RESUMEN
Cyclic nigerosyl nigerose (CNN) is a cyclic tetrasaccharide that exhibits properties distinct from other conventional cyclodextrins. Herein, we demonstrate that treatment of B16 melanoma with CNN results in a dose-dependent decrease in melanin synthesis, even under conditions that stimulate melanin synthesis, without significant cytotoxity. The effects of CNN were prolonged for more than 27 days, and were gradually reversed following removal of CNN. Undigested CNN was found to accumulate within B16 cells at relatively high levels. Further, CNN showed a weak but significant direct inhibitory effect on the enzymatic activity of tyrosinase, suggesting one possible mechanism of hypopigmentation. While a slight reduction in tyrosinase expression was observed, tyrosinase expression was maintained at significant levels, processed into a mature form, and transported to late-stage melanosomes. Immunocytochemical analysis demonstrated that CNN treatment induced drastic morphological changes of Pmel17-positive and LAMP-1-positive organelles within B16 cells, suggesting that CNN is a potent organelle modulator. Colocalization of both tyrosinase-positive and LAMP-1-positive regions in CNN-treated cells indicated possible degradation of tyrosinase in LAMP-1-positive organelles; however, that possibility was ruled out by subsequent inhibition experiments. Taken together, this study opens a new paradigm of functional oligosaccharides, and offers CNN as a novel hypopigmenting molecule and organelle modulator.
Asunto(s)
Ciclodextrinas/farmacología , Glucanos/farmacología , Hipopigmentación/patología , Melanoma Experimental/patología , Animales , Western Blotting , Línea Celular Tumoral , Glucosamina/farmacología , Inmunohistoquímica , Lisosomas/efectos de los fármacos , Lisosomas/metabolismo , Melaninas/biosíntesis , Melanoma Experimental/metabolismo , Melanosomas/efectos de los fármacos , Melanosomas/metabolismo , Ratones , Monofenol Monooxigenasa/metabolismo , Presión Osmótica , Estrés Fisiológico/efectos de los fármacosRESUMEN
Our group recently demonstrated that simultaneous administration of trehalose with a high-fat diet (HFD) suppresses adipocyte hypertrophy and mitigates insulin resistance in mice. For the present study, we hypothesized that similar effects of trehalose would be observed in mice with previously-established obesity. Obese mice were fed a HFD and drinking water containing 0.3 or 2.5% (weight/volume) trehalose or distilled water (DW) ad libitum for 8 wk. After 7 wk intake of a HFD and trehalose, fasting serum insulin levels and homeostasis model assessment-insulin resistance (HOMA-IR) in the 0.3% Tre/HFD group were significantly lower than those in the DW/HFD group (p<0.05). After 8 wk of treatment, mesenteric adipocytes in the 0.3% Tre/HFD group showed significantly less hypertrophy than those in the DW/HFD group. Mechanistic analysis indicated that levels of high molecular weight (HMW) adiponectin in the serum of the 0.3% Tre/HFD group were significantly higher than those in the DW/HFD group. The expression levels of insulin receptor substrate-1 (IRS-1) and insulin receptor substrate-2 (IRS-2) messenger RNA (mRNA) in muscle were also significantly increased by trehalose intake. Our data therefore suggest that administration of trehalose to obese mice mitigates insulin resistance by suppressing adipocyte hypertrophy and increasing serum HMW adiponectin, resulting in upregulation of IRS-1, and IRS-2 expression in muscle. These results further suggest that trehalose is a functional saccharide that may be used to prevent the progression of insulin resistance.
Asunto(s)
Suplementos Dietéticos , Hiperinsulinismo/dietoterapia , Resistencia a la Insulina , Grasa Intraabdominal/patología , Síndrome Metabólico/prevención & control , Obesidad/fisiopatología , Trehalosa/uso terapéutico , Adiponectina/agonistas , Adiponectina/sangre , Adiponectina/metabolismo , Adiposidad , Animales , Dieta Alta en Grasa/efectos adversos , Femenino , Regulación de la Expresión Génica , Hiperinsulinismo/etiología , Hiperinsulinismo/patología , Hiperinsulinismo/fisiopatología , Hipertrofia , Proteínas Sustrato del Receptor de Insulina/agonistas , Proteínas Sustrato del Receptor de Insulina/genética , Proteínas Sustrato del Receptor de Insulina/metabolismo , Grasa Intraabdominal/metabolismo , Síndrome Metabólico/etiología , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/metabolismo , Obesidad/etiología , Distribución Aleatoria , Trehalosa/administración & dosificaciónRESUMEN
Hepatocyte growth factor (HGF) is useful as a potential therapeutic agent for hepatic and renal fibrosis and cardiovascular diseases through inducing proliferation of epithelial and endothelial cells. HGF inducers may also be useful as therapeutic agents for these diseases. However, there have been no reports on induction of HGF production by plant extracts or juices. An extract of bitter melon (Momordica charantia L.) pulp markedly induced HGF production. There was a time lag of 72 h before induction of HGF production after the extract addition. Its stimulatory effect was accompanied by upregulation of HGF gene expression. Increases in mitogen-activated protein kinases (MAPKs) were observed from 72 h after the extract addition. Inhibitors of MAPKs suppressed the extract-induced HGF production. The extract also stimulated cell proliferation. Both activities for induction of HGF production and cell proliferation were eluted together in a single peak with 14,000 Da on gel filtration. The results indicate that bitter melon pulp extract induced HGF production and cell proliferation of human dermal fibroblasts and suggest that activation of MAPKs is involved in the HGF induction. Our findings suggest potential usefulness of the extract for tissue regeneration and provide an insight into the molecular mechanism underlying the wound-healing property of bitter melon.
Asunto(s)
Cucurbitaceae/metabolismo , Fibroblastos/metabolismo , Regulación de la Expresión Génica , Factor de Crecimiento de Hepatocito/biosíntesis , Piel/citología , Proliferación Celular , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Modelos Biológicos , Fosforilación , Extractos Vegetales/metabolismo , Piel/metabolismo , Sales de Tetrazolio/farmacología , Tiazoles/farmacología , Factores de TiempoRESUMEN
Carboxylic acids have various biological activities and play critical roles in cellular metabolic pathways such as the tricarboxylic acid (TCA) cycle. It has been shown that some carboxylic acids induce cell proliferation and production of cytokines or growth factors. However, there have been no reports on effects of carboxylic acids on hepatocyte growth factor (HGF) expression. In this study, we found that only maleic acid among various carboxylic acids examined markedly induced HGF production from human dermal fibroblasts. Maleic acid also induced HGF production from human lung fibroblasts and neuroblastoma cells. The stimulatory effect was accompanied by upregulation of HGF gene expression. Increase in phosphorylation of extracellular signal-regulated protein kinase (ERK) and c-Jun N-terminal kinase (JNK) but not in phosphorylation of p38 was observed from 6 h and up to 24 h after maleic acid addition. The ERK kinase inhibitor PD98059 and the JNK inhibitor SP600125 potently inhibited maleic acid-induced HGF production, while the p38 inhibitor SB203580 did not significantly inhibit the production. The protein synthesis inhibitor cycloheximide completely inhibited upregulation of HGF mRNA induced by maleic acid but superinduced HGF mRNA expression upregulated by 12-O-tetradecanoylphorbol 13-acetate (TPA). These results suggest that maleic acid indirectly induced HGF expression from human dermal fibroblasts through activation of ERK and JNK and that de novo protein synthesis is required for maleic acid-induced upregulation of HGF mRNA.
Asunto(s)
Fibroblastos/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Factor de Crecimiento de Hepatocito/genética , Maleatos/farmacología , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Células Cultivadas , Fibroblastos/citología , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Fosforilación , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Hepatocyte growth factor (HGF), which is produced by surrounding stromal cells, including fibroblasts and endothelial cells, has been shown to be a significant factor responsible for cancer cell invasion mediated by tumor-stromal interactions. We found in this study that the anti-tumor agent valproic acid (VPA), a histone deacetylase (HDAC) inhibitor, strongly inhibited tumor-stromal interaction. VPA inhibited HGF production in fibroblasts induced by epidermal growth factor (EGF), platelet-derived growth factor, basic fibroblast growth factor, phorbol 12-myristate 13-acetate (PMA) and prostaglandin E(2) without any appreciable cytotoxic effect. Other HDAC inhibitors, including butyric acid and trichostatin A (TSA), showed similar inhibitory effects on HGF production stimulated by various inducers. Up-regulations of HGF gene expression induced by PMA and EGF were also suppressed by VPA and TSA. Furthermore, VPA significantly inhibited HGF-induced invasion of HepG2 hepatocellular carcinoma cells. VPA, however, did not affect the increases in phosphorylation of MAPK and Akt in HGF-treated HepG2 cells. These results demonstrated that VPA inhibited two critical processes of tumor-stromal interaction, induction of fibroblastic HGF production and HGF-induced invasion of HepG2 cells, and suggest that those activities serve for other anti-tumor mechanisms of VPA besides causing proliferation arrest, differentiation, and/or apoptosis of tumor cells.
Asunto(s)
Carcinoma Hepatocelular/metabolismo , Comunicación Celular/efectos de los fármacos , Fibroblastos/metabolismo , Factor de Crecimiento de Hepatocito/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Receptores de Factores de Crecimiento/metabolismo , Células del Estroma/metabolismo , Ácido Valproico/administración & dosificación , Línea Celular , Relación Dosis-Respuesta a Droga , Fibroblastos/efectos de los fármacos , Humanos , Proteínas Proto-Oncogénicas c-met , Transducción de Señal/efectos de los fármacosRESUMEN
Hepatocyte growth factor (HGF) stimulates the proliferation of hepatocytes and biliary epithelial cells and protects hepatocytes from apoptosis induced by various stimuli. In view of HGF induction by interferons, substances used for the treatment of chronic hepatitis C, this study was conducted to determine whether ursodeoxycholic acid (UDCA), which is widely used for the treatment of cholestatic liver diseases, modulates HGF production. UDCA did not induce HGF production in human dermal fibroblasts, but it potently inhibited phorbol-12-myristate-13-acetate (PMA)- and cholera-toxin-induced HGF production without affecting cell viability. The inhibitory effects of UDCA were as potent as those of transforming growth factor-beta1 and dexamethasone. Up-regulations of HGF gene expression induced by PMA and cholera toxin were also inhibited by UDCA. Moreover, UDCA dose-dependently inhibited high constitutive HGF production by MRC-5 cells without decreasing cell viability. Deoxycholate, chenodeoxycholate, taurochenodeoxycholate and glycochenodeoxycholate also inhibited cholera-toxin-induced HGF production at non-cytotoxic doses. UDCA, however, had no apparent effect on PMA-induced phosphorylation of mitogen-activated protein kinase, which is crucial for HGF induction by PMA. These results indicate that non-cytotoxic doses of UDCA inhibited constitutive and induced HGF production and suggest that UDCA supplemented with HGF or HGF inducers could have a more potential therapeutic effect.