Regulatory Element-binding Protein Research Articles

Abstract 2077: Exploring the multifaceted efficacy of de novo lipogenesis inhibitor in cancer therapy

Abstract Cancer cells frequently exhibit altered lipid metabolism to sustain their rapid growth and proliferation.Sterol Regulatory Element-Binding Proteins (SREBPs) are key transcription factors that orchestratelipid biosynthesis and uptake in cells. Dysregulation of SREBPs is a common feature of many cancertypes, making them attractive targets for therapeutic intervention. In this study, we present acomprehensive exploration of the potential and rationale for targeting lipid metabolism in cancerthrough the use of a small molecule inhibitor that targets activation of SREBP. We illustrate that oursmall molecule inhibitor (SMI) exhibits potent anticancer activity and effectively targets SREBPsignaling in a variety of prostate cancer models as well as in non-prostate cancer models that arelipogenesis driven. Our novel SMI induces apoptosis and reduces PCa cell proliferation, suppressescell invasion and migration. Furthermore, we illustrate that in addition to suppressing the transcriptionfactor activity of SREBPs, these novel SMIs are highly effective in suppressing both the full length andvariant androgen receptor and their signaling in prostate cancer cells. Notably, in cell and organoidmodels, these novel inhibitors exhibit synergistic anticancer activity in combination with the ARsignaling inhibitor, Enzalutamide. The anticancer activity of these inhibitors extend beyond ARdependent prostate cancer and are also effective in organoid models of neuroendocrine PCa, aparticularly drug-resistant and aggressive PC subtype. Lastly, we illustrate excellent pharmacokineticsand potent anti-tumor effects of these novel SREBP activation inhibitors using in vivo castration-resistant prostate cancer CDX tumors. Given the urgent need for novel targets in PCa, our first-in-class inhibitor represents a promising therapeutic strategy with the potential to effectively targetdysregulated lipogenesis in cancer patients. Citation Format: Dallin Lowder, Jin Na Shin, Maria E. Ruiz-Echartea, Jenny Deng, Aleksandra Rusin, Darlene Skapura, Michael Ittmann, Salma Kaochar. Exploring the multifaceted efficacy of de novo lipogenesis inhibitor in cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2077.

Tricin-enriched Zizania latifolia ameliorates non-alcoholic fatty liver disease through AMPK-dependent pathways.

This study aimed to identify and elucidate the mechanism underlying the protective effect of tricin-enriched Zizania latifolia (Z. latifolia) extract (ETZL) against free fatty acid (FFA)-induced lipid accumulation in vitro and non-alcoholic fatty liver disease (NAFLD) induced by a high-fat diet and fructose diet (HFD/F) in vivo. ETZL treatment significantly lowered body weight gain and decreased adipose tissue, lipid, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels in HFD/F-fed mice. ETZL acted on phosphorylated acetyl-CoA carboxylase (ACC) and anti-peroxisome proliferator-activated receptor α (PPARα) by activating the adenosine monophosphate-activated protein kinase (AMPK) pathway and inhibiting sterol regulatory element-binding proteins-1 (SREBP)/fatty acid synthase (FAS) signaling to inhibit de novo adipogenesis and increase fatty acid oxidation. In addition, treatment with ETZL increased nuclear factor erythroid-2-related factor 2 (Nrf2) levels to activate the antioxidant pathway. FFA-induced oxidative stress and fatty acid accumulation in HepG2 cells confirmed the improvement in fat accumulation through the AMPK and Nrf2 pathway activities of ETZL. These results suggest that ETZL ameliorates NAFLD by regulating lipid metabolism and defending against oxidative stress via AMPK-dependent pathways.

A novel deformable liposomal hydrogel loaded with a SREBP-1-inhibiting polypeptide for reducing sebum synthesis in golden hamster model

Excessive sebum is the major factor involved in the pathophysiology of seborrheic diseases. Chemical medicines can result in mild to severe side effects. Polypeptides with much less side effects make them ideal for reducing sebum synthesis. Sterol regulatory element-binding proteins-1 (SREBP-1) is necessary for the biosynthesis of sterols. A SREBP-1-inhibiting polypeptide (SREi), which competitively inhibits the ubiquitination of Insig-1 so as to suppress the activation of SREBP-1 was selected as an active ingredient and formulated into skin topical preparations. The SREi anionic deformable liposomes contained sodium deoxycholate (SDCh) at the concentration of 4.4 mg/mL (SREi-ADL3) and SREi-ADL3 in 0.3% (w/v) carbomer hydrogel (SREi-ADL3-GEL) were prepared and characterized. The SREi-ADL3 presented a high entrapment efficiency of 92.62 ± 6.32%, a particle size of 99.54 ± 7.56 nm and a surface charge of -19.18 ± 0.45 mV. SREi-ADL3-GEL exhibited a sustained release behavior, a higher stability, a much more cellular uptake ability and transdermal absorption. In vivo golden hamster model confirmed that SREi-ADL3-GEL presented the strongest inhibitory effect on sebaceous gland growth and sebum synthesis by down-regulating the mRNA and protein expression of SREBP-1, fatty acid synthase (FAS) and acetyl–coenzyme A carboxylase 1 (ACC1). As confirmed by histological analysis, only a small amount of sebaceous gland lobes with the lightest staining intensity and the smallest dyeing area could be observed in the SREi-ADL3-GEL group. Taken together, SREi-ADL3-GEL displayed potential applications in sebum excessive production related diseases.

The Effect of Guisangyou Tea on Abnormal Lipid Metabolism in Mice Induced by High-Fat Diet.

This study was aimed to investigate the effect of Guisangyou tea (GSY tea) in improving abnormal lipid metabolism in mice with obesity induced by a high-fat diet (HFD). The results showed that intervention of the water extract of GSY tea (WE) decreased serum levels of lipids, and positively regulated the related antioxidant enzyme activities and the inflammatory factors in the serum and liver. In the liver, the mRNA and protein expression levels of sterol regulatory element-binding proteins-1 (SREBP-1), stearoyl-CoA desaturase-1 (SCD-1), fatty acid synthase (FASN), and acetyl CoA carboxylase (ACC) related to lipid synthesis were downregulated, and the mRNA and protein expression levels of the farnesoid X receptor (FXR) and small heterodimer partner (SHP) related to bile acid production were upregulated. The results demonstrate that GSY tea can improve abnormal lipid metabolism in obese mice by improving the body's antioxidant capacity, regulating the inflammatory state, and reducing the synthesis of lipids and increasing the production of bile acids. GSY tea can be processed and utilized as a safe and effective resource for improving abnormal lipid metabolism.

Malic Enzyme 1 as a Novel Anti-Ferroptotic Regulator in Hepatic Ischemia/Reperfusion Injury.

Ferroptosis has been linked to the pathogenesis of hepatic injury induced by ischemia/reperfusion (I/R). However, themechanistic basis remains unclear. In this study, by using a mouse model of hepatic I/R injury, it is observed that glutathione (GSH) and cysteine depletion are associated with deficiency of the reducing power ofnicotinamide adenine dinucleotide phosphate (NADPH). Genes involved in maintaining NADPH homeostasis are screened, and it is identified that I/R-induced hepatic ferroptosis is significantly associated with reduced expression and activity of NADP+ -dependent malic enzyme 1 (Me1). Mice with hepatocyte-specific Me1 gene deletion exhibit aggravated ferroptosis and liver injury under I/R treatment; while supplementation with L-malate, the substrate of ME1, restores NADPH and GSH levels and eventually inhibits I/R-induced hepatic ferroptosis and injury. A mechanistic study further reveals that downregulation of hepatic Me1 expression is largely mediated by the phosphatase and tensin homologue (PTEN)-dependent suppression of the mechanistic target of rapamycin/sterol regulatory element-binding protein 1 (mTOR/SREBP1) signaling pathway in hepatic I/R model. Finally, PTEN inhibitor, mTOR activator, or SREBP1 over-expression all increase hepatic NADPH, block ferroptosis, and protect liver against I/R injury. Taken together, the findings suggest that targeting ME1 may provide new therapeutic opportunities for I/R injury and other ferroptosis-related hepatic conditions.

TGF-β1 promotes SCD1 expression via the PI3K-Akt-mTOR-SREBP1 signaling pathway in lung fibroblasts

BackgroundLung fibroblast activation is associated with airway remodeling during asthma progression. Stearoyl-CoA desaturase 1 (SCD1) plays an important role in the response of fibroblasts to growth factors. This study aimed to explore the effects of SCD1 on fibroblast activation induced by transforming growth factor-β1 (TGF-β1) and the role of the phosphatidylinositol-3-kinase-AKT serine-threonine protein kinase-mechanistic target of rapamycin (PI3K-Akt-mTOR) pathway on the regulation of SCD1 expression in airway remodeling.MethodsFemale C57BL/6 mice were sensitized and challenged with house dust mites to generate a chronic asthma model. The inhibitor of SCD1 was injected i.g. before each challenge. The airway hyper-responsiveness to methacholine was evaluated, and airway remodeling and airway inflammation were assessed by histology. The effects of SCD1 on fibroblast activation were evaluated in vitro using an SCD1 inhibitor and oleic acid and via the knockdown of SCD1. The involvement of the PI3K-Akt-mTOR-sterol regulatory element-binding protein 1 (SREBP1) pathway in lung fibroblasts was investigated using relevant inhibitors.ResultsThe expression of SCD1 was increased in fibroblasts exposed to TGF-β1. The inhibition of SCD1 markedly ameliorated airway remodeling and lung fibroblast activation in peripheral airways. The knockdown or inhibition of SCD1 resulted in significantly reduced extracellular matrix production in TGF-β1-treated fibroblasts, but this effect was reversed by the addition of exogenous oleic acid. The PI3K-Akt-mTOR-SREBP1 pathway was found to be involved in the regulation of SCD1 expression and lung fibroblast activation.ConclusionsThe data obtained in this study indicate that SCD1 expression contributes to fibroblast activation and airway remodeling and that the inhibition of SCD1 may be a therapeutic strategy for airway remodeling in asthma.

The miR-216/miR-217 Cluster Regulates Lipid Metabolism in Laying Hens With Fatty Liver Syndrome via PPAR/SREBP Signaling Pathway.

Fatty liver syndrome (FLS), a common metabolic disease in laying hens, caused by excessive hepatic fat deposition is a bottleneck in the poultry industry. However, no specific therapeutic methods have been developed. Evidence suggests that microRNAs (miRNAs) are essential for liver lipid metabolism and homeostasis, providing strong evidence for targeting miRNAs as a potential treatment option for liver diseases. However, the roles of miRNAs in the pathogenesis of FLS remain unclear. In present study, RNA-sequencing was performed to discern the expression patterns of miRNAs in normal and fatty livers of laying hens. In total, 12 dysregulated miRNAs (2 down-regulated and 10 up-regulated) were detected between the normal and fatty livers. Functional enrichment analysis showed the potential impacts of the dysregulated miRNAs on lipid metabolism. Notably, miR-216a/b and miR-217-5p, which belong to the miR-216/miR-217 cluster, were up-regulated in the sera and livers of FLS chickens, as well as free fatty acid (FFA)-induced LMH cells. Oil-red O staining revealed that up-regulation of the miR-216/miR-217 cluster induced lipid accumulation in FFA-induced LMH cells. Furthermore, the dual luciferase gene reporter assay and RT-qPCR analysis demonstrated that 3-hydroxyacyl-CoA dehydratase 2, F-box protein 8, and transmembrane 9 superfamily member 3 (TM9SF3) were directly targeted by miR-216a/b and miR-217-5p, respectively, and suppressed in the fatty livers of laying hens. Moreover, overexpression of the miR-216/miR-217 cluster or reduction in TM9SF3 levels led to activation of the proliferator-activated receptor/sterol regulatory-element binding protein (PPAR/SREBP) pathway. Overall, these results demonstrate that the miR-216/miR-217 cluster regulates lipid metabolism in laying hens with FLS, which should prove helpful in the development of new interventional strategies.

AGR2-induced cholesterol synthesis drives lovastatin resistance that is overcome by combination therapy with allicin.

Anterior gradient 2 (AGR2), a protein disulfide isomerase (PDI), is a multifunctional protein under physiological and pathological conditions. In this study we investigated the roles of AGR2 in regulating cholesterol biogenesis, lipid-lowering efficiency of lovastatin as well as in protection against hypercholesterolemia/statin-induced liver injury. We showed that AGR2 knockout significantly decreased hepatic and serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in mice with whole-body or hepatocyte-specific Agr2-null mutant, compared with the levels in their wild-type littermates fed a normal chow diet (NCD) or high-fat diet (HFD). In contrast, mice with AGR2 overexpression (Agr2/Tg) exhibited an increased cholesterol level. Mechanistic studies revealed that AGR2 affected cholesterol biogenesis via activation of AKT/sterol regulatory element-binding protein-2 (SREBP2), to some extent, in a PDI motif-dependent manner. Moreover, elevated AGR2 led to a significant decrease in the lipid-lowering efficacy of lovastatin (10 mg· kg-1· d-1, ip, for 2 weeks) in mice with hypercholesterolemia (hyperCho), which was validated by results obtained from clinical samples in statin-treated patients. We showed that lovastatin had limited effect on AGR2 expression, but AGR2 was inducible in Agr2/Tg mice fed a HFD. Further investigations demonstrated that drug-induced liver toxicity and inflammatory reactions were alleviated in hypercholesterolemic Agr2/Tg mice, suggesting the dual functions of AGR2 in lipid management and hyperCho/statin-induced liver injury. Importantly, the AGR2-reduced lipid-lowering efficacy of lovastatin was attenuated, at least partially, by co-administration of a sulfhydryl-reactive compound allicin (20 mg· kg-1· d-1, ip, for 2 weeks). These results demonstrate a novel role of AGR2 in cholesterol metabolism, drug resistance and liver protection, suggesting AGR2 as a potential predictor for selection of lipid-lowering drugs in clinic.

Research on the mechanism of hypoxia promoting the migration of lung adenocarcinoma A549 cells

Objective: To investigate the mechanism that hypoxia promotes the migration of lung adenocarcinoma A549 cells. Methods: A549 cells were cultured and cells that knockdown of acetyl-CoA carboxylase 1 (ACC1) were obtained by transfection with lentivirus, and cells that knockdown of sterol regulatory element-binding proteins-1 (SREBP-1) were obtained by treated with si-RNA. A549 cells were treated with hypoxia combined with hypoxia inducible factor-1α (HIF-1α) inhibitor PX-478 (25 μmol); Hypoxia combined with linoleic acid (LA) (20 μmol) treated A549 cells with ACC1 knockdown, and A549 cells with SREBP-1 knockdown were treated by hypoxia. Transwell migration assay was used to detect cell migration. Western blot was conducted to detect HIF-1α, ACC1 and epithelial mesenchymal transition (EMT) related proteins, Vimentin, E-Cadherin and SREBP-1; Real-time fluorescent quantitative polymerase chain reaction (RT-qPCR) was performed to detect the changes of ACC1 and SREBP-1 mRNA in A549 cells after hypoxia and HIF-1α inhibitor PX-478 (25 μmol) treatment. Each experiment was repeated three times. Results: Compared with the normoxic control group, hypoxia promoted the migration of A549 cells (P＜0.01), and up-regulated the expressions of ACC1, HIF-1α (all P＜0.01) and SREBP-1 (P＜0.05). PX-478 (25 μmol) inhibited the migration of A549 cells induced by hypoxia and down-regulated the expression of SREBP-1 (all P＜0.05). ACC1 mRNA and SREBP-1 mRNA levels were increased after hypoxia treatment of A549 cells (all P＜0.05). The levels of ACC1 mRNA and SREBP-1 mRNA were decreased after A549 cells treated with hypoxia combined with PX-478 (25 μmol) for 24 h (P＜0.05, P＜0.01). Knockdown of SREBP-1 in A549 cells was obtained by transfection with si-RNA. Transwell migration assay showed the number of cell migration in si-SREBP-1 group was less than that in normoxia control group (P＜0.01). The si-SREBP-1 group and the si-NC group were treated with hypoxia. Compared with the control group, the number of cell migration in the si-SREBP-1 group was decreased (P＜0.01), however, the difference was not statistically significant compared with the normoxia si-SREBP-1 group (P＞0.05). Western blot showed that the expression of ACC1 in the si-SREBP-1 group was lower than that in the control group (P＜0.01). Compared with the control group, the expression of ACC1 was decreased after si-SREBP-1 group treated with hypoxia (P＜0.01). Knockdown of ACC1 inhibited the migration of A549 cells (P＜0.05). After knockdown of ACC1, the migration number of A549 cells under normoxia and 5% O2 conditions had no significant difference (P＞0.05). Application of LA under hypoxia condition rescued ACC1-knockdown induced inhibitory effect on hypoxia-promoted A549 cell migration (P＜0.05). Conclusion: Hypoxia promotes migration of lung adenocarcinoma A549 cells by regulating fatty acid metabolism through HIF-1α/SREBP-1/ACC1 pathway.

Taurine prevents high-fat diet-induced-hepatic steatosis in rats by direct inhibition of hepatic sterol regulatory element-binding proteins and activation of AMPK.

This study investigated if the protective effect of taurine against high fat diet-induced hepatic steatosis involves modulating the hepatic activity of 5' AMP-activated protein kinase (AMPK) and levels/activity of the sterol regulatory element-binding proteins-1/2 (SREBP1/2). Rats were divided into four groups (n=12/group) as (a) STD, fed standard diet (3.85kcal/g); (b) STD+taurine (500mg/kg); (c) HFD, fed HFD (4.73kcal/g); and (d) HFD+taurine. All treatments were conducted for 12weeks. Independent of food intake or modulating glucose or insulin levels, taurine administration to STD and HFD-fed rats significantly lowered weekly weight gain and the accumulation of the retroperitoneal, visceral and subcutaneous fats. In both groups, taurine also reduced serum and hepatic levels of triglycerides and cholesterol and reduced hepatic mRNA and protein levels of fatty acid synthase (FAS), acetyl CoA carboxylase-1 (ACC-1), HMG-CoA-reductase and HMG-CoA synthetase. In control rats only, taurine reduced hepatic levels of mature forms of sterol regulatory element-binding proteins (SREBP)-1/2. In HFD-fed rats, taurine reduced SREBP-1/2 precursor and mature forms in the livers of HFD-fed rats. Besides, taurine significantly increased levels of glutathione (GSH), the activity of superoxide dismutase (SOD), and the activity of AMPK and its downstream β-oxidation genes including peroxisome proliferator-activated receptor-α (PPAR-α) and carnitine palmitoyltransferase (CPT-1) in the livers of both the control and HFD-fed rats. In conclusion, taurine protects against HFD-induced hepatic steatosis stimulating antioxidant levels, and concomitant stimulating hepatic β-oxidation and suppressing lipid synthesis, mediated by activation of AMPK and suppression of SREBP-1.

Sorghum (Sorghum bicolor) Extract Affects Plasma Lipid Metabolism and Hepatic Macrophage Infiltration in Diabetic Rats

Background: Chronic hyperglycemia is known to be a high-risk factor for progressive chronic liver diseases, such as abnormal lipid metabolism. The activation of AMP-activated protein kinase (AMPK) has a beneficial effect on dyslipidemia. Polyphenols derived from various plants are involved in AMPK activation. Objective: We investigated the effects of polyphenol-containing sorghum (Sorghum bicolor) extract (SE) on plasma lipid metabolism and macrophage infiltration, and measured the expression and phosphorylation of AMPK and acetyl-CoA carboxylase (ACC) in diabetic rat livers. Methods: Streptozotocin-induced diabetic rats received 0, 50, or 250 mg/kg of SE orally for 4 weeks. Blood chemistry, total and phosphorylated protein levels of AMPK and ACC, sterol regulatory element- binding protein-1c (SREBP-1c) mRNA and protein levels, and macrophage infiltration in the livers were examined. Results: Plasma glucose and triacylglycerol levels, which were increased in the untreated diabetic rats, were significantly lower in the 250 mg/kg SE-treated diabetic rats. AMPK and ACC phosphorylation levels were significantly increased in the 250 mg/kg SE-treated diabetic rats compared with those in the untreated rats. There was no difference in the hepatic expression of SREBP-1c between the diabetic rat groups. Macrophage infiltration in the liver was suppressed by 250 mg/kg of SEtreatment. Conclusion: These data suggest that SE treatment may affect plasma lipid metabolism and chronic inflammation by upregulating phosphorylation of AMPK and ACC in diabetic rat livers.

Schaftoside alleviates HFD-induced hepatic lipid accumulation in mice via upregulating farnesoid X receptor

Ethnopharmacology relevanceThe farnesoid X receptor (FXR) is a therapeutic target of for the treatment of non-alcoholic fatty liver disease (NAFLD) owing to its regulatory role in lipid homeostasis. Schaftoside (SS) is a bioactive compound of Herba Desmodii Styracifolii, which has traditionally been used to treat hepatitis and cholelithiasis. However, the potential hepatoprotective effect of SS against NAFLD and the underlying mechanisms remain unknown. Aim of the studyWe investigated whether SS could improve NAFLD-induced liver injury by decreasing lipid accumulation via the activation of FXR signalling. Materials and methodsIn vivo, the effects of SS on high-fat diet (HFD)-induced lipid accumulation in the liver of mice were evaluated by serum biochemical parameters and histopathological analysis. In vitro, the intracellular triglyceride (TG) level and Oil Red O staining were used to evaluate the lipid removal ability of SS in Huh-7 cells or FXR knockout mouse primary hepatocytes (MPHs) induced by oleic acid (OA). Moreover, FXR/sterol regulatory element-binding protein 1 (SREBP1) mRNA and protein expression levels were detected. ResultsSS reduced HFD-induced lipid accumulation in the liver, as indicated by decreased aspartate aminotransferase (AST), cholesterol (Ch), and TG levels in serum and TG levels in liver tissue, and subsequently resulting in attenuation of liver histopathological injury. Gene expression profiles demonstrated that SS dose-dependently prevented HFD-induced decrease of FXR expression and inversely inhibited SREBP1 expression in the nucleus. Furthermore, SS significantly suppressed excessive TG accumulation and decreased intracellular TG level in Huh-7 cells or MPHs via the upregulation of FXR and inhibition of SREBP1 expression in the nucleus. ConclusionOur results suggest that SS ameliorates HFD-induced NAFLD by the decrease of lipid accumulation via the control of FXR-SREBP1 signalling.

Promoter methylation of SREBP-2 in circulating leukocytes correlates with relevant risk factors in patients with coronary artery disease

Objective To examine the correlation between the promoter methylation of Sterol regulatory-element binding protein-2 (SREBP-2) and miR-33a expression as well as serum markers in patients with coronary artery disease (CAD). Methods The case-control study. 100 participants who underwent coronary angiography from August 2017 to April 2018 in TaiheHospital, Hubei University of Medicine, were recruited in this study.The methylation level of two fragments, including 12 CpG sites in the promoter region of SREBP-2, have been detected by pyrosequencing in 50 patients with coronary artery disease (CAD) and 50 non-CAD controls. Serum miR-33a level and a panel of 15 CAD related biomarkers were examined by qPCR and routine biochemistry methods. Results Methylation level of one CpG site (F1-4 loci) in SREBP-2 promoter region were significant higher in CAD patients than in controls(4.56%±0.70% vs 3.54%±0.72%, t=-3.864, P<0.001); methylation level of F1-4 site was negatively correlates with the serum miR-33a levels and high-density lipoprotein cholesterol (HDL-C) levels(r=-0.318, P=0.001; r=-0.225, P=0.024, respectively). Furthermore, F1-4 hypermethylation was an independent risk factor of CAD, independent of age, gender, histories of hypertension, hyperlipidemia, and diabetes(OR=2.452, 95%CI=1.398-4.299, P=0.002). Conclusion These results suggest that DNA methylation and miRNA might cooperate to regulate the lipid metabolism in CAD. Key words: Coronary disease; Sterol regulatory element binding protein 2; DNA methylation; microRNAs; Lipid metabolism disorders

MEX3C regulates lipid metabolism to promote bladder tumorigenesis through JNK pathway.

Purpose: Bladder cancer (BC) is the most common urinary cancer among men with a high rate of deaths despite the improved medical technology and treatment. Recent evidence demonstrated that Mex-3 RNA-Binding Family Member C (MEX3C) plays various roles in different biological activities, but its molecular mechanisms underlying the pathogenesis of BC remain unclear yet. The aim of this research was to explore the expression patterns of MEX3C and its biological functions in human BC.Materials and methods: The Cancer Genome Atlas (TCGA) and Oncomine databases were jointly used to analyze the expression of MEX3C in BC and its correlation with the clinicopathological features, while real-time PCR and immunohistochemistry analysis were used to verify the predicted results. Wound-healing assay, Matrigel invasion assay, BODIPY staining and Western blot analysis were used in a cell model to assess the effect of MEX3C on the lipid metabolism, invasion and migration of BC and its mechanisms.Results: MEX3C was highly expressed in BC tissues and cells compared with their normal counterparts, and its expression was positively correlated with the clinicopathological features, especially the invasiveness phenotype. Overexpression of MEX3C accumulated lipid droplets and promoted cell adhesion, invasion and migration. We further demonstrated that MEX3C regulated lipid metabolism and promoted tumor development and progression through activation of JNK signaling and upregulating the JNK downstream protein levels of sterol regulatory element-binding proteins-1, fatty acid synthase and acetyl-CoA carboxylase-1.Conclusion: Here we identified MEX3C as a new oncogene to promote bladder tumorigenesis by regulating lipid metabolism through Mitogen-activated protein kinase/c-Jun N-terminal kinase (MAPK/JNK) pathway. These findings suggest a new role of MEX3C in promoting BC tumorigenesis and provide a novel biomarker or molecular target for diagnosis or treating BC.

Expression of mammalian target of rapamycin (mTOR), nuclear factor-κ B (NF-κB) and sterol regulatory element binding protein 2 (SREBP2) in serum and placentas among gravidas with preeclampsia

Objective To investigate the level of mammalian target of rapamycin (mTOR) in serum and the expression of mTOR, nuclear factor-κ B (NF-κB) and sterol regulatory element binding protein 2 (SREBP2) in placenta among gravidas with preeclampsia. Methods From August 2015 to August 2017, 60 gravidas including 40 with severe preeclampsia (SPE) and 20 with mild preeclampsia (MPE) who underwent regular prenatal care and delivered by caesarean section were selected from the Second Xiangya Hospital of Central South University. According to the ratio of 2∶1, 30 gravidas who delivered through caesarean section due to cephalopelvic disproportion, abnormal fetal position or social factors during the same period were enrolled as the control group. Peripheral blood samples were obtained to determine the concentrations of serum mTOR, high density lipoprotein-cholesterol (HDL-C), low density lipoprotein-cholesterol (LDL-C), triglyceride (TG) and total cholesterol (TC) by enzyme linked immunosorbent assay (ELISA). The expression of mTOR, phospho-mTOR (p-mTOR), NF-κB and SREBP2 in placenta were measured by Western blot. Clinical datas were statistically analyzed using one-way ANOVA, Bonferroni or Dunnett's T3 test, and Pearson's correlation analysis. Results (1) The serum levels of mTOR and LDL-C in the SPE and MPE group were both higher than that in the control group [mTOR:(11 765.56±1 698.95) and (8 278.56±1 106.59) vs (4 366.19±716.43) pg/ml; LDL-C:(7.81±1.90) and (4.11±0.75) vs (2.42±0.45) mmol/L, all P<0.05].Furthermore the serum levels of mTOR and LDL-C in the SPE group were both higher than those in the MPE group (both P<0.05). The serum level of HDL-C in the SPE and MPE group were lower than that in the control group [(0.36±0.12) and (0.85±0.11) vs (1.33±0.16) mmol/L, both P<0.05], and that in the SPE group was lower than that in the MPE group (P<0.05). Women in the SPE group showed higher TG level when comparing with the MPE and control group [(46.19±18.92) vs (35.55±6.54) and (33.24±9.78) nmol/L, both P<0.05], while the TC levels in the SPE and MPE group were higher than that in the control group[(24.72±7.17) and (21.83±4.19) vs (16.32±3.88) nmol/L, both P<0.05]. (2) The placental expressions of mTOR, p-mTOR, NF-κB and SREBP2 protein in the SPE and MPE group were higher compared with that in the control group [mTOR: (0.52±0.09) and (0.38±0.08) vs (0.24±0.05); p-mTOR: (0.42±0.08) and (0.26±0.05) vs (0.14±0.03); NF-κB: (0.58±0.10) and (0.36±0.05) vs (0.21±0.03); SREBP2: (0.52±0.08) and (0.33±0.05) vs (0.20±0.05); all P<0.05], and those expressions of the SPE group also higher comparing with the MPE group. Otherwise the p-mTOR/mTOR ratios in the SPE group and MPE group were higher than that in the control group [(0.75±0.10) and (0.69±0.14) vs (0.59±0.13), both P<0.05]. (3) Pearson's correlation analysis showed that serum level of mTOR and placental expressions of mTOR and p-mTOR in the SPE group were positively correlated with serum LDL-C (r=0.682, 0.584 and 0.504, all P<0.05), TG (r=0.612, 0.658 and 0.422, all P<0.05) , while serum level of mTOR and placental expressions of mTOR in the SPE group were positively correlated with TC (r=0.598 and 0.452, all P<0.05), but were negatively correlated with serum HDL-C (r=-0.375, -0.442 and -0.390, all P<0.05). The NF-κB expression in placenta of the SPE group was significantly positively correlated with the mTOR expression in placenta and serum LDL-C (r=0.375 and 0.391, both P<0.05). Moreover, in the SPE group, the SREBP2 level in placenta was significantly positively correlated with placental expression of mTOR and serum TC level (r=0.364 and 0.392, both P<0.05). (4) In the MPE group, mTOR level in serum and levels of mTOR and p-mTOR in placenta were significantly positively correlated with serum LDL-C (r=0.813, 0.641 and 0.465, all P<0.05), TG (r=0.646, 0.529 and 0.502, all P<0.05) and TC (r=0.558, 0.482 and 0.483, all P<0.05), while the level of serum mTOR was negatively correlated with the level of serum HDL-C (r=-0.606, P<0.05). The NF-κB level in placenta in MPE group was positively correlated with the mTOR in placenta and the serum LDL-C (r=0.458 and 0.595, both P<0.05), while the SREBP2 level in placenta was significantly positively correlated with mTOR in placenta and serum TC (r=0.580, 0.560, respectively; both P<0.05) in the MPE group. Conclusions mTOR, NF-κB and SREBP2 may play important roles in the onset and development of preeclampsia by interfering lipid metabolism. Key words: Pre-eclampsia; Mammalian torget of rapamycin; Nuclear factor-κB; Sterol regulatory element binding protein 2

Disrupting cholesterol esterification by bitter melon suppresses triple-negative breast cancer cell growth.

Triple negative breast cancer (TNBC) is aggressive with a worse prognosis. We have recently shown that bitter melon extract (BME) treatment was more effective in inhibition of TNBC tumor growth in mouse models as compared to ER positive breast tumor growth. Aberrant dysregulation of lipid metabolism is associated with breast cancer progression, however, anti-cancer mechanism of BME linking lipid metabolism in breast cancer growth remains unexplored. Here, we observed that accumulation of esterified cholesterol was reduced in BME treated TNBC cell lines as compared to control cells. We next evaluated expression levels of acyl-CoA: cholesterol acyltransferase 1 (ACAT-1) in TNBC cells treated with BME. Our results demonstrated that BME treatment inhibited ACAT-1 expression in TNBC cells. Subsequently, we found that sterol regulatory element-binding proteins-1 and -2, and FASN was significantly reduced in BME treated TNBC cell lines. Low-density lipoprotein receptor was also downregulated in BME treated TNBC cells as compared to control cells. We further demonstrated that BME feeding reduced tumor growth in TNBC mammospheres implanted into NSG mice, and inhibits ACAT-1 expression. To our knowledge, this is the first report demonstrating BME suppresses TNBC cell growth through ACAT-1 inhibition, and have potential for additional therapeutic regimen against human breast cancer.

Colocynth (Citrullus colocynthis) Flesh Extract Suppresses Adipogenesis by Down-Regulating Adipogenic Transcription Factors and Their Target Genes in 3T3-L1 Preadipocytes

Citrullus colocynthis, a member of the Cucurbitaceae family, is widely distributed in North Africa. The fruits are recognized for their wide range of medicinal uses and promising pharmaceutical potential. The present study aimed to investigate the anti-obesity effect of the ethanol extract of colocynth flesh (FCEE) in 3T3-L1 cells following treatment at different doses. The viability of 3T3-L1 preadipocytes was analyzed via MTT assay and triglycerides were stained with Oil red O to assess lipid accumulation. Additionally, adipogenesis-related gene expression was quantified via qRT-PCR. FCEE (0 - 150 μg/mL) dose-dependently suppressed intracellular triglyceride accumulation during the adipogenesis by 23% and 66% at 100 and 150 μg/mL, respectively, but did not affect cell viability. Analysis of the time-dependence of the effect of FCEE demonstrated that the greatest anti-adipogenic activity was observed during the early stages of differentiation. FCEE also decreased GPDH activity in a dose-dependent manner, with 98% decrease observed at 150 μg/mL. In addition, at same range of FCEE concentrations, the main transcription factors, including CCAAT/enhancer binding protein α (C/EBPα), peroxisome proliferator activated receptor γ (PPARγ), and sterol regulatory element-binding protein 1c (SREBP-1c), were downregulated by 90%, 89%, and 89%, respectively at 150 μg/mL. As these are the master regulators of adipogenesis. The inhibition of their downstream target genes was also observed. Colocynth may be useful in the treatment of obesity owing to its powerful effects on fat, which result in changes to adipocyte differentiation and fat mobilization.

Natural Extracts Abolished Lipid Accumulation in Cells Harbouring non-favourable PNPLA3 genotype

Background & aims. G-allele of PNPLA3 (rs738409) favours triglycerides accumulation and steatosis. In this study, we examined the effect of quercetin and natural extracts from mushroom and artichoke on reducing lipid accumulation in hepatic cells.Material and methods. Huh7.5 cells were exposed to oleic acid (OA) and treated with quercetin and extracts to observe the lipid accumulation, the intracellular-TG concentration and the LD size. Sterol regulatory element binding proteins-1 (SREBP-1), peroxisome proliferator-activated receptor (PPARα-γ) and cholesterol acyltransferase (ACAT) gene expression levels were analysed.Results. Quercetin decreased the intracellular lipids, LD size and the levels of intracellular-TG through the down-regulation of SREBP-1c, PPARγand ACAT1 increasing PPARα. The natural-extracts suppressed OA-induced lipid accumulation and the intracellular-TG. They down-regulate the hepatic lipogenesis through SREBP-1c, besides the activation of lipolysis through the increasing of PPARα expression.Conclusions. Quercetin and the aqueous extracts decrease intracellular lipid accumulation by down-regulation of lipogenesis and up-regulation of lipolysis.

JNK Activation of BIM Promotes Hepatic Oxidative Stress, Steatosis, and Insulin Resistance in Obesity.

The members of the BCL-2 family are crucial regulators of the mitochondrial pathway of apoptosis in normal physiology and disease. Besides their role in cell death, BCL-2 proteins have been implicated in the regulation of mitochondrial oxidative phosphorylation and cellular metabolism. It remains unclear, however, whether these proteins have a physiological role in glucose homeostasis and metabolism in vivo. In this study, we report that fat accumulation in the liver increases c-Jun N-terminal kinase-dependent BCL-2 interacting mediator of cell death (BIM) expression in hepatocytes. To determine the consequences of hepatic BIM deficiency in diet-induced obesity, we generated liver-specific BIM-knockout (BLKO) mice. BLKO mice had lower hepatic lipid content, increased insulin signaling, and improved global glucose metabolism. Consistent with these findings, lipogenic and lipid uptake genes were downregulated and lipid oxidation enhanced in obese BLKO mice. Mechanistically, BIM deficiency improved mitochondrial function and decreased oxidative stress and oxidation of protein tyrosine phosphatases, and ameliorated activation of peroxisome proliferator-activated receptor γ/sterol regulatory element-binding protein 1/CD36 in hepatocytes from high fat-fed mice. Importantly, short-term knockdown of BIM rescued obese mice from insulin resistance, evidenced by reduced fat accumulation and improved insulin sensitivity. Our data indicate that BIM is an important regulator of liver dysfunction in obesity and a novel therapeutic target for restoring hepatocyte function.

Changes of suppressors of cytokine signaling-3 and sterol regulatory element binding proteins-1c pathway in steatosis HepG2/HepG2.2.15 cells

Objective To investigate the effects of HepG2 and HepG2.2.15 cells steatosis on the mRNA and protein expressions of suppressors of cytokine signaling-3(SOCS-3) and sterol regulatory element binding proteins (SREBP-1c). Methods The cell model of chronic hepatitis B (CHB) combined with nonalcoholic fatty liver disease (NAFLD) was successfully constructed using an oleic acid-induced HepG2 and HepG2.2.15 cells steatosis. Cells were divided into HepG2 cell control group (HepG2 cell control group), HepG2.2.15 cell control group (HepG2.2.15 cell control group), HepG2 cell steatosis group (HepG2 cell steatosis group) and HepG2.2.15 cell steatosis group (HepG2.2.15 cell steatosis group). The expression levels of SOCS-3 and SREBP-1c mRNA were detected by real-time quantitative polymerase chain reaction (PCR). Changes in protein expressions of SOCS-3 and SREBP-1c were measured by western blot. Results SOCS-3 mRNA expression level in HepG2.2.15 cell control group was significantly lower than that in HepG2 cell control group (P<0.01). The level in HepG2 cell steatosis group was also significantly lower than that in HepG2 cell control group (P<0.01). However, the level of SOCS-3 mRNA in HepG2.2.15 cell steatosis group was lower than HepG2.2.15 cell control group with no statistical significance (P=0.173). There was interaction between cells and steatosis (F=25.547, P<0.01). The expression of SREBP-1c mRNA in HepG2.2.15 cell control group was significantly lower than that in HepG2 cell control group (P<0.01), and was significantly higher in HepG2.2.15 cell steatosis group than that in HepG2.2.15 cell control group (P<0.01). There was no significant difference between HepG2 cell steatosis group and HepG2 cell control group (P=1.000). There was interaction between cells and steatosis (F=5.04, P<0.05). Western blot analysis showed that protein levels of SOCS-3 and SREBP-1c in steatosis cells at 48 h and 72 h were significantly higher than those in non-alcoholic steatosis cells. Conclusions Protein expressions of SOCS-3 and SREBP-1c are up-regulated in both steatosis groups. Factorial analysis shows that there is interaction between cells and steatosis. HBV gene could inhibit SOCS-3 mRNA expression and promote the expression of SREBP-1c mRNA in steatosis cells. Key words: Hepatitis B; Fatty liver; SOCS-3; SREBP-1c