scholarly journals Isolation of Resveratrol from Vitis Viniferae Caulis and Its Potent Inhibition of Human Tyrosinase

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Jiaa Park ◽  
Yong Chool Boo
Keyword(s):  
Potent Inhibitor ◽  
Melanin Synthesis ◽  
Coumaric Acid ◽  
Solvent Fractionation ◽  
Skin Whitening ◽  
Hek 293 ◽  
293 Cells ◽  
Potent Inhibition ◽  
Rate Limiting ◽  
Potential Use

Tyrosinase (TYR) catalyzes rate-limiting reactions of cellular melanin synthesis, and its inhibitors are of commercial interest as potential skin whitening agents. However, the limited availability of human TYR makes the screening of TYR inhibitors difficult. To overcome this hurdle, we transformed nonmelanocytic human embryonic kidney (HEK) 293 cells to express human TYR constitutively. Using these cells as a source of human TYR, the ethanolic extracts of 52 medicinal plants grown in Korea were tested for human TYR activity, and the extract of Vitis Viniferae Caulis (dried stems of the grape tree,Vitis viniferaL.) was found to inhibit human TYR activity potently. An active compound was isolated from this extract by solvent fractionation followed by liquid column chromatography and identified as resveratrol by spectroscopic and chromatographic analyses. Resveratrol was determined to be a highly potent inhibitor of human TYR (IC50=0.39 μg mL−1) as compared with p-coumaric acid (IC50=0.66 μg mL−1) and arbutin (IC50>100 μg mL−1) and inhibited melanin synthesis by human epidermal melanocytes at subtoxic concentrations. This study suggests that resveratrol and resveratrol-containing extracts of Vitis Viniferae Caulis have a potential use as skin whitening agents.

scholarly journals Removal of Pollutants in Mine Wastewater by a Non-Cytotoxic Polymeric Bioflocculant from Alcaligenes faecalis HCB2

2019 ◽  
Vol 16 (20) ◽  
pp. 4001 ◽  
Author(s):  
Tsolanku Sidney Maliehe ◽  
Albertus Kotze Basson ◽  
Nkosinathi Goodman Dlamini
Keyword(s):  
Oxygen Demand ◽  
Alcaligenes Faecalis ◽  
Inoculum Size ◽  
Hek 293 ◽  
Flocculation Activity ◽  
293 Cells ◽  
Mass Proportion ◽  
Mine Wastewater ◽  
The Fourier Transform ◽  
Potential Use

Bioflocculation is a physicochemical technique often employed to efficiently remove colloidal water pollutants. Consequently, in this study, a bioflocculant was produced, characterised and applied to remove pollutants in mine wastewater. The maximum flocculation activity of 92% was recorded at 30 °C, pH 9.0 when maltose and urea were used as energy sources and 72 h of fermentation at the inoculum size of 1% (v/v). K+ proved to be a favourable cation. The bioflocculant yield of 4 g/L was obtained. Scanning electron microscopy illustrated a hexagonal-like structure of the bioflocculant. It is composed of carbohydrates and proteins in mass proportion of 88.6 and 9.5%, respectively. The Fourier transform infrared spectrum revealed the presence of hydroxyl, amide and amino functional groups. More than 73% of the bioflocculant was obtained after exposure to 600 °C using the thermogravimetric analyser. Human embryonic kidney 293 (HEK 293) cells exhibited 95% viability after being treated with 200 µg/µL of the bioflocculant. The flocculation mechanisms were proposed to be as a result of a double layer compression by K+, chemical reactions and bridging mechanism. The removal efficiencies of 59, 72, and 75% on biological oxygen demand, chemical oxygen demand and sulphur, were obtained respectively. Thus, the bioflocculant have potential use in wastewater treatment.

scholarly journals PKA phosphorylation of HERG protein regulates the rate of channel synthesis

2009 ◽  
Vol 296 (5) ◽  
pp. H1244-H1254 ◽  
Author(s):  
Jian Chen ◽  
Jakub Sroubek ◽  
Yamini Krishnan ◽  
Yan Li ◽  
Jinsong Bian ◽  
...  
Keyword(s):  
Herg Channel ◽  
Synthesis Rate ◽  
Protein Synthesis Rate ◽  
Protein Abundance ◽  
Hek 293 ◽  
Channel Protein ◽  
Disease States ◽  
293 Cells ◽  
Acute Changes ◽  
Rate Limiting

Acute changes in cAMP and protein kinase A (PKA) signaling can regulate ion channel protein activities such as gating. Effects on channels due to chronic PKA signaling, as in stress or disease states, are less understood. We examined the effects of prolonged PKA activity on the human ether- a- go- go-related gene (HERG) K+ channel in stably transfected human embryonic kidney (HEK)293 cells. Sustained elevation of cAMP by either chlorophenylthiol (CPT)-cAMP or forskolin increased the HERG channel protein abundance two- to fourfold within 24 h, with measurable difference as early as 4 h. The cAMP-induced augmentation was not due to changes in transcription and was specific for HERG compared with other cardiac K+ channels (Kv1.4, Kv1.5, Kir2.1, and KvLQT1). PKA activity was necessary for the effect on HERG protein and did not involve other cAMP signaling pathways. Direct PKA phosphorylation of the HERG protein was responsible for the cAMP-induced augmentation. Enhanced abundance of HERG protein was detected in endoplasmic reticulum-enriched, Golgi, and plasma membrane without significant changes in trafficking rates or patterns. An increase in the K+ current density carried by the HERG channel was also observed, but with a delay, suggesting that traffic to the surface is rate-limiting traffic. Acceleration of the HERG protein synthesis rate was the primary factor in the cAMP/PKA effect with lesser effects on protein stability. These results provide evidence for a novel mechanism whereby phosphorylation of a nascent protein dictates its rate of synthesis, resetting its steady-state abundance.

General lysosomal hydrolysis can process prorenin accurately

2014 ◽  
Vol 307 (5) ◽  
pp. R505-R513 ◽  
Author(s):  
Lucie K. Xa ◽  
Marie-Josée Lacombe ◽  
Chantal Mercure ◽  
Claude Lazure ◽  
Timothy L. Reudelhuber
Keyword(s):  
Renin Angiotensin System ◽  
Lysosomal Hydrolases ◽  
Angiotensin System ◽  
Hek 293 ◽  
Dense Granules ◽  
Amino Terminal ◽  
Active Renin ◽  
Rate Limiting Step ◽  
293 Cells ◽  
Rate Limiting

Renin, an aspartyl protease that catalyzes the rate-limiting step of the renin-angiotensin system, is first synthesized as an inactive precursor, prorenin. Prorenin is activated by the proteolytic removal of an amino terminal prosegment in the dense granules of the juxtaglomerular (JG) cells of the kidney by one or more proteases whose identity is uncertain but commonly referred to as the prorenin-processing enzyme (PPE). Because several extrarenal tissues secrete only prorenin, we tested the hypothesis that the unique ability of JG cells to produce active renin might be explained by the existence of a PPE whose expression is restricted to JG cells. We found that inducing renin production by the mouse kidney by up to 20-fold was not associated with the concomitant induction of candidate PPEs. Because the renin-containing granules of JG cells also contain several lysosomal hydrolases, we engineered mouse Ren1 prorenin to be targeted to the classical vesicular lysosomes of cultured HEK-293 cells, where it was accurately processed and stored. Furthermore, we found that HEK cell lysosomes hydrolyzed any artificial extensions placed on the protein and that active renin was extraordinarily resistant to proteolytic degradation. Altogether, our results demonstrate that accurate processing of prorenin is not restricted to JG cells but can occur in classical vesicular lysosomes of heterologous cells. The implication is that renin production may not require a specific PPE but rather can be achieved by general hydrolysis in the lysosome-like granules of JG cells.

scholarly journals Increase of 5-HT levels is induced both in mouse brain and HEK-293 cells following their exposure to a non-viral tryptophan hydroxylase construct

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Emiliano Tesoro-Cruz ◽  
Leticia Manuel-Apolinar ◽  
Norma Oviedo ◽  
Sandra Orozco-Suárez ◽  
Minerva Crespo Ramírez ◽  
...  
Keyword(s):  
Brain Function ◽  
Tryptophan Hydroxylase ◽  
Impulsive Behavior ◽  
Hek 293 ◽  
Hek 293 Cells ◽  
293 Cells ◽  
Serotonin Deficiency ◽  
Rate Limiting ◽  
The Brain

AbstractTryptophan hydroxylase type 2 (Tph2) is the rate-limiting enzyme for serotonin (5-HT) biosynthesis in the brain. Dysfunctional Tph2 alters 5-HT biosynthesis, leading to a deficiency of 5-HT, which could have repercussions on human behavior. In the last decade, several studies have associated polymorphisms of the TPH2 gene with suicidal behavior. Additionally, a 5-HT deficiency has been implicated in various psychiatric pathologies, including alcoholism, impulsive behavior, anxiety, and depression. Therefore, the TPH2 gene could be an ideal target for analyzing the effects of a 5-HT deficiency on brain function. The aim of this study was to use the construct pIRES-hrGFP-1a-Tph2-FLAG to treat CD1-male mice and to transfect HEK-293-cells and then to evaluate whether this treatment increases 5-HT production. 5-HT levels were enhanced 48 h post-transfection, in HEK-293 cells. Three days after the ocular administration of pIRES-hrGFP-1a-Tph2-FLAG to mice, putative 5-HT production was significantly higher than in the control in both hypothalamus and amygdala, but not in the brainstem. Further research will be needed on the possible application of this treatment for psychiatric diseases involving a Tph2 dysfunction or serotonin deficiency.

scholarly journals Species-Specific Regulation of TRPM2 by PI(4,5)P2 via the Membrane Interfacial Cavity

2021 ◽  
Vol 22 (9) ◽  
pp. 4637
Author(s):  
Daniel Barth ◽  
Andreas Lückhoff ◽  
Frank J. P. Kühn
Keyword(s):  
Amino Acid Residues ◽  
Hek 293 ◽  
Complete Inactivation ◽  
293 Cells ◽  
Activation Mechanisms ◽  
Specific Regulation ◽  
Species Specific ◽  
Inside Out ◽  
Positively Charged

The human apoptosis channel TRPM2 is stimulated by intracellular ADR-ribose and calcium. Recent studies show pronounced species-specific activation mechanisms. Our aim was to analyse the functional effect of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), commonly referred to as PIP2, on different TRPM2 orthologues. Moreover, we wished to identify the interaction site between TRPM2 and PIP2. We demonstrate a crucial role of PIP2, in the activation of TRPM2 orthologues of man, zebrafish, and sea anemone. Utilizing inside-out patch clamp recordings of HEK-293 cells transfected with TRPM2, differential effects of PIP2 that were dependent on the species variant became apparent. While depletion of PIP2 via polylysine uniformly caused complete inactivation of TRPM2, restoration of channel activity by artificial PIP2 differed widely. Human TRPM2 was the least sensitive species variant, making it the most susceptible one for regulation by changes in intramembranous PIP2 content. Furthermore, mutations of highly conserved positively charged amino acid residues in the membrane interfacial cavity reduced the PIP2 sensitivity in all three TRPM2 orthologues to varying degrees. We conclude that the membrane interfacial cavity acts as a uniform PIP2 binding site of TRPM2, facilitating channel activation in the presence of ADPR and Ca2+ in a species-specific manner.

scholarly journals Preliminary Study on Pasta Samples Characterized in Antioxidant Compounds and Their Biological Activity on Kidney Cells

Nutrients ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 1131 ◽  
Author(s):  
Federico Di Marco ◽  
Francesco Trevisani ◽  
Pamela Vignolini ◽  
Silvia Urciuoli ◽  
Andrea Salonia ◽  
...  
Keyword(s):  
Indoleacetic Acid ◽  
Antioxidant Properties ◽  
Antioxidant Compounds ◽  
Kidney Cells ◽  
Hek 293 ◽  
293 Cells ◽  
High Concentrations ◽  
Β Carotene ◽  
Egg Pasta

Pasta is one of the basic foods of the Mediterranean diet and for this reason it was chosen for this study to evaluate its antioxidant properties. Three types of pasta were selected: buckwheat, rye and egg pasta. Qualitative–quantitative characterization analyses were carried out by HPLC-DAD to identify antioxidant compounds. The data showed the presence of carotenoids such as lutein and polyphenols such as indoleacetic acid, (carotenoids from 0.08 to 0.16 mg/100 g, polyphenols from 3.7 to 7.4 mg/100 g). To assess the effect of the detected metabolites, in vitro experimentation was carried out on kidney cells models: HEK-293 and MDCK. Standards of β-carotene, indoleacetic acid and caffeic acid, hydroalcoholic and carotenoid-enriched extracts from samples of pasta were tested in presence of antioxidant agent to determine viability variations. β-carotene and indoleacetic acid standards exerted a protective effect on HEK-293 cells while no effect was detected on MDCK. The concentrations tested are likely in the range of those reached in body after the consumption of a standard pasta meal. Carotenoid-enriched extracts and hydroalcoholic extracts showed different effects, observing rescues for rye pasta hydroalcoholic extract and buckwheat pasta carotenoid-enriched extract, while egg pasta showed milder dose depending effects assuming pro-oxidant behavior at high concentrations. The preliminary results suggest behaviors to be traced back to the whole phytocomplexes respect to single molecules and need further investigations.

scholarly journals Electron Paramagnetic Resonance Gives Evidence for the Presence of Type 1 Gonadotropin-Releasing Hormone Receptor (GnRH-R) in Subdomains of Lipid Rafts

Molecules ◽  
2021 ◽  
Vol 26 (4) ◽  
pp. 973
Author(s):  
Tilen Koklič ◽  
Alenka Hrovat ◽  
Ramon Guixà-González ◽  
Ismael Rodríguez-Espigares ◽  
Damaris Navio ◽  
...  
Keyword(s):  
Lipid Rafts ◽  
Order Parameter ◽  
Spin Probe ◽  
Gonadotropin Releasing Hormone ◽  
Paramagnetic Resonance ◽  
Hek 293 ◽  
293 Cells ◽  
Gonadotropin Releasing Hormone Receptor ◽  
Electron Paramagnetic

This study investigated the effect of type 1 gonadotropin releasing hormone receptor (GnRH-R) localization within lipid rafts on the properties of plasma membrane (PM) nanodomain structure. Confocal microscopy revealed colocalization of PM-localized GnRH-R with GM1-enriched raft-like PM subdomains. Electron paramagnetic resonance spectroscopy (EPR) of a membrane-partitioned spin probe was then used to study PM fluidity of immortalized pituitary gonadotrope cell line αT3-1 and HEK-293 cells stably expressing GnRH-R and compared it with their corresponding controls (αT4 and HEK-293 cells). Computer-assisted interpretation of EPR spectra revealed three modes of spin probe movement reflecting the properties of three types of PM nanodomains. Domains with an intermediate order parameter (domain 2) were the most affected by the presence of the GnRH-Rs, which increased PM ordering (order parameter (S)) and rotational mobility of PM lipids (decreased rotational correlation time (τc)). Depletion of cholesterol by methyl-β-cyclodextrin (methyl-β-CD) inhibited agonist-induced GnRH-R internalization and intracellular Ca2+ activity and resulted in an overall reduction in PM order; an observation further supported by molecular dynamics (MD) simulations of model membrane systems. This study provides evidence that GnRH-R PM localization may be related to a subdomain of lipid rafts that has lower PM ordering, suggesting lateral heterogeneity within lipid raft domains.

scholarly journals Characteristics and requirements of basal autophagy in HEK 293 cells

Autophagy ◽  
2013 ◽  
Vol 9 (9) ◽  
pp. 1407-1417 ◽  
Author(s):  
Patience Musiwaro ◽  
Matthew Smith ◽  
Maria Manifava ◽  
Simon A. Walker ◽  
Nicholas T. Ktistakis
Keyword(s):  
Hek 293 ◽  
Hek 293 Cells ◽  
293 Cells

scholarly journals Using kICS to Reveal Changed Membrane Diffusion of AQP-9 Treated with Drugs

Membranes ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 568
Author(s):  
Jakob L. Kure ◽  
Thommie Karlsson ◽  
Camilla B. Andersen ◽  
B. Christoffer Lagerholm ◽  
Vesa Loitto ◽  
...  
Keyword(s):  
Diffusion Coefficient ◽  
Plasma Membrane ◽  
Membrane Proteins ◽  
Actin Polymerization ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Membrane Diffusion ◽  
Hek 293 ◽  
293 Cells ◽  
Aquaporin 9

The formation of nanodomains in the plasma membrane are thought to be part of membrane proteins regulation and signaling. Plasma membrane proteins are often investigated by analyzing the lateral mobility. k-space ICS (kICS) is a powerful image correlation spectroscopy (ICS) technique and a valuable supplement to fluorescence correlation spectroscopy (FCS). Here, we study the diffusion of aquaporin-9 (AQP9) in the plasma membrane, and the effect of different membrane and cytoskeleton affecting drugs, and therefore nanodomain perturbing, using kICS. We measured the diffusion coefficient of AQP9 after addition of these drugs using live cell Total Internal Reflection Fluorescence imaging on HEK-293 cells. The actin polymerization inhibitors Cytochalasin D and Latrunculin A do not affect the diffusion coefficient of AQP9. Methyl-β-Cyclodextrin decreases GFP-AQP9 diffusion coefficient in the plasma membrane. Human epidermal growth factor led to an increase in the diffusion coefficient of AQP9. These findings led to the conclusion that kICS can be used to measure diffusion AQP9, and suggests that the AQP9 is not part of nanodomains.

Halothane Inhibition of Recombinant Cardiac L-type Ca2+ Channels Expressed in HEK-293 Cells

2005 ◽  
Vol 103 (6) ◽  
pp. 1156-1166 ◽  
Author(s):  
Kevin J. Gingrich ◽  
Son Tran ◽  
Igor M. Nikonorov ◽  
Thomas J. Blanck
Keyword(s):  
Charge Transfer ◽  
Calcium Channels ◽  
Dependent Manner ◽  
Current Time ◽  
Hek 293 ◽  
Hek 293 Cells ◽  
293 Cells ◽  
Dependent Inactivation ◽  
Voltage Dependent

Background Volatile anesthetics depress cardiac contractility, which involves inhibition of cardiac L-type calcium channels. To explore the role of voltage-dependent inactivation, the authors analyzed halothane effects on recombinant cardiac L-type calcium channels (alpha1Cbeta2a and alpha1Cbeta2aalpha2/delta1), which differ by the alpha2/delta1 subunit and consequently voltage-dependent inactivation. Methods HEK-293 cells were transiently cotransfected with complementary DNAs encoding alpha1C tagged with green fluorescent protein and beta2a, with and without alpha2/delta1. Halothane effects on macroscopic barium currents were recorded using patch clamp methodology from cells expressing alpha1Cbeta2a and alpha1Cbeta2aalpha2/delta1 as identified by fluorescence microscopy. Results Halothane inhibited peak current (I(peak)) and enhanced apparent inactivation (reported by end pulse current amplitude of 300-ms depolarizations [I300]) in a concentration-dependent manner in both channel types. alpha2/delta1 coexpression shifted relations leftward as reported by the 50% inhibitory concentration of I(peak) and I300/I(peak)for alpha1Cbeta2a (1.8 and 14.5 mm, respectively) and alpha1Cbeta2aalpha2/delta1 (0.74 and 1.36 mm, respectively). Halothane reduced transmembrane charge transfer primarily through I(peak) depression and not by enhancement of macroscopic inactivation for both channels. Conclusions The results indicate that phenotypic features arising from alpha2/delta1 coexpression play a key role in halothane inhibition of cardiac L-type calcium channels. These features included marked effects on I(peak) inhibition, which is the principal determinant of charge transfer reductions. I(peak) depression arises primarily from transitions to nonactivatable states at resting membrane potentials. The findings point to the importance of halothane interactions with states present at resting membrane potential and discount the role of inactivation apparent in current time courses in determining transmembrane charge transfer.

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