was carried out on three groups of male Wistar rats received control diet, hypercaloric choline-deficient diet (HCCD) or HCCD with LA (HCCD+LA). In the liver of rats the expression of acetyl-CoA carboxylase 1 (Acaca), fatty acid synthase (Fasn), stearoyl-CoA desaturase (Scd) and transcriptional regulators sterol-regulatory element-binding protein SREBP (Srebf1), carbohydrate response element-binding protein ChREBP (Mlxipl), peroxisome proliferator-activated receptor alpha PPARα (Ppara), heme oxygenase-1 HO-1 (Hmox1) and nuclear erythroid 2-related factor 2 Nrf2 (Nrf2), cytochrome P450 1A1 CYP1A1 (CYP1A1) and aryl hydrocarbon receptor AhR (AhR), cytochrome P450 2E1 CYP2E1 (CYP2E1) genes were evaluated. Supplementation of HCCD with LA led to an even greater than in HCCD group decrease in Scd gene expression by 88% (p&lt;0.05), as well as a marked suppression of Ppara and Mlxipl by 37% (p&lt;0.05) and 27% (p&lt;0.05), to an 80% (p&lt;0.05) elevation of Hmox1 gene expression relative to the HCCD group, had no pronounced effect on CYP1A1 and AhR gene expression but resulted in a 38% (p&lt;0.05) suppression of CYP2E1 expression compared with that slightly elevated by HCCD. In conclusion, the current study showed that LA reduces lipogenesis de novo, restores the expression of Hmox1 gene of the antioxidant enzyme HO-1, diminished by the HCCD diet, and decreases HCCD-induced level of the expression of CYP2E1 gene of CYP2E1 enzyme, a potential source of free radicals.

Alveolar type II epithelial cell FASN maintains lipid homeostasis in experimental COPD.

Alveolar epithelial type II (AEC2) cells strictly regulate lipid metabolism to maintain surfactant synthesis. Loss of AEC2 cell function and surfactant production are implicated in the pathogenesis of the smoking-related lung disease chronic obstructive pulmonary disease (COPD). Whether smoking alters lipid synthesis in AEC2 cells and whether altering lipid metabolism in AEC2 cells contributes to COPD development are unclear. In this study, high-throughput lipidomic analysis revealed increased lipid biosynthesis in AEC2 cells isolated from mice chronically exposed to cigarette smoke (CS). Mice with a targeted deletion of the de novo lipogenesis enzyme, fatty acid synthase (FASN), in AEC2 cells (FasniΔAEC2) exposed to CS exhibited higher bronchoalveolar lavage fluid (BALF) neutrophils, higher BALF protein, and more severe airspace enlargement. FasniΔAEC2 mice exposed to CS had lower levels of key surfactant phospholipids but higher levels of BALF ether phospholipids, sphingomyelins, and polyunsaturated fatty acid-containing phospholipids, as well as increased BALF surface tension. FasniΔAEC2 mice exposed to CS also had higher levels of protective ferroptosis markers in the lung. These data suggest that AEC2 cell FASN modulates the response of the lung to smoke by regulating the composition of the surfactant phospholipidome.

FAM120A couples SREBP-dependent transcription and splicing of lipogenesis enzymes downstream of mTORC1

The mechanistic target of rapamycin complex 1 (mTORC1) is a master regulator of cell growth that stimulates macromolecule synthesis through transcription, RNA processing, and post-translational modification of metabolic enzymes. However, the mechanisms of how mTORC1 orchestrates multiple steps of gene expression programs remain unclear. Here, we identify family with sequence similarity 120A (FAM120A) as a transcription co-activator that couples transcription and splicing of de novo lipid synthesis enzymes downstream of mTORC1-serine/arginine-rich protein kinase 2 (SRPK2) signaling. The mTORC1-activated SRPK2 phosphorylates splicing factor serine/arginine-rich splicing factor 1 (SRSF1), enhancing its binding to FAM120A. FAM120A directly interacts with a lipogenic transcription factor SREBP1 at active promoters, thereby bridging the newly transcribed lipogenic genes from RNA polymerase II to the SRSF1 and U1-70K-containing RNA-splicing machinery. This mTORC1-regulated, multi-protein complex promotes efficient splicing and stability of lipogenic transcripts, resulting in fatty acid synthesis and cancer cell proliferation. These results elucidate FAM120A as a critical transcription co-factor that connects mTORC1-dependent gene regulation programs for anabolic cell growth.

Effect of intraductal drug delivery of orexin receptor antagonists into lactating rat mammary gland on milk cholesterol metabolism by regulating Fas and Hmgcr genes

In recent years, many studies have demonstrated that the system of orexin plays a pivotal role in regulating lipogenesis enzymes. However, its effect on the mammary glands is not entirely known. This study answers the question of whether intra-ductal injection of orexin antagonists (OX1RA and OX2RA) into the mammary glands can result in the expression of fatty acid synthase (Fas) and HMG-CoA reductase (Hmgcr) genes and the secretion of cholesterol in lactating female rats or not. To this end, 42 Lactating rats were randomly divided into experimental groups including a control group and groups receiving OX1RA and OX2RA intraductal (with doses of 5, 10, and 20 µg/kg, i.duc). Milk samples were collected for cholesterol testing. Using specific primers for each gene, the target genes were measured via real-time PCR. Data differences were considered significant with P &lt;0.05. PCR exhibited that the injection of orexin antagonists significantly reduced Fas and Hmgcr gene expression. Moreover, the injection of antagonists significantly reduced milk cholesterol. Intra-mammary injection of orexin antagonists reduces milk cholesterol levels by affecting the expression of Fas and Hmgcr genes.

Abstract 414: Targeting lipid metabolism to improve efficacy of braf-targeted therapy in colorectal cancer

Abstract Background: Aberrant lipid metabolism is associated with poor prognosis in colorectal cancer (CRC). Fatty acid synthase (FASN), a key enzyme of lipogenesis, is overexpressed and a therapeutic target in CRC. Overexpression of CD36, a fatty acid translocase, plays a pro-tumorigenic role in CRC. BRAFV600E is the mutation occurring in about 10-15% of CRC cases. BRAF-targeted therapy is effective, but quickly developed resistance is an issue. Our preliminary data show that development of resistance to BRAF-targeted therapy is associated with an increase in expression of FASN, CD36, accumulation of triglycerides (TGs), and mitochondrial respiration. Therefore, our central hypothesis is that inhibition of lipid metabolism will sensitize CRC cells to BRAF inhibitors and overcome acquired resistance. Methods: We established HT29 cells and primary PT130 and PT2449pt cells resistant to PLX8394, a novel BRAF inhibitor. IC50 curves, PrestoBlue viability, CytoSelect™ 24-Well Cell Invasion, and Triglyceride Assays, Seahorse XF analysis, western blot, and confocal microscopy were used to evaluate differences between parental and resistant cells. RNA-seq and lipid analysis were used to evaluate changes in gene expression and lipids levels. Combination of PLX8394 and TVB3664 (FASN inhibitor) was tested on cell viability in parental and resistant cells. Results: PLX8394 resistant cells have a higher IC50, an increased proliferation, and invasion compared to parental cells. An increase in invasion of resistant cells is associated with a decrease in E-cadherin. RNA sequencing shows a significant increase in FASN expression. Western blot on resistant cells confirms upregulation of FASN, CD36, and other lipogenic markers. Consistently, resistant cells have an increase in levels of triglycerides as compared to parental cells. Seahorse XF Cell Mito Stress Test shows that resistant cells forgo the Warburg effect and instead rely more heavily on oxidative phosphorylation. To further support our hypothesis, FASN shRNA knockdown of FASN in parental HT29 cells is more susceptible to PLX8394 treatment compared to control. We also observed the synergetic effect of PLX8394 and TVB3664 treatment on cell viability in parental cells, but not in resistant cells. Conclusion: Our study demonstrates that resistance to BRAF inhibitors is associated with a significant increase in proliferation, metastasis, and upregulation of lipid metabolism. We show that combination of FASN and BRAF inhibitors has a combinational effect on inhibiting cell viability in parental but not resistant cells, suggesting that the addition of FASN inhibitor to the standard regimen for BRAFV600E mutation positive patients can improve efficacy of these therapies. Additional screening of lipid metabolism-targeted therapies in combination with standard BRAF regimens are needed to develop novel and more efficacious strategies for CRC patients with BRAF mutations. Citation Format: Mariah E. Geisen, Dang he, Chi Wang, Jill M. Kolesar, Yekaterina Zaytseva. Targeting lipid metabolism to improve efficacy of braf-targeted therapy in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 414.

Swietenine Alleviates Nonalcoholic Fatty Liver Disease in Diabetic Mice via Lipogenesis Inhibition and Antioxidant Mechanisms.

Our previous studies have reported the effect of swietenine (a major bioactive component of Swietenia macrophylla seeds) in reversing and potentiating the effect of metformin in hyperglycemia and hyperlipidaemia in diabetic rats. Moreover, we reported that the anti-inflammatory effect of swietenine is mediated via the activation of nuclear factor erythroid 2-related factor 2 (Nrf2). This study evaluated the effect of swietenine and its mechanisms in nonalcoholic fatty liver disease (NAFLD) in high-fat diet/streptozotocin-induced diabetic mice. The effect was assessed by determining blood biochemical parameters (glucose, cholesterol, triglycerides, alanine transaminase (ALT), asparate transaminase (AST), alkaline phosphatase (ALP), glutathione (GSH), total antioxidant capacity (TAC), and malondialdehyde (MDA)) and liver biochemical parameters (liver index, cholesterol, and triglycerides). Hepatic lipid accumulation (initial causative factor in NAFLD) was determined by oil-O-red staining. Gene expression (qPCR) and immunohistochemical studies were performed to elucidate the mechanism of swietenine's effect in NAFLD. The critical regulators (genes and proteins) involved in lipogenesis (ACLY, ACC1, FASN, SREBP1c, and ChREBPβ) and oxidative stress (Nrf2, NQO-1 and HO-1) pathways were determined. In mice fed with a high-fat diet followed by streptozotocin injection, the liver cholesterol, triglycerides, and lipids were elevated. These increases were reversed by the oral administration of swietenine, 80 mg/kg body weight, on alternate days for eight weeks. Gene expression and immunohistochemical studies showed that swietenine reversed the elevated levels of crucial enzymes of lipogenesis (ACLY, ACC1 and FASN) and their master transcription factors (SREBP1c and ChREBPβ). Furthermore, swietenine activated the Nrf2 antioxidant defense mechanism, as evidenced by the upregulated levels of Nrf2, NQO-1, and HO-1. It is concluded that swietenine shows beneficial effects in diabetes-induced NAFLD via inhibiting lipogenesis and activating the Nrf2 pathway.

Investigating dual inhibition of ACC and CD36 for the treatment of non-alcoholic fatty liver disease in mice.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide. Dysregulation in hepatic lipid metabolism, including increased fatty acid uptake and de novo lipogenesis (DNL), is a hallmark of NAFLD. Here, we investigated dual inhibition of the fatty acid transporter fatty acid translocase (FAT/CD36), and acetyl-CoA carboxylase (ACC), the rate-limiting enzyme in DNL, for the treatment of NAFLD in mice. Mice with hepatic CD36 deletion (Cd36LKO) and wildtype littermates were fed a high fat diet for 12 weeks and treated daily with either oral administration of an ACC inhibitor (GS-834356, Gilead Sciences, ACCi) or vehicle for 8 weeks. Neither CD36 deletion or ACC inhibition impacted body composition, energy expenditure, or glucose tolerance. Cd36LKO mice had elevated fasting plasma insulin, suggesting mild insulin resistance. Whole-body fatty acid oxidation was significantly decreased in Cd36LKO mice. Liver triglyceride content was significantly reduced in mice treated with ACCi, however, CD36 deletion caused an unexpected increase in liver triglycerides. This was associated with upregulation of genes and proteins of DNL, including ACC, and decreased liver triglyceride secretion ex vivo. Overall, these data confirm the therapeutic utility of ACC inhibition for steatosis resolution but indicate that inhibition of CD36 is not an effective treatment for NAFLD in mice.

QSAR Study on Diacylgycerol Acyltransferase-1 (DGAT-1) Inhibitor as Anti-diabetic using PSO-SVM Methods

Diabetes mellitus is a chronic disease that can occurred to anyone. Up until now, there are no specific drugs that have been found which can completely cure diabetes. One of the possible steps to treat diabetes mellitus is by inhibiting the growth of Diacylglycerol Acyltransferase-1 (DGAT-1) enzyme. This study aims to build a QSAR model on DGAT-1 inhibitors as anti-diabetic using Particle Swarm Optimization (PSO) and Support Vector Machine (SVM). Acyl-CoA: DGAT1 is a mikrosomal enzyme in lipogenesis which is increased in metabolically active cells to meet nutrient requirements. Microsomal-enzymes-that have an-important in the triglyceride-synthesis process-of 1,2-diacylglycerol by-catalyzing-acyl-coa-dependent-acylations as anti-diabetics. The dataset used in this study consists of 228 samples containing molecular structures and their inhibitor activities. We reduce the number of features by removing features with a standard deviation less than the threshold value, followed by the PSO algorithm. The best-predicted result is obtained through the implementation of SVM with RBF kernel, with the score of and are 0.75 and 0.67, respectively.&#x0D;