Ivy Leaf Dry Extract EA 575® Has an Inhibitory Effect on the Signalling Cascade of Adenosine Receptor A2B.

Ivy leaf dry extract EA 575® is used to improve complaints of chronic inflammatory bronchial diseases and acute inflammation of the respiratory tract accompanied by coughing. Its mechanism of action has so far been explained by influencing ß2-adrenergic signal transduction. In the present study, we investigated a possible influence on adenosine receptor A2B (A2BAR) signalling, as it has been described to play a significant and detrimental role in chronic inflammatory airway diseases. The influence of EA 575® on A2BAR signalling was assessed with measurements of dynamic mass redistribution. Subsequently, the effects on A2BAR-mediated second messenger cAMP levels, ß-arrestin 2 recruitment, and cAMP response element (CRE) activation were examined using luciferase-based HEK293 reporter cell lines. Lastly, the impact on A2BAR-mediated IL-6 release in Calu-3 epithelial lung cells was investigated via the Lumit™ Immunoassay. Additionally, the adenosine receptor subtype mediating these effects was specified, and A2BAR was found to be responsible. The present study demonstrates an inhibitory influence of EA 575® on A2BAR-mediated general cellular response, cAMP levels, ß-arrestin 2 recruitment, CRE activation, and IL-6 release. Since these EA 575®-mediated effects occur within a time frame of several hours of incubation, its mode of action can be described as indirect. The present data are the first to describe an inhibitory effect of EA 575® on A2BAR signalling. This may offer an explanation for the beneficial clinical effects of the extract in adjuvant asthma therapy.

Ivy leaves dry extract EA 575® mediates biased ß2-adrenergic receptor signaling.

BACKGROUND: ß2-adrenergic receptor (ß2-AR) stimulation activates the G protein/cAMP pathway, which is opposed by the GRK2/ß-arrestin 2 pathway. The latter is undesirable in the treatment of respiratory diseases. HYPOTHESIS/PURPOSE: EA 575® is capable of mediating a biased ß2-adrenergic signaling pathway. METHODS: The impact of the ivy leaves dry extract EA 575® on ß2-adrenergic signaling was tested in a dynamic mass redistribution assay in HEK wild-type and in HEK ß-arrestin knock-out cells. cAMP formation and recruitment of ß-arrestin 2 were investigated using GloSensor™ and PathHunter® assays, respectively. NFκB transcriptional activity was determined in both HEK wild-type as well as HEK ß-arrestin knock-out cells. RESULTS: EA 575® inhibits the recruitment of ß-arrestin 2 and thereby enhances G protein/cAMP signaling under ß2-stimulating conditions, as evidenced by a corresponding increase in cAMP formation. While ß2-AR-mediated inhibition of NFκB transcriptional activity is ß-arrestin-dependent, EA 575® leads to significant inhibition of NFκB transcriptional activity in ß-arrestin knock-out cells and thus via a ß-arrestin-independent signaling pathway. CONCLUSION: EA 575® is the first active phytopharmaceutical ingredient for which biased ß2-adrenergic activation has been described. This shift towards G protein/cAMP signaling provides the molecular basis for the clinically proven efficacy of EA 575® in the treatment of lower respiratory tract diseases. In this light, EA 575® could potentially reduce ß-arrestin-mediated adverse effects in new combinatorial therapeutic approaches.

Effect of Melatonin on the Reduction of Hepatic Steatosis and Intestinal Leptin Expression in Zebrafish Exposed to Fructose.

Melatonin is a hormone related to circadian rhythms and has potential clinical applications. Our objectives were to verify the effect of melatonin on the liver of zebrafish exposed to fructose and evaluate the expression of appetite-related genes (leptin, ghrelin, and melanocortin receptor 4 [MC4R]). Animals were divided into three groups: control (CG, n = 25), fructose (FG, n = 25), and fructose+melatonin (FMG, n = 25). The study was carried out in 8 weeks. FG and FMG were exposed to 2% fructose and FMG treated with 1 µM of melatonin. Histological liver studies and gene expression analyses of Leptin, Ghrelin, and MC4R (liver and intestines) were performed. FG developed hepatic steatosis, which did not occur with CG and FMG. Genetic expression of hepatic leptin and MC4R did not show significant difference among the groups. Animals exposed to fructose (FG) presented an increased expression of intestinal leptin compared to those administered with melatonin. Animals exposed to fructose gained weight and developed an important hepatic steatosis, but melatonin reduced significantly the hepatic damage. Intestinal leptin showed increased expression in the group exposed to fructose.

Effect of dietary crude glycerin on the productive performance of Nile tilapia fingerlings.

The aim of this study was to evaluate the effect of different crude glycerin levels in the diet of Nile tilapia fingerlings (mean initial weight 0.32 ± 0.06 g, n = 450) on growth performance parameters, whole-body composition, blood glucose and liver morphology. Crude glycerin was tested at six different levels (0, 4, 8, 12, 16, and 20%) in diets containing 30% digestible protein and 3,000 kcal kg-1 digestible energy. After 37 days of feeding, the inclusion of crude glycerin resulted in positive effects on final weight, visceral fat, weight gain, feed conversion, specific growth rate and feed intake. The different treatments did not influence fillet yield, glycemia, survival and hepatosomatic index, but intermediate levels of inclusion decreased the area of hepatocytes. Regarding fish body composition, significant differences were found in moisture and ash contents, with no changes in crude protein and total lipid. The inclusion of crude glycerin in the Nile tilapia diet improves growth performance without negatively affecting survival rate and glycemia of fingerlings.

A comparative study on the influence of an ivy preparation and an ivy/thyme combination on the ß2-adrenergic signal transduction.

The ß2-adrenergic receptor (ß2AR) is relevant for surfactant formation in alveolar type 2 cells and reduction of intracellular calcium concentration in bronchial muscle cells and thus for secretolytic and bronchospasmolytic effects. Herbal medicinal products that affect the ß2AR system are used to treat common cold and bronchitis accompanied with mucus covered and narrowed airways. The present work compares the influence of an ivy preparation and an ivy/thyme combination on the ß2-adrenergic signal transduction. For receptor binding studies and characterization of the lateral mobility of ß2AR we have used single molecule detection by fluorescence correlation spectroscopy and single particle tracking. For the determination of both the second messenger cAMP and the internalization of ß2AR we have generated luciferase based reporter cell lines, which produce a cAMP-dependent luciferase in the cytosol and express ß2AR with extracellular luciferase moiety in the plasma membrane. While both preparations increased the ß2AR binding, a significant increase of the cAMP level was observed only for the ivy preparation, which can be explained by the inhibited internalization of HiBiT-tagged ß2AR under stimulating conditions. In contrast, isoprenaline-mediated internalization of HiBiT-tagged ß2AR of ivy/thyme combination pre-treated cells was not inhibited. Cells comparatively pre-treated with a thyme preparation did not show inhibition of ß2AR internalization either. Furthermore, SNAP-tagged ß2AR of ivy preparation pre-treated cells, which were not internalized after isoprenaline stimulation, showed a redistribution from fast-to-slowly diffusing ß2AR. A corresponding redistribution of these receptors was not observed after pre-treatment with both the ivy/thyme combination and the thyme preparation. Comparable to the ivy/thyme combination, no decrease in the intratrack transitioning probability ratio (p23/p32) for fast and slow diffusing ß2AR was found for the thyme preparation, which, however, significantly decreased for control cells and for pre-treatment with the ivy preparation under stimulating conditions. It can therefore be concluded that the thyme fluid extract fraction in the ivy/thyme combination may have in part a negative effect on the ß2-adrenergic signal transduction.

Hepatoprotective Effect of Probiotic Lactobacillus rhamnosus GG Through the Modulation of Gut Permeability and Inflammasomes in a Model of Alcoholic Liver Disease in Zebrafish.

Objective: Alcoholic liver disease (ALD) is among the leading causes of death from liver disease. Among the factors involved in its pathogenesis are inflammation and increased intestinal permeability. The aim of this study was to assess the effect of Lactobacillus rhamnosus GG (LGG) on hepatic lipid accumulation, activation of inflammasomes, and gut permeability markers in experimental model of ALD with zebrafish.Methods: An experiment was conducted to assess the effective LGG dose capable of promoting intestinal colonization. Animals were divided into three groups (n = 64/group): ethanol group (E), ethanol + probiotic group (EP), and control group (C). Groups E and EP were exposed to 0.5% ethanol concentration for 28 days. At the end of this period, animals were euthanized, and livers were collected for Oil Red staining and assessment of the inflammasome system. Intestines were collected for evaluation of gut permeability markers.Results: The dose of 1.55 × 106 UFC LGG/fish/d promoted intestinal colonization. Group EP presented lower hepatic lipid accumulation, lower il-1ß expression, and higher cldn15a expression when compared to group E.Conclusions: Supplementation with LGG was protective for hepatic steatosis in ALD model. In addition, LGG influenced the modulation of the inflammatory response and markers of gut permeability, improving the gut barrier structure.

Alcoholic extract of propolis in Nile tilapia post-larvae and fingerlings' diets: effects on production performance, body composition and intestinal histology.

The aim of this study was to evaluate the production performance of Nile tilapia post-larvae and fingerlings fed with increasing levels of alcoholic extract of propolis into diets. In Experiment 1, 1800 post-larvae were distributed in 30 tanks, in a completely randomized design with five treatments composed by the inclusion of 0, 1, 2, 3 and 4 g of dry propolis/kg of feed, and six replicates. In experiment 2, 1600 fingerlings were distributed in the same system and designed as experiment 1. No significant effect was observed between treatments, for final weight, total and standard length, survival, and intestinal villus height. However, the propolis extract inclusion of 1 g/kg in Nile tilapia post-larvae and fingerlings' feed resulted in a better body condition factor and higher body protein deposition (p<0.05). The condition factor is an estimate for the future growth of the animals, possibly the fish treated with propolis extract will present better growth, survival and greater reproductive potential rates. The results of the present study demonstrate that alcoholic extract of propolis improves the nutritional condition of Nile tilapia post-larvae and fingerlings, potentially leading to increased productivity in subsequent stages, as well as leading to improvement in fingerlings muscle deposition.

Lactobacillus rhamnosus GG Effect on Behavior of Zebrafish During Chronic Ethanol Exposure.

Ethanol is a widely consumed drug, which acts on the central nervous system to induce behavioral alterations ranging from disinhibition to sedation. Recent studies have produced accumulating evidence for the therapeutic role of probiotic bacteria in behavior. We aimed to investigate the effect of Lactobacillus rhamnosus GG (LGG) on the behavior of adult zebrafish chronically exposed to ethanol. Adult wild-type zebrafish were randomly divided into four groups, each containing 15 fish. The following groups were formed: Control (C), received unsupplemented feed during the trial period; Probiotic (P), fed with feed supplemented with LGG; Ethanol (E), received unsupplemented feed and 0.5% of ethanol directly added to the tank water; and Probiotic+Ethanol (P+E), group under ethanol exposure (0.5%) and fed with LGG supplemented feed. After 2 weeks of exposure, the novel tank test was used to evaluate fish behavior, which was analyzed using computer-aided video tracking. LGG alone did not alter swimming behavior of the fish. Ethanol exposure led to robust behavioral effects in the form of reduced anxiety levels, as indicated by increased vertical exploration and more time spent in the upper region of the novel tank. The group exposed to ethanol and treated with LGG behaved similarly to animals exposed to ethanol alone. Taken together, these results show that zebrafish behavior was not altered by LGG per se, as seen in murine models. This was the first study to investigate the effects of a probiotic diet on behavior after a chronic ethanol exposure.

Effects of Lactobacillus rhamnosus GG on hepatic and serum lipid profiles in zebrafish exposed to ethanol.

Zebrafish is a powerful tool in pharmacological research and useful to identify new therapies. Probiotics can offer therapeutic options in alcoholic liver disease. This study was done in two independent experiments: first, we confirmed the intestinal colonization of probiotic Lactobacillus rhamnosus GG (LGG) after ethanol exposure. Second, four groups were performed: control (C), probiotic (P), ethanol (E), and probiotic+ethanol (P+E). Liver histology, hepatocytes morphometry, hepatic and serum lipid quantifications were conducted in second experiment. During 4 weeks, P and P+E groups were fed with LGG supplemented feed; E and C unsupplemented. E and P+E groups received 0.5% of ethanol added into tank water. Zebrafish exposed to ethanol (E group) presented intense liver steatosis after 28 days in contrast to the almost normalized liver histology of P+E group at the same period. Liver morphometry showed a significant enlargement of hepatocytes of E group after 4 weeks (p<0.0001). Serum triglycerides decreased in P+E group compared with C, P (p<0.001), and E (p=0.004), after 14 and 28 days similarly. Serum cholesterol was also decreased by LGG; P group decreased compared with C and E after 14 days (p=0.002 and p=0.007, respectively) and P+E group decreased significantly compared with E and C groups (p<0.0001) after 28 days. Hepatic triglycerides were reduced in P+E group after 28 days compared to E (p=0.006). The persistence of LGG in zebrafish intestines was demonstrated. LGG decreased serum levels of triglycerides and cholesterol and improved hepatic steatosis.

Composição química e digestibilidade aparente da energia e nutrientes da farinha de resíduos da indústria de filetagem de tilápias, para a tilápia do Nilo (Oreochromis niloticus) / Apparent energy digestibility and meal nutrients from tilapia filleting industrial waste for the Nile tilapia (Oreochromis niloticus L. )

O trabalho foi realizado com objetivo de determinar os coeficientes de digestibilidade aparente (CD) da matéria seca (MS), proteína bruta (PB), energia bruta (EB) e a disponibilidade dos aminoácidos (AA), fósforo (P) e cálcio (Ca) da farinha de resíduos da indústria de filetagem de tilápias (FT) para a tilápia do Nilo. Foram utilizadas 60 tilápias com peso e comprimento médio de 80,69±15,97g e 15,97±1,87cm, respectivamente. Os animais foram submetidos à metodologia de coleta de fezes por sedimentação em tanques afunilados. A MS, PB e EB da FT apresentaram CD de 83,55, 88,13 e 84,74%, respectivamente. Apresentando valores de proteína e energia digestíveis de 44,39% e 3799,02Kcal de ED kg-1. A FT apresenta CDa médio dos AA de 91,60% com CDas variando de 67,90% para a Fen até 100% para a Arg e a Cis. O CD do P foi de 70,44 por cento apresentando 1,94% de P disponível, o CDa do Ca da FT é de 51,10%apresentando 4,02% de Ca disponível. A FT é um alimento protéico com bons índices de disponibilidade aparente de seus nutrientes e pode ser utilizado na alimentação de tilápias com a aplicação destes índices visando formular rações de custo mínimo que atendam às exigências da espécie.

Coefficients of apparent digestibility (CD) of dry matter (DM), crude protein (CP), crude energy (CE) and the availability of amino-acids (AA), phosphorus (P) and calcium (Ca) of meals from tilapia filleting industrial waste (FT) for the Nile tilapia were determined. Sixty tilapias, mean weight and length 80.69±15.97g and 15.97±1.87cm respectively, underwent feces collection by sedimentation in cone-bottom tanks. DM, CP and BE of FT had a CD of 83.55; 88.13 and 84.74% respectively, with digestible protein and energy rates equivalent to 44.39% and 3799.02 Kcal of ED kg-1. FT had average CD of AA of 91.60%, with CD varying from 67.90% for Fen up to 100% for Arg and Cis. Whereas CD of P was 70.44% with 1.94% of available P, CD of Ca of FT amounts to 51.10 percent with 4.02% of Ca available. FT is a protein feed with a good index of apparent availability of its nutrients and may be used in feed of tilapia. Above indexes will produce low cost feeds that also meet the requirements of the fish.

Sensibilidade antimicrobiana de bactérias isoladas de Jundiá (Rhamdia quelen) / Antimicrobial sensibility of bacterial isolates from Jundiá (Rhamdia quelen)

Objetivando o delineamento do perfil de sensibilidade dos agentes bacterianos causadores de enfermidades em peixes, 51 isolados bacterianos provenientes de Jundiá e pertencentes aos gêneros Acinetobacter spp. (8), Aeromonas spp. (15), Edwardsiella spp. (2), Enterobacter spp. (2), Klebsiella spp. (1), Plesiomonas spp. (5), Pseudomonas spp. (1), Staphylococcus spp.(11) e Vibrio spp. (6) foram testados frente aos antimicrobianos utilizados no tratamento de enfermidades em peixes. Dos 51 isolados bacterianos obtidos de exemplares de Jundiá (Rhamdia quelen) 51 (100 por cento) foram sensíveis a gentamicina, 49 (96,08 por cento) ao sulfazotrim, 47 (92,16 por cento) ao cloranfenicol, 43 (84,31 por cento), a tetraciclina, 43 (84,31 por cento) ao ácido nalidíxico, 31 (60,78 por cento) à nitrofurantoina, 22 (43,14 por cento) à eritromicina, 22 (43,14 por cento) à ampicilina, 15 (29,41 por cento) à espiramicina, 13 (25,50 por cento) à colistina e 5 (3 por cento) foram sensíveis a penicilina G. Com exceção de um isolado do gênero Staphylococcus spp., as bactérias analisadas no presente estudo foram resistentes a um ou mais agentes antimicrobianos testados. O conhecimento do perfil de sensibilidade das bactérias envolvidas em processos infecciosos nos peixes permitirá aos técnicos à adoção de uma antimicrobianoterapia racional, que contribuirá para o controle das enfermidades em Rhamdia quelen, sem causar grandes riscos à saúde pública e ao meio ambiente.(AU)

Aiming the evaluation of sensitivity profiles of pathogen bacteria responsible for fish diseases, 51 bacterial isolates from Jundiá (Rhamdia quelen) belonging to the genus Acinetobacter spp. (8), Aeromonas spp. (15), Edwardsiella spp. (2), Enterobacter spp. (2), Klebsiella spp. (1), Plesiomonas spp. (5), Pseudomonas spp. (1), Staphylococcus spp. (11), and Vibrio spp. (6), were tested against antimicrobial agents used for treatment of bacterial fish diseases. All samples were processed at the Laboratory of Bacteriology, Department of Preventive Veterinary Medicine, UFSM. From 51 bacteria isolated from jundiá fishes (Rhamdia quelen) 51 (100 percent) were sensitive to gentamycin, 49 (96,08 percent) to sulphazotrin, 47(92,16 percent) to chloramphenicol, 43 (84,31 percent) to tetracylin, 43 (84,31 percent) to naldixic acid, 31 (60,78 percent) to nitrofurantoin, 22 (43,14 percent) to erytromycin, 22 (43,14 percent) to ampicillin, 15 (29,41 percent) spiramycin, 13 (25,50 percent) to cholystin, and 5 (3 percent) to penicillin G. With exception of an isolate of Staphylococcus spp., the bacteria analyzed in the present study were resistant to one or more antimicrobial agents tested. Knowledge of the sensitivity profile of bacteria involved in infectious processes in fish will allow rational antimicrobial treatment that will contribute to the control of fish diseases in Rhamdia quelen without causing great risks to public health and the environment.(AU)

Polissacarídeos não amiláceos na nutrição de peixes - revisão / Non-starchpolysaccharides in fish nutrition - A Review

Os polissacarídeos não amiláceos são componentes das paredes celulares presentes em alimentos de origem vegetal comumente encontrados em rações para peixes. Estes elementos são geralmente a celulose, hemicelulose e pectinas, os quais podem diminuir o desempenho animal, dependendo de sua concentração. Não podem ser degradados por enzimas endógenas, porém o são, numa extensão variável, por microorganismos presentes em seu trato digestivo. Estes elementos afetam a digestibilidade dos nutrientes e modificação no tempo de permanência do alimento no trato digestivo. O maior problema apresentado por esta categoria de componentes é a sua ampla variabilidade e conseqüentes efeitos sobre o desempenho animal. A utilização de enzimas exógenas e técnicas de processamento dos alimentos têm sido adotadas para reduzir os efeitos negativos dos polissacarídeos não amilásceos.