Ommochromes from the Compound Eyes of Insects: Physicochemical Properties and Antioxidant Activity.

The objective of this study was screening of ommochromes from the compound eyes of insects and comparison of their antioxidant properties. Ommochromes were isolated in preparative quantities from insects of five different families: Stratiomyidae, Sphingidae, Blaberidae, Acrididae, and Tenebrionidae. The yield of ommochromes (dry pigment weight) was 0.9-5.4% of tissue wet weight depending on the insect species. Isolated pigments were analyzed by high-performance liquid chromatography and represented a mixture of several ommochromes of the ommatin series. The isolated ommochromes displayed a pronounced fluorescence with the emission maxima at 435-450 nm and 520-535 nm; furthermore, the emission intensity increased significantly upon ommochrome oxidation with hydrogen peroxide. The ommochromes produced a stable EPR signal consisting of a singlet line with g = 2.0045-2.0048, width of 1.20-1.27 mT, and high concentration of paramagnetic centers (> 1017 spin/g dry weight). All the investigated ommochromes demonstrated high antiradical activity measured from the degree of chemiluminescence quenching in a model system containing luminol, hemoglobin, and hydrogen peroxide. The ommochromes strongly inhibited peroxidation of the photoreceptor cell outer segments induced by visible light in the presence of lipofuscin granules from the human retinal pigment epithelium, as well as suppressed iron/ascorbate-mediated lipid peroxidation. The obtained results are important for understanding the biological functions of ommochromes in invertebrates and identifying invertebrate species that could be used as efficient sources of ommochromes for pharmacological preparations to prevent and treat pathologies associated with the oxidative stress development.

Implication of Integrin α2ß1 in Proliferation and Invasion of Human Breast Carcinoma and Melanoma Cells: Noncanonical Function of Akt Protein Kinase.

Blocking the expression of integrin α2ß1, which was accomplished by transduction of α2-specific shRNA, resulted in significant inhibition of proliferation and clonal activity in human MCF-7 breast carcinoma and SK-Mel-147 melanoma cells. Along with these changes, deprivation of α2ß1 caused a sharp decrease in melanoma cell invasion in vitro. Analysis of integrin-mediating signal pathways that control cell behavior revealed a significant increase in activity of Akt protein kinase in response to depletion of α2ß1. The increase in Akt activity that accompanies a suppressive effect on cell invasion contradicts well-known Akt function aimed at stimulation of tumor progression. This contradiction could be explained by the "reversed" (noncanonical) role played by Akt in some cells that consists in suppression rather than promotion of invasive phenotype. To test this suggestion, the effects of Akt inhibitors on invasive activity of SK-Mel-147 cells were investigated. If the above suggestion is true, then inhibition of Akt in cells depleted of α2ß1 should result in the restoration of their invasive activity. It appeared that treatment with LY294002, which inhibits all Akt isoforms (Akt1, Akt2, Akt3), not only failed to restore the invasive phenotype of melanoma cells but further attenuated their invasive activity. However, treatment of the cells with an Akt1-specific inhibitor significantly increased their invasion. Thus, the stimulating effect of α2ß1 integrin on invasion of melanoma cells is realized through a mechanism based on inhibition of one of the Akt isoforms, which in these cells exhibits a noncanonical function consisting in suppression of invasion.

Paramagnetics melanin and Mn2+ in black soldier fly Hermetia illucens.

Larva, prepupa (last instar larva), pupa, and an empty shell of pupa after hatching of the black soldier fly Hermetia illucens contain eumelanin, an active synthesis of which is observed at the prepupal stage, which is probably due to the release of prepupa from the feed substrate thickness to the open space for pupation. It was shown for the first time that prepupa contains high quantities of the magnetically active form of manganese Mn2+. This fact indicates that Mn2+ stimulates the copper-containing tyrosinase-the key enzyme in the synthesis of melanin in the period of migration and adaptation of the insect to the solar radiation.

Characteristics of lipid fractions of larvae of the black soldier fly Hermetia illucens.

The lipid fraction of larvae of the black soldier fly Hermetia illucens was shown to contain lauric acid (38.43 wt %) and its esters, azelaic and sebacic acids, and azelaic acid dibutyl ester. The dominant compound in the group of identified glycerides was lauric acid monoglyceride (0.70 wt %). Glycerides were also represented by triglycerides and diglycerides of lauric acid. Sterols were represented primarily by phytosterols (over 75%), the major of which was alpha-sitosterol (45%). The identified lipid complex composition is apparently determined by the biological characteristics of the fly Hermetia illucens and ensures antibacterial defence of larvae and stability of lipids at changing ambient temperature.

[Implication of integrin alpha5beta1 signal pathways in proliferation and apoptosis of MCF-7/Dox human breast carcinoma cells]. / Razlichiia mezhdu integrinom al'fa5beta1i retseptorom EGRF v signal'nykh putiakh, kontroliruiushchikh proliferatsiiu i apoptoz kletokMCF-7/Dox kartsinomy molochnoi zhelezy cheloveka.

In MCF-7/Dox human breast carcinoma cells, down-regulation of integrin alpha5beta1 and inhibition of epidermal growth factor receptor (EGFR) markedly reduced rates of cell proliferation. Mitotic cycle analysis showed that alpha5beta1 down-regulation resulted in cell cycle arrest at the S phase, followed by a significant increase in the population of apoptotic cells (subG1 population). Inhibition of EGFR activity also caused cell cycle arrest at the S-phase but without any increase in the subG1 population. Down-regulation of alpha5beta1 and EGFR inhibition resulted in a significant decrease of cell content of the active (phosphorylated) forms of FAK and Erk protein kinases. The data obtained suggest that alpha5beta1 integrin is implicated in cell growth control via inhibition of apoptotic cell death and through EGFR activation.

[BLOCKING SENSORIMOTOR DISORDERS IN STRIATIC DOPAMINERGIC DEFICIENT MICE BY THE EXTRACTS OF "REMEDY BEETLE" ALPHITOBIUS DIAPERINUS].

Five groups of C57Bl/6JSto mice (groups 1-5) were tested on a vertical pole 2 weeks since subcutaneous administration of proneurotoxin MPTP (40 mg per kg b. w.) to groups 2-5. Group 1 served as control. It is known that this MPTP dosage forms within 2 weeks sensorimotor disturbances similar to the initial stage of Parkinson's disease. One week before MPTP injection and during all the period after it (2 weeks) animals of groups 3-5 got one of the three extracts of biomass homogenate of the «remedy beetle¼ Alphitobius diaperinus as an additive to their food (8 g per 1 kg of food). The extracts were as follows: water (N 1), water-methanol (N 2) and water-methanol one after solid-phase extraction (N 3). All extracts were immobilized on a phytocarrier. Animals of groups 1 & 2 used ordinary food. Sensorimotor disturbances which could be clearly seen in the 2nd group of mice (MPTP) were absent in group 1 (control) as well as groups 3-5 which received an «antidote¼ with their food as one of the extracts of homogenate of the beetle biomass. The extract N 3 demonstrated maximal efficiency of all.

Features of the Effect of a Complex Probiotic with Bacillus Bacteria and the Larvae of Hermetia illucens Biomass on Mozambique Tilapia (Oreochromis mossambicus O. niloticus) and Russian Sturgeon (Acipenser gueldenstaedti) Fry.

A study of the effect of a complex probiotic preparation with bacteria of the genus Bacillus and a microdose of larvae of the fly Hermetia illucens on the physiological. parameters of fish has been carried out. It was established that introduction of the complex into artificial feeds for tilapia and Russian sturgeon juveniles during their incubation in recirculating aquaculture systems improves the digestion efficiency and increases the rate of fish growth and body mass accumulation. In tilapia, a decrease in the cholesterol level was revealed, while in sturgeon, an increase in the hemoglobin content was found. It was noted that a probiotic with biologically active agents H. illucens had a stimulating effect on the development of sexual gonads in female sturgeon fry, promoted an increase in the ovary mass, and provided for the development of eggs without morphological disturbances.

Influence of fucoidans and their derivatives on antitumor and phagocytic activity of human blood leucocytes.

The immunotropic activity of structurally different fucoidans and their derivatives towards isolated immune blood cells, effectors of innate immune system, was studied. The most potent effect was observed for high molecular weight fucoidan CF from the alga Chordaria flagelliformis, whose backbone is built of (1→3)-linked units of α-L-fucopyranose, and branches included residues of α-D-glucuronic acid and α-L-fucofuranose. This compound at the concentration of 0.05 mg/ml potentiated phagocytosis of Saccharomyces cerevisiae and Lactobacillus acidophilus by neutrophils, increasing relative quantity of phagocytes as well as their effectiveness. Along with this, 14% increase in the concentration of membrane-bound integrin CD11c molecules was observed. The systemic effect of CF at the dose of 0.01 mg/mouse i.p. led to potentiation of cytotoxic activity of spleen mononuclear leucocytes towards melanoma cells of line B16 by 1.9-fold and towards chronic myelogenous leukemia cells of line K-562 by 1.7-fold. These results indicate that fucoidan CF can stimulate anti-infective and antitumor activity of effectors of the innate immune system via CD11c integrins.

[Special effects of a complex probiotic containing cellulolytic bacteria Cellulomonas on actively growing rabbits].

It was shown that the association of probiotic bacteria of the genuses Bacillus and Cellulomonas form biolayers on the surface of beet marc particles. The positive effect of a fodder additive that contained the biolayer on the basis of a phytomatrix on the growth and development of young rabbits was shown. Feeding of animals with a mixed fodder that contained 0.1% preparation resulted in stimulation of digestion of all components of the food. Among other components of the mixed fodder, cellulose was digested most effectively. An increase in the biomass of symbiotic bacteria and enzymatic activity in the blindgut chymus was also observed. The positive nitrogen balance demonstrated an increase in the nitrogen content in animals and a decrease of its losses with excretion. The mechanism of response of the rabbit's organism to introduction of the complex probiotic preparation into the digestive tract is discussed.

[Implication of integrin alpha5beta1 in human breast carcinoma apoptosis and drug resistance].

Doxorubicin-resistant MCF-7Dox line, which is a derivative of the drug-sensitive MCF-7 human breast carcinoma line, differs from the latter by a strongly reduced expression of the alpha2beta1 integrin and a highly increased expression of the alpha5beta1 receptor. Silencing of this integrin in the MCF-7Dox cells by transfection with alpha5-specific siRNA markedly stimulated anoikis and increased sensitivity of the cells to doxorubicin. Alpha5beta1 silencing also leads to significant inhibition of the activity of kinases Akt and Erk2 in MCF-7Dox cells. Our results suggest that integrins alpha5beta1-induced signals, controlling distinct aspects of cell behavior, are conducted through the common signal pathways.

[Anaerobic solid-phase fermentation of plant substrates by Bacillus subtilis].

Solid-phase growth of Bacillus subtilis 8130 on cellulose-rich plant substrates (presscakes or pulp) under hypoxic conditions was accompanied by cellulose depolymerization, protein hydrolysis, and degradation of other plant components, including some processes of mixed-type carbohydrate fermentation. The bacterial fermentation yielded propionic, butyric, and hexanoic acids and butyric acid derivatives. The bacterial metabolism and fermentation degree can be characterized by the proportions of fatty acids in the reaction mixture. The product of sea buckthorn cake fermentation has a good sorption quality.

[Anticoagulant activity of fucoidans from brown algae].

The anticoagulant activity of polysaccharide fucoidans from 11 species of brown algae was studied. The anticoagulant activity was measured by the activated partial thromboplastin time (APTT), prothrombin time and thrombin time. Inhibitory action of fucoidans varied significantly from one species to another. Fucoidans from Laminaria saccharina and Fucus distichus showed high anticoagulant activities, while fucoidans from Cladosiphon okamuranus and Analipus japonicus were almost inactive. The fucoidan inhibitory effect on thrombin and factor Xa in the presence or in the absence of natural thrombin inhibitor, antithrombin III (AT III), was also studied. In contrast to the best studied anticoagulant heparin the most of the fucoidans inhibited thrombin in the absence of AT III. In the presence of AT III inhibitory effect of fucoidans was increased considerably. Unlike heparin, the effect of fucoidans on factor Xa was very weak in the presence of AT III and was not observed in the absence of AT III. The correlation between the anticoagulant activities of this series of fucoidans and their anti-inflammatory action, studied by us earlier, was not found. It is expected that two these types of fucoidan activities depend on different structural features of fucoidans. These findings show the possibility to obtain fucoidans with high anti-inflammatory action and with low anticoagulant activity. Anticoagulant activity of the fucoidans did not depend on the content of fucose, the other neutral sugars and sulfates in the preparations, and also on the structure of the backbone of molecule. Taken together, these results indicate on prospects of fucoidan study as potential therapeutic agents.

Implication of alpha5beta1 integrin in invasion of drug-resistant MCF-7/ADR breast carcinoma cells: a role for MMP-2 collagenase.

Expression of alpha5beta1 integrin in the drug-resistant MCF-7/ADR breast carcinoma cells was inhibited by treatment of these cells with alpha5-specific siRNA. The decrease of alpha5beta1 expression resulted in a sharp decrease of expression of MMP-2 collagenase and inhibition of invasion activity of these cells in vitro. Similar decrease of invasion was also observed during inhibition of MMP-2 expression by treatment of these cells with MMP-2-specific siRNA. Inhibition of alpha5beta1 expression was also accompanied by significant decrease in cell content of active (phosphorylated) forms of signal protein kinases Akt and Erk1/2. Inhibition of activity of these kinases by treatment of cells with PI-3K/Akt-specific inhibitor LY294002 or Erk-specific inhibitor PD98059 resulted in inhibition of MMP-2 expression and the decrease of invasion in vitro. These data suggest that alpha5beta1 controls invasion ability of these cells by regulating expression of MMP-2, which involves PI-3K and Erk1/2 protein kinase signaling.

[A study of the mechanisms of probiotic effect of Bacillus subtilis 8130 strain].

The wild-type Bacillus subtilis strain 8130 secreted metabolites that stimulated two to three times the growth of the test cultures of lactic acid bacteria. It exhibited endoglucanase activity that depended on the composition of nutrient medium. The addition of the product of two-stage culturing of B. subtilis 8130 to the diet of pigs (0.2% of fodder weight) made it possible to increase the daily weight gain by 19% and decrease the consumption of mixed fodder by 10%. Digestion of protein, fat, and other organic compounds increased by 3-4% and cellulose by 12%. It was shown that B. subtilis 8130 is a probiotic with targeted action stimulating digestion (primarily the digestion of cellulose). The enrichment of a dry-beer pellet with the product of solid-phase fermentation by bacillus (1 x 10(8) cells per gram dry pellet) allowed the pellet to entered into the diet of a calf (6% of the weight of fodder with probiotic), causing additional weight gain by 12% and a 10% economy of fodder consumption.

The role of beta1 integrin subfamily in anchorage-dependent apoptosis of breast carcinoma cells differing in multidrug resistance.

Integrin expression was investigated in MCF-7 human breast adenocarcinoma line and in the MCF-7Dox line, which was selected from MCF-7 by a resistance to multiple antitumor drugs (MDR). We have shown that acquisition of MDR was accompanied by a drastically reduced expression of some integrins of the beta1-subfamily (alpha2beta1, alpha3beta1, alpha6beta1) and of alpha vbeta5 intergin in the adenocarcinoma cells. In contrast, expression of alpha5beta1 integrin was markedly increased in the MDR cells. Along with multiple antitumor drug resistance, MCF-7Dox cells demonstrate elevated resistance to anchorage-dependent apoptosis (anoikis) and enhanced in vitro invasive activity. To elucidate the implication of beta1-integrins in the above phenotypic modifications, the effect of beta1-integrin signaling was assayed. Stimulation of beta1-mediated signaling was accomplished by treating of the cells with antibodies to the beta1-subunit common for members of the beta1-subfamily. These data show that activation of beta1-integrin signaling markedly upregulated anoikis of the adenocarcinoma cells.

Monomeric and multimeric blockers of selectins: comparison of in vitro and in vivo activity.

The potency of the oligosaccharides SiaLe(x), SiaLe(a), HSO(3)Le(x), and HSO(3)Le(a), their conjugates with polyacrylamide (PAA, 40 kD), and other monomeric and polymeric selectin inhibitors has been compared with that of the polysaccharide fucoidan. The following assay systems were used: 1) a 96-well assay based either on the use of recombinant E-, P-, and L-selectins or an analogous assay with natural P-selectin isolated from human platelets; 2) a platelet-based P-selectin cell assay; and 3) a rat model of peritoneal inflammation. IC(50) values for the neoglycoconjugate SiaLe(a)-PAA were 6, 40, and 85 microM for recombinant E-, P-, and L-selectins, respectively; all monomeric inhibitors were about two orders of magnitude weaker. PAA-conjugates, containing as a ligand tyrosine-O-sulfate (sTyr) in addition to one of the sialylated oligosaccharides, were the most potent synthetic blockers in vitro. Compared with fucoidan, the most potent known P- and L-selectin blocker, the bi-ligand glycoconjugate HSO(3)Le(a)-PAA-sTyr displayed similar inhibitory activity in vitro towards L-selectin and about ten times lower activity towards P-selectin. All of the tested synthetic polymers displayed a similar ability to inhibit neutrophil extravasation in the peritonitis model (in vivo) at 10 mg/kg. The data provide evidence that monomeric SiaLe(x) is considerably more effective as a selectin blocker in vivo than in vitro, whereas the opposite is true for fucoidan and the bi-ligand neoglycoconjugate HSO(3)Le(a)-PAA-sTyr.