Dermatomycosis in lower limbs of diabetic patients followed by podiatry consultation

BackgroundDiabetic patients are particularly susceptible to fungal infections due to modifications that occur in their immunological system. These modifications compromise natural defences, such as skin and nails, especially from lower limbs.AimsAssessing the presence of dermatomycosis in lower limbs of Portuguese diabetic patients followed on Podiatry consultation. Determination of possible predisposing factors and the most frequent fungal species associated with the cases are included in the study.MethodsA six-month prospective study was carried out in 163 diabetic patients with signs and symptoms of dermatomycosis followed by Podiatry at the Portuguese Diabetes Association in Lisbon. Samples from the skin and/or nails of the lower limbs were collected and demographic and clinical data of those patients were recorded.ResultsTrichophyton rubrum was the most frequently isolated dermatophyte (12.1%), followed by Trichophyton mentagrophytes (7.7%) and Trichophyton tonsurans (4.4%). Our study showed positive associations between type 2 diabetes and the presence of dermatomycosis in the studied population (p = 0.013); this association was also shown between the occurrence of dermatomycosis and the localization of the body lesion (p = 0.000). No other predisposing factor tested was positively associated with infection (p > 0.05).ConclusionsData on superficial fungal infections in diabetic patients are scarce in Portugal. This study provides information on the characterization of dermatomycosis in lower limbs of diabetic patients.     

Cystic fibrosis transmembrane conductance regulator contributes to reacidification of alkalinized lysosomes in RPE cells

The role of the cystic fibrosis transmembrane conductance regulator (CFTR) in lysosomal acidification has been difficult to determine. We demonstrate here that CFTR contributes more to the reacidification of lysosomes from an elevated pH than to baseline pH maintenance. Lysosomal alkalinization is increasingly recognized as a factor in diseases of accumulation, and we previously showed that cAMP reacidified alkalinized lysosomes in retinal pigmented epithelial (RPE) cells. As the influx of anions to electrically balance proton accumulation may enhance lysosomal acidification, the contribution of the cAMP-activated anion channel CFTR to lysosomal reacidification was probed. The antagonist CFTR(inh)-172 had little effect on baseline levels of lysosomal pH in cultured human RPE cells but substantially reduced the reacidification of compromised lysosomes by cAMP. Likewise, CFTR activators had a bigger impact on cells whose lysosomes had been alkalinized. Knockdown of CFTR with small interfering RNA had a larger effect on alkalinized lysosomes than on baseline levels. Inhibition of CFTR in isolated lysosomes altered pH. While CFTR and Lamp1 were colocalized, treatment with cAMP did not increase targeting of CFTR to the lysosome. The inhibition of CFTR slowed lysosomal degradation of photoreceptor outer segments while activation of CFTR enhanced their clearance from compromised lysosomes. Activation of CFTR acidified RPE lysosomes from the ABCA4(-/-) mouse model of recessive Stargardt's disease, whose lysosomes are considerably alkalinized. In summary, CFTR contributes more to reducing lysosomal pH from alkalinized levels than to maintaining baseline pH. Treatment to activate CFTR may thus be of benefit in disorders of accumulation associated with lysosomal alkalinization.     

Acidic Nanoparticles Are Trafficked to Lysosomes and Restore an Acidic Lysosomal pH and Degradative Function to Compromised ARPE-19 Cells

Lysosomal enzymes function optimally in acidic environments, and elevation of lysosomal pH can impede their ability to degrade material delivered to lysosomes through autophagy or phagocytosis. We hypothesize that abnormal lysosomal pH is a key aspect in diseases of accumulation and that restoring lysosomal pH will improve cell function. The propensity of nanoparticles to end up in the lysosome makes them an ideal method of delivering drugs to lysosomes. This study asked whether acidic nanoparticles could traffic to lysosomes, lower lysosomal pH and enhance lysosomal degradation by the cultured human retinal pigmented epithelial cell line ARPE-19. Acidic nanoparticles composed of poly (DL-lactide-co-glycolide) (PLGA) 502 H, PLGA 503 H and poly (DL-lactide) (PLA) colocalized to lysosomes of ARPE-19 cells within 60 min. PLGA 503 H and PLA lowered lysosomal pH in cells compromised by the alkalinizing agent chloroquine when measured 1 hr. after treatment, with acidification still observed 12 days later. PLA enhanced binding of Bodipy-pepstatin-A to the active site of cathepsin D in compromised cells. PLA also reduced the cellular levels of opsin and the lipofuscin-like autofluorescence associated with photoreceptor outer segments. These observations suggest the acidification produced by the nanoparticles was functionally effective. In summary, acid nanoparticles lead to a rapid and sustained lowering of lysosomal pH and improved degradative activity.     

SPECTROSCOPIC DETERMINATION OF THE TOXICITY,ABSORPTION, REDUCTION,AND TRANSLOCATION OF Cr(VI) IN TWO MAGNOLIOPSIDA SPECIES

Hexavalent chromium is a contaminant highly mobile in the environment that is toxic for plants at low concentrations. In this work, the physiological response of Convolvulus arvensis and Medicago truncatula plants to Cr(VI) treatments was compared. C. arvensis is a potential Cr hyperaccumulator well adapted to semiarid conditions that biotransform Cr(VI) to the less toxic Cr(III). M. truncatula is a model plant well adapted to semiarid conditions with a well studied genetic response to heavy metal stress. The results demonstrated that C. arvensis is more tolerant to Cr toxicity and has a higher Cr translocation to the leaves. The inductively coupled plasma optical emission spectroscopy results showed that C. arvensis plants treated with 10 mg Cr(VI) L^–1 accumulated 1512, 210, and 131 mg Cr kg^–1 in roots, stems, and leaves, respectively. While M. truncatula plants treated with the same Cr(VI) concentration accumulated 1081, 331, and 44 (mg Cr kg^–1) in roots, stems, and leaves, respectively. Enzymatic assays demonstrated that Cr(VI) decreased ascorbate peroxidase activity and increased catalase activity in M. truncatula, while an opposite response was found in C. arvensis. The x-ray absorption spectroscopy studies showed that both plant species reduced Cr(VI) to the less toxic Cr(III).     

Exploiting cross-reactivity to neutralize two different scorpion venoms with one single chain antibody fragment

We report the optimization of a family of human single chain antibody fragments (scFv) for neutralizing two scorpion venoms. The parental scFv 3F recognizes the main toxins of Centruroides noxius Hoffmann (Cn2) and Centruroides suffusus suffusus (Css2), albeit with low affinity. This scFv was subjected to independent processes of directed evolution to improve its recognition toward Cn2 (Riaño-Umbarila, L., Juárez-González, V. R., Olamendi-Portugal, T., Ortíz-León, M., Possani, L. D., and Becerril, B. (2005) FEBS J. 272, 2591–2601) and Css2 (this work). Each evolved variant showed strong cross-reactivity against several toxins, and was capable of neutralizing Cn2 and Css2. Furthermore, each variant neutralized the whole venoms of the above species. As far as we know, this is the first report of antibodies with such characteristics. Maturation processes revealed key residue changes to attain expression, stability, and affinity improvements as compared with the parental scFv. Combination of these changes resulted in the scFv LR, which is capable of rescuing mice from severe envenomation by 3 LD₅₀ of freshly prepared whole venom of C. noxius (7.5 μg/20 g of mouse) and C. suffusus (26.25 μg/20 g of mouse), with surviving rates between 90 and 100%. Our research is leading to the formulation of an antivenom consisting of a discrete number of human scFvs endowed with strong cross-reactivity and low immunogenicity.     

Structural basis of neutralization of the major toxic component from the scorpion Centruroides noxius Hoffmann by a human-derived single-chain antibody fragment

It has previously been reported that several single-chain antibody fragments of human origin (scFv) neutralize the effects of two different scorpion venoms through interactions with the primary toxins of Centruroides noxius Hoffmann (Cn2) and Centruroides suffusus suffusus (Css2). Here we present the crystal structure of the complex formed between one scFv (9004G) and the Cn2 toxin, determined in two crystal forms at 2.5 and 1.9 Å resolution. A 15-residue span of the toxin is recognized by the antibody through a cleft formed by residues from five of the complementarity-determining regions of the scFv. Analysis of the interface of the complex reveals three features. First, the epitope of toxin Cn2 overlaps with essential residues for the binding of β-toxins to its Na⁺ channel receptor site. Second, the putative recognition of Css2 involves mainly residues that are present in both Cn2 and Css2 toxins. Finally, the effect on the increase of affinity of previously reported key residues during the maturation process of different scFvs can be inferred from the structure. Taken together, these results provide the structural basis that explain the mechanism of the 9004G neutralizing activity and give insight into the process of directed evolution that gave rise to this family of neutralizing scFvs.     

Reciprocal neural influences upon the period length of the ERG circadian rhythm in the crayfish

Abstract

The relationship between left and right circadian oscillations in the ERG of the crayfishProcambarus bouvieri have been studied in intact animals and in animals with cerebral ganglion damage. Except in split‐brain preparations, the ERG oscillations of both sides were always in phase. Surgical bisection of the cerebral ganglion, also results in a shortening of the period length; this suggests that ERG circadian pacemakers normally receive a bilateral input of light stimulation. The neural connections between the eyestalks at the protocerebrum seem to have the most important role in the synchronization of the ERG oscillations of both sides.     

Accounting for multiple ecosystem services in a simulation of land‐use decisions: Does it reduce tropical deforestation?

Conversion of tropical forests is among the primary causes of global environmental change. The loss of their important environmental services has prompted calls to integrate ecosystem services (ES) in addition to socio‐economic objectives in decision‐making. To test the effect of accounting for both ES and socio‐economic objectives in land‐use decisions, we develop a new dynamic approach to model deforestation scenarios for tropical mountain forests. We integrate multi‐objective optimization of land allocation with an innovative approach to consider uncertainty spaces for each objective. These uncertainty spaces account for potential variability among decision‐makers, who may have different expectations about the future. When optimizing only socio‐economic objectives, the model continues the past trend in deforestation (1975–2015) in the projected land‐use allocation (2015–2070). Based on indicators for biomass production, carbon storage, climate and water regulation, and soil quality, we show that considering multiple ES in addition to the socio‐economic objectives has heterogeneous effects on land‐use allocation. It saves some natural forest if the natural forest share is below 38%, and can stop deforestation once the natural forest share drops below 10%. For landscapes with high shares of forest (38%–80% in our study), accounting for multiple ES under high uncertainty of their indicators may, however, accelerate deforestation. For such multifunctional landscapes, two main effects prevail: (a) accelerated expansion of diversified non‐natural areas to elevate the levels of the indicators and (b) increased landscape diversification to maintain multiple ES, reducing the proportion of natural forest. Only when accounting for vascular plant species richness as an explicit objective in the optimization, deforestation was consistently reduced. Aiming for multifunctional landscapes may therefore conflict with the aim of reducing deforestation, which we can quantify here for the first time. Our findings are relevant for identifying types of landscapes where this conflict may arise and to better align respective policies.     

The role of bioenergy for global deep decarbonization: CO2 removal or low‐carbon energy?

Bioenergy is expected to have a prominent role in limiting global greenhouse emissions to meet the climate change target of the Paris Agreement. Many studies identify negative emissions from bioenergy generation with carbon capture and storage (BECCS) as its key contribution, but assume that no other CO₂ removal technologies are available. We use a global integrated assessment model, TIAM‐UCL, to investigate the role of bioenergy within the global energy system when direct air capture and afforestation are available as cost‐competitive alternatives to BECCS. We find that the presence of other CO₂ removal technologies does not reduce the pressure on biomass resources but changes the use of bioenergy for climate mitigation. While we confirm that when available BECCS offers cheaper decarbonization pathways, we also find that its use delays the phase‐out of unabated fossil fuels in industry and transport. Furthermore, it displaces renewable electricity generation, potentially increasing the likelihood of missing the Paris Agreement target. We found that the most cost‐effective solution is to invest in a basket of CO₂ removal technologies. However, if these technologies rely on CCS, then urgent action is required to ramp up the necessary infrastructure. We conclude that a sustainable biomass supply is critical for decarbonizing the global energy system. Since only a few world regions carry the burden of producing the biomass resource and store CO₂ in geological storage, adequate international collaboration, policies and standards will be needed to realize this resource while avoiding undesired land‐use change.     

Optimization and evaluation of surface-enhanced laser-desorption/ionization time-of-flight mass spectrometry for protein profiling of cerebrospinal fluid

Cerebrospinal fluid (CSF) potentially carries an archive of peptides and small proteins relevant to pathological processes in the central nervous system (CNS) and surrounding brain tissue. Proteomics is especially well suited for the discovery of biomarkers of diagnostic potential in CSF for early diagnosis and discrimination of several neurodegenerative diseases. ProteinChip surface-enhanced laser-desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) is one such approach which offers a unique platform for high throughput profiling of peptides and small proteins in CSF. In this study, we evaluated methodologies for the retention of CSF proteins < 20 kDa in size, and identify a strategy for screening small proteins and peptides in CSF. ProteinChip array types, along with sample and binding buffer conditions, and matrices were investigated. By coupling the processing of arrays to a liquid handler reproducible and reliable profiles, with mean peak coefficients of variation < 20%, were achieved for intra- and inter-assays under selected conditions. Based on peak m/z we found a high degree of overlap between the tested array surfaces. The combination of CM10 and IMAC30 arrays was sufficient to represent between 80–90% of all assigned peaks when using either sinapinic acid or α-Cyano-4-hydroxycinnamic acid as the energy absorbing matrices. Moreover, arrays processed with SPA consistently showed better peak resolution and higher peak number across all surfaces within the measured mass range. We intend to use CM10 and IMAC30 arrays prepared in sinapinic acid as a fast and cost-effective approach to drive decisions on sample selection prior to more in-depth discovery of diagnostic biomarkers in CSF using alternative but complementary proteomic strategies.