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The endometrial cycle in response to hormonal stimulation is essential for implantation. The female has endometrium that repeats this cycle through about half of a lifetime. The cycle includes three phases, proliferative, secretory, and menstrual, and each phase has distinct characteristics. The endometrial stromal cells (EnSCs) in each phase also have specialized characteristics, including cell cycle, morphologies, and cellular metabolic state. So we hypothesized that the cells in each phase have unique mitochondrial morphologies because they are generally linked to cellular metabolic state. To investigate the metabolic characteristics in each phase, we investigated the mitochondrial morphologies by transmission electron microscopy, oxygen consumption rate (OCR), and intracellular adenosine triphosphate (ATP) production. The decidualized EnSCs have shorter mitochondria than those in the proliferative phase. Besides, they also displayed distinct intracellular structural characteristics compared with the proliferative phase, such as ribosome-rich endoplasmic reticulum and increased formation of vesicles. OCR and luminescent ATP detection assay revealed that the basal respiration and ATP production in the decidualized EnSCs were lower than those in the proliferative phase. Thus, we concluded that morphological and intracellular structural changes were induced during the decidualization. Moreover, the decreased mitochondrial length was shown to correlate with decreased dependency on oxidative phosphorylation and ATP concentration in EnSCs.

Adult stem cells age during long-term in vitro culture, and neural stem cells (NSCs), which can self-renew and differentiate into neurons and glial cells, also display reduced differentiation potential after repeated passaging. However, the mechanistic details underlying this process remain unclear. In this study, we found that long-term in vitro culture of NSCs resulted in aging-related upregulation of inflammatory- and endoplasmic reticulum (ER) stress-related genes, including the proinflammatory cytokines interleukin (IL)1β and IL6, the senescence-associated enzyme matrix metallopeptidase 13 (MMP13), and the ER stress-responsive transcription factor activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP). However, the cyclic and transient induction of four reprogramming factors (POU domain, class 5, transcription factor 1, also known as octamer-binding transcription factor 4; SRY [sex determining region Y]-box 2; Kruppel-like factor 4; and myelocytomatosis oncogene; collectively referred to as OSKM) can inhibit NSC aging, as indicated by the decreased expression of the inflammatory and ER stress-related genes. We used ROSA-4F NSCs, which express OSKM from only one allele, to minimize the potential for full reprogramming or tumor formation during NSC rejuvenation. We expect that this novel rejuvenation method will enhance the potential of NSCs as a clinical approach to the treatment of neurological diseases.

Sarcopenia is prevalent as the aging population grows. Therefore, the need for supplements for the elderly is increasing. This study aimed to investigate the efficacy and mechanism of a

The incidence of sarcopenic obesity, muscle atrophy induced by obesity, has steadily increased and is emerging as a health problem. Although the anti-obesity effect of Codonopsis lanceolata (CL) is known, its efficacy against sarcopenic obesity has not been studied.We aimed to investigate the effect of CL on sarcopenic obesity and the changes in the related mechanisms to confirm the potential of CL as an effective natural therapeutic agent for sarcopenic obesity.C57BL/6 mice were fed a high-fat diet (HFD) for 9 weeks, and CL was administered for 6 weeks with HFD feeding. Body weight and grip strength were measured twice a week. After sacrifice, muscle fiber histological analysis, blood lipid analysis, muscle triglyceride extraction, western blot, and real-time PCR were performed. High-performance liquid chromatography (HPLC)-electrospray ionization (ESI)-mass spectrometry (MS) analysis and in vitro experiments using C2C12 cells were performed to verify the main and active compounds of CL. Confluent C2C12 cells were differentiated for 4 days, and then the main compound of CL was co-treated with palmitic acid for 24 h.CL reduced body weight, mass of three fat tissues (epididymal fat, mesenteric fat, and perirenal fat), adipocyte cross-sectional area (CSA), and improved insulin signaling. Simultaneously, CL improved grip strength, mass of three muscle tissues (quadriceps, gastrocnemius, and soleus), and muscle fiber CSA. These results were due to the recovery of both the phosphatidylinositol-3-kinase (PI3K)/ protein kinase B (Akt) signaling pathway and lipid metabolisms in skeletal muscle. Lipids accumulated in skeletal muscle interrupt the PI3K/Akt pathway, but CL reduced intramyocellular triglyceride concentration by restoring gene expression of factors related to triglyceride synthesis and fatty acid oxidation. Therefore, the activated PI3K/Akt pathway enhanced muscle protein synthesis by increasing phosphorylation of ribosomal protein S6 kinase 1 and eIF4E-binding protein 1 and suppressed muscle protein degradation by decreasing expression of muscle ring finger-1 and muscle atrophy F-box protein. In addition, tangshenoside I (TS) was verified as the main compound of CL by HPLC-ESI-MS analysis, and its efficacy of inhibiting myotube atrophy and lipid accumulation in myotubes was confirmed, verifying that TS is an active compound.CL is an effective natural material for sarcopenic obesity that suppresses muscle atrophy by inhibiting the accumulation of lipids in skeletal muscle through restoration of impaired PI3K/Akt pathway and lipid metabolism.

Sarcopenia, defined as loss of muscle mass and strength with age, becomes a public health concern as the elderly population increases. This study aimed to determine whether the mixture of soluble whey protein hydrolysate (WPH) andIn the first experiment, C57BL/6 mice were hindlimb immobilized for one-week and then administered WPH 800 mg/kg, GBE 100 mg/kg, WPH 800 mg/kg+ GBE 100 mg/kg mixture, andIn the first experiment, we confirmed the synergetic effect of WPH and GBE on muscle mass and identified that GBE was more effective on the protein synthesis side, and WPH tended to be slightly more effective for protein degradation. In the second experiment, among three different ratios, the WPH 800 mg/kg+ GBE 100 mg/kg was most effective for muscle mass and strength. The mixtures activated muscle protein synthesis via PI3K/Akt/mTORc1 pathway and inhibited muscle protein degradation via suppressing ubiquitin-proteasome system (UPS) and autophagy-lysosome system (ALS), and these effects were more GBE dose-dependent than WPH.The WPH and GBE mixture having a synergetic effect is a potential agent to prevent sarcopenia.

The native hepatocellular cancer (HCC) microenvironment is characterized by more hypoxic, hypoglycemic, and acidic conditions than those used in standard cell culture. This study aimed to investigate whether HCC cells cultured in more native conditions have an altered phenotype and drug sensitivity compared to those cultured in standard conditions. Six HCC cell lines were cultured in "standard" (21% O2, 25 mM glucose) or more "native" (1% O2, 5 mM glucose, 10 mM lactate) conditions. Cells were assessed for growth rates, cell cycle distribution, relevant metabolite and protein levels, genome-wide gene expression, mitochondrial DNA sequence and sensitivity to relevant drugs. Many differences in cellular and molecular phenotypes and drug sensitivity were observed between the cells. HCC cells cultured in native conditions had slower doubling times, increased HK2 and GLUT, lower PHDA and ATP levels, and mutations in mitochondrial DNA. Thirty-one genes, including the hypoxia-associated NDRG1, were differentially expressed between the cells. HCC patients in The Cancer Genome Atlas (TCGA) with tumors with a high score based on these 31 genes had a poorer prognosis than those with a low score (p = 0.002). From 90 comparisons of drug sensitivity, increased resistance and sensitivity for cells cultured in native conditions was observed in 14 (16%) and 8 (9%) comparisons respectively. In conclusion, cells cultured in more native conditions can have a more glycolytic and aggressive phenotype and varied drug sensitivity to those cultured in standard conditions, and may provide new insights to understanding tumor biology and drug development.

Sarcopenia is prevalent as the aging population grows. Therefore, the need for supplements for the elderly is increasing. This study aimed to investigate the efficacy and mechanism of a Panax ginseng berry extract (GBE) and soluble whey protein hydrolysate (WPH) mixture on a sarcopenia-related muscular deterioration in aged mice. Ten-month-old male C57BL/6J mice were administered three different doses of the GBE + WPH mixture for 8 weeks; 700 mg/kg, 900 mg/kg, and 1100 mg/kg. Grip strength, serum inflammatory cytokines level, and mass of muscle tissues were estimated. The deteriorating function of aging muscle was investigated via protein or gene expression. Grip strength and mass of three muscle tissues were increased significantly in a dose-dependent manner, and increased anti-inflammatory cytokine alleviated systemic inflammatory state. The mixture resolved the imbalance of muscle protein turnover through activation of the PI3K/Akt pathway and increased gene expression of the muscle regeneration-related factors, while decreasing myostatin, which interferes with muscle protein synthesis and regeneration. Furthermore, we confirmed that increased mitochondria number in muscle with the improvement of mitochondrial biogenesis. These physiological changes were similar to the effects of exercise.

Acquired resistance (AQR) to drug treatment occurs frequently in cancer patients and remains an impediment to successful therapy. The aim of this study was to gain insight into how AQR arises following the application of PI3K/mTOR inhibitors. H1975 lung cancer cells with EGFR T790M mutations that confer resistance to EGFR inhibitors underwent prolonged treatment with the PI3K/mTOR inhibitor, BEZ235. Monoclonal cells with stable and increased resistance to BEZ235 were obtained after 8 months treatment. These AQR clones showed class-specific resistance to PI3K/mTOR inhibitors, reduced G1 cell cycle arrest and impedance of migration following PI3K/mTOR inhibition, reduced PTEN expression and increased Akt and S6RP phosphorylation. Transcriptome analysis revealed the AQR clones had increased expression of the metabolite transporters SLC16A9 and SLC16A7, suggestive of altered cell metabolism. Subsequent experiments revealed that AQR clones possess features consistent with elevated glycolysis, including increased levels of glucose, lactate, glutamine, glucose dependence, GLUT1 expression, and rates of post-glucose extracellular acidification, and decreased levels of reactive oxygen species and rates of oxygen consumption. Combination treatment of BEZ235 with the glycolysis inhibitor 3-bromopyruvate was synergistic in AQR clones, but only additive in parental cells. DNA sequencing revealed the presence of a mitochondrial DNA (mtDNA) MT-C01 variant in AQR but not parental cells. Depletion of mitochondrial DNA in parental cells induced resistance to BEZ235 and other PI3K/mTOR inhibitors, and was accompanied by increased glycolysis. The results of this study provide the first evidence that a metabolic switch associated with mtDNA mutation can be an underlying mechanism for AQR.

During embryonic development, cells undergo changes in gene expression, signaling pathway activation/inactivation, metabolism, and intracellular organelle structures, which are mediated by mitochondria. Mitochondria continuously switch their morphology between elongated tubular and fragmented globular via mitochondrial fusion and fission. Mitochondrial fusion is mediated by proteins encoded byMfn1,Mfn2, andOpa1, whereas mitochondrial fission is mediated by proteins encoded byFis1andDnm1L. Here, we investigated the expression patterns of mitochondria-related genes during the differentiation of mouse embryonic stem cells (ESCs). Pluripotent ESCs maintain stemness in the presence of leukemia inhibitory factor (LIF) via the JAK-STAT3 pathway but lose pluripotency and differentiate in response to the withdrawal of LIF. We analyzed the expression levels of mitochondrial fusion- and fission-related genes during the differentiation of ESCs. We hypothesized that mitochondrial fusion genes would be overexpressed while the fission genes would be downregulated during the differentiation of ESCs. Though the mitochondria exhibited an elongated morphology in ESCs differentiating in response to LIF withdrawal, only the expression ofMfn2was increased and that ofDnm1Lwas decreased as expected, the other exceptions beingMfn1,Opa1, andFis1. Next, by comparing gene expression and mitochondrial morphology, we proposed an index that could precisely represent mitochondrial changes during the differentiation of pluripotent stem cells by analyzing the expression ratios of three fusion- and two fission-related genes. Surprisingly, increasedMfn2/Dnm1Lratio was correlated with elongation of mitochondria during the differentiation of ESCs. Moreover, application of this index to other specialized cell types revealed that neural stems cells (NSCs) and mouse embryonic fibroblasts (MEFs) showed increasedMfn2/Dnm1Lratio compared to ESCs. Thus, we suggest that theMfn2/Dnm1Lratio could reflect changes in mitochondrial morphology according to the extent of differentiation.

Background: The aim of the present study was to compare levels of fetuin-A, osteoprotegerin (OPG), and heat shock protein (HSP)70 according to the stage of chronic kidney disease (CKD), as well as to evaluate the association between serum fetuin-A, OPG, and HSP70 concentrations with respect to vascular stiffness and calcification in hemodialysis (HD) patients. Methods: We measured fetuin-A, OPG, and HSP70 in 35 healthy controls, 35 patients with stage 3 CKD, 35 patients with stage 4 CKD, and 81 HD patients. Using these data, we studied the association of fetuin-A, OPG, and HSP70 with clinical, biochemical, and vascular measures in HD patients. Results: Levels of OPG and HSP70 were higher and fetuin-A was lower in HD patients than in healthy controls. The cardio-ankle vascular index (CAVI) showed a positive correlation with OPG (r = 0.248, p = 0.040) and the OPG/fetuin-A ratio (r = 0.260, p = 0.031). The ankle-brachial index (ABI) showed a negative correlation with OPG (r = -0.245, p = 0.031) and the OPG/fetuin-A ratio (r = -0.267, p = 0.018). Intima-media thickness (IMT) showed a positive correlation with OPG (r = 0.273, p = 0.014) and the OPG/fetuin-A ratio (r = 0.269, p = 0.015). On stepwise multiple linear regression analyses, only the logarithmic function of the OPG/fetuin-A ratio was independently associated with CAVI (β = 13.325, SE = 6.247, p = 0.038). Conclusions: Our results demonstrate that OPG and the OPG/fetuin-A ratio are correlated with increased vascular stiffness and IMT in HD patients. In addition, the OPG/fetuin-A ratio was independently associated with vascular stiffness in HD patients.

Hepatocellular carcinoma (HCC) is an endemic disease globally, with a 5-year survival rate of 15%. Current treatment of late-stage HCC involves the use of chemotherapy and sorafenib, a multi-kinase inhibitor. Metabolic studies have indicated HCC has a high glycolysis rate, and is surrounded by a hypoglycemic microenvironment. Given the need for therapeutic options for HCC and its glycolytic nature, this study explored the activity of novel GLUT1 inhibitors in varied glucose concentrations. To assess target inhibition, [3H] deoxy-D-glucose (2DG) uptake assays were performed in HEK293 cells stably over-expressing GLUT1-4 individually. Six HCC cell lines were cultured in media with standard high glucose (HG:25mM) and low glucose/lactic acidosis (LGLA: 5mM glucose/20mM lactic acid) to mimic the hypoglycemic microenvironment. IC50s were assessed using the SRB assay, and other phenotypes using standard methods. Four novel GLUT1 inhibitors (IOM1-4) from the same chemical series were identified from diversity screening. The compounds exhibited differential potencies to GLUT proteins at the sub-μM level. The inhibitors reduced the uptake of 2DG, the extrusion of lactate, and increased apoptosis. In HCC cells, IC50s were lowest with IOM1 and 2 and highest with IOM4 (Table 1). HEP3B cells displayed exceptional sensitivity to the inhibitors. The ranking of potency according to the IC50s correlated with the 2DG uptake studies. There was no correlation between GLUT protein levels and IC50s. Significant differences in sensitivity to inhibitors were observed in cells cultured in HG and LGLA conditions. Cells cultured in LGLA had consistently lower glucose uptake compared those in HG conditions. In LGLA conditions, cells with low levels of reactive oxygen species (ROS) were more sensitive to GLUT1 inhibitors compared to cells with high ROS levels. In conclusion, GLUT1 inhibition can be influenced by microenvironment glucose levels and could be a strategy for HCC treatment. IC50 values (uM) for respective inhibitors in respective HCC cell linesCELL LINESInhibitorCultureC3AHEP3BHUH7PLCSKHEP1SNU449IOM1HG>5.00.8±0.040.3±0.06>5.01.4±0.451.7±0.38IOM1LGLA>5.00.2±0.134.0±1.20>5.00.1±0.021.6±0.28IOM2HG4.7±0.445.00.1±0.080.2±0.093.2±1.290.1±0.021.3±0.42IOM3HG>5.00.2±0.040.9±0.222.3±0.251.1±0.641.6±0.40IOM3LGLA>5.05.00.4±0.092.3±1.13IOM4HG>5.02.8±0.354.3±1.494.6±1.404.7±0.85>5.0IOM4LGLA>5.01.4±0.493.8±1.10>5.03.1±0.56>5.0 Citation Format: Bhaskar Bhattacharya, Sanamerjit S. Mann, Min Ji Han, Sarah HH Low, Gim Hwa Tan, Barry E. McGuinness, Sarah C. Trewick, Phillip M. Cowley, Alan Wise, Richie Soong. Identification and activity of novel GLUT1 inhibitors in hepatocellular carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1283.

Digital storytelling is the process in which stories are expressed in digital media, which is invaluable due to the limitless potential in the field of advertisement and education. An intriguing digital storytelling is to have engaging and high sensual stimulating animations that contain the narration required for the desired result. For this study, we deal with energy drinks which have recently had a high attention and rapidly growing markets. In particular, we focus on Jeju specific energy drinks along with a narration for hand-held devices such as mobile phones by implementing interactive animations which include a touch interface and a narration involved.