Discovery of a Remarkable Methyl Shift Effect in the Vanilloid Activity of Triterpene Amides

As part of a study on triterpenoid conjugates, the dietary pentacyclic triterpenoids oleanolic (2a) and ursolic acids (3a) were coupled with vanillamine, and the resulting amides (2b and 3b, respectively) were assayed for activity on the vanilloid receptor TRPV1. Despite a structural difference limited to the location of a methyl group in their conformationally rigid pentacyclic core, oleanoloyl vanillamide dramatically outperformed ursoloyl vanillamide in terms of potency (EC50 = 35 ± 2 nM for 2b and 5.4 ± 2.3 μM for 3b). Using molecular docking and dynamics, this difference was translated into distinct accommodation modes at the TRPV1 vanillyl ligand pocket, suggesting a critical role of a C-H πphenyl interaction between the triterpenoid C-29 methyl and Phe591 of TRPV1. Because the molecular mechanisms underlying the activation process of transient receptor channels (TRPs) remain to be fully elucidated, the observation of spatially restricted structure-activity information is of significant relevance to identify the molecular detail of TRPV1 ligand gating

Comparison of Antibiotic Resistance Profile and Biofilm Production of Staphylococcus aureus Isolates Derived from Human Specimens and Animal-Derived Samples

Background: The diusion of antimicrobial resistance is a significant concern for public health worldwide. Staphylococcus aureus represents a paradigm microorganism for antibiotic resistance in that resistant strains appear within a decade after the introduction of new antibiotics. Methods: Fourteen S. aureus isolates from human specimens and twenty-one from samples of animal origin, were compared for their antimicrobial resistance and biofilm capability. In addition, they were characterized at the molecular level to detect the antimicrobial resistance mecA gene and genes related with enterotoxin, toxin, and biofilm production. Results: Both phenotypic and molecular analysis showed main dierences among human- and animal-derived isolates. Among the human-derived isolates, more multidrug-resistant isolates were detected and mecA gene, enterotoxin, and toxin genes were more prevalent. Dierent genes involved in biofilm production were detected with bap present only in animal-derived isolates and sasC present in both isolates, however, with a higher prevalence in the human-derived isolates. Biofilm capability was higher in human-derived isolates mainly associated to the sasC gene. Conclusions: The overall results indicate that human S. aureus isolates are more virulent and resistant than the isolates of animal origin randomly selected with no infection anamnesis. This study confirms that selection for more virulent and resistant S. aureus strains is related to the clinical practice

Interactions of GFAP with ceftriaxone and phenytoin: SRCD and molecular docking and dynamic simulation

Abstract Background GFAP is the major intermediate filament protein in mature astrocytes. Its increased expression and aggregation was firstly associated to Alexander's disease, and successively in different neurological diseases including scrapie, Alzheimer's and Creutzfeld–Jacob diseases. Recently, ceftriaxone a multi-potent β-lactam antibiotic able to overcome the blood–brain barrier, successfully eliminated the cellular toxic effects of misfolded mutated GFAP, similarly to phenytoin sodium, in a cellular model of Alexander's disease and inhibited α-synuclein aggregation protecting PC12 cells from the exposure to 6-hydroxydopamine. Methods In this study, synchrotron radiation circular dichroism spectroscopy has been used to obtain structural information about the GFAP-ceftriaxone (phenytoin) interactions, while computational methods allowed the identification of the relevant putative binding site of either ceftriaxone or phenytoin on the dimer structure of GFAP, permitting to rationalize the spectroscopic experimental results. Results We found that GFAP exhibited enhanced stability upon the addition of two equivalents of each ligands with ceftriaxone imparting a more spontaneous interactions and a more ordered complex system than phenytoin. Conclusions SRCD data and MD models indicate a stronger protective effect of ceftriaxone in neurological disorders characterized by an increased production and polymerization of GFAP. General significance This result, in addition to our previous works in which we documented that ceftriaxone interacts with α-synuclein inhibiting its pathological aggregation and that a cyclical treatment with this molecule in a patient with adult-onset Alexander's disease halted, and partly reversed, the progression of neurodegeneration, suggests the possibility of a chaperone-like effect of ceftriaxone on protein involved in specific neurodegenerative diseases

Antibiotic Resistance Profiling, Analysis of Virulence Aspects and Molecular Genotyping of Staphylococcus aureus Isolated in Sicily, Italy

Staphylococcus aureus is the major cause of foodborne diseases worldwide. In this retrospective study, 84 S. aureus strains were characterized. The collection comprises 78 strains isolated during 1998 and 2014 from dairy products and tissue samples from livestock bred for dairy production in Sicily. One isolate was obtained from a pet (dog), one from an exotic animal (a circus elephant), and four human isolates were obtained during a severe food poisoning outbreak that occurred in Sicily in 2015. All the strains were characterized by pulsed-field gel electrophoresis (PFGE), for antibiotic resistance and presence of toxin genes. PFGE results showed 10 different pulsotypes, with three relatively frequent and three unique. The antibiotic resistance profiling showed that penicillin G (35.7%) and tetracycline (20.2%) resistance is largely spread. Most isolates contained at least one toxin gene making them a potential threat for public health. Enterotoxin sec gene was observed in 28.6% and seg in 23.8% of the strains, respectively; the human isolates were the only ones to concurrently harbor both seg and sei genes. In addition, 24 isolates were randomly selected and analyzed by multilocus sequence typing. Interestingly, the analysis showed the presence of 12 sequence types (STs), of which 6 were novel. One of them, ST700, was detected in 29% of the isolates and was found to be spread throughout Sicily. ST700 has been present in the island for almost 16 years (1998-2014) and it shows no host preference since it was isolated from different ruminant species. Four human isolates shared both the pulsotype (PT10) and the sequence type (ST9), as well as the virulence genes (seg-sei); this observation suggests that the isolates originated from a single clone, although they were obtained from two different individuals

Insights into the Structural Basis of the GADD45β-mediated Inactivation of the JNK Kinase, MKK7/JNKK2

NF-kappaB/Rel factors control programmed cell death (PCD), and this control is crucial to oncogenesis, cancer chemoresistance, and antagonism of tumor necrosis factor (TNF) alpha-induced killing. With TNFalpha, NF-kappaB-mediated protection involves suppression of the c-Jun-N-terminal kinase (JNK) cascade, and we have identified Gadd45beta, a member of the Gadd45 family, as a pivotal effector of this activity of NF-kappaB. Inhibition of TNFalpha-induced JNK signaling by Gadd45beta depends on direct targeting of the JNK kinase, MKK7/JNKK2. The mechanism by which Gadd45beta blunts MKK7, however, is unknown. Here we show that Gadd45beta is a structured protein with a predicted four-stranded beta-sheet core, five alpha-helices, and two acidic loops. Association of Gadd45beta with MKK7 involves a network of interactions mediated by its putative helices alpha3 and alpha4 and loops 1 and 2. Whereas alpha3 appears to primarily mediate docking to MKK7, loop 1 and alpha4-loop 2 seemingly afford kinase inactivation by engaging the ATP-binding site and causing conformational changes that impede catalytic function. These data provide a basis for Gadd45beta-mediated blockade of MKK7, and ultimately, TNFalpha-induced PCD. They also have important implications for treatment of widespread diseases

Elongation of the Hydrophobic Chain as a Molecular Switch:Discovery of Capsaicin Derivatives and Endogenous Lipids as Potent Transient Receptor Potential Vanilloid Channel 2 Antagonists

The transient receptor potential vanilloid type-2 (TRPV2) protein is a nonselective Ca2+ permeable channel member of the TRPV subfamily, still considered an orphan TRP channel due to the scarcity of available selective and potent pharmacological tools and endogenous modulators. Here we describe the discovery of novel synthetic long-chain capsaicin derivatives as potent TRPV2 antagonists in comparison to the totally inactive capsaicin, the role of their hydrophobic chain, and how the structure-activity relationships of such derivatives led, through a ligand-based approach, to the identification of endogenous long-chain fatty acid ethanolamides or primary amides acting as TRPV2 antagonists. Both synthetic and endogenous antagonists exhibited differential inhibition against known TRPV2 agonists characterized by distinct kinetic profiles. These findings represent the first example of both synthetic and naturally occurring TRPV2 modulators with efficacy in the submicromolar/low-micromolar range, which will be useful for clarifying the physiopathological roles of this receptor, its regulation, and its targeting in pathological conditions.We gratefully acknowledge financial support from Universitat de Lleida, Ministerio de Educación, Cultura y Deporte and Banco Santander (Programa UdL-Impuls). The authors are grateful to the Serveis Cientifictècnics (SCT) of the Universitat de Lleida for providing us with spectroscopic and chromatographic facilities. We acknowledge Dr. Alberto Minassi, Dipartimento di Scienze del Farmaco, Universitàdel Piemonte Orientale, Novara, Italy, for the kind gift of olvanil

Combination of the Systemin peptide with the beneficial fungus Trichoderma afroharzianum T22 improves plant defense responses against pests and diseases

Trichoderma spp. are among the most widely used plant beneficial fungi in agriculture. Its interaction with the plant triggers resistance responses by the activation of Induced Systemic Resistance mediated by Jasmonic acid and Ethylene and/or Systemic Acquired Resistance, which involves Salicylic acid, with the consequent control of a wide range of plant parasites. However, the benefit they can confer to plants may be reduced or nullified by environmental conditions or fungal ecological fitness. A novel approach to enhance their effectiveness in plant defense is to combine them with bioactive molecules including plant-derived compounds. Here, we show that plant treatment with Trichoderma afroharzianum (strain T22) and Systemin, a tomato peptide active in triggering plant defense, confers protection against the fungal pathogens Fusarium oxysporum, Botrytis cinerea and the insect pest Tuta absoluta. The observed defensive response was associated with an increase of Jasmonic acid and related metabolites and a decrease of Salicili acid

Serum epidermal growth factor predicts cognitive functions in early, drug-naive Parkinson's disease patients

Epidermal growth factor (EGF) has been proposed as a candidate biomarker for cognitive impairment in Parkinson's disease (PD). We aimed to assess the relationship between serum EGF and cognitive functions in early, drug-naive PD patients and evaluate the predictive value of EGF on cognitive functions in a 2-year follow-up study. Serum EGF was measured in 65 early, drug-naive PD patients, that underwent a comprehensive neuropsychological battery. Motor symptoms were assessed by means of the Unified Parkinson's Disease Rating Scale, Part III (UPDRS-III). Neuropsychological evaluation was repeated after 2 years. Spearman's rank correlation was used to assess the relationship between serum EGF levels and neuropsychological variables. Linear regression analysis was used to evaluate the relationship between EGF and neuropsychological scores as well as other variables (age, gender, UPDRS-III, levodopa equivalent dose, and type of treatment at follow-up) potentially affecting cognitive performance. Variation over time in cognitive scores was analyzed using repeated-measures ANOVA. At baseline, EGF was the only significant variable associated with performance on semantic fluency (R (2) = 0.131; p = 0.005). EGF levels (p = 0.025), together with UPDRS-III (p = 0.009) and age (p = 0.011), were associated with performance on frontal assessment battery (R (2) = 0.260). At 2-year follow-up, EGF was the only significant variable to predict performance on semantic fluency (R (2) = 0.147; p = 0.025) and color naming task of Stroop color-word test (R (2) = 0.121; p = 0.044). Serum EGF levels are related to frontal and temporal cognitive functions in early, drug-naive PD patients and predict performance on frontal and posterior cognitive functions at 2-year follow-up. EGF is proposed as a potential serum biomarker for early cognitive impairment in PD

Fishing for Targets of Alien Metabolites: A Novel Peroxisome Proliferator-Activated Receptor (PPAR) Agonist from a Marine Pest

Although the chemical warfare between invasive and native species has become a central problem in invasion biology, the molecular mechanisms by which bioactive metabolites from invasive pests influence local communities remain poorly characterized. This study demonstrates that the alkaloid caulerpin (CAU)—a bioactive component of the green alga Caulerpa cylindracea that has invaded the entire Mediterranean basin—is an agonist of peroxisome proliferator-activated receptors (PPARs). Our interdisciplinary study started with the in silico prediction of the ligand-protein interaction, which was then validated by in vivo, ex vivo and in vitro assays. On the basis of these results, we candidate CAU as a causal factor of the metabolic and behavioural disorders observed in Diplodus sargus, a native edible fish of high ecological and commercial relevance, feeding on C. cylindracea. Moreover, given the considerable interest in PPAR activators for the treatment of relevant human diseases, our findings are also discussed in terms of a possible nutraceutical/pharmacological valorisation of the invasive algal biomasses, supporting an innovative strategy for conserving biodiversity as an alternative to unrealistic campaigns for the eradication of invasive pest