Ionophore Antibiotic Research Articles

Salinomycin and Sulforaphane Exerted Synergistic Antiproliferative and Proapoptotic Effects on Colorectal Cancer Cells by Inhibiting the PI3K/Akt Signaling Pathway in vitro and in vivo.

BackgroundBoth salinomycin (SAL) and sulforaphane (SFN) exert their antitumorigenic effects in various types of cancer We investigated whether combining salinomycin (SAL, an antibiotic ionophore) with sulforaphane (SFN, a phytochemical) exerted synergistic antiproliferative and proapoptotic activities in colorectal cancer (CRC) cells in vitro and in vivo by evaluating the proliferative and apoptotic responses of two CRC cell lines.Materials and MethodsThe combination index (CI) was calculated using the Chou-Talalay method, and the effects of the synergistic combination (CI<1) of lower doses of SAL and SFN were selected for further studies. Anti-tumor effect of the combination of SAL and SFN was tested both in vitro and in vivo.ResultsCotreatment effectively inhibited proliferation, migration and invasion and enhanced apoptosis. The xenograft model also showed similar results. Furthermore, we evaluated the molecular mechanism behind SAL- and SFN-mediated CRC cell apoptosis. The combination treatment induced apoptosis in Caco-2 and CX-1 cells by inhibiting the PI3K/Akt pathway, which increased the expression of the tumor suppressor protein p53. The treatment also decreased the expression of the survival protein Bcl-2 and increased the expression of the proapoptotic protein Bax, which increased the Bax/Bcl-2 ratio, as well as enhanced poly ADP-ribose polymerase (PARP) cleavage. Upon inhibiting the PI3K/Akt pathway with LY294002 prior to cotreatment, we detected enhanced PARP cleavage compared to that in the cotreatment only group.ConclusionWe investigated whether the combination of SAL and SFN had antiproliferative and proapoptotic effects in CRC cells both in vitro and in vivo. Cotreatment also significantly decreased migration and invasion compared to that of the control and SAL or SFN monotherapies. This novel combination of SAL and SFN might provide a potential strategy to treat CRC.

Sorption and abiotic transformation of monensin by iron and manganese oxides

Monensin, an ionophore antibiotic, is commonly administered as a feed additive to cattle and poultry. A large percentage of the administered dose is excreted in animal waste, which is often applied to agricultural fields as fertilizer. The objective of this work is to gain insight into the fate of monensin in soil by investigating the interactions between monensin and common soil minerals, including sorption and transformation to unmonitored partial oxidation products. Batch sorption experiments across varying conditions (i.e., pH, ionic strength) and desorption experiments (i.e., methanol, PO43−, methyl tert-butyl ether) were used to determine the extent to which a selection of common redox-active soil minerals [birnessite (δ-MnO2), goethite (α-FeOOH), hematite (α-Fe2O3)] can bind and transform monensin. Monensin was bound by hematite (pH < 7.5, up to 7.5 mmol kg−1), goethite (pH < 7.5, up to 3.4 mmol kg−1), and birnessite (pH < 7, up to 0.1 mmol kg−1). Combined sorption and transformation were the greatest for hematite and the lowest for birnessite. Sorption to hematite was more reversible than to goethite. Each desorption from goethite recovered <10% of sorbed monensin, whereas desorption from hematite recovered up to 69% of sorbed monensin, dependent on the solution. The potential for iron and manganese (hydr)oxides to abiotically transform monensin through reductive dissolution to partial oxidation products was evaluated by mass spectral analysis following sorption experiments. Additionally, the dominant sorption mechanism was inferred through ATR-FTIR spectroscopy, via examination of the carboxylate peak separation differences, on goethite and hematite to be bridging bidentate.

Novel Salinomycin Analogs Show Improved Selectivity Towards Breast Cancer Stem Cells

Breast cancer tumor metastasis is the leading cause of mortality among women. Cancer stem cells (CSCs) are the tumor‐initiating cells which drive initiation, progression, metastasis, and tumor reoccurrence. Their resistance to conventional chemo‐ and radiotherapies and their ability to survive such treatment enables tumor reestablishment. Thus, therapeutic strategies targeting CSCs hold great potential for novel advances in cancer treatment. Salinomycin (SAL) is a naturally occurring polyether ionophore antibiotic with anti‐cancer activity towards various types of tumor cells, but its toxicity towards normal cells limits broad application. A library of 21 novel SAL analogs was screened to identify compounds with improved selectivity especially against breast cancer stem cells. SAL analogs were either single modified with ester or amide in the C1 position or double‐modified C20‐oxo derivatives. Eight single‐ and two double‐modified derivatives were more potent (IC50 range of 1.13 ± 0.19 to 3.93 ± 0.39 μM) towards the breast cancer cell line MDA‐MB‐231 compared to SAL (IC50 of 5.81 ± 1.50). These derivatives induced DNA fragmentation suggestive of apoptotic cell death. Further, most of these derivatives had improved selectivity toward MDA‐MB‐231 cells compared to SAL when also tested against normal breast epithelial MCF10A cells. In addition, a butyl ester derivative of SAL, chosen based on increased potency and selectivity versus SAL, significantly reduced the colony‐forming efficiency of MDA‐MB‐231 cells in association with selective loss of the CD44+/CD24−/low stem‐cell‐like subpopulation. Select derivatives were next screened against the NCI‐60 Human Tumor Cell Line Panel. A double‐modified analog of C20‐oxo‐SAL with hydroxamic acid was found to be more potent than SAL towards all 6 breast cancer cell lines in the panel as well as other tumor types especially leukemia. Studies are underway to test the analogs in three dimensional culture using breast cancer organoids derived from both cell lines and patients. The present findings highlight the therapeutic potential of SAL analogs towards breast cancer and breast cancer stem cells and support further research and clinical development of these compounds.Support or Funding InformationThe present study was funded by grants (to AMM and TCC) from the Arkansas Breast Cancer Research Program. Testing was performed by the Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute, http://dtp.cancer.gov.

A SOX2 Reporter System Identifies Gastric Cancer Stem-Like Cells Sensitive to Monensin.

Gastric cancer remains a serious health burden with few therapeutic options. Therefore, the recognition of cancer stem cells (CSCs) as seeds of the tumorigenic process makes them a prime therapeutic target. Knowing that the transcription factors SOX2 and OCT4 promote stemness, our approach was to isolate stem-like cells in human gastric cancer cell lines using a traceable reporter system based on SOX2/OCT4 activity (SORE6-GFP). Cells transduced with the SORE6-GFP reporter system were sorted into SORE6+ and SORE6– cell populations, and their biological behavior characterized. SORE6+ cells were enriched for SOX2 and exhibited CSC features, including a greater ability to proliferate and form gastrospheres in non-adherent conditions, a larger in vivo tumor initiating capability, and increased resistance to 5-fluorouracil (5-FU) treatment. The overexpression and knockdown of SOX2 revealed a crucial role of SOX2 in cell proliferation and drug resistance. By combining the reporter system with a high-throughput screening of pharmacologically active small molecules we identified monensin, an ionophore antibiotic, displaying selective toxicity to SORE6+ cells. The ability of SORE6-GFP reporter system to recognize cancer stem-like cells facilitates our understanding of gastric CSC biology and serves as a platform for the identification of powerful therapeutics for targeting gastric CSCs.

Regulatory Patterns of Crp on Monensin Biosynthesis in Streptomyces cinnamonensis

Monensin, produced by Streptomyces cinnamonensis, is a polyether ionophore antibiotic widely used as a coccidiostat and a growth-promoting agent in agricultural industry. In this study, cyclic AMP receptor protein (Crp), the global transcription factor for regulation of monensin biosynthesis, was deciphered. The overexpression and antisense RNA silencing of crp revealed that Crp plays a positive role in monensin biosynthesis. RNA sequencing analysis indicated that Crp exhibited extensive regulatory effects on genes involved in both primary metabolic pathways and the monensin biosynthetic gene cluster (mon). The primary metabolic genes, including acs, pckA, accB, acdH, atoB, mutB, epi and ccr, which are pivotal in the biosynthesis of monensin precursors malonyl-CoA, methylmalonyl-CoA and ethylmalonyl-CoA, are transcriptionally upregulated by Crp. Furthermore, Crp upregulates the expression of most mon genes, including all PKS genes (monAI to monAVIII), tailoring genes (monBI-monBII-monCI, monD and monAX) and a pathway-specific regulatory gene (monRI). Enhanced precursor supply and the upregulated expression of mon cluser by Crp would allow the higher production of monensin in S. cinnamonensis. This study gives a more comprehensive understanding of the global regulator Crp and extends the knowledge of Crp regulatory mechanism in Streptomyces.

Improved performance of Eimeria-infected chickens fed corn expressing a single-domain antibody against interleukin-10.

Antimicrobial resistance is a significant challenge for human and animal health, and developing effective antibiotic-free treatments is a strategy to help mitigate microbial resistance. The global poultry industry faces growing challenges from Eimeria-induced coccidiosis, a serious enteric disease of chickens that currently requires treatment using ionophore antibiotics. Eimeria stimulates interleukin-10 (IL-10) expression in the small intestine and caecum of infected chickens, suppressing their immune response and facilitating disease progression. Single-domain antibodies raised from llamas immunized with chicken IL-10 (cIL-10) were developed that bind cIL-10 in vitro, block cIL-10 receptor binding and induce interferon gamma (IFN-γ) secretion from cIL-10-repressed primary chicken splenocytes. Single-domain antibodies expressed in transgenic corn demonstrated significant accumulation in phenotypically normal plants. When fed to Eimeria-challenged chickens, the transgenic corn significantly improved body weight gain (equal to that of salinomycin-treated animals), normalized the feed conversion ratio (to the same level as uninfected control animals), lowered E. tenella lesion scores to those of salinomycin-treated control animals, and reduced oocyst counts below those of infected untreated control animals. Here, we propose that transgenic corn may have a role in reducing the use of antibiotics in poultry production and maintaining animal health and productivity, and may contribute to efforts against global antimicrobial resistance.

Determination of the efficacy of monensin capsule (continuous release capsule) on metabolic parameters in transition dairy cows

Monensin is an ionophore antibiotic that changes the population of rumen bacteria. The aim of this study was to determine the effects of monensin controlled capsule administered 3 wk before calving on blood metabolites (serum beta-hydroxybutiric acid (BHBA), cholesterol (CHOL), urea, creatinine (CREA), gamma glutamyl transferase (GGT), aspartate aminotransferase (AST), total protein (TP), phosphorus (P), magnesium (Mg+), calcium (Ca++), glucose (GLU) and plasma non-esterified fatty acids (NEFA) concentration) and milk yield in the pre and post-calving term. 50 Holstein-Friesian cows were selected from the same flock. Blood samples were taken 3 weeks before the expected calving date and during weeks 1, 2, 4, 6 and 8 post calving. Cows were divided into two group: a study group (Group M, n:40) and a control group (Group C, n: 10). The group M orally received a cylindrical device (Kexxtone, ELANCO Animal Health, Guelph, ON, Canada) and the group C received no treatment. BHBA and NEFA was lower (P˂0,.05) in group M compared with group C in the 6 and 8 week post-calving. GLU was higher (P˂0.05) in group M compared with group C in the 4 week post-calving. Urea was higher (P˂0.05) in group M compared with group C in the 1 week post-calving. There was no statistically difference between two groups (P˂0.05) in milk yields. Although milk yield was similar in both groups, the change in concentration of BHBA, NEFA, GLU and urea showed that monensin CRC could be as a glycogenic precursor.

Outbreak of monensin poisoning in cattle due to supplementation error

ABSTRACT: Iatrogenic intoxications occur in domestic animals. Mortality was observed in a group of calves, reaching 20 deaths until the day of the technical visit, with or without sudden clinical signs, on a farm in the city of Pires do Rio, State of Goiás, Brazil. Deaths after the visit were not accounted systematically. Clinical signs included muscle weakness, ataxia, recumbency, bilateral jugular distention, and death. Suspected diagnosis of poisoning by ionophore antibiotics was made based on clinical and laboratory findings from nine animals, which included high plasma CK and LDH levels, as well as on anatomopathological findings of a recently dead calf, which showed myocardial and skeletal muscle degeneration and necrosis. Monensin overdose (25.5 µg/kg in skeletal muscle and 209.4 µg/kg in the liver) was detected by liquid-chromatography and mass-spectrophotometry analysis, contributing to the confirmation of ionophore poisoning diagnosis.

PSX-28 Narasin inclusion and different levels of hay change the cecal parameters in feedlot lambs

Abstract Narasin is an antibiotic ionophore that may change rumen fermentation; however, the effects on the post-rumen are unknown. The objective of this study was to determinate the effects of different levels of hay and narasin inclusion on the cecal parameters of lambs. Forty-four lambs were allotted in a randomized complete block design, defined by initial BW, in a 2x2 factorial treatment arrangement. The first factor was forage (coastcross hay) inclusion (10 or 20%, DM basis) and the second factor was narasin (0 or 13 ppm). The experimental diets were isonitrogenous (17.4%, DM basis) and the experiment lasted 112 days. At the end of the experimental period, the lambs were slaughtered without fasting, and the cecal fluid was collected to determinate pH and short chain fatty acid (SCFA). Data were analyzed using the MIXED procedure of SAS. The effects were considered significant when P &amp;lt; 0.05. There was no interaction between forage content and narasin inclusion. Narasin increased the molar concentration of propionate (N0: 19.4; N13: 21.4 mM/100mM; P = 0.02) and decreased isobutyrate (N0: 1.6; N13: 0.7 mM/100mM; P = 0.05) and acetate:propionate ratio (N0: 3.4; N13: 2.9; P = 0.03). The valerate was greater in diets with 10% hay when compared of diets 20% (H10: 2.0; H20: 1.3 mM/100mM; P &amp;lt; 0.01). The experimental diets not affect the molar proportion of acetate (62.7±1.3 mM/100mM), butyrate (12.9 ± 0.7 mM/100mM), isovalerate (1.3 ± 0.5 mM/100mM) total SCFA concentration (68.6 ± 8.5 mM) and cecal pH (6.2 ± 0.14). Therefore, the inclusion of 13 ppm of narasin changes the cecal fermentation in high concentrate diets. In addition, the inclusion of 10 or 20% of hay has a little influence on cecal fermentation.

Comparative study on the effects of salinomycin, monensin and meso-2,3-dimercaptosuccinic acid on the concentrations of lead, calcium, copper, iron and zinc in lungs and heart in lead-exposed mice

Background and aimEnvironmental lead (Pb) exposure damages the lungs and is a risk factor for death from cardiovascular disease. Pb induces toxicity by a mechanism, which involves alteration of the essential elements homeostasis. In this study we compare the effects of salinomycin (Sal), monensin (Mon) and meso-2,3-dimercaptosuccinic acid (DMSA) on the concentrations of lead (Pb), calcium (Ca), copper (Cu), iron (Fe) and zinc (Zn) in the lungs and heart of lead-exposed mice. MethodsSixty days old male ICR mice were divided into five groups: control (Ctrl) – untreated mice obtained distilled water for 28 days; Pb-intoxicated group (Pb) – exposed to 80 mg/kg body weight (BW) Pb(NO3)2 during the first 14 days of the experimental protocol; DMSA-treated (Pb + DMSA) – Pb-exposed mice, subjected to treatment with an average daily dose of 20 mg/kg BW DMSA for two weeks; Monensin-treated (Pb + Mon) – Pb-exposed mice, obtained an average daily dose of 20 mg/kg BW tetraethylammonium salt of monensic acid for 14 days; Pb + Sal - Pb-exposed mice, treated with an average daily dose of 20 mg/kg BW tetraethylammonium salt of salinomycinic acid for two weeks. On the 29th day of the experiment the samples (lungs and heart) were taken for atomic absorption analysis. ResultsThe results revealed that exposure of mice to Pb for 14 days significantly increased the concentration of the toxic metal in both organs and elevated the cardiac concentrations of Ca, Cu and Fe compared to untreated mice. Pb exposure diminished the lung concentrations of Ca and Zn compared to that of untreated controls. DMSA, monensin and salinomycin decreased the concentration of Pb in the lungs and heart. Among the tested chelating agents, only salinomycin restored the cardiac Fe concentration to normal control values. ConclusionThe results demonstrated the potential application of polyether ionophorous antibiotic salinomycin as antidote for treatment of Pb-induced toxicity in the lungs and heart. The possible complexation of the polyether ionophorous antibiotics with Ca(II) and Zn(II), which can diminish the endogenous concentrations of both ions in the lungs should be taken into account.

Correction the blood biochemical parameters in transition dairy cows by hop cones and vitamin E supplement

In the prevention and treatment of cow ketosis, the regulation of glucose and fatty acids is the main focus, while ammonia intoxication is largely ignored. The intensity of formation of the rumen ammonia can be reduced by ionophore antibiotic monensin. Hop cones contain a number of biologically active components including phytoionoрhores: prenylated polyphenols lupulon, humulon and their derivatives. Therefore, hop cones can be considered as a potential substitute for antibiotics-ionophores. Vitamin E, fed to ruminants in large quantities, stimulates the cellulosolytic bacteria of the rumen. Accordingly, co-feeding cows with hops and vitamin E supplements can reduce ammonia formation without inhibition the carbohydrate fermentation in the rumen. The experiment used two groups of dry Ukrainian dairy black-and-white breed cows with productivity 6–7 thousand kg of milk for previous lactation, 10 animals per each group. The trial was performed during the last 3 weeks of dry period and the first 3 weeks after calving. Animals were fed a standard balanced diet containing: grass silage, corn silage, barley grain, corn grain, soybean meal, molasses, salt, mineral and vitamin premix. The first group was the control. The diet of second group was supplemented with 300 mg of α-tocopherol acetate (0.6 g of Rovimix E-50) and 1g of dry hop cones per kg of DM. Prior to calving, the tested feed additive reduced the concentration of lipid peroxidation products (P &lt; 0.05) in the blood plasma without affection other parameters. More significant changes were detected after calving. An increase in glucose concentration (P &lt; 0.05), triacylglycerol (P &lt; 0.05), cholesterol esters (P &lt; 0.05) and a decrease in the concentration of NEFA (P &lt; 0.05) were found in the blood of cows of the experimental group in a week after parturition. One month after calving, differences between control and experimental blood plasma parameters were significantly less pronounced. Therefore, addition of 300 mg of α-tocopherol acetate and 1 g/kg of dry hop cones per kg of dry matter of the diet during the transition period stimulates liver glucose synthesis and reduces the rate of fatty acid release from adipose tissue. The specified feed additive can be used to prevent metabolic disorders in early-lactation dairy cows.

Behavior of ionophore antibiotics in aquatic environments in Argentina: The distribution on different scales in water courses and the role of wetlands in depuration

We studied for the first time three ionophore anticoccidial drugs: monensin (MON), lasalocid (LAS), and salinomycin (SAL) as emerging pollutants originating from animal and plant husbandry in surface waters (n = 89) in one of the most extensive hydrological basins in South América (Del Plata basin). The soluble fraction of ionophores was pretreated by solid-phase extraction and analyzed by LC-MS/MS at a limit of detection of 1.7 ng·L−1. A statistical approach noted the need to report parameters calculated by methods based on the number of observations and the censorship percentage over substitution methods for more precise estimations of environmental data with a high percentage of left-censored data. Water collectors adjacent to intensive-husbandry facilities, placed in direct runoffs from animal excreta, or in wastewater emissions contained median concentrations of MON and SAL approximately 70 times higher than those found in regional tributaries and main courses of 5 sub-basins of the pampas and mesopotamic regions, thus exhibiting a relevance to other similar agricultural pollutants widely reported as pesticides. Chemical speciation of these compounds in surface water was characterized especially for MON and SAL, where the pH and chemical oxygen demand of the natural water body was associated with the concentration of the soluble fraction. The concentrations in abundant rivers such as the Gualeguay deliver a contribution to a natural wetland such as the Paraná-River delta, which registered only one sample with a [MON] ≤ the limit of quantification. Since wetlands possess a limited removal capability, these affluent contributions recorded strongly indicate that attention must be paid to the development of guidelines involving quality criteria for assessing the impact of ionophore antibiotics on such ecosystems.

EVALUATION OF THE EFFECT OF FOUR ESSENTIAL OILS AS POTENTIAL ALTERNATIVES FOR MONENSIN ON RUMEN FERMENTATION CHARACTERISTICS AND NUTRIENT DEGRADABILITY

The objective of this study was to evaluate the effect of four essential oils (EOs) from guava (Psidium guajava), citronella (Cymbopogon nardus), lemongrass (Cymbopogon citratus), and geranium (Pelargonium graveolens) on gas production and rumen fermentation in vitro as a natural substitute for the ionophore antibiotic monensin. These EOs are chemically characterized by Gas Chromatography-Mass and evaluated in vitro at four different concentrations (0, 15, 30, and 45 µl per 45 ml buffered rumen fluid) regarding their effects on gas production and rumen fermentation characteristics and were compared to those of monensin. Compared to the negative control, monensin significantly depressed gas production and truly degraded dry matter (TDDM) but enhanced propionate production. All EOs except P. graveolens significantly decreased gas production with increasing concentrations. TDDM was significantly reduced with C. citratus (at 45 µl) and P. graveolens (at 30 and 45 µl). No significant change was detected in the ammonia nitrogen concentration with all assayed EOs except C. nardus and C. Citrus. Compared to monensin and the negative control, C. nardus and C. Citrus reduced the ammonia concentration at high levels. High levels of all tested EOs significantly reduced protozoa counts. The EOs of C. citratus (at 45 µl) and P. graveolens (at 30 and 45 µl) also significantly increased the acetate proportion. Moreover, the acetate to propionate ratio was significantly increasedby30 µl P. graveolens. The results of the current study concluded that the tested EOs, except P. graveolens, efficiently diminished gas production with a similar potency to monensin. Furthermore, they exceed the monensin in their ability to reduce the ammonia nitrogen concentration and protozoa count without adversely affecting volatile fatty acid levels. But, they were less effective than monensin in modifying ruminal volatile fatty acid profile especially propionate and acetate to propionate ratio. Hence, P. guajava, C. nardus, and C. citratus EOs could be a safe and promising rumen manipulator.

Effects of Moringa oleifera extracts and monensin on performance of growing rabbits

This study was designed to assess the effects of Moringa oleifera (moringa) leaf ethanolic extract (MLEE), moringa root ethanolic extract (MREE), and ionophore antibiotic monensin on growth performance, hematological variables, blood plasma metabolites and antioxidant indicators, digestibility, and cecal fermentation of growing rabbits. A total of 96 growing rabbits weaned at 35 d of age were randomly assigned to 4 treatments with 12 male and 12 female rabbits/treatment. The first treatment was the control (no additives), the second treatment was a basal diet containing 10 mg monensin/kg dry matter (DM), and the third and fourth treatments were MLEE and MREE, respectively, at concentration of 250 mg/L drinking water. From 35 to 77 d of age, the rabbits were housed in groups (8 cages/treatment, 3 rabbits/cage), and growth performance variables including body weight (BW), feed consumption, BW gain, and feed conversion ratio were recorded. From 78 to 93 d of age, after the end of the growth performance trial, the rabbits were housed individually. At 87 d of age, blood samples were collected from 10 rabbits (5 males and 5 females) that were selected randomly to determine hematological variables and blood plasma metabolites and antioxidant indicators. From 87 to 93 d of age, 8 male rabbits/treatment were selected randomly to determine digestibility, N balance, and cecal fermentation activity. The major chemical compounds in MLEE were α-linolenic acid, glycerine, ascorbic acid, and diethyl phthalate, whereas ascorbic acid, S-phenylmercapturic acid, 2-myristynoyl pantetheine, and 8,11,14-docosatrienoic acid, methyl ester were major compounds in MREE. Both the MLEE and MREE treatments improved (P < 0.05) final BW and overall feed conversion ratios compared to the monensin and control treatments. From d 64 to 77 of age, the monensin treatment decreased (P < 0.05) feed consumption compared to the other treatments. None of the treatments affected hematological variables, but the monensin, MLEE, and MREE treatments increased (P < 0.05) plasma albumin concentration. The monensin and MREE treatments decreased (P < 0.05) plasma globulin concentration compared to the MLEE and control treatments. Compared to the control, the monensin, MLEE, and MREE treatments increased (P < 0.05) plasma total antioxidant capacity by 66.17% but decreased (P < 0.05) both the plasma malondialdehyde concentration by 24.96% and reduced glutathione activity by 25.96%. The MLEE treatment increased (P < 0.05) the apparent digestibility of acid detergent fiber (6.49%) and hemicellulose (13.53%) compared to the other treatments. The MLEE and MREE treatments increased (P < 0.05) apparent cellulose digestibility and body N retention compared to the monensin and control treatments. The lowest (P < 0.05) cecal total volatile fatty acid concentration was observed with the monensin treatment, while the greatest (P < 0.05) was with the MLEE treatment. Compared to the control, the monensin and MLEE treatments decreased the molar proportions of butyrate (72.20 and 31.10%, respectively; P < 0.05) and increased the molar proportions of propionate (140.70 and 67.80%, respectively; P < 0.05). In conclusion, MLEE or MREE could be used at concentration of 250 mg/L drinking water to promote growth of weaned rabbits.

Competitive electrochemical immunosensor for maduramicin detection by multiple signal amplification strategy via hemin@Fe-MIL-88NH2/AuPt

Maduramicin (MD) is a type of monoglycoside polyether ionophore antibiotic that can effectively treat coccidiosis and facilitate animal growth. However, its extensive and excessive use brings potential risk to human health. Herein, an electrochemical immunosensor based on indirect competitive format was fabricated for analysis of MD residue in eggs by a multiple signal amplification system. Initially, Au nanoparticles were deposited onto glassy carbon electrode surface to load the coating antigen MD-BSA and to improve conductivity. Then the signal amplification platform was constructed by encapsulating hemin into Fe-MIL-88 NH2 metal-organic frameworks (hemin@MOFs), and then the obtained composites were decorated with AuPt nanoparticles. The synthesized hemin@MOFs/AuPt was not only used as a signal amplification mediator, but also utilized as a carrier for immobilization of horseradish peroxidase-conjugated affinipure goat anti-mouse antibody (Ab2-HRP) and horseradish peroxidase (HRP). The constructed hemin@MOFs/AuPt-Ab2-HRP bioconjugates could effectively amplify the current signal since hemin@MOFs, AuPt and HRP all exhibited high catalytic activity towards the hydrogen peroxide. Moreover, the established immunosensor showed high sensitivity and stability during the detection procedure. With the synergistic catalytic effect of hemin@MOFs, AuPt and HRP, a wide detection range of 0.1–50 ng mL−1 and a low detection limit of 0.045 ng mL−1 were achieved (S/N = 3), respectively. Ultimately, the developed method displayed excellent performance in practical applications, providing a promising probability to detect other veterinary drug residues to guarantee food safety.

Prediction of Transformation Products of Monensin by Electrochemistry Compared to Microsomal Assay and Hydrolysis

The knowledge of transformation pathways and identification of transformation products (TPs) of veterinary drugs is important for animal health, food, and environmental matters. The active agent Monensin (MON) belongs to the ionophore antibiotics and is widely used as a veterinary drug against coccidiosis in broiler farming. However, no electrochemically (EC) generated TPs of MON have been described so far. In this study, the online coupling of EC and mass spectrometry (MS) was used for the generation of oxidative TPs. EC-conditions were optimized with respect to working electrode material, solvent, modifier, and potential polarity. Subsequent LC/HRMS (liquid chromatography/high resolution mass spectrometry) and MS/MS experiments were performed to identify the structures of derived TPs by a suspected target analysis. The obtained EC-results were compared to TPs observed in metabolism tests with microsomes and hydrolysis experiments of MON. Five previously undescribed TPs of MON were identified in our EC/MS based study and one TP, which was already known from literature and found by a microsomal assay, could be confirmed. Two and three further TPs were found as products in microsomal tests and following hydrolysis, respectively. We found decarboxylation, O-demethylation and acid-catalyzed ring-opening reactions to be the major mechanisms of MON transformation.

Maduramicin inactivation of Akt impairs autophagic flux leading to accumulated autophagosomes-dependent apoptosis in skeletal myoblast cells.

It has been clinically documented that maduramicin (Mad), a polyether ionophore antibiotic widely used in the control of coccidiosis in poultry worldwide, can elicit skeletal muscle degeneration, heart failure, and even death in animals and humans, if improperly used. Here, we show that Mad induced apoptosis dose-dependently, which was associated with impaired autophagic flux in skeletal myoblast (C2C12 and L6) cells. This is supported by the findings that Mad treatment resulted in increase of autophagosomes with a concomitant elevation of LC3-II and p62 in the cells. Also, Mad increased co-localization of mCherry and GFP tandem-tagged LC3 puncta in the cells, suggesting a blockage of autophagic flux. Furthermore, addition of chloroquine (CQ) strengthened the basic and Mad-enhanced LC3-II and p62 levels, autophagosome formation and cell apoptosis, whereas pretreatment with rapamycin alleviated the effects in the cells exposed to Mad. Moreover, we noticed that Mad treatment inactivated Akt dose-dependently. Inhibition of Akt with inhibitor X potentiated Mad-induced decrease in phosphorylated Akt, and increases in LC3-II and p62 levels, autophagosome formation and cell apoptosis, whereas ectopic expression of constitutively active Akt rendered resistance to these events. Collectively, these results indicate that Mad inactivation of Akt impairs autophagic flux leading to accumulated autophagosomes-dependent apoptosis in skeletal myoblast cells. Our findings suggest that manipulation of Akt activity to improve autophagic flux is a promising strategy against Mad-induced myotoxicity.

Measurements of Ion Binding to Lipid-Hosted Ionophores by Affinity Chromatography.

The binding affinity between antibiotic ionophores and alkali ions within supported lipid bilayers was evaluated using affinity chromatography. We used zonal elution and frontal analysis methods in nanovolume liquid chromatography to characterize the binding selectivity of the carrier and channel ionophores valinomycin and gramicidin A within different phosphatidylcholine bilayers. Distinct binding sensitivity to the lipid phase, both in affinity and selectivity, is observed for valinomycin, whereas gramicidin is less sensitive to changes in a membrane environment, behavior that is consistent with ion binding occurring within the interior of an established channel. There is good agreement between the chromatographic retention and the reported binding selectivity measured by other techniques. Surface potential near the binding site affects ion retention and the apparent association binding constants, but not the binding selectivity or enthalpy measurements. A model accounting for the surface potential contributions of retained ions during frontal analyses yields values close to intrinsic binding constants for gramicidin A (KA for K+ between 70 and 120 M-1) using reasonable estimates of the initial potential that is postulated to arise from the underlying silica.

Methane and Constipation-predominant Irritable Bowel Syndrome: Entwining Pillars of Emerging Neurogastroenterology.

Functional gut disorders have long known to cause depravity in quality of life. Among the group of these heterogeneous disorders, irritable bowel syndrome (IBS) has been known to affect a large chunk of our population. IBS is not as simple as it sounds. Caused by a multitude of factors, the heterogeneity of this disorder has laid the foundation for research and the new principles of neurogastroenterology. Dysbiosis and methane production are one of the forthcoming factors that are currently under investigation. Down the road of exclusive enteric anaerobic fermentation of polysaccharides, methane is produced. It was considered to be an inert gas in the past, with little to no role in gut activity but now it is established that it has an impressive role in the etiology of constipation-predominant IBS (C-IBS). Acting as a neurotransmitter, it is known to affect ileal and colonic transit time, which has currently been shown in animal studies. Many laxatives, ionophore antibiotics, drugs like rifamixin and neomycin have been targeted against this very principle. Lately, lovastatin has emerged as a potential pharmacologic therapy to devoid the gut of methane without disrupting the gut niche in itself and has shown promise in relieving the symptoms of C-IBS. The goal of this article is to compile and assemble the literature available on IBS and the neuromodulation of methane to teach physicians and research scientists about the current age of gastroenterology and the growing need to emphasize the role of methane in the symptomatology of functional gut disorders like C-IBS.

Identification of dinactin, a macrolide antibiotic, as a natural product-based small molecule targeting Wnt/β-catenin signaling pathway in cancer cells

Despite the fact that hyper-activation of Wnt/β-catenin signaling pathway has been seen in many cancers, including liver, colorectal and lung carcinoma, no small molecule inhibitors are available that specifically target this pathway. In this study, we analyzed the impact of dinactin (DA), an antibiotic ionophore produced by Streptomycesspecies, as an effective small molecule targeting Wnt/β-catenin signaling pathway in cancer cells. We performed MTT assays to investigate cell viability and proliferation after exposure to small molecules. Protein expression analysis was carried out by western blotting. Top-Flash reporter assays were used to score for β-catenin signaling and cell cycle analysis was carried out by flow cytometry. In the first set of experiments, DA was seen to selectively inhibit the proliferation of HCT-116 and HepG2 cancer cells, unlike HEK-293 cells (a lowtumorigenic cell line), in apoptosis-independent manner. Further, DA was seen to block the G1/S progression and decrease the expression of cyclin D1 in cancer cells. Since cyclin D1 is the downstream target gene of Wnt/β-catenin signaling, we examined the impact of DA on TCF-dependent β-catenin activity using Top-Flash reporter assay. Interestingly, DA significantly decreased Top-Flash activity at lower nano-molar concentrations when compared with salinomycin in HCT-116 and HepG2 cells. We report the identification of dinactin as a natural product-based small molecule that effectively blocks the Wnt/β-catenin signaling pathway in cancer cells at nano-molar concentration. We anticipate that DA could be developed as a novel drug for anti-cancer therapy and for the management of neuropathic pain.