Measuring healthy ageing: current and future tools.

Human ageing is a complex, multifactorial process characterised by physiological damage, increased risk of age-related diseases and inevitable functional deterioration. As the population of the world grows older, placing significant strain on social and healthcare resources, there is a growing need to identify reliable and easy-to-employ markers of healthy ageing for early detection of ageing trajectories and disease risk. Such markers would allow for the targeted implementation of strategies or treatments that can lessen suffering, disability, and dependence in old age. In this review, we summarise the healthy ageing scores reported in the literature, with a focus on the past 5 years, and compare and contrast the variables employed. The use of approaches to determine biological age, molecular biomarkers, ageing trajectories, and multi-omics ageing scores are reviewed. We conclude that the ideal healthy ageing score is multisystemic and able to encompass all of the potential alterations associated with ageing. It should also be longitudinal and able to accurately predict ageing complications at an early stage in order to maximize the chances of successful early intervention.

Mixed circuit training acutely reduces arterial stiffness in patients with chronic stroke: a crossover randomized controlled trial.

PURPOSE: Investigate whether a single bout of mixed circuit training (MCT) can elicit changes in arterial stiffness in patients with chronic stroke. Second, to assess the between-day reproducibility of post-MCT arterial stiffness measurements. METHODS: Seven participants (58 ± 12 years) performed a non-exercise control session (CTL) and two bouts of MCT on separate days in a randomized counterbalanced order. The MCT involved 3 sets of 15 repetition maximum for 10 exercises, with each set separated by 45-s of walking. Brachial-radial pulse wave velocity (br-PWV), radial artery compliance (AC) and reflection index (RI1,2) were assessed 10 min before and 60 min after CTL and MCT. Ambulatory arterial stiffness index (AASI) was calculated from 24-h recovery ambulatory blood pressure monitoring. RESULTS: Compared to CTL, after 60 min of recovery from the 1st and 2nd bouts of MCT, lower values were observed for br-PWV (mean diff = - 3.9 and - 3.7 m/s, respectively, P < 0.01; ICC2,1 = 0.75) and RI1,2 (mean diff = - 16.1 and - 16.0%, respectively, P < 0.05; ICC2,1 = 0.83) concomitant with higher AC (mean diff = 1.2 and 1.0 × 10-6 cm5/dyna, respectively, P < 0.01; ICC2,1 = 0.40). The 24-h AASI was reduced after bouts of MCT vs. CTL (1st and 2nd bouts of MCT vs. CTL: mean diff = - 0.32 and - 0.29 units, respectively, P < 0.001; ICC2,1 = 0.64). CONCLUSION: A single bout of MCT reduces arterial stiffness during laboratory (60 min) and ambulatory (24 h) recovery phases in patients with chronic stroke with moderate-to-high reproducibility. TRIAL REGISTRATION: Ensaiosclinicos.gov.br identifier RBR-5dn5zd.

Machine learning approach for automatic recognition of tomato-pollinating bees based on their buzzing-sounds.

Bee-mediated pollination greatly increases the size and weight of tomato fruits. Therefore, distinguishing between the local set of bees-those that are efficient pollinators-is essential to improve the economic returns for farmers. To achieve this, it is important to know the identity of the visiting bees. Nevertheless, the traditional taxonomic identification of bees is not an easy task, requiring the participation of experts and the use of specialized equipment. Due to these limitations, the development and implementation of new technologies for the automatic recognition of bees become relevant. Hence, we aim to verify the capacity of Machine Learning (ML) algorithms in recognizing the taxonomic identity of visiting bees to tomato flowers based on the characteristics of their buzzing sounds. We compared the performance of the ML algorithms combined with the Mel Frequency Cepstral Coefficients (MFCC) and with classifications based solely on the fundamental frequency, leading to a direct comparison between the two approaches. In fact, some classifiers powered by the MFCC-especially the SVM-achieved better performance compared to the randomized and sound frequency-based trials. Moreover, the buzzing sounds produced during sonication were more relevant for the taxonomic recognition of bee species than analysis based on flight sounds alone. On the other hand, the ML classifiers performed better in recognizing bees genera based on flight sounds. Despite that, the maximum accuracy obtained here (73.39% by SVM) is still low compared to ML standards. Further studies analyzing larger recording samples, and applying unsupervised learning systems may yield better classification performance. Therefore, ML techniques could be used to automate the taxonomic recognition of flower-visiting bees of the cultivated tomato and other buzz-pollinated crops. This would be an interesting option for farmers and other professionals who have no experience in bee taxonomy but are interested in improving crop yields by increasing pollination.

Biocontamination and diversity of epilithic bacteria and fungi colonising outdoor stone and mortar sculptures.

Microbial communities colonising outdoor sculptures form intricate and dynamic ecosystems, which can accelerate the deterioration processes of the artworks and pose challenges to their conservation. In this study, the bacterial and fungal communities colonising the surfaces of five contemporary outdoor sculptures were characterised by high-throughput sequencing. The sculptures, made of marble, granite, Ançã limestone and mortar, are in urban parks and squares in the district of Porto, Portugal. The analysis of the microbial populations revealed great taxonomic diversity and species richness, including in well-preserved sculptures showing few visible traces of contamination. Proteobacteria, namely the genera Pseudomonas and Sphingomonas, were the core taxa common to all the sculptures, while Massilia and Aureobasidium were dominant only in granite. An abundance of pigment-producing microorganisms, such as Deinococcus, Methylobacterium, Rhodotorula and Sporobolomyces, was also found in granite. These are relevant taxonomic groups that can negatively impact stone and mortar artworks. The study was complemented with colourimetric analyses and bioluminescence assays to measure the adenosine triphosphate (ATP) content of samples collected from specific contaminated areas of the sculptures. The characterisation of the microbiomes of sculptures can provide further knowledge on the deterioration risks of this type of artwork in the region and help outline future targeted conservation strategies. KEY POINTS: • Rich and abundant microbiomes expose sculptures' vulnerability to deterioration. • Well-preserved sculptures are at risk of deterioration by pigment-producing taxa. • ATP and colourimetry quickly identified the most relevant contaminated areas.

Higher Cortical Facilitation and Serum BDNF Are Associated with Increased Sensitivity to Heat Pain and Reduced Endogenous Pain Inhibition in Healthy Males.

Background: Although the brain-derived neurotrophic factor (BDNF) has been intensively investigated in animal models of chronic pain, its role in human pain processing is less understood. Objective: To study the neurophysiology of BDNF modulation on acute experimental pain, we performed a cross-sectional study. Methods: We recruited 20 healthy male volunteers (19-40 years old) and assessed their serum BDNF levels, quantitative sensory testing, and cortical excitability parameters using transcranial magnetic stimulation. Results: Linear regression models demonstrated that the BDNF (ß = -5.245, P = 0.034) and intracortical facilitation (ß = -3.311, P = 0.034) were inversely correlated with heat pain threshold (adjusted R2 = 44.26). The BDNF (ß = -3.719, P ≤ 0.001) was also inversely correlated with conditioned pain modulation (adjusted R2 = 56.8). Conclusions: Our findings indicate that higher serum BDNF and intracortical facilitation of the primary motor cortex are associated with increased sensitivity to heat pain and high serum BDNF with reduced pain inhibition during noxious heterotopic stimulation.

A diffusion-based connectivity map of the GPi for optimised stereotactic targeting in DBS.

BACKGROUND: The GPi (globus pallidus internus) is an important target nucleus for Deep Brain Stimulation (DBS) in medically refractory movement disorders, in particular dystonia and Parkinson's disease. Beneficial clinical outcome critically depends on precise electrode localization. Recent evidence indicates that not only neurons, but also axonal fibre tracts contribute to promoting the clinical effect. Thus, stereotactic planning should, in the future, also take the individual course of fibre tracts into account. OBJECTIVE: The aim of this project is to explore the GPi connectivity profile and provide a connectivity-based parcellation of the GPi. METHODS: Diffusion MRI sequences were performed in sixteen healthy, right-handed subjects. Connectivity-based parcellation of the GPi was performed applying two independent methods: 1) a hypothesis-driven, seed-to-target approach based on anatomic priors set as connectivity targets and 2) a purely data-driven approach based on k-means clustering of the GPi. RESULTS: Applying the hypothesis-driven approach, we obtained five major parcellation clusters, displaying connectivity to the prefrontal cortex, the brainstem, the GPe (globus pallidus externus), the putamen and the thalamus. Parcellation clusters obtained by both methods were similar in their connectivity profile. With the data-driven approach, we obtained three major parcellation clusters. Inter-individual variability was comparable with results obtained in thalamic parcellation. CONCLUSION: The three parcellation clusters obtained by the purely data-driven method might reflect GPi subdivision into a sensorimotor, associative and limbic portion. Clinical and physiological studies indicate greatest clinical DBS benefit for electrodes placed in the postero-ventro-lateral GPi, the region displaying connectivity to the thalamus in our study and generally attributed to the sensorimotor system. Clinical studies relating DBS electrode positions to our GPi connectivity map would be needed to complement our findings.

The role of the pallidothalamic fibre tracts in deep brain stimulation for dystonia: A diffusion MRI tractography study.

BACKGROUND: Deep Brain Stimulation (DBS) of the Globus pallidus internus (GPi) is gold standard treatment in medically refractory dystonia. Recent evidence indicates that stimulation effects are also due to axonal modulation and affection of a fibre network. For the GPi, the pallidothalamic tracts are known to be the major motor efferent pathways. The aim of this study is to explore the anatomic vicinity of these tracts and DBS electrodes in dystonia applying diffusion tractography. METHODS: Diffusion MRI was acquired in ten patients presenting for DBS for dystonia. We applied both a conventionally used probabilistic tractography algorithm (FSL) as well as a probabilistic streamline tracking approach, based on constrained spherical deconvolution and particle filtering with anatomic priors, to the datasets. DBS electrodes were coregistered to the diffusion datasets. RESULTS: We were able to delineate the pallidothalamic tracts in all patients. Using the streamline approach, we were able to distinguish between the two sub-components of the tracts, the ansa lenticularis and the fasciculus lenticularis. Clinically efficient DBS electrodes displayed a close anatomic vicinity pathway of the pallidothalamic tracts, and their course was consistent with previous tracer labelling studies. Although we present only anatomic data, we interpret these findings as evidence of the possible involvement of fibre tracts to the clinical effect in DBS. Electrophysiological intraoperative recordings would be needed to complement our findings. In the future, a clear and individual delineation of the pallidothalamic tracts could optimize the stereotactic process of optimal electrode localization. Hum Brain Mapp 38:1224-1232, 2017. © 2016 Wiley Periodicals, Inc.

Immunoproteomics of Plasmodium falciparum-infected red blood cell membrane fractions.

BACKGROUND: The surface of infected red blood cells (iRBCs) has been widely investigated because of the molecular complexity and pathogenesis mechanisms involved. Asymptomatic individuals are important in the field because they can perpetuate transmission as natural reservoirs and present a challenge for diagnosing malaria because of their low levels of circulating parasites. Recent studies of iRBC antibody recognition have shown that responses are quantitatively similar in symptomatic and asymptomatic infections, but no studies have characterised the plasmodial proteins targeted by this response. OBJECTIVES: Our main objective was to identify Plasmodium falciparum proteins associated with iRBC ghosts recognised by antibodies in the sera of symptomatic and asymptomatic individuals in the Brazilian Amazon. METHODS: We collected symptomatic and asymptomatic sera from patients residing in the Brazilian Amazon and P. falciparum iRBC ghosts to identify the proteins involved in natural antibody recognition by 2D-electrophoresis, western blotting, and high- resolution mass spectrometry. FINDINGS: 2D gel-based immunoproteome analysis using symptomatic and asymptomatic sera identified 11 proteins with at least one unique peptide, such as chaperones HSP70-1 and HSP70-x, which likely are components of the secretion machinery/PTEX translocon. PfEMP1 is involved in antigenic variation in symptomatic infections and we found putative membrane proteins whose functions are unknown. MAIN FINDINGS: Our results suggest a potential role of old and new proteins, such as antigenic variation proteins, iRBC remodelling, and membrane proteins, with no assigned functions related to the immune response against P. falciparum, providing insights into the pathogenesis, erythrocyte remodelling, and secretion machinery important for alternative diagnosis and/or malaria therapy.

The urogenital microbiome in chronic kidney disease patients on peritoneal dialysis.

INTRODUCTION AND OBJECTIVES: Diabetes, dyslipidemia, older age, gender, urinary tract infections, and recent antibiotic intake have been associated with a decrease in the urobiome richness and other fluctuations in this microbiome. Gut and blood microbiome have been reported to be altered in patients with chronic kidney disease (CKD), and specifically in peritoneal dialysis (PD) patients. Still, there are currently no studies describing the urogenital microbiome in CKD-PD patients. In this study we characterized the urobiome profile in 46 PD patients and analyzed its clinical and inflammatory parameters. MATERIALS AND METHODS: Mid-stream urine, fecal and blood samples were collected from 46 patients undergoing PD at Centro Hospitalar Universitário de São João (CHUSJ) in Porto, Portugal. Exclusion criteria were age under 18 years old, inability to give informed consent, history of infection in the last three months, and antibiotic intake in the last three months. The microbiome communities were analyzed by amplification and sequencing of the V3-V4 region of the bacterial 16S rRNA gene. Correlations with the patients' clinical data and inflammatory profile were performed. RESULTS: CKD-PD patients presented a unique urobiome profile dominated by Bacillota, Actinomycetota and Pseudomonadota and characterized by a lower Shannon diversity than fecal and blood microbiome. The taxonomic profiles of urogenital samples were organized in multiple subtypes dominated by populations of Lactobacillus, Staphylococcus, Streptococcus, Gardnerella, Prevotella, Escherichia-Shigella, being similar to other non-PD-CKD patients. Gender, sCD14, residual diuresis and history of peritonitis were significantly associated to variations in the urobiome. Although not reaching statistical significance, diabetes and the time on PD also showed association with particular taxonomic groups. Depletion of Gardnerella, Staphylococcus, Corynebacterium, Lactobacillus or Dermabacter populations correlated with CKD-PD patients with history of diabetes, history of peritonitis and altered levels of sCD14. CONCLUSIONS: Our results highlight urogenital microbiome as a potential partner and/or marker in the overall health state of CKD-PD patients.

Influence of exercise order on the number of repetitions, oxygen uptake, and rate of perceived exertion during strength training in younger and older women.

The study investigated the effect of resistance exercise order on the number of repetitions, oxygen uptake (VO(2max)) and rate perceived exertion (RPE) in younger (YG: N = 10; 22 ± 2 years; VO(2peak) 42.2 ± 2.9 ml·kg(-1)·min(-1)) and older (EG: N = 8; 69 ± 7 years; VO(2peak)22.7 ± 2.5 ml·kg(-1)·min(-1)) women. The subjects performed 3 sets of each exercise until fatigue using 10 repetition maximum in 2 sequences of opposite order: (a) sequence A (SEQA): bench press (BP), machine shoulder press (SP), pulley triceps extension (TE); (b) sequence B (SEQB): TE-SP-BP. The VO(2) was assessed during the exercises, rest intervals, and 20 minutes after sequences (postexercise oxygen consumption [EPOC]). The number of repetitions decreased in both groups (p < 0.05) throughout sets. No difference (p > 0.22) was found between sequences for total VO(2) (exercise sequences + EPOC) in YG (SEQA = 25.41 ± 6.51 L vs. SEQB = 24.81 ± 4.08 L) and EG (SEQA = 26.45 ± 5.24 L vs. SEQB = 26.91 ± 4.62 L). In both groups, the VO(2) within the sequences was higher during BP when it was placed at the end of SEQB, the same occurring with TE in SEQA (p < 0.05). The VO(2) within sequences and RPE were higher in SEQB compared with SEQA (p < 0.05) in EG but not in YG. In conclusion, the exercise order did not affect total VO(2). The VO(2) within a given sequence was always lower when an exercise was performed first as compared with last regardless of the exercise order. Accumulated fatigue reflected by the VO(2) within sequences and RPE was influenced by the exercise order only in EG, suggesting that to prevent early fatigue, resistance training sessions in this group should preferably progress from large toward small-muscle group exercises.

Acute effect of proprioceptive neuromuscular facilitation stretching on the number of repetitions performed during a multiple set resistance exercise protocol.

It has been proposed that fatigue during strength exercise is negatively influenced by prior proprioceptive neuromuscular facilitation (PNF) stretching. However, it is possible that the effects of PNF on muscle endurance are affected by stretching duration. This study investigated the influence of PNF on the number of repetitions of the leg curl exercise performed with multiple sets and submaximal load. Nineteen men (age 25 ± 1 years, weight 75.8 ± 4.2 kg, height 178.1 ± 3.8 cm, 10-repetition maximum [RM] 78.3 ± 6.9 kg) performed 4 sets of leg curl with 10RM load with and without previous PNF (3 sets of hip flexion either with knees extended or flexed, duration ~2.5 minutes). The total number of repetitions decreased along sets in both situations (38.6% in control and 41.0% in PNF sessions, p < 0.001). However, no difference between control and PNF was detected for the number of repetitions in each set (first set, p = 0.330; second set, p = 0.072; third set, p = 0.061; fourth set, p = 0.150). In conclusion, the number of repetitions performed in multiple sets of the leg curl was not decreased by prior PNF stretching. Therefore, it appears that a moderate level of PNF could be used before resistance exercise with a minimal negative effect.

Corrigendum: In a zebrafish biomedical model of human Allan-Herndon-Dudley syndrome impaired MTH signaling leads to decreased neural cell diversity.

[This corrects the article DOI: 10.3389/fendo.2023.1157685.].

In a zebrafish biomedical model of human Allan-Herndon-Dudley syndrome impaired MTH signaling leads to decreased neural cell diversity.

Background: Maternally derived thyroid hormone (T3) is a fundamental factor for vertebrate neurodevelopment. In humans, mutations on the thyroid hormones (TH) exclusive transporter monocarboxylic acid transporter 8 (MCT8) lead to the Allan-Herndon-Dudley syndrome (AHDS). Patients with AHDS present severe underdevelopment of the central nervous system, with profound cognitive and locomotor consequences. Functional impairment of zebrafish T3 exclusive membrane transporter Mct8 phenocopies many symptoms observed in patients with AHDS, thus providing an outstanding animal model to study this human condition. In addition, it was previously shown in the zebrafish mct8 KD model that maternal T3 (MTH) acts as an integrator of different key developmental pathways during zebrafish development. Methods: Using a zebrafish Mct8 knockdown model, with consequent inhibition of maternal thyroid hormones (MTH) uptake to the target cells, we analyzed genes modulated by MTH by qPCR in a temporal series from the start of segmentation through hatching. Survival (TUNEL) and proliferation (PH3) of neural progenitor cells (dla, her2) were determined, and the cellular distribution of neural MTH-target genes in the spinal cord during development was characterized. In addition, in-vivo live imaging was performed to access NOTCH overexpression action on cell division in this AHDS model. We determined the developmental time window when MTH is required for appropriate CNS development in the zebrafish; MTH is not involved in neuroectoderm specification but is fundamental in the early stages of neurogenesis by promoting the maintenance of specific neural progenitor populations. MTH signaling is required for developing different neural cell types and maintaining spinal cord cytoarchitecture, and modulation of NOTCH signaling in a non-autonomous cell manner is involved in this process. Discussion: The findings show that MTH allows the enrichment of neural progenitor pools, regulating the cell diversity output observed by the end of embryogenesis and that Mct8 impairment restricts CNS development. This work contributes to the understanding of the cellular mechanisms underlying human AHDS.

Development of a Novel Phagomagnetic-Assisted Isothermal DNA Amplification System for Endpoint Electrochemical Detection of Listeria monocytogenes.

The hitherto implemented Listeria monocytogenes detection techniques are cumbersome or require expensive non-portable instrumentation, hindering their transposition into on-time surveillance systems. The current work proposes a novel integrated system resorting to loop-mediated isothermal amplification (LAMP), assisted by a bacteriophage P100-magnetic platform, coupled to an endpoint electrochemical technique, towards L. monocytogenes expeditious detection. Molybdophosphate-based optimization of the bacterial phagomagnetic separation protocol allowed the determination of the optimal parameters for its execution (pH 7, 25 °C, 32 µg of magnetic particles; 60.6% of specific capture efficiency). The novel LAMP method targeting prfA was highly specific, accomplishing 100% inclusivity (for 61 L. monocytogenes strains) and 100% exclusivity (towards 42 non-target Gram-positive and Gram-negative bacteria). As a proof-of-concept, the developed scheme was successfully validated in pasteurized milk spiked with L. monocytogenes. The phagomagnetic-based approach succeeded in the selective bacterial capture and ensuing lysis, triggering Listeria DNA leakage, which was efficiently LAMP amplified. Methylene blue-based electrochemical detection of LAMP amplicons was accomplished in 20 min with remarkable analytical sensitivity (1 CFU mL-1). Hence, the combined system presented an outstanding performance and robustness, providing a 2.5 h-swift, portable, cost-efficient detection scheme for decentralized on-field application.

Reprogramming iPSCs to study age-related diseases: Models, therapeutics, and clinical trials.

The unprecedented rise in life expectancy observed in the last decades is leading to a global increase in the ageing population, and age-associated diseases became an increasing societal, economic, and medical burden. This has boosted major efforts in the scientific and medical research communities to develop and improve therapies to delay ageing and age-associated functional decline and diseases, and to expand health span. The establishment of induced pluripotent stem cells (iPSCs) by reprogramming human somatic cells has revolutionised the modelling and understanding of human diseases. iPSCs have a major advantage relative to other human pluripotent stem cells as their obtention does not require the destruction of embryos like embryonic stem cells do, and do not have a limited proliferation or differentiation potential as adult stem cells. Besides, iPSCs can be generated from somatic cells from healthy individuals or patients, which makes iPSC technology a promising approach to model and decipher the mechanisms underlying the ageing process and age-associated diseases, study drug effects, and develop new therapeutic approaches. This review discusses the advances made in the last decade using iPSC technology to study the most common age-associated diseases, including age-related macular degeneration (AMD), neurodegenerative and cardiovascular diseases, brain stroke, cancer, diabetes, and osteoarthritis.

Automatic acoustic recognition of pollinating bee species can be highly improved by Deep Learning models accompanied by pre-training and strong data augmentation.

Introduction: Bees capable of performing floral sonication (or buzz-pollination) are among the most effective pollinators of blueberries. However, the quality of pollination provided varies greatly among species visiting the flowers. Consequently, the correct identification of flower visitors becomes indispensable to distinguishing the most efficient pollinators of blueberry. However, taxonomic identification normally depends on microscopic characteristics and the active participation of experts in the decision-making process. Moreover, the many species of bees (20,507 worldwide) and other insects are a challenge for a decreasing number of insect taxonomists. To overcome the limitations of traditional taxonomy, automatic classification systems of insects based on Machine-Learning (ML) have been raised for detecting and distinguishing a wide variety of bioacoustic signals, including bee buzzing sounds. Despite that, classical ML algorithms fed by spectrogram-type data only reached marginal performance for bee ID recognition. On the other hand, emerging systems from Deep Learning (DL), especially Convolutional Neural Networks (CNNs), have provided a substantial boost to classification performance in other audio domains, but have yet to be tested for acoustic bee species recognition tasks. Therefore, we aimed to automatically identify blueberry pollinating bee species based on characteristics of their buzzing sounds using DL algorithms. Methods: We designed CNN models combined with Log Mel-Spectrogram representations and strong data augmentation and compared their performance at recognizing blueberry pollinating bee species with the current state-of-the-art models for automatic recognition of bee species. Results and Discussion: We found that CNN models performed better at assigning bee buzzing sounds to their respective taxa than expected by chance. However, CNN models were highly dependent on acoustic data pre-training and data augmentation to outperform classical ML classifiers in recognizing bee buzzing sounds. Under these conditions, the CNN models could lead to automating the taxonomic recognition of flower-visiting bees of blueberry crops. However, there is still room to improve the performance of CNN models by focusing on recording samples for poorly represented bee species. Automatic acoustic recognition associated with the degree of efficiency of a bee species to pollinate a particular crop would result in a comprehensive and powerful tool for recognizing those that best pollinate and increase fruit yields.

How can we modulate aging through nutrition and physical exercise? An epigenetic approach.

The World Health Organization predicts that by 2050, 2.1 billion people worldwide will be over 60 years old, a drastic increase from only 1 billion in 2019. Considering these numbers, strategies to ensure an extended "healthspan" or healthy longevity are urgently needed. The present study approaches the promotion of healthspan from an epigenetic perspective. Epigenetic phenomena are modifiable in response to an individual's environmental exposures, and therefore link an individual's environment to their gene expression pattern. Epigenetic studies demonstrate that aging is associated with decondensation of the chromatin, leading to an altered heterochromatin structure, which promotes the accumulation of errors. In this review, we describe how aging impacts epigenetics and how nutrition and physical exercise can positively impact the aging process, from an epigenetic point of view. Canonical histones are replaced by histone variants, concomitant with an increase in histone post-translational modifications. A slight increase in DNA methylation at promoters has been observed, which represses transcription of previously active genes, in parallel with global genome hypomethylation. Aging is also associated with deregulation of gene expression - usually provided by non-coding RNAs - leading to both the repression of previously transcribed genes and to the transcription of previously repressed genes. Age-associated epigenetic events are less common in individuals with a healthy lifestyle, including balanced nutrition, caloric restriction and physical exercise. Healthy aging is associated with more tightly condensed chromatin, fewer PTMs and greater regulation by ncRNAs.

Purification, biochemical characterization and antifungal activity of a new lipid transfer protein (LTP) from Coffea canephora seeds with α-amylase inhibitor properties.

BACKGROUND: A growing number of cysteine-rich antimicrobial peptides (AMPs) have been isolated from plants and particularly from seeds. It has become increasingly clear that these peptides, which include lipid transfer proteins (LTPs), play an important role in the protection of plants against microbial infection. METHODS: Peptides from Coffea canephora seeds were extracted in Tris-HCl buffer (pH 8.0), and chromatographic purification of LTP was performed by DEAE and reverse-phase HPLC. The purified peptide was submitted to amino acid sequence, antimicrobial activity and mammalian α-amylase inhibitory analyses. RESULTS: The purified peptide of 9kDa had homology to LTPs isolated from different plants. Bidimensional electrophoresis of the 9kDa band showed the presence of two isoforms with pIs of 8.0 and 8.5. Cc-LTP(1) exhibited strong antifungal activity, against Candida albicans, and also promoted morphological changes including the formation of pseudohyphae on Candida tropicalis, as revealed by electron micrograph. Our results show that Cc-LTP(1) interfered in a dose-dependent manner with glucose-stimulated, H(+)-ATPase-dependent acidification of yeast medium and that the peptide permeabilized yeast plasma membranes to the dye SYTOX green, as verified by fluorescence microscopy. Interestingly, we also showed for the first time that the well characterized LTP(1) family, represented here by Cc-LTP(1), was also able to inhibit mammalian α-amylase activity in vitro. CONCLUSIONS AND GENERAL SIGNIFICANCE: In this work we purified, characterized and evaluated the in vitro effect on yeast of a new peptide from coffee, named Cc-LPT1, which we also showed, for the first time, the ability to inhibit mammalian α-amylase activity.

Four stanniocalcin genes in teleost fish: structure, phylogenetic analysis, tissue distribution and expression during hypercalcemic challenge.

Stanniocalcin (STC), first isolated from the corpuscles of Stannius (CS) of teleost fishes and a systemic regulator of mineral metabolism, is present in all vertebrates as two isoforms, STC1 and STC2, encoded by separate genes. Here we show that the genome of Tetraodon nigroviridis, and other teleosts, possess duplicate genes for each STC isoform, designated stc1-a and -b, and stc2-a and -b. Stc1-a was cloned from CS, stc2-a from muscle and the two novel cDNAs, stc1-b and stc2-b, from brain. However, stc2-b was isolated as a conjoined (read-through) transcript with bod1 (bi-orientation defective 1, or FAM44B), and two additional alternative conjoined transcripts were also isolated. The predicted STC products shared the typical vertebrate 10 conserved cysteine residues and N-linked glycosylation motifs, in addition to specific features. Gene structure was generally conserved with four exons and three introns with the exception of stc1-a which gained an extra intron in exon three, originating one extra exon. Gene order and synteny is also maintained across vertebrates and the cpeb4 gene identified in the homologue region of the chordate Ciona was linked to vertebrate stc2 but not stc1. Immunohistochemistry in different species revealed that STC1-A was found only in CS and in a few cells in kidney. STC1-B had a restricted expression and was more prominent in the gills. STC2-A was detected in a variety of tissues, including pituitary, with most abundant immunoreaction in kidney cells and gill rakers and the CS was negative. Expression of stc1-a in CS of Tetraodon was 15-fold (p<0.05) up-regulated 2 h after transfer from 2.9 mM Ca(2+) to 10 mM Ca(2+) water and down-regulated after 12 hours to 11-fold lower than 2.9 mM Ca(2+) fish (p<0.05). With the exception of stc1-a in CS, low expression levels and high individual variation were generally found for the expression of stc transcripts in kidney and gills, with no statistically significant changes in response to the hypercalcemic shock. In conclusion, both stc1 and stc2 genes are represented by paralogues in teleosts genomes and the analysis performed suggests that only stc1-a in the CS is involved in extracellular calcium regulation. The widespread distribution of stcs in fish tissues supports pleiotropic roles.