Specific Inhibitory Compounds Research Articles

Pointing Out Opportunities to Increase Grassland Pastures Productivity via Microbial Inoculants: Attending the Society’s Demands for Meat Production with Sustainability

Estimates are that land area occupied by grass pasture far exceeds that of other crops; at least half are at some stage of degradation. The use of elite plant-growth-promoting microorganisms (PGPM) as inoculants represents an important strategy to achieve qualitative and quantitative improvements in forage biomass, increasing the productivity and sustainability of livestock production. Several studies have reported the benefits of PGPM in grass pastures, with an emphasis on bacteria of the genera Azospirillum, Pseudomonas, Bacillus, rhizobia, and on arbuscular mycorrhiza fungi (AMF). The main grasses studied are Urocholoa (syn. Brachiaria), Megathyrsus (syn. Panicum), Paspalum, Cynodon, and Agropyron. Several microbial processes associated with improvements in root and shoot growth, nutrient content in biomass, and other benefits have been described. Promotion of plant growth has been associated with the synthesis of phytohormones and enzymes regulating several steps of plant development, nutrient mineralization by release of microbial molecules, biological nitrogen fixation, nutrient uptake facilitation by means of molecules such as exopolysaccharides, amongst others. Outstanding benefits of increased root growth, resulting in higher uptake of water and nutrients, either by phytohormones released by bacteria or by expanding root surface by AMF, have been reported. Biocontrol is another important property of PGPM, by a variety of mechanisms, including the synthesis of antimicrobial molecules, lytic enzymes, siderophores, and the release of specific inhibitory compounds such as hydrogen cyanide. Although improvements in forage management can enhance microbial performance, as shown for AMF, in general, inoculation with elite strains positively impacts growth parameters. Globally, the use of microbial inoculants has significantly increased in the past few years, but their application is still modest on grass pastures, which are generally degraded and would need special use of microbial inoculants for reclamation. Efforts towards increasing the use of PGPM in pastures can have deep positive environmental, economic, and social impacts worldwide.

The identification of small molecule inhibitors of the plant inositol phosphorylceramide synthase which demonstrate herbicidal activity

Resistance to 157 different herbicides and 88% of known sites of action has been observed, with many weeds resistant to two or more modes. Coupled with tighter environmental regulation, this demonstrates the need to identify new modes of action and novel herbicides. The plant sphingolipid biosynthetic enzyme, inositol phosphorylceramide synthase (IPCS), has been identified as a novel, putative herbicide target. The non-mammalian nature of this enzyme offers the potential of discovering plant specific inhibitory compounds with minimal impact on animals and humans, perhaps leading to the development of new non-toxic herbicides. The best characterised and most highly expressed isoform of the enzyme in the model-dicot Arabidopsis, AtIPCS2, was formatted into a yeast-based assay which was then utilized to screen a proprietary library of over 11,000 compounds provided by Bayer AG. Hits from this screen were validated in a secondary in vitro enzyme assay. These studies led to the identification of a potent inhibitor that showed selectivity for AtIPCS2 over the yeast orthologue, and activity against Arabidopsis seedlings. This work highlighted the use of a yeast-based screening assay to discover herbicidal compounds and the status of the plant IPCS as a novel herbicidal target.

Identification and characterisation of ataxia telangiectasia mutated (ATM) in regulation of oxidative stress-induced vascular endothelial cells senescence

BackgroundAging is known to be a major cardiovascular risk factor. Recent studies provide evidence for a critical role of cellular senescence in the process of vascular aging including hypertension and...

Acceleration of Enterococcus faecalis Biofilm Formation by Aggregation Substance Expression in an Ex Vivo Model of Cardiac Valve Colonization

Infectious endocarditis involves formation of a microbial biofilm in vivo. Enterococcus faecalis Aggregation Substance (Asc10) protein enhances the severity of experimental endocarditis, where it has been implicated in formation of large vegetations and in microbial persistence during infection. In the current study, we developed an ex vivo porcine heart valve adherence model to study the initial interactions between Asc10+ and Asc10− E. faecalis and valve tissue, and to examine formation of E. faecalis biofilms on a relevant tissue surface. Scanning electron microscopy of the infected valve tissue provided evidence for biofilm formation, including growing masses of bacterial cells and the increasing presence of exopolymeric matrix over time; accumulation of adherent biofilm populations on the cardiac valve surfaces during the first 2–4 h of incubation was over 10-fold higher than was observed on abiotic membranes incubated in the same culture medium. Asc10 expression accelerated biofilm formation via aggregation between E. faecalis cells; the results also suggested that in vivo adherence to host tissue and biofilm development by E. faecalis can proceed by Asc10-dependent or Asc10-independent pathways. Mutations in either of two Asc10 subdomains previously implicated in endocarditis virulence reduced levels of adherent bacterial populations in the ex vivo system. Interference with the molecular interactions involved in adherence and initiation of biofilm development in vivo with specific inhibitory compounds could lead to more effective treatment of infectious endocarditis.

Ataxia Telangiectasia Mutated (ATM)-mediated DNA Damage Response in Oxidative Stress-induced Vascular Endothelial Cell Senescence

Oxidative stress regulates dysfunction and senescence of vascular endothelial cells. The DNA damage response and its main signaling pathway involving ataxia telangiectasia mutated (ATM) have been implicated in playing a central role in mediating the actions of oxidative stress; however, the role of the ATM signaling pathway in vascular pathogenesis has largely remained unclear. Here, we identify ATM to regulate oxidative stress-induced endothelial cell dysfunction and premature senescence. Oxidative stress induced senescence in endothelial cells through activation/phosphorylation of ATM by way of an Akt/p53/p21-mediated pathway. These actions were abrogated in cells in which ATM was knocked down by RNA interference or inhibited by specific inhibitory compounds. Furthermore, the in vivo significance of this regulatory pathway was confirmed using ATM knock-out mice in which induction of senescent endothelial cells in the aorta in a diabetic mouse model of endothelial dysfunction and senescence was attenuated in contrast to pathological changes seen in wild-type mice. Collectively, our results show that ATM through an ATM/Akt/p53/p21-dependent signaling pathway mediates an instructive role in oxidative stress-induced endothelial dysfunction and premature senescence.

Mitogen-activated protein (MAP) kinases mediate PMMA-induction of osteoclasts

Inflammatory osteolysis induced by implant-derived wear debris is associated with infiltration of various cell-types to the implant–bone interface leading to abundant secretion of pro-inflammatory cytokines and activation of proteinases that together lead to propagation of the localized inflammatory response and periprosthetic bone erosion. Tumor necrosis factor family members are considered to be direct mediators of inflammation and osteolysis. These cytokines exert their osteoclastic effects via activation of the transcription factor NF-κB and certain MAP kinases, including c-Jun, Erks and p38, all known to be essential for the development of osteoclasts. We have recently documented that the osteoclastogenic cytokines TNF and RANKL play a pivotal role in the development of inflammatory osteolysis. We have also found that PMMA particles stimulate osteoclastogenesis, at least in part, by induction of RANKL, TNF, and by activation of the transcription factor NF-κB. More importantly, our data indicate that inhibitors of the osteoclastogenic factors, TNF and RANKL abrogate particle-induced osteoclastogenesis. In the current study, we investigated if PMMA particles activate MAP kinases, and the potential role of these kinases as mediators of osteolysis. Using kinase assays, we show that in osteoclast precursors, PMMA particles markedly and rapidly activate p38 and ERK MAP kinases. This activation was specific, evident by complete blockade with specific inhibitory compounds. Similarly, we show that PMMA particles activate the JNK pathway, which is known to be involved in inflammatory and osteoclastogenic events. We also show that p38 MAP kinase regulates PMMA-activation of NF-κB, thus providing a possible mechanism for particle action in osteoclast precursors. Finally, we provide evidence that specific inhibitors of MAP kinases are capable of inhibiting PMMA-stimulated osteoclastogenesis. These data provide evidence that MAP kinases are potent mediators of particle-induced osteoclastogenesis.

Amyloids and Other Abnormal Assembly Processes FASEB Summer Conference Copper Mountain, Colorado June 11-16, 2000

The amyloid field has, over the past few years, experienced a rapid growth in a number of areas. The intent of this meeting was to examine the complete amyloid pathway from crystal structures to animal models and the creation of novel methods of treating these disorders. This involved bringing together different disciplines, ideas and approaches which examined amyloidogenic proteins from unique perspectives. The first keynote address was given by D. Cleveland (UC San Diego) who reviewed the aggregation pathway of superoxide dismutase (SOD) as it relates to amyotrophic lateral sclerosis (ALS) or Lou Gehrig's disease. This address provided insights into all aspects of the abnormal accumulation of SOD from the effects of mutations on protein folding to the recapitulation of these deposits in transgenic mice. The second keynote was delivered by J. Kelly (Scripps) who dealt with the complexities of amyloid assembly as seen from a protein folding viewpoint and addressed the question of whether or not it is possible to design specific inhibitory compounds. Using the transthyretin (TTR) model, it was demonstrated that aggregation could be monitored by techniques such as utracentrifugation and that molecules which maintained a stable native conformation were capable of reducing fibril formation. The combination of these two addresses encapsulated the overall goal of this meeting.

EFFECT OF GRAPEFRUIT JUICE AND OTHER JUICES ON P-GLYCOPROTEIN FUNCTION OF DRUG ABSORPTION IN THE INTESTINE

We investigated the inhibitory effect of components in grapefruit juice (GFJ) and orange juice (OJ) on the uptake of vinblastine, a substrate of P-glycoprotein, by Caco-2 cells. Significantly increased uptake of [3H] vinblastine at the steady state by Caco-2 cells was observed in the presence of GFJ and OJ. This suggests that the both juices involve compounds inhibiting P-gp function. We examined the effect of three organic solvent extracts, ethyl acetate, diethyl ether and methylene chrolide, of GO and OJ on the uptake of [3H] vinblastine. The inhibitory effect was shown more in ethyl acetate extracts than the other two extracts, in the both case of GFJ and OJ. Furthermore, the ethyl acetate extracts of GFJ and OJ were fractionated by cosmosil column chromatography with stepwise methanol solution. Inhibitory effect of GFJ on P-gp function was maximun in the fraction eluted with 60% methanol solution, while OJ's was maximun in 70% methanol solution. Each fraction contained different component. Moreover, inhibitory effect of GFJ on CYP3A4 was maximun in the fraction eluted with 80% methanol solution. This study suggested the existence of unknown specific inhibitory compounds in GFJ and OJ for the P-gp, but not for CYP3A4 function.

Anaerobic treatment applications and fundamentals: substrate specificity during phase separation

ABSTRACT: The effect of phase separation on anaerobic biodegradation was evaluated in terms of thermodynamic principles. Individual components of common wastewaters were considered, and results indicate that the response of different substrates to phase‐separated anaerobic treatment is diverse. A key element is the response of soluble substrates to acidification when syntrophic relationships are eliminated and biological reactions that require syntrophic relationships are altered. The acidification of carbohydrates and some proteins is thermodynamically favorable. In contrast, the acidification of fatty acids and aromatics is not thermodynamically favorable unless a sink for reducing equivalents is available. Reported data are consistent with this analysis and phase separation has been observed to enhance the treatment of carbohydrate wastewaters. In addition, the potential for detoxification of specific inhibitory compounds under acidifying conditions has been observed. Whether or not treatment of certain substrates will be enhanced by phase separation should be considered in the choice of reactor design, recognizing that various reactor designs exhibit different degrees of phase separation. This paper presents an overview on how phase‐separated treatment affects the biodegradation of specific substrates and relates this to the selection of reactor designs.

Relevance of Spontaneous Activity to Urinary Bladder Function: An in Vitro and in Vivo Study

The presence and functional significance (if any) of spontaneous activity in the normal urinary bladder during filling is a controversial subject. One model used by many investigators to study spontaneous activity has been isolated urinary bladder smooth muscle strips. Although spontaneous activity is a property commonly observed in isolated urinary bladder strip preparations, the in vitro whole bladder preparation (rabbit) is devoid of spontaneous activity. Additionally, under normal conditions the in vivo rabbit bladder does not display spontaneous activity during the filling phases of micturition.The present study compares the spontaneous activity of isolated smooth muscle strips, the whole bladder preparation, and the catheterized in vivo bladder (rabbit). The results are as follows:1) The spontaneous activity (frequency and amplitude) of isolated strips is extremely variable among strips of the same bladder. 2) Spontaneous activity is not affected by the following specific inhibitory compounds: tetrodotoxin, atropine, phentolamine, propranolol and hexamethonium. This indicates that spontaneous activity observed in isolated strips is myogenic in nature and not dependent on the activation of specific autonomic receptors. 3) The in vitro whole bladder preparation shows no spontaneous activity at any volume or pressure unless longitudinal tension is applied. 4) The spontaneous activity of the whole bladder subjected to longitudinal tension is not affected by the same compounds mentioned above. 5) Spontaneous activity of the in vivo bladder is absent at low intravesical volumes and pressures. Spontaneous activity develops upon reaching a critical pressure. However, this activity is completely inhibited by intravenous ganglionic blockade (hexamethonium). In the presence of hexamethonium, the in vivo bladder is devoid of spontaneous activity at any volume or pressure, thus the in vivo “spontaneous activity” is mediated through neuronal reflexes.It is concluded that under normal circumstances the rabbit bladder is devoid of myogenic spontaneous activity and that the spontaneous activity observed in isolated strips is directly related to longitudinal stretch. Since under normal conditions the bladder is not subjected to longitudinal stretch, the spontaneous activity observed in the isolated strip studies has little physiological significance under normal conditions, but could help explain the pathophysiology of certain dysfunctions during the filling stage of micturition.

Factors Affecting Nitrification

Experiments were conducted to determine the effects of various factors upon the rate of nitrate production by a stock culture of nitrifying bacteria. Actively metabolizing heterotrophic bacteria were found to have no effect upon the rate of nitrate production by the autotrophs. Under some circumstances, however, heterotrophs released materials into the medium which appeared to stimulate the autotrophs. Common domestic sewage was also found to have no effect upon the rate of nitrate production. When samples were removed from batch activated sludge reactors during the period of substrate removal and mixed with the autotrophs, no deleterious effects upon the rate of nitrate production were observed. The results suggest that unless specific inhibitory compounds are present in a wastewater, neither organic matter nor heterotrophic bacteria will have a deleterious effect upon the activity of autotrophic nitrifiers.