Role of Oxidative Stress in the Etiology of Parkinson’s Disease: Advanced Therapeutic Products

Russian Journal of Bioorganic Chemistry - The etiology of Parkinson’s disease (PD) is not exactly known. However, oxidative stress was identified as a key factor that initiates and...     

Isolation of expressed in E. coli human interferon beta1b (Ser17) by ion-exchange chromatography

A method for isolation of interferon beta1b (Serl7) from inclusion bodies, comprising the steps of solution and reduction of protein from the inclusion bodies, refolding, chromatography on DEAE-Sepharose, chromatography on SP-Sepharose, concentrating, desalting and addition of stabilizers. The solution of reduced protein was diluted with pH 8.0 buffer of 50 mM Tris-HCl, 25 microM CuCl2 and 0.5% Twin 20 for refolding. We used gradient of pH (from 9.3 upto 11.3) for elution of interferon-beta from cation-exchange column. We concentrated of eluate and then desalted on the Sephadex G-50 column with 1 mM NaOH. Then the protein solution was neutralized with mannitol and Na-phosphate. Obtained preparation of interferon-beta was pure by gel-electrophoresis and by HPLC analysis, and had practically indentical level of antiproliferative activity with well-known preparation of Betaferone. Thus we show the possibility of isolation and obtaining of pure and active interferone-beta by ion-exchange chromatography in the presence of non-ion detergent Twin 20. We believe this method for interferon betalb preparation is perspective for scaling and using in the develop of industrial technology for production of this preparation.     

Interactions of β-lactoglobulin with serotonin and arachidonyl serotonin

β‐Lactoglobulin (β‐LG) is a lipocalin, which is the major whey protein of cow's milk and the milk of other mammals. However, it is absent from human milk. The biological function of β‐LG is not clear, but its potential role in carrying fatty acids through the digestive tract has been suggested. β‐LG has been found in complexes with lipids such as butyric and oleic acids and has a high affinity for a wide variety of compounds. Serotonin (5‐hydroxytryptamine, 5‐HT), an important compound found in animals and plants, has various functions, including the regulation of mood, appetite, sleep, muscle contraction, and some cognitive functions such as memory and learning. In this study, the interaction of serotonin and one of its derivatives, arachidonyl serotonin (AA‐5HT), with β‐LG was investigated using circular dichroism (CD) and fluorescence intensity measurements. These two ligands interact with β‐LG forming equimolar complexes. The binding constant for the serotonin/β‐LG interaction is between 10⁵ and 10⁶ M⁻¹, whereas for the AA‐5HT/β‐LG complex it is between 10⁴ and 10⁵ M⁻¹ as determined by measurements of either protein or ligand fluorescence. The observed binding affinities were higher in hydroethanolic media (25% EtOH). The interactions between serotonin/β‐LG and AA‐5HT/β‐LG may compete with self‐association (micellization) of both the ligand and the protein. According to far‐ and near‐UV CD results, these ligands have no apparent influence on β‐LG secondary structure, however they partially destabilize its tertiary structure. Their binding by β‐LG may be one of the peripheral mechanisms of the regulation of the content of serotonin and its derivatives in the bowel of milk‐fed animals. © 2011 Wiley Periodicals, Inc. Biopolymers 95: 871–880, 2011.     

Effects of N-Arachidonoyl Dopamine,Haloperidol, and Their Mixtures on Regeneration of Freshwater Hydra attenuata

N-Arachidonoyl dopamine and haloperidol, both separately and in different combinations, inhibit regeneration of the gastral and basal regions of hydra. In addition, both substances induce stable anomalies of morphogenesis in the form of outgrowths and additional tentacles in gastral regenerates. In the presence of both substances at different combinations, anomalies either do not appear altogether, or exist for a short time, thus suggesting the normalization pf morphogenesis. Possible mechanisms underlying the effects of these substances are discussed.     

Neuroprotective properties of endocannabinoids N-arachidonoyl dopamine and N-docosahexaenoyl dopamine examined in neuronal precursors derived from human pluripotent stem cells

Neuroprotective properties of endocannabinoids N-arachidonoyl dopamine (NADA) and N-docosahexaenoyl dopamine (DHDA) were examined in neuronal precursor cells differentiated from human induced pluripotent stem cells and subjected to oxidative stress. Both compounds exerted neuroprotective activity, which was enhanced by elevating the concentration of the endocannabinoids within the 0.1–10 μM range. However, both agents at 10 μM concentration showed a marked toxic effect resulting in death of ~30% of the cells. Finally, antagonists of cannabinoid receptors as well as the receptor of the TRPV1 endovanilloid system did not hamper the neuroprotective effects of these endocannabinoids.     

Synthesis of N-acyl derivatives of Pro-Gly-Pro-Leu peptide: Proteolytic stability in vitro and effects on mouse macrophage cells RAW264.7

Acetyl, oleoyl, arachidonoyl, and docosahexaenoyl derivatives of the Pro-Gly-Pro-Leu peptide with a chemical purity of 99.8% were synthesized. The degradation kinetics of the Pro-Gly-Pro-Leu derivatives under the action of leucine aminopeptidase, nasal mucus, and microsomal fraction of the brain and blood of rats was studied. It was shown that the N-acyl derivatives of Pro-Gly-Pro-Leu proved to be more resistant to the action of leucine aminopeptidase and other enzyme systems. The study of the cytotoxic and anti-inflammatory activity of preparations on the mouse macrophage cell line RAW264.7 showed that acylation with oleic and arachidonic acid makes the peptide cytotoxic with LC50 in the range of 70–15 μM and gives it anti-inflammatory properties with EC50 of 32 and 36 μM, respectively.     

Investigation of peptide stability upon hydrolysis by of fragments of the organs of the gastrointestinal tract of rats

The hydrolytic stability of a range of cyclic tripeptides, including the therapeutically important dalargin and stemokin, as well as peptides modified by ibuprofen and aspirin, has been studied. The first two experimental systems used utilized purified enzymes (pepsin, trypsin, and chymotrypsin), while the second one utilized fragments of the stomach and small intestine of rats. The linear peptides containing only L-amino acid residues were shown to be hydrolyzed by stomach and intestine fragments, although some of these peptides were resistant to hydrolysis by individual enzymes. The peptides containing D-amino acid residues and cyclic peptides were stable under all of the conditions used, but the peptides modified by aspirin lost the acetyl group of the aspirin moiety in acidic media, this process being accelerated in the presence of pepsin.     

Hydrolysis of anandamide and eicosapentaenoic acid ethanolamide in mouse splenocytes

The hydrolysis of anandamide has been studied in mouse splenocytes using tritiated anandamide analogs labeled in the acyl- or ethanolamide parts of the molecule. [3H]Anandamide undergoes rapid (t(1/2) = 2.5 min) uptake and hydrolysis, yielding ethanolamine and arachidonic acid. The anandamide hydrolysis in splenocytes is sensitive to inhibition by phenylmethylsulfonyl fluoride, and it is assumed that the observed activity is due to fatty acid amide hydrolase, which inactivates anandamide in central and peripheral tissues. Eicosapentaenoic acid ethanolamide and the 15-hydroxy-derivative of anandamide are shown to be amidohydrolase substrates as well. The fatty acids derived from hydrolytic cleavage of acylethanolamines undergo rapid oxidation by splenocyte lipoxygenase, yielding the corresponding 12-hydroxy-derivatives. Oxygenated ethanolamide derivatives were not found. The data suggest that polyenoic fatty acid ethanolamides are metabolic precursors of eicosanoids in splenocytes and that amide bond hydrolysis is the key point in switching of biological activity spectra between endocannabinoids and oxylipins.