3,5-dicaffeoylquinic acid lowers 3T3-L1 mitotic clonal expansion and adipocyte differentiation by enhancing heme oxygenase-1 expression

Adipogenesis is a complex process in which cell commitment and mitotic clonal expansion (MCE) are in-sequence crucial events leading to terminal adipocyte differentiation. The molecules able to block some key signals in this cascade can hamper adipogenesis becoming promising agents to counteract hyperplasia and hypertrophy of adipose tissue. Mono- and di-caffeoylquinic acid isomers are biologically active polyphenols, displaying in vitro and in vivo antioxidant, hepatoprotective, anti-diabetic and anti-obesity properties. Among these isomers, 3,5-dicaffeoylquinic acid (DCQA) has been reported to inhibit lipid accumulation in adipose cells more successfully than others. Thus, we investigated DCQA effects and molecular mechanisms on 3T3-L1 pre-adipocytes induced to differentiate with a hormonal cocktail (MDI). Oil Red O incorporation assessed that DCQA pre-treatment inhibited lipid accumulation in 3T3-L1 cells induced to differentiate for 10 days. At this time, an increased phosphorylation of both AMP-activated kinase and acetyl-CoA carboxylase, as well as a strong decrease in fatty acid synthase protein level, were registered by immunoblotting, thereby suggesting that DCQA treatment can reduce fatty acid anabolism in 3T3-L1 adipocytes. Furthermore, BrdU incorporation assay, performed 48 h after hormonal stimulation, revealed that DCQA treatment was also able to hinder the 3T3-L1 cell proliferation during the MCE, which is an essential step in the adipogenic process. Thus, we focused our attention on early signals triggered by the differentiation stimuli. In the first hours after hormonal cocktail administration, the activation of ERK1/2 and Akt kinases, or CREB and STAT3 transcription factors, was not affected by DCQA pre-treatment. Whereas 24 h after MDI induction, DCQA pre-treated cells showed increased level of the transcription factor Nrf2, that induced the expression of the antioxidant enzyme heme oxygenase 1 (HO-1). In control samples, the expression level of HO-1 was reduced 24 h after MDI induction in comparison with the higher amount of HO-1 protein found at 2 h. The HO-1 decrease was functional by allowing reactive oxygen species to boost and allowing cell proliferation induction at the beginning of MCE phase. Instead, in DCQA-treated cells the HO-1 expression was maintained at high levels for a further 24 h; in fact, its expression decreased only 48 h after MDI stimulation. The longer period in which HO-1 expression remained high led to a delay of the MCE phase, with a subsequent inhibition of both C/EBP-α expression and adipocyte terminal differentiation. In conclusion, DCQA counteracting an excessive adipose tissue expansion may become an attractive option in obesity treatment

PPAR-α Contributes to the Anti-Inflammatory Activity of Verbascoside in a Model of Inflammatory Bowel Disease in Mice

The previous results suggest that peroxisome proliferator-activated receptor-alpha (PPAR)-α, an intracellular transcription factor activated by fatty acids, plays a role in control of inflammation. There is persuasive epidemiological and experimental evidence that dietary polyphenols have anti-inflammatory activity. In this regard, it has been demonstrated that verbascoside (VB) functions as intracellular radical scavenger and reduces the microscopic and macroscopic signs of experimental colitis. With the aim to characterize the role of PPAR-α in VB-mediated anti-inflammatory activity, we tested the efficacy of VB in an experimental model of inflammatory bowel disease induced by dinitrobenzene sulfonic acid, comparing mice lacking PPAR-α (PPAR-αKO) with wild type (WT) mice. Results indicate that VB-mediated anti-inflammatory activity is weakened in PPAR-αKO mice, compared to WT controls, especially in the inhibition of neutrophil infiltration, intestinal permeability and colon injury. These results indicate that PPAR-α can contribute to the anti-inflammatory activity of VB in inflammatory bowel disease

Extracts obtained from hoodia gordonii cells lines, their preparation and use

The present invention refers to the preparation and use of the extracts for nutritional, pharmacological and cosmetic purposes. Said extracts are obtained from cell cultures of Hoodia gordonii IRB HGORD 42 having accession number DSMZ: DMS 17433. Furthermore, the present invention concerns the preparation and use of said extracts, for the production of drugs or nutritional or cosmetic substances, such extracts possessing appetite suppressant properties

Phenylpropanoid glycosides from plant cell cultures induce heme oxygenase 1 gene expression in a human keratinocyte cell line by affecting the balance of NRF2 and BACH1 transcription factors.

Phenylpropanoids have several highly significant biological properties in both plants and animals. Four phenylpropanoid glycosides (PPGs), verbascoside (VB), forsythoside B (FB), echinacoside (EC) and campneoside I (CP), were purified and tested for their capability to activate NRF2 and induce phase II cyto-protective enzymes in a human keratinocyte cell line (HaCaT). All four substances showed similar strong antioxidant and radical-scavenging activities as determined by diphenylpicrylhydrazyl assay. Furthermore, in HaCaT cells, FB and EC are strong activators of NRF2, the nuclear transcription factor regulating many phase II detoxifying and cytoprotective enzymes, such as heme oxygenase 1 (HMOX1). InHaCaT cells, FB and EC (200 microM) induced nuclear translocation of NRF2 protein after 24 h and reduced nuclear protein levels of BACH1, a repressor of the antioxidant response element. FB and EC greatly HMOX1mRNA levels by more than 40-fold in 72 h. Cytoplasmic HMOX1 protein levels were also increased at 48 h after treatment. VB was less active compared to FB and EC, and CP was slightly active only at later times of treatment.We suggest that hydroxytyrosol (HYD) could be a potential bioactivemetabolite of PPGs since HYD, in equimolar amounts to PGGs, is able to both activateHO-1 transcription andmodifyNrf2/Bach1 nuclear protein levels. This is in agreementwith the poor activity of CP,which contains aHYDmoietymodified by an O-methyl group. In conclusion, FB and EC fromplant cell culturesmay provide long-lasting skin protection by induction of phase II cytoprotective capabilities

Effects of potassium dichromate on ATP content of mammalian cells cultured in vitro.

In order to elucidate the mechanism of the cytotoxic activity of hexavalent chromium (Cr(VI)), the alterations of intracellular ATP levels induced by potassium dichromate in cultured hamster fibroblasts (BHK line) have been studied. Two kinds of treatment procedures were adopted: (1) BHK cell suspensions were exposed to 0.05--1.00 mM K2Cr2O7 in Hanks' balanced salt solution (BSS) for up to 180 min and ATP concentrations were determined immediately after the exposure to Cr(VI). A decrease of ATP content was observed with 0.25--12.00 mM K2Cr2O7 but only in the case of the highest dose was it related in a linear fashion to the duration of the treatment. (2) Cells were preincubated in BSS for 30 min with 0.05--1.00 mM dichromate. They were then reincubated in Eagle's minimal essential medium (MEM) for up to 180 min and ATP was measured at different time points. Immediately after the exposure to chromium all the treated cultures showed a depletion of ATP content. However while the cells treated with 0.25--0.25 mM dichromate rapidly resumed ATP levels very similar to that of the control, no recovery was detected in cells treated with 0.50 and 1.0 mM K2Cr2O7, even after 180 min. The observed effects have been attributed to the oxidizing activity of Cr(VI), which subtracts electrons from electron donors involved in metabolic pathways producing ATP, and to the ability of Cr(III), deriving from Cr(VI) reduction, to form stable coordination complexes with ATP precursors and enzymes involved in ATP synthesis

Metabolomics analysis reveals that the accumulation of specific secondary metabolites in Echinacea angustifolia cell cultured in vitro can be controlled by light

Echinacea angustifolia cell suspension culturesare usually grown and maintained in the dark, but we alsoexposed cells to light for one culture cycle (14 days) andthen compared the metabolomes of dark-grown and illu-minated cells by liquid chromatography\u2013mass spectrome-try. Among 256 signals, we putatively identified 159molecules corresponding to 56 different metabolites plustheir fragments, adducts and isotopologs. The E. angustifoliametabolome consisted mainly of caffeic acid deriva-tives, comprising (a) caffeic acid conjugated with tartaric,quinic and hexaric acids; and (b) caffeic acid conjugatedwith hydroxytyrosol glycosides (e.g., echinacoside, ver-bascoside and related molecules). Many of these metabo-lites have not been previously described in E. angustifolia,which currently lacks detailed metabolic profiles. Exposureto light significantly increased the levels of certain caffeicacid derivatives (particularly caffeoylquinic acids andhydroxytyrosol derivatives lacking rhamnose residues) andreduced the level of hydroxytyrosol derivatives withrhamnose residues, revealing that light specifically inhibitsthe rhamnosylation of caffeoyl phenylethanoid glycosides.These results are significant because they suggest that the metabolic profile of cell cultures can be manipulated bycontrolling simple environmental variables such as illu-mination to modulate the levels of potentially therapeuticcompounds

Activity and Stability Studies of Verbascoside, a Novel Antioxidant, in Dermo-Cosmetic and Pharmaceutical Topical Formulations

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