Si Young Cho

Identification of Mouse Prp19p as a Lipid Droplet-associated Protein and Its Possible Involvement in the Biogenesis of Lipid Droplets

Prp19p is an integral component of the heteromeric protein complex (the NineTeen complex) in the nucleus, and it is essential for the structural integrity of NineTeen complex and its subsequent activation of the spliceosome. We identified Prp19p, which has never been reported in relation to any function outside of the nucleus, as a member of proteins associated with lipid droplets. Down-regulation of Prp19p expression with RNA interference in 3T3-L1 cells repressed lipid droplet formation with the reduction in the level of expression of perilipin and S3-12. The levels of expression of SCD1 (stearoyl-CoA desaturase-1), DGAT-1 (acyl-CoA diacylglycerol acyltransferase-1), and glycerol-3-phosphate acyltransferase were also reduced in Prp19p down-regulated cells, and a significant decrease in triglycerides was observed. Unlike perilipin, which is one of the most extensively studied lipid droplet-associated proteins, Prp19p is not essential for cAMP- and hormone-sensitive lipase-dependent lipolysis pathways, even though Prp19p is a component of the lipid droplet phospholipid monolayer, and down-regulation of Prp19p represses fat accretion significantly. These results suggest that Prp19p or Prp19-interacting proteins during lipid droplet biogenesis in adipocytes may be considered as another class of potential targets for attacking obesity and obesity-related problems.

Transcriptional activation of Cidec by PPARγ2 in adipocyte

Cidec is a lipid droplet-associated protein, which inhibits lipolysis, leading to the accumulation of triglycerides in adipocytes. However, the transcriptional regulation of Cidec in adipocyte remains unknown. In the present study we investigated that the mouse Cidec transcript is regulated by PPARgamma2. After the differentiation of adipocyte, the expression pattern of Cidec was similar to that of PPARgamma2. In the presence of a PPARgamma agonist, the level of Cidec mRNA was highly increased. In addition, putative PPRE sites were identified in the Cidec promoter. By chromatin immunoprecipitation assay and reporter assay, we observed the binding of PPARgamma2 to the promoter of Cidec. Gel shift assay and the mutagenesis study were showed that the -219/-207 region of the Cidec promoter could function as a PPRE of the Cidec promoter. These results suggest that PPARgamma2 is required for the transcriptional activity of Cidec during adipogenesis, which could be contributed to understand the molecular mechanism of lipid droplet formation in adipocytes.

Proteomic analysis of mitochondrial proteins of basal and lipolytically (isoproterenol and TNF‐α)‐stimulated adipocytes

The regulation of adipocyte lipolysis is increasingly believed to influence insulin resistance, in a process that may be associated with mitochondrial dysfunction. However, the molecular basis of the relationship between mitochondrial protein expression, lipolytic responsiveness, and insulin resistance remains unknown. A set of proteins that shows altered abundances in the mitochondria of untreated and treated 3T3‐L1 adipocytes with TNF‐α or isoproterenol was identified. These include the proteins associated with energy production, including fatty acid oxidation, TCA cycle, and oxidative phosphorylation. Proteins associated with oxidative stress dissipation were down‐regulated in lipolytically stimulated adipocytes. Lipolytic stimulation with isoproterenol and TNF‐α, which is also a potent proinflammatory cytokine, showed some noticeable differences in mitochondrial protein expression. For example, isoproterenol markedly enhanced the expression of prohibitin which is involved in the integrity of mitochondria but TNF‐α did not. These results provide valuable information on mitochondrial dysfunction associated with oxidative stress induced by lipolytic stimulation. J. Cell. Biochem. 106: 257–266, 2009.

Chitooligosaccharide Induces Mitochondrial Biogenesis and Increases Exercise Endurance through the Activation of Sirt1 and AMPK in Rats

By catabolizing glucose and lipids, mitochondria produce ATPs to meet energy demands. When the number and activity of mitochondria are not sufficient, the human body becomes easily fatigued due to the lack of ATP, thus the control of the quantity and function of mitochondria is important to optimize energy balance. By increasing mitochondrial capacity? it may be possible to enhance energy metabolism and improve exercise endurance. Here, through the screening of various functional food ingredients, we found that chitooligosaccharide (COS) is an effective inducer of mitochondrial biogenesis. In rodents, COS increased the mitochondrial content in skeletal muscle and enhanced exercise endurance. In cultured myocytes, the expression of major regulators of mitochondrial biogenesis and key components of mitochondrial electron transfer chain was increased upon COS treatment. COS-mediated induction of mitochondrial biogenesis was achieved in part by the activation of silent information regulator two ortholog 1 (Sirt1) and AMP-activated protein kinase (AMPK). Taken together, our data suggest that COS could act as an exercise mimetic by inducing mitochondrial biogenesis and enhancing exercise endurance through the activation of Sirt1 and AMPK.

An Ethanol Extract of Artemisia iwayomogi Activates PPARδ Leading to Activation of Fatty Acid Oxidation in Skeletal Muscle

Although Artemisia iwayomogi (AI) has been shown to improve the lipid metabolism, its mode of action is poorly understood. In this study, a 95% ethanol extract of AI (95EEAI) was identified as a potent ligand of peroxisome proliferator-activated receptorδ (PPARδ) using ligand binding analysis and cell-based reporter assay. In cultured primary human skeletal muscle cells, treatment of 95EEAI increased expression of two important PPARδ-regulated genes, carnitine palmitoyl-transferase-1 (CPT1) and pyruvate dehydrogenase kinase isozyme 4 (PDK4), and several genes acting in lipid efflux and energy expenditure. Furthermore, 95EEAI stimulated fatty acid oxidation in a PPARδ-dependent manner. High-fat diet-induced obese mice model further indicated that administration of 95EEAI attenuated diet-induced obesity through the activation of fatty acid oxidation in skeletal muscle. These results suggest that a 95% ethanol extract of AI may have a role as a new functional food material for the prevention and/or treatment of hyperlipidermia and obesity.

Transcriptional activation of melanocortin 2 receptor accessory protein by PPARγ in adipocytes

Adrenocorticotropic hormone (ACTH) in rodents decreases lipid accumulation and body weight. Melanocortin receptor 2 (MC2R) and MC2R accessory protein (MRAP) are specific receptors for ACTH in adipocytes. Peroxisome proliferator-activated receptor γ (PPARγ) plays a role in the transcriptional regulation of metabolic pathways such as adipogenesis and β-oxidation of fatty acids. In this study we investigated the transcriptional regulation of MRAP expression during differentiation of 3T3-L1 cells. Stimulation with ACTH affected lipolysis in murine mature adipocytes via MRAP. Putative peroxisome proliferator response element (PPRE) was identified in the MRAP promoter region. In chromatin immunoprecipitation and reporter assays, we observed binding of PPARγ to the MRAP promoter. The mutagenesis experiments showed that the -1209/-1198 region of the MRAP promoter could function as a PPRE site. These results suggest that PPARγ is required for transcriptional activation of the MRAP gene during adipogenesis, which contributes to understanding of the molecular mechanism of lipolysis in adipocytes.

Ginseng Berry and its Biological Effects as a Natural Phytochemical

A recent trial in the development of new medications and immune-modulatory agents is to search for candidates among natural products because they have relatively low toxicities in clinical applications. Ginseng has been used as a traditional medicine in Asia and has demonstrated efficacy against various human diseases, such as viral infectious diseases, diabetes, atherosclerosis, and cancer. Recent studies and clinical cases have enhanced the interest in the potential biological effects of the ginseng berry, an association that appears to be due to the phytochemical content of this fruit. The ginseng berry has various bioactivities, such as anti-diabetic, anti-oxidation, anti-neuro degeneration, anti-inflammation, anti-cancer, and enhancement of sexual function bioactivities. Moreover, syringaresinol, the effective anti-aging component of the ginseng berry, has the ability to stimulate longevity via gene activation. Further molecular and clinical studies are necessary to uncover the numerous bioactive substances in the ginseng berry that contribute to public health.

Mild Mitochondrial Uncoupling Prevents Premature Senescence in Human Dermal Fibroblasts

TO THE EDITOR Mitochondrial uncoupling, an increased permeability of the inner membrane to protons not coupled to ATP synthesis, dissipates mitochondrial membrane potential. Mild mitochondrial uncoupling is believed to prolong lifespan of certain model organisms by reducing the production of reactive oxygen species (ROS) and preventing oxidant damage (Brand, 2000). Indeed, long-lived Caenorhabditis elegans strains have a lower membrane potential compared with wild-type strains (Lemire et al., 2009). Overexpression of the uncoupling protein UCP1 in murine skeletal muscle offers protection from age-related damage and disease (Gates et al., 2007). In addition, uncouplers extend the lifespan of yeast (Longo et al., 1999) and mice (Caldeira da Silva et al., 2008), suggesting that mild mitochondrial uncoupling might mitigate at least some deleterious aspects of aging. However, little is known about the effect of mitochondrial uncoupling on the aging of human skin. In this current study, we investigated the protective effect of mild mitochondrial uncoupling against oxidative stressinduced premature senescence in human dermal fibroblasts (HDFs). Oxidative stress is a pivotal mechanism leading to skin aging (Masaki, 2010). Accumulation of ROS elicits premature cellular senescence and deleterious alteration of collagen homeostasis, which can contribute to the development of characteristics of aged skin such as coarse, rough, and wrinkled appearance (Varani et al., 2006; Velarde et al., 2012). HDFs undergo cellular senescence by addition of 200mM H2O2 for 2 hours and prolonged subculture (Chen and Ames, 1994; Ido et al., 2012), which can be detected by changes in cellular morphology and senescence-associated b-galactosidase (SA-bgal) staining at 12 days after addition of H2O2 (Figure 1a). In order to determine whether mild mitochondrial uncoupling could inhibit oxidative stress–induced premature senescence, we pretreated HDFs with 60 nM carbonyl cyanide (P-trifluoromethoxy)-phenylhydrazone (FCCP) for 30 minutes before addition of H2O2. In a pilot study, the effective concentration of FCCP that would reduce the ROS production and mitochondrial membrane potential but have no discernible effect on cell viability was determined (Supplementary Figures S1 and S2 online). We found that cells pretreated with FCCP showed delayed senescence (Figure 1a), increased proliferative capacity (Figure 1b), reduced levels of oxidants (Figure 1c), and decreased expression levels of senescence-associated molecular markers, p21 and p16 (Figure 1d), compared with untreated senescent HDFs. Next, we investigated the uncoupling effect triggered by FCCP on collagen homeostasis in premature senescent HDFs. Aberrant collagen homeostasis, a prominent feature of aged human skin, Accepted article preview online 20 August 2013; published online 19 September 2013 Abbreviations: FCCP, carbonyl cyanide (p-trifluoromethoxy)-phenylhydrazone; HDF, human dermal fibroblasts; JNK, c-Jun N-terminal kinases; MMP-1, Matrix metalloproteinase-1; SA-bgal, senescenceassociated b-galactosidase SY Cho et al. Critical Role of Mitochondrial Uncoupling in Aging of Human Skin

Effects of Chitooligosaccharide Lactate Salt on Activity of Acetaldehyde Dehydrogenase

Chitooligosaccharides (COS), a kind of oligosaccharide made from chitin or chitosan, have been used a popular remedy for hangovers. In this study we investigated the in vitro effect of COS lactate salt on ethanol-induced cytotoxicity and the in vivo effect of short-term COS lactate salt feeding on ethanol-induced hangover. Pretreatment of HepG2 cells with COS lactate salt significantly reduced ethanol-induced cytotoxicity and suppressed generation of reactive oxygen species. In addition, COS lactate salt dose-dependently increased acetaldehyde dehydrogenase (ALDH) activity in vitro and reversed the ALDH inhibition induced by daidzin. Furthermore, oral administration of COS lactate salt (200 mg/kg) for 5 days significantly decreased the blood levels of alcohol and acetaldehyde in ethanol-treated mice. It was also demonstrated that hepatic mitochondrial ALDH activity was significantly increased in COS lactate salt-treated mice. Taken together, these findings indicate that COS lactate salt may have efficacy for the management of alcoholic hangovers.