Expression Of Sterol Response Element-binding Protein-1 Research Articles

Alterations in IL-4, IL-10 and IFN-γ levels synergistically decrease lipid content and protein expression of FAS and mature SREBP-1 in human sebocytes

When anti-acne alternatives from dietary and plant sources are ingested, systemic alterations of interleukin (IL)-4, IL-10, IL-12 and interferon (IFN)-γ, individually or simultaneously, are induced at a 0.1-10.0-fold (×) range of normal physiological concentrations (1×). However, little is known about the effects of these cytokines on excess sebum, a pathophysiological factor of acne development. In this study, human sebocytes were treated with 0.1-10.0× of IL-4, IL-10, IL-12 and IFN-γ for 3 or 5days to elucidate the effects on lipid content. Treatment with individual cytokines decreased the lipid content at specific concentrations rather than in a concentration-dependent manner. Specifically, 5.0× of IL-4, 5.0× of IFN-γ (5.0IFN), and 0.5×, 5.0× and 10.0× of IL-10 for 3days, and 0.5× of IL-4 (0.5IL4) for 5days decreased lipid content to 87.6-93.0% of the control. Treatment with other concentrations of IL-4, IL-10 and IFN-γ, and 0.1-10.0× of IL-12 did not alter lipid content. Combined treatment with 0.5IL4, 5.0IFN and 0.5× of IL-10 for 3 or 5days decreased the lipid content more than each individual treatment. However, this effect was more evident after 3days, in parallel with decreased levels of triglycerides, cholesterol esters and free fatty acids, the major lipid compositions of sebocytes, and decreased protein expression of fatty acid synthase (FAS) and mature sterol response element-binding protein-1 (SREBP-1), the lipogenesis-related factors, without altered cell proliferation. We demonstrated that suppressed IL-4 and IL-10 with enhanced IFN-γ synergistically decreased lipid content and protein expression of FAS and mature SREBP-1 in human sebocytes.

Palmitate-Induced SREBP1 Expression and Activation Underlies the Increased BACE 1 Activity and Amyloid Beta Genesis.

Numerous cross-sectional and longitudinal studies have implicated saturated fat-enriched diets in the etio-pathogenesis of Alzheimer's disease (AD). Emerging evidence shows that saturated fat-enriched diets, such as palmitate-enriched diets, increase amyloid-beta (Aβ) production, the histopathological hallmark of AD. However, the molecular mechanisms that underlie the deleterious effects of palmitate-enriched diets in the augmentation of Aβ genesis are yet to be characterized. Sterol response element binding protein 1 (SREBP1) is a transcription factor that is modulated by saturated fatty acids, such as palmitate, and consequently regulates the expression of genes that code for proteins involved in almost all facets of lipid metabolism. Herein, we determined the role of changes in SREBP1 expression and transcriptional activity in the palmitate-induced effects on Aβ genesis and BACE1 expression, the enzyme that catalyzes the rate-limiting step in Aβ biosynthesis. We demonstrate that palmitate-induced SREBP1 activation directly regulates BACE1 expression at the transcriptional level in the mouse hippocampus and mouse Neuro-2a (N2a) neuroblastoma cells. Chromatin immunoprecipitation (ChIP) studies show that palmitate increases the binding of SREBP1 to the Bace1 promoter region in the mouse hippocampus and mouse N2a neuroblastoma cells. Ectopic expression of the dominant negative SREBP1 mutant and knocking-down SREBP1 expression significantly reduced the palmitate-induced increase in BACE1 expression and subsequent Aβ genesis in mouse N2a neuroblastoma cells. Our study unveils SREBP1 activation as a novel molecular player in the palmitate-induced upregulation of BACE1 expression and subsequent Aβ genesis.

O-linked N-acetylglucosamine transferase enhances secretory clusterin expression via liver X receptors and sterol response element binding protein regulation in cervical cancer.

O-linked N-acetylglucosamine transferase (OGT) expression is increased in various cancer types, indicating the potential importance of O-GlcNAcylation in tumorigenesis. Secretory clusterin (sCLU) is involved in cancer cell proliferation and drug resistance, and recently, liver X receptors (LXRs) and sterol response element binding protein-1 (SREBP-1) were reported to regulate sCLU transcription. Here, we found that sCLU is significantly increased in cervical cancer cell lines, which have higher expression levels of O-GlcNAc and OGT than keratinocytes. OGT knockdown decreased expression of LXRs, SREBP-1 and sCLU through hypo-O-GlcNAcylation of LXRs. Additionally, treatment with Thiamet G, O-GlcNAcase OGA inhibitor, increased expression of O-GlcNAcylation and sCLU, and high glucose increased levels of LXRs, SREBP-1 and sCLU in HeLa cells. Moreover, OGT knockdown induced G0/G1 phase cell cycle arrest and late apoptosis in cisplatin-treated HeLa cells, and decreased viability compared to OGT intact HeLa cells. Taken together, these findings suggest that OGT, O-GlcNAcylated LXRs, and SREBP-1 increase sCLU expression in cervical cancer cells, which contributes to drug resistance.

PALMITATE INDUCES BACE1 EXPRESSION AND ACTIVITY BY INDUCING STEROL RESPONSE ELEMENT BINDING PROTEIN 1 EXPRESSION AND ACTIVATION IN THE MOUSE HIPPOCAMPUS AND HUMAN SH-SY5Y NEUROBLASTOMA CELLS

The etiology of Alzheimer's disease (AD) is multifactorial and poorly defined and characterized by a dynamic interplay of a myriad of risk variables that include dietary and environmental factors. Epidemiological studies have implicated saturated fat-enriched diets in the etio-pathogenesis of AD. The aspartyl protease beta-site amyloid precursor protein cleaving enzyme 1 (BACE1) catalyzes the rate limiting step in amyloid-beta (Aβ) production, a peptide that serves as a core instigator of pathophysiological neurodegenrative processes in AD. Emerging evidence from our laboratory studies and other contemporary studies have shown that saturated fat-enriched diets, such as palmitate-enriched diets, increase BACE1 activity and enhance Aβ production. Our preliminary studies have shown that a palmitate-enriched diet increases the expression and activity of Sterol Response Element binding protein 1 (SREBP1) in the mouse hippocampus. In this study, we determined the role of SREBP1 activation in the palmitate-induced increase in BACE1 expression, activity, and subsequent Aβ genesis. We fed nine-month old C57BL/6J mice a palmitate-enriched diet or the respective control-chow diet for three months and determined the extent to which increased SREBP1 expression and activity plays a role in the palmitate-enriched diet-induced increase in BACE1 expression, activity, and Aβ genesis in the hippocampus. We also determined the role SREBP1 activation in the induction of BACE1 expression, activity, and Aβ genesis in exogenous palmitate-treated human SH-SY5Y neuroblastoma cells and delineated the underlying mechanisms involved. Our ChIP analysis and luciferase reporter assays show that palmitate-induced SREBP1 activation directly regulates BACE1 expression at the transcriptional level. Knocking-down or ectopic expression of the dominant negative SREBP1 mutant significantly mitigated the palmitate-induced BACE1 expression and subsequent Aβ genesis. Our study elucidates a novel signaling pathway and SREBP1 as a unique downstream effector of palmitate-induced up-regulation in BACE1 expression and subsequent Aβ genesis.

Long-term administration of DHEA prevents fat deposition in rats fed a high-fat diet

The effects of dehydroepiandrosterone (DHEA) on lipid metabolism and lipogenic gene mRNA expression in rats subjected to a high-fat diet were determined. Totally 75 rats were randomly divided into 5 groups: control group 1 fed a normal diet (NCG), and groups 2–5 fed a high-fat diet with 0 (HCG), 25 (HLG), 50 (HMG), 100 (HHG) mg DHEA per kg body weight via gavage once a day for 8 weeks, respectively. DHEA significantly decreased body weight in HMG group as compared with HCG group (P < 0.05). Hepatic triglyceride and total cholesterol contents were decreased in HMG and HHG groups (P < 0.05), and hepatic lipase activity in HMG group was higher (P < 0.01) than in HCG group. Fatty acid synthesis (FAS) mRNA level was decreased in HLG and HHG groups (P < 0.01), and sterol response element binding protein-1 (SREBP-1) mRNA level was decreased in HMG and HHG groups when compared with HCG group (P < 0.01). Acyl-CoA oxidase (ACO) and liver carnitine palmitoyl transferase-1 (LCPT-1) mRNA abundance was decreased in HLG and HHG groups (P < 0.01), whereas hormone sensitive lipase (HSL) mRNA level was increased in HMG group as compared with HCG group (P < 0.05). These results indicated that long-term administration of DHEA reduced the synthesis of endogenous triglycerides by inhibiting SREBP-1 and FAS expression, and augmented the lipolysis of exogenous triglycerides through enhancing HSL expression, which eventually led to reduced fat deposition in rats fed a high-fat diet.

Expression of Protease-Activated Receptor-2 in SZ95 Sebocytes and its Role in Sebaceous Lipogenesis, Inflammation, and Innate Immunity

Protease-activated receptor-2 (PAR-2) functions as innate biosensor for proteases and regulates numerous functions of the skin. However, the expression and physiological role of PAR-2 in sebocytes remain to be elucidated. Here, we identified PAR-2 expression in SZ95 sebocytes at both mRNA and protein levels. Intracellular Ca(2+) mobilization by PAR-2 agonist peptide (PAR-2 AP) or Propionibacterium acnes (P. acnes) culture supernatant was detected, indicating that P. acnes is a potent activator of PAR-2 on sebocytes. The small interfering RNA (siRNA)-mediated PAR-2 knockdown in sebocytes resulted in defective differentiation and lipogenesis. PAR-2 AP treatment enhanced lipogenesis and sterol response element-binding protein-1 (SREBP-1) expression, suggesting a role of PAR-2 in the differentiation and lipogenesis of sebocytes. Moreover, PAR-2 AP induced cytokines and human β-defensin-2 (hBD-2) transcription in sebocytes. PAR-2 expression was increased in sebaceous glands of acne lesions. PAR-2 silencing by siRNA abrogated the increase in sebaceous lipogenesis and SREBP-1 expression by P. acnes supernatant. PAR-2 knockdown also inhibited the P. acnes supernatant-induced expression of cytokines and hBD-2. In conclusion, PAR-2 is expressed in SZ95 sebocytes and mediates differentiation, lipogenesis, inflammation, and innate immunity in response to P. acnes. Therefore, PAR-2 might be a therapeutic target for sebaceous gland disorders such as acne.

Ezetimibe suppresses cholesterol accumulation in lipid-loaded vascular smooth muscle cells in vitro via MAPK signaling.

To investigate the mechanisms of anti-atherosclerotic action of ezetimibe in rat vascular smooth muscle cells (VSMCs) in vitro. VSMCs of SD rats were cultured in the presence of Chol:MβCD (10 μg/mL) for 72 h, and intracellular lipid droplets and cholesterol levels were evaluated using Oil Red O staining, HPLC and Enzymatic Fluorescence Assay, respectively. The expression of caveolin-1, sterol response element-binding protein-1 (SREBP-1) and ERK1/2 were analyzed using Western blot assays. Translocation of SREBP-1 and ERK1/2 was detected with immunofluorescence. Treatment with Chol:MβCD dramatically increased the cellular levels of total cholesterol (TC), cholesterol ester (CE) and free cholesterol (FC) in VSMCs, which led to the formation of foam cells. Furthermore, Chol:MβCD treatment significantly decreased the expression of caveolin-1, and stimulated the expression and nuclear translocation of SREBP-1 in VSMCs. Co-treatment with ezetimibe (3 μmol/L) significantly decreased the cellular levels of TC, CE and FC, which was accompanied by elevation of caveolin-1 expression, and by a reduction of SREBP-1 expression and nuclear translocation. Co-treatment with ezetimibe dose-dependently decreased the expression of phosphor-ERK1/2 (p-ERK1/2) in VSMCs. The ERK1/2 inhibitor PD98059 (50 μmol/L) altered the cholesterol level and the expression of p-ERK1/2, SREBP-1 and caveolin-1 in the same manner as ezetimibe did. Ezetimibe suppresses cholesterol accumulation in rat VSMCs in vitro by regulating SREBP-1 and caveolin-1 expression, possibly via the MAPK signaling pathway.

Conjugated linoleic acid-induced milk fat reduction associated with depressed expression of lipogenic genes in lactating Holstein mammary glands

The efficacy of conjugated linoleic acid (CLA) in diet supplements for milk fat reduction is well documented in several species. However, the mechanisms by which fatty acids regulate mammary lipogenesis remain largely unknown, especially with regard to gene expression of enzyme and regulators. In this study, 8 Holstein dairy cows in their mid-lactation period were randomly divided into 2 groups. Control cows received a Ca salt of palm oil fatty acid dietary supplement, and those in the CLA group were fed Ca salts of CLA (Ca-CLA), all in a dose of approximately 200 g∙cow(-1)∙day(-1) for 14 days. The milk yield was recorded daily, and protein, lactose, and fat in the milk were quantified every 3 days for 2 weeks. Fatty acids in the milk were analyzed with gas-liquid chromatography. Measurement of messenger RNA levels of the main lipogenic genes of lipoprotein lipase, acetyl-coenzyme A (CoA) carboxylase, fatty acid synthase, stearoyl-CoA desaturase, and transcription factors such as sterol response element binding protein 1 (SREBP1) and peroxisome proliferator-activated receptor γ was performed in biopsy samples of mammary tissue on the last day. The results indicated that dietary Ca-CLA caused a continuous reduction of milk fat (P < 0.01) with no effect on milk yield, milk protein, and lactose. The fatty acid profile in the milk from the CLA group differed from that from controls, and the yield of milk fatty acid decreased (P < 0.01) with Ca-CLA supplementation. The depressed expression of lipogenic genes (lipoprotein lipase, acetyl-CoA carboxylase, fatty acid synthase, and stearoyl-CoA desaturase) demonstrated inhibition of fatty acid de novo synthesis and uptake in the mammary gland of the CLA group. Furthermore, the gene expression of transcription factor SREBP1 was also downregulated (P < 0.01), but peroxisome proliferator-activated receptor γ was unchanged, suggesting that SREBP1 may play a key role in the regulation of lipogenic gene expression in the lactating mammary gland.

IGF-1 Induces SREBP-1 Expression and Lipogenesis in SEB-1 Sebocytes via Activation of the Phosphoinositide 3-Kinase/Akt Pathway

Understanding the factors that regulate sebum production is important in identifying therapeutic targets for acne therapy. Insulin and IGF-1 stimulate sebaceous gland lipogenesis. IGF-1 increases expression of sterol response element-binding protein-1 (SREBP-1), a transcription factor that regulates numerous genes involved in lipid biosynthesis. SREBP-1 expression, in turn, stimulates lipogenesis in sebocytes. The goal of this study was to identify the intracellular signaling pathway(s) that transduces the lipogenic signal initiated by IGF-1. Sebocytes were treated with IGF-1 and assayed for activation of the phosphoinositide 3-kinase (PI3-K) pathway and of the three major arms of the mitogen-activated protein kinase (MAPK) pathway (MAPK/extracellular signal-regulated kinase (ERK), p38 MAPK, and stress-activated protein kinase/c-Jun-N terminal kinase). IGF-1 activated the MAPK/ERK and PI-3K pathways. Using specific inhibitors of each pathway, we found that the increase in expression of SREBP-1 induced by IGF-1 was blocked in the presence of the PI3-K inhibitor but not in the presence of the MAPK/ERK inhibitor. Furthermore, inhibition of the PI3-K pathway also blocked the IGF-1-induced transcription of SREBP target genes and sebocyte lipogenesis. These data indicate that IGF-1 transmits its lipogenic signal in sebocytes through activation of Akt. Specific targeted interruption of this pathway in the sebaceous gland could be a desirable approach to reducing sebum production and improving acne.

Expression of Lipogenic Factors Galectin-12, Resistin, SREBP-1, and SCD in Human Sebaceous Glands and Cultured Sebocytes

The transcription factors CCAAT enhancer-binding protein alpha, beta, and delta, and peroxisome proliferator-activated receptor gamma are known to be crucial to the differentiation of adipocytes and are expressed in sebaceous gland cells. As lipogenesis is key to both adipocyte and sebocyte differentiation we hypothesize that sebocytes follow a similar program of differentiation to adipocytes. We have investigated the expression of known adipogenic factors resistin, galectin-12, sterol response-element-binding protein-1 (SREBP-1) and stearoyl-CoA desaturase in the immortalized sebaceous gland cell line SZ95 and whole skin. Reverse transcriptase-PCR analysis showed the expression of galectin-12, resistin, SREBP-1, and stearoyl-CoA desaturase mRNAs in SZ95 sebocytes. Immunoreactivity was observed for galectin-12 and SREBP-1 in the nuclei and resistin in the cytoplasm of basal sebocytes, and stearoyl CoA desaturase in the cytoplasm of basal and luminal sebocytes of human scalp skin. Expression of galectin-12, resistin, and SREBP-1 in SZ95 sebocytes was confirmed by Western blot analysis. These data provide further evidence that pathways of differentiation in adipocytes and sebocytes could be similar and therefore further understanding of sebaceous gland differentiation and lipogenesis and potential therapies for sebaceous gland disorders may be obtained from our knowledge of adipocyte differentiation.

Essential Role in Vivo of Upstream Stimulatory Factors for a Normal Dietary Response of the Fatty Acid Synthase Gene in the Liver

In the liver, transcription of several genes encoding lipogenic and glycolytic enzymes, in particular the gene for fatty acid synthase (FAS), is known to be stimulated by dietary carbohydrates. The molecular dissection of the FAS promoter pointed out the critical role of an E box motif, located at position -65 with respect to the start site of transcription, in mediating the glucose- and insulin-dependent regulation of the gene. Upstream stimulatory factors (USF1 and USF2) and sterol response element binding protein 1 (SREBP1) were shown to be able to interact in vitro with this E box. However, to date, the relative contributions of USFs and SREBP1 ex vivo remain controversial. To gain insight into the specific roles of these factors in vivo, we have analyzed the glucose responsiveness of hepatic FAS gene expression in USF1 and USF2 knock-out mice. In both types of mouse lines, defective in either USF1 or USF2, induction of the FAS gene by refeeding a carbohydrate-rich diet was severely delayed, whereas expression of SREBP1 was almost normal and insulin response unchanged. Therefore, USF transactivators, and especially USF1/USF2 heterodimers, seem to be essential to sustain the dietary induction of the FAS gene in the liver.