Exploring the biocontrol potential of fungal endophytes from an Andean Colombian Paramo ecosystem

We studied the diversity and biocontrol potential of 100 fungal endophytes isolated from Espeletia spp., endemic plant species from the Paramo in the Andean mountain range. Our sample was genotypically highly diverse at all ITS similarity levels. The antagonistic properties of these isolates were tested against common crop pathogens in Colombia, including Pectobacterium carotovorum, Ralstonia solanacearum, Pseudomonas syringae, Xanthomonas campestris, Rhizoctonia solani, Botrytis cinerea, Fusarium oxysporum, and Phytophthora infestans. All endophytic isolates were able to significantly inhibit the growth of at least one of the plant pathogens tested (P?<?0.05). Three main types of endophyte/pathogen interactions were observed. However, only those endophytes that produced an evident inhibition halo were further studied using their crude extracts to confirm that the inhibitory effect was due to the production of endophytic bioactive metabolites. From these experiments, nine promising isolates were selected for co-inoculation tests with R. solani in tomato plants. The isolates identified as Aureobasidium pullulans and Paraconiothyrium sporulosum not only protected the plants against this pathogen but also allowed them to exhibit similar growth and development as the uninoculated control. This work explores new alternatives for disease management without the application of chemical pesticides.     

Leaf functional traits of Neotropical savanna trees in relation to seasonal water deficit

The seasonal savannas (cerrados) of Central Brazil are characterized by a large diversity of evergreen and deciduous trees, which do not show a clear differentiation in terms of active rooting depth. Irrespective of the depth of the root system, expansion of new foliage in deciduous species occurs at the end of the dry season. In this study, we examined a suite of leaf traits related to C assimilation, water and nutrients (N, P) in five deciduous and six evergreen trees that were among the dominant families of cerrado vegetation. Maximum CO₂ assimilation on a mass basis (A_mass) was significantly correlated with leaf N and P, and specific leaf area (SLA; leaf area per unit of leaf mass). The highest leaf concentrations of both nutrients were measured in the newly mature leaves of deciduous species at the end of the dry period. The differences in terms of leaf N and P between evergreen and deciduous species decreased during the wet season. Deciduous species also invested less in the production of non-photosynthetic leaf tissues and produced leaves with higher SLA and maintained higher water use efficiency. Thus, deciduous species compensated for their shorter leaf payback period by maintaining higher potential payback capacity (higher values of A_mass) and lower leaf construction costs (higher SLA). Their short leafless period and the capacity to flush by the end of the dry season may also contribute to offset the longer payback period of evergreen species, although it may involve the higher cost of maintaining a deep-root system or a tight control of plant water balance in the shallow-rooted ones.     

Engineering of Saccharomyces cerevisiae for efficient anaerobic alcoholic fermentation of L-arabinose

For cost-effective and efficient ethanol production from lignocellulosic fractions of plant biomass, the conversion of not only major constituents, such as glucose and xylose, but also less predominant sugars, such as l-arabinose, is required. Wild-type strains of Saccharomyces cerevisiae, the organism used in industrial ethanol production, cannot ferment xylose and arabinose. Although metabolic and evolutionary engineering has enabled the efficient alcoholic fermentation of xylose under anaerobic conditions, the conversion of l-arabinose into ethanol by engineered S. cerevisiae strains has previously been demonstrated only under oxygen-limited conditions. This study reports the first case of fast and efficient anaerobic alcoholic fermentation of l-arabinose by an engineered S. cerevisiae strain. This fermentation was achieved by combining the expression of the structural genes for the l-arabinose utilization pathway of Lactobacillus plantarum, the overexpression of the S. cerevisiae genes encoding the enzymes of the nonoxidative pentose phosphate pathway, and extensive evolutionary engineering. The resulting S. cerevisiae strain exhibited high rates of arabinose consumption (0.70 g h(-1) g [dry weight](-1)) and ethanol production (0.29 g h(-1) g [dry weight](-1)) and a high ethanol yield (0.43 g g(-1)) during anaerobic growth on l-arabinose as the sole carbon source. In addition, efficient ethanol production from sugar mixtures containing glucose and arabinose, which is crucial for application in industrial ethanol production, was achieved.     

Pre-clinical and clinical significance of heparanase in Ewing's sarcoma

Heparanase is an endoglycosidase that specifically cleaves heparan sulphate side chains of heparan sulphate proteoglycans, activity that is strongly implicated in cell migration and invasion associated with tumour metastasis, angiogenesis and inflammation. Heparanase up-regulation was documented in an increasing number of human carcinomas, correlating with reduced post-operative survival rate and enhanced tumour angiogenesis. Expression and significance of heparanase in human sarcomas has not been so far reported. Here, we applied the Ewing's sarcoma cell line TC71 and demonstrated a potent inhibition of cell invasion in vitro and tumour xenograft growth in vivo upon treatment with a specific inhibitor of heparanase enzymatic activity (compound SST0001, non-anticoagulant N-acetylated, glycol split heparin). Next, we examined heparanase expression and cellular localization by immunostaining of a cohort of 69 patients diagnosed with Ewing's sarcoma. Heparanase staining was noted in all patients. Notably, heparanase staining intensity correlated with increased tumour size (P = 0.04) and with patients' age (P = 0.03), two prognostic factors associated with a worse outcome. Our study indicates that heparanase expression is induced in Ewing's sarcoma and associates with poor prognosis. Moreover, it encourages the inclusion of heparanase inhibitors (i.e. SST0001) in newly developed therapeutic modalities directed against Ewing's sarcoma and likely other malignancies.     

Striatal pre- and postsynaptic profile of adenosine A(2A) receptor antagonists

Striatal adenosine A(2A) receptors (A(2A)Rs) are highly expressed in medium spiny neurons (MSNs) of the indirect efferent pathway, where they heteromerize with dopamine D(2) receptors (D(2)Rs). A(2A)Rs are also localized presynaptically in cortico-striatal glutamatergic terminals contacting MSNs of the direct efferent pathway, where they heteromerize with adenosine A(1) receptors (A(1)Rs). It has been hypothesized that postsynaptic A(2A)R antagonists should be useful in Parkinson's disease, while presynaptic A(2A)R antagonists could be beneficial in dyskinetic disorders, such as Huntington's disease, obsessive-compulsive disorders and drug addiction. The aim or this work was to determine whether selective A(2A)R antagonists may be subdivided according to a preferential pre- versus postsynaptic mechanism of action. The potency at blocking the motor output and striatal glutamate release induced by cortical electrical stimulation and the potency at inducing locomotor activation were used as in vivo measures of pre- and postsynaptic activities, respectively. SCH-442416 and KW-6002 showed a significant preferential pre- and postsynaptic profile, respectively, while the other tested compounds (MSX-2, SCH-420814, ZM-241385 and SCH-58261) showed no clear preference. Radioligand-binding experiments were performed in cells expressing A(2A)R-D(2)R and A(1)R-A(2A)R heteromers to determine possible differences in the affinity of these compounds for different A(2A)R heteromers. Heteromerization played a key role in the presynaptic profile of SCH-442416, since it bound with much less affinity to A(2A)R when co-expressed with D(2)R than with A(1)R. KW-6002 showed the best relative affinity for A(2A)R co-expressed with D(2)R than co-expressed with A(1)R, which can at least partially explain the postsynaptic profile of this compound. Also, the in vitro pharmacological profile of MSX-2, SCH-420814, ZM-241385 and SCH-58261 was is in accordance with their mixed pre- and postsynaptic profile. On the basis of their preferential pre- versus postsynaptic actions, SCH-442416 and KW-6002 may be used as lead compounds to obtain more effective antidyskinetic and antiparkinsonian compounds, respectively.     

<Emphasis Type="Italic">Slc11a1</Emphasis> (<Emphasis Type="Italic">Nramp1</Emphasis>) alleles interact with acute inflammation loci to modulate wound-healing traits in mice

Lines of mice were obtained by selective breeding for maximum (AIRmax) or minimum (AIRmin) acute inflammation. They present distinct neutrophil influx and show frequency disequilibrium of the solute carrier family 11a member 1 (Slc11a1) alleles. This gene is involved in ion transport at the endosomes within macrophages and neutrophils, interfering in their activation. Homozygous AIRmax and AIRmin sublines for the Slc11a1 gene were produced to examine the interaction of this gene with the acute inflammatory loci. The present work investigated wound-healing traits in AIRmax and AIRmin mice, in F₁ and F₂ intercrosses, and in Slc11a1 sublines. Two-millimeter ear punches were made in the mice and hole closure was measured during 40 days. AIRmax mice demonstrated significant tissue repair while AIRmin mice did not. Significant differences between the responses of male and female mice were also observed. Wound-healing traits demonstrated a correlation with neutrophil influx in F₂ populations. AIRmax ^SS showed higher ear-wound closure than AIRmax ^RR mice, suggesting that the Slc11a1 S allele favored ear tissue repair. QTL analysis has detected two inflammatory loci modulating ear wound healing on chromosomes 1 and 14. These results suggest the involvement of the acute inflammation modifier QTL in the wound-healing phenotype.     

Interleukin-17 acts independently of TNF-alpha under arthritic conditions

The proinflammatory T cell cytokine IL-17 is a potent inducer of other cytokines such as IL-1 and TNF-alpha. The contribution of TNF in IL-17-induced joint inflammation is unclear. In this work we demonstrate using TNF-alpha-deficient mice that TNF-alpha is required in IL-17-induced joint pathology under naive conditions in vivo. However, overexpression of IL-17 aggravated K/BxN serum transfer arthritis to a similar degree in TNF-alpha-deficient mice and their wild-type counterparts, indicating that the TNF dependency of IL-17-induced pathology is lost under arthritic conditions. Also, during the course of the streptococcal cell wall-induced arthritis model, IL-17 was able to enhance inflammation and cartilage damage in the absence of TNF. Additional blocking of IL-1 during IL-17-enhanced streptococcal cell wall-induced arthritis did not reduce joint pathology in TNF-deficient mice, indicating that IL-1 is not responsible for this loss of TNF dependency. These data provide further understanding of the cytokine interplay during inflammation and demonstrate that, despite a strong TNF dependency under naive conditions, IL-17 acts independently of TNF under arthritic conditions.     

MDC1 maintains genomic stability by participating in the amplification of ATM-dependent DNA damage signals

MDC1 functions in checkpoint activation and DNA repair following DNA damage. To address the physiological role of MDC1, we disrupted the MDC1 gene in mice. MDC1-/- mice recapitulated many phenotypes of H2AX-/- mice, including growth retardation, male infertility, immune defects, chromosome instability, DNA repair defects, and radiation sensitivity. At the molecular level, H2AX, MDC1, and ATM form a positive feedback loop, with MDC1 directly mediating the interaction between H2AX and ATM. MDC1 binds phosphorylated H2AX through its BRCT domain and ATM through its FHA domain. Through these interactions, MDC1 accumulates activated ATM flanking the sites of DNA damage, facilitating further ATM-dependent phosphorylation of H2AX and the amplification of DNA damage signals. In the absence of MDC1, many downstream ATM signaling events are defective. These results suggest that MDC1, as a signal amplifier of the ATM pathway, is vital in controlling proper DNA damage response and maintaining genomic stability.     

Aging exacerbates negative remodeling and impairs endothelial regeneration after balloon injury

Many older patients, because of their high prevalence of coronary artery disease, are candidates for percutaneous coronary interventions (PCI), but the effects of vascular aging on restenosis after PCI are not yet well understood. Balloon injury to the right carotid artery was performed in adult and old rats. Vascular smooth muscle cell (VSMC) proliferation, apoptotic cell death, together with Akt induction, telomerase activity, p27kip1, and endothelial nitric oxide synthase (eNOS) expression was assessed in isolated arteries. Neointima hyperplasia and vascular remodeling along with endothelial cell regeneration were also measured after balloon injury. Arteries isolated from old rats exhibited a significant reduction of VSMC proliferation and an increase in apoptotic death after balloon injury when compared with adult rats. In the vascular wall of adult rats, balloon dilation induced Akt phosphorylation, and this was barely present in old rats. In arteries from old rats, Akt-modulated cell cycle check points like telomerase activity and p27kip1 expression were decreased and increased, respectively, compared with adults. After balloon injury, old rats showed a significant reduction of neointima formation and an increased vascular negative remodeling compared with adults. These results were coupled by a marked delay in endothelial regeneration in aged rats, partially mediated by a decreased eNOS expression and phosphorylation. Interestingly, chronic administration of L-arginine prevented negative remodeling and improved reendothelialization after balloon injury in aged animals. A decreased neointimal proliferation, an impaired endothelial regeneration, and an increase in vascular remodeling after balloon injury were observed in aged animals. The molecular mechanisms underlying these responses seem to be a reduced Akt and eNOS activity.     

Arrhythmias after tetralogy of fallot repair

Tetralogy of Fallot is the most common cyanotic congenital heart disease, with a good outcome after total surgical correction. In spite of a low perioperative mortality and a good quality of life, late sudden death remains a significant clinical problem, mainly related to episodes of sustained ventricular tachycardia and ventricular fibrillation. Fibro-fatty substitution around infundibular resection, intraventricular septal scar, and patchy myocardial fibrosis, may provide anatomical substrates of abnormal depolarization and repolarization causing reentrant ventricular arrhythmias. Several non-invasive indices based on classical examination such as ECG, signal-averaging ECG, and echocardiography have been proposed to identify patients at high risk of sudden death, with hopeful results. In the last years other more sophisticated invasive and non-invasive tools, such as heart rate variability, electroanatomic mapping and cardiac magnetic resonance added a relevant contribution to risk stratification. Even if each method per se is affected by some limitations, a comprehensive multifactorial clinical and investigative examination can provide an accurate risk evaluation for every patient.