The illusion of moral decline.

Anecdotal evidence indicates that people believe that morality is declining1,2. In a series of studies using both archival and original data (n = 12,492,983), we show that people in at least 60 nations around the world believe that morality is declining, that they have believed this for at least 70 years and that they attribute this decline both to the decreasing morality of individuals as they age and to the decreasing morality of successive generations. Next, we show that people's reports of the morality of their contemporaries have not declined over time, suggesting that the perception of moral decline is an illusion. Finally, we show how a simple mechanism based on two well-established psychological phenomena (biased exposure to information and biased memory for information) can produce an illusion of moral decline, and we report studies that confirm two of its predictions about the circumstances under which the perception of moral decline is attenuated, eliminated or reversed (that is, when respondents are asked about the morality of people they know well or people who lived before the respondent was born). Together, our studies show that the perception of moral decline is pervasive, perdurable, unfounded and easily produced. This illusion has implications for research on the misallocation of scarce resources3, the underuse of social support4 and social influence5.

Do conversations end when people want them to?

Do conversations end when people want them to? Surprisingly, behavioral science provides no answer to this fundamental question about the most ubiquitous of all human social activities. In two studies of 932 conversations, we asked conversants to report when they had wanted a conversation to end and to estimate when their partner (who was an intimate in Study 1 and a stranger in Study 2) had wanted it to end. Results showed that conversations almost never ended when both conversants wanted them to and rarely ended when even one conversant wanted them to and that the average discrepancy between desired and actual durations was roughly half the duration of the conversation. Conversants had little idea when their partners wanted to end and underestimated how discrepant their partners' desires were from their own. These studies suggest that ending conversations is a classic "coordination problem" that humans are unable to solve because doing so requires information that they normally keep from each other. As a result, most conversations appear to end when no one wants them to.

Chemical-Vapor-Deposition-Synthesized Two-Dimensional Non-Stoichiometric Copper Selenide (ß-Cu2-xSe) for Ultra-Fast Tetracycline Hydrochloride Degradation under Solar Light.

The high concentration of antibiotics in aquatic environments is a serious environmental issue. In response, researchers have explored photocatalytic degradation as a potential solution. Through chemical vapor deposition (CVD), we synthesized copper selenide (ß-Cu2-xSe) and found it an effective catalyst for degrading tetracycline hydrochloride (TC-HCl). The catalyst demonstrated an impressive degradation efficiency of approximately 98% and a reaction rate constant of 3.14 × 10-2 min-1. Its layered structure, which exposes reactive sites, contributes to excellent stability, interfacial charge transfer efficiency, and visible light absorption capacity. Our investigations confirmed that the principal active species produced by the catalyst comprises O2- radicals, which we verified through trapping experiments and electron paramagnetic resonance (EPR). We also verified the TC-HCl degradation mechanism using high-performance liquid chromatography-mass spectrometry (LC-MS). Our results provide valuable insights into developing the ß-Cu2-xSe catalyst using CVD and its potential applications in environmental remediation.

Ericoid shrub encroachment shifts aboveground-belowground linkages in three peatlands across Europe and Western Siberia.

In northern peatlands, reduction of Sphagnum dominance in favour of vascular vegetation is likely to influence biogeochemical processes. Such vegetation changes occur as the water table lowers and temperatures rise. To test which of these factors has a significant influence on peatland vegetation, we conducted a 3-year manipulative field experiment in Linje mire (northern Poland). We manipulated the peatland water table level (wet, intermediate and dry; on average the depth of the water table was 17.4, 21.2 and 25.3 cm respectively), and we used open-top chambers (OTCs) to create warmer conditions (on average increase of 1.2°C in OTC plots compared to control plots). Peat drying through water table lowering at this local scale had a larger effect than OTC warming treatment per see on Sphagnum mosses and vascular plants. In particular, ericoid shrubs increased with a lower water table level, while Sphagnum decreased. Microclimatic measurements at the plot scale indicated that both water-level and temperature, represented by heating degree days (HDDs), can have significant effects on the vegetation. In a large-scale complementary vegetation gradient survey replicated in three peatlands positioned along a transitional oceanic-continental and temperate-boreal (subarctic) gradient (France-Poland-Western Siberia), an increase in ericoid shrubs was marked by an increase in phenols in peat pore water, resulting from higher phenol concentrations in vascular plant biomass. Our results suggest a shift in functioning from a mineral-N-driven to a fungi-mediated organic-N nutrient acquisition with shrub encroachment. Both ericoid shrub encroachment and higher mean annual temperature in the three sites triggered greater vascular plant biomass and consequently the dominance of decomposers (especially fungi), which led to a feeding community dominated by nematodes. This contributed to lower enzymatic multifunctionality. Our findings illustrate mechanisms by which plants influence ecosystem responses to climate change, through their effect on microbial trophic interactions.

Divergent effects of oxytocin on "mind-reading" in healthy males.

The neuropeptide oxytocin (OT) has been associated with a broad range of human behaviors, particularly in the domain of social cognition, and is being discussed to play a role in a range of psychiatric disorders. Studies using the Reading The Mind In The Eyes Test (RMET) to investigate the role of OT in mental state recognition reported inconsistent outcomes. The present study applied a randomized, double-blind, cross-over design, and included measures of serum OT. Twenty healthy males received intranasal placebo or OT (24 IU) before performing the RMET. Frequentist and Bayesian analyses showed that contrary to previous studies (Domes et al., 2007; Radke & de Bruijn, 2015), individuals performed worse in the OT condition compared to the placebo condition (p = 0.023, Cohen's d = 0.55, 95% confidence interval [CI] [0.08, 1.02], BF10 = 6.93). OT effects did not depend on item characteristics (difficulty, valence, intensity, sex) of the RMET. Furthermore, OT serum levels did not change after intranasal OT administration. Given that similar study designs lead to heterogeneous outcomes, our results highlight the complexity of OT effects and support evidence that OT might even interfere with social cognitive abilities. However, the Bayesian analysis approach shows that there is only moderate evidence that OT influences mind-reading, highlighting the need for larger-scale studies considering the discussed aspects that might have led to divergent study results.

Tips From the Top: Do the Best Performers Really Give the Best Advice?

Everyone knows that if you want to learn how to do something, you should get advice from people who do it well. But is everyone right? In a series of studies (N = 8,693), adult participants played a game after receiving performance advice from previous participants. Although advice from the best-performing advisors was no more beneficial than advice from other advisors, participants believed that it had been-and they believed this despite the fact that they were told nothing about their advisors' performance. Why? The best performers did not give better advice, but they did give more of it, and participants apparently mistook quantity for quality. These studies suggest that performing and advising may often be unrelated skills and that in at least some domains, people may overvalue advice from top performers.

Higher Ultrasonic Frequency Liquid Phase Exfoliation Leads to Larger and Monolayer to Few-Layer Flakes of 2D Layered Materials.

Among the most reliable techniques for exfoliation of two-dimensional (2D) layered materials, sonication-assisted liquid-phase exfoliation (LPE) is considered as a cost-effective and straightforward method for preparing graphene and its 2D inorganic counterparts at reasonable sizes and acceptable levels of defects. Although there were rapid advances in this field, the effect and outcome of the sonication frequency are poorly understood and often ignored, resulting in a low exfoliation efficiency. Here, we demonstrate that simple mild bath sonication at a higher frequency and low power positively contributes to the thickness, size, and quality of the final exfoliated products. We show that monolayer graphene flakes can be directly exfoliated from graphite using ethanol as a solvent by increasing the frequency of the bath sonication from 37 to 80 kHz. The statistical analysis shows that ∼77% of the measured graphene flakes have a thickness below three layers with an average lateral size of 13 µm. We demonstrate that this approach works for digenite (Cu9S5) and silver sulfide (Ag2S), thus indicating that this exfoliation technique can be applied to other inorganic 2D materials to obtain high-quality few-layered flakes. This simple and effective method facilitates the formation of monolayer/few layers of graphene and transition metal chalcogenides for a wide range of applications.

A method for high-content functional imaging of intracellular calcium responses in gelatin-immobilized non-adherent cells.

Functional imaging of the intracellular calcium concentration [Ca2+]i using fluorescent indicators is a powerful and frequently applied method for assessing various biological questions in vitro, including ion channel function and intracellular signaling in homeostasis and disease. In functional [Ca2+]i imaging experiments, the fluorescence intensity of single cells is typically recorded during application of a chemical stimulus, i.e. by exchange of modified extracellular media, exposure to drugs and/or ligands. The concomitant mechanical perturbation caused by the perfusion of different solution during experimentation severely hinders calcium imaging in non-adherent cells, including peripheral immune cells, as cells in suspension are dislocated by turbulent flow during chemical stimulation. The quantitative analysis, involving time-courses of intracellular fluorescence signal changes, necessitates cells to remain at the same position throughout the experiment. To prevent dislocation of cells during solution exchange, and to enable imaging as well as analysis of Ca2+ responses in immune cells, a gelatin-based method for immobilization of non-adherent cells was developed. Gelatin has been a long-serving material for cell immobilization, e.g. in 3D bio-printing of cells and has thus, also been employed in the context of this study. To demonstrate the applicability of the established method for functional Ca2+ imaging in gelatin-immobilized suspension cells, a proof-of-concept study was conducted using human peripheral blood model cell lines (Jurkat/T-lymphocytes and THP-1/monocytes), Ca2+ indicators (Fluo-4 and Fura-2) and two different fluorescence microscopy rigs. The data presented that the established methodology is applicable for studying Ca2+ signaling by in vitro high-content functional imaging of [Ca2+]i in suspension cells, including but not restricted to human immune cells.

Impaired Transmigration of Myeloid-Derived Suppressor Cells across Human Sinusoidal Endothelium Is Associated with Decreased Expression of CD13.

Human monocytic myeloid-derived suppressor cells (MO-MDSCs) within the hepatic compartment suppress inflammation and impair immune surveillance in liver cancer. It is currently not known whether recruitment of MO-MDSCs from blood via hepatic sinusoidal endothelium (HSEC) contributes to their enrichment within the hepatic compartment. We compared the transmigratory potential of MO-MDSCs and monocytes after adhesion to hepatic endothelial monolayers in flow-based assays that mimic in vivo shear stress in the sinusoids. Despite comparable binding to HSEC monolayers, proportionally fewer MO-MDSCs underwent transendothelial migration, indicating that the final steps of extravasation, where actin polymerization plays an important role, are impaired in MO-MDSCs. In this article, we found reduced levels of CD13 on MO-MDSCs, which has recently been reported to control cell motility in monocytes, alongside reduced VLA-4 expression, an integrin predominantly involved in adherence to the apical side of the endothelium. CD13 and VLA-4 blocking and activating Abs were used in flow-based adhesion assays, live-cell imaging of motility, and actin polymerization studies to confirm a role for CD13 in impaired MO-MDSC transmigration. These findings indicate that CD13 significantly contributes to tissue infiltration by MO-MDSCs and monocytes, thereby contributing to the pathogenesis of hepatic inflammation.

When fairness matters less than we expect.

Do those who allocate resources know how much fairness will matter to those who receive them? Across seven studies, allocators used either a fair or unfair procedure to determine which of two receivers would receive the most money. Allocators consistently overestimated the impact that the fairness of the allocation procedure would have on the happiness of receivers (studies 1-3). This happened because the differential fairness of allocation procedures is more salient before an allocation is made than it is afterward (studies 4 and 5). Contrary to allocators' predictions, the average receiver was happier when allocated more money by an unfair procedure than when allocated less money by a fair procedure (studies 6 and 7). These studies suggest that when allocators are unable to overcome their own preallocation perspectives and adopt the receivers' postallocation perspectives, they may allocate resources in ways that do not maximize the net happiness of receivers.

Structural changes at the myrtenol backbone reverse its positive allosteric potential into inhibitory GABAA receptor modulation.

GABAA receptors are ligand-gated anion channels that form pentameric arrangements of various subunits. Positive allosteric modulators of GABAA receptors have been reported as being isolated either from plants or synthesized analogs of known GABAA receptor targeting drugs. Recently, we identified monoterpenes, e.g. myrtenol as a positive allosteric modulator at α1ß2 GABAA receptors. Here, along with pharmacophore-based virtual screening studies, we demonstrate that scaffold modifications of myrtenol resulted in the loss of modulatory activity. Two independent approaches, fluorescence-based compound analysis and electrophysiological recordings in whole-cell configurations were used for analysis of transfected cells. C-atoms 1 and 2 of the myrtenol backbone were identified as crucial to preserve positive allosteric potential. A modification at C-atom 2 and lack of the hydroxyl group at C-atom 1 exhibited significantly reduced GABAergic currents at α1ß2, α1ß2γ, α2ß3, α2ß3γ and α4ß3δ receptors. This effect was independent of the γ2 subunit. A sub-screen with side chain length and volume differences at the C-atom 1 identified two compounds that inhibited GABAergic responses but without receptor subtype specificity. Our combined approach of pharmacophore-based virtual screening and functional readouts reveals that side chain modifications of the bridged six-membered ring structure of myrtenol are crucial for its modulatory potential at GABAA receptors.

Transposable Elements Mediate Adaptive Debilitation of Flagella in Experimental Escherichia coli Populations.

Although insertion sequence (IS) elements are generally considered genomic parasites, they can mediate adaptive genetic changes in bacterial genomes. We discovered that among 12 laboratory-evolved Escherichia coli populations, three had experienced at least six different IS1-mediated deletions of flagellar genes. These deletions all involved the master flagellar regulator flhDC, and as such completely incapacitate motility. Two lines of evidence strongly suggest that these deletions were adaptive in our evolution experiment: (1) parallel evolution in three independent populations is highly unlikely just by chance, and (2) one of these deletion mutations swept to fixation within ~1000 generations, which is over two million times faster than expected if this deletion was instead selectively neutral and thus evolving by genetic drift. Because flagella are energetically expensive to synthesize and operate, we suspect that debilitating their construction conferred a fitness advantage in our well-stirred evolution experiment. These findings underscore the important role that IS elements can play in mediating adaptive loss-of-function mutations in bacteria.

The Novelty Penalty.

People often tell each other stories about their past experiences. But do they tell the right ones? Speakers and listeners predicted that listeners would enjoy hearing novel stories (i.e., stories about experiences the listeners had never had) more than familiar stories (i.e., stories about experiences the listeners had already had). In fact, listeners enjoyed hearing familiar stories much more than novel ones (Studies 1 and 2). This did not happen because the familiar and novel stories differed in their content or delivery (Study 3). Rather, it happened because human speech is riddled with informational gaps, and familiar stories allow listeners to use their own knowledge to fill in those gaps (Study 4). We discuss reasons why novel stories are more difficult to tell, and why familiar stories are more enjoyable to hear, than either speakers or listeners expect.

Optimizing neuronal differentiation of human pluripotent NT2 stem cells in monolayer cultures.

Human pluripotent embryonal carcinoma (NT2) cells are increasingly considered as a suitable model for in vitro developmental toxicity and neurotoxicity (DT/DNT) studies as they undergo neuronal differentiation upon stimulation with retinoic acid (RA) and allow toxicity testing at different stages of maturation. However, differentiation of NT2 cells is not straightforward. There are different protocols available in the literature reporting varying results with regard to differentiation efficiency, expression of neuronal markers and morphological characteristics of differentiated cells. Yet, the efficiency of available protocols has not been systematically compared. To address this question, we quantified the number and size of cell cluster formed during differentiation using published and modified protocols and analyzed the abundance of neuronal and non-neuronal expression markers using immunocytochemistry. In the course of the experiments we observed that differentiation results strongly depend on the cell density at differentiation-initiation as well as on the type of used cell culture plastic ware. Based on those observations and the results from our comparative analysis, we created our own optimized and robust protocol that reproducibly reveals differentiated cells with high yield. We conclude that our method may be superior to differentiation of NT2 cells for systematic in vitro-based primary screening for developmental toxicants and neurotoxicants at different stages of maturation over previous protocols used. Our approach will also contribute to reduce animal testing in the context of the 3Rs.

Modulation of Oxygen Content in Graphene Surfaces Using Temperature-Programmed Reductive Annealing: Electron Paramagnetic Resonance and Electrochemical Study.

The oxidation level and properties of reduced graphene oxides (rGOs) were fine-tuned using temperature-programmed reductive annealing. rGOs were annealed at different temperatures (from 500 to 1000 °C) in hydrogen to modulate their oxidation levels. The surface of the rGOs was fully characterized using electron paramagnetic resonance backed by Raman, X-ray diffraction, and chemical analysis measurements. These experiments were used to study the changes in the surface of the rGO, its surface functionalities, and its defects as a function of the reduction temperature. In addition, electrochemical measurements to quantify the oxidation level of the rGOs offer a simple tool to correlate the properties of rGOs with their structure. Finally, we explored the effect of different levels of reduction on conductivity, capacitance, and surface reactivity. This research offers simple methodological techniques and routes to control and characterize the oxidation level of bulk quantities of rGO.

Clinical symptoms, signs and tests for identification of impending and current water-loss dehydration in older people.

BACKGROUND: There is evidence that water-loss dehydration is common in older people and associated with many causes of morbidity and mortality. However, it is unclear what clinical symptoms, signs and tests may be used to identify early dehydration in older people, so that support can be mobilised to improve hydration before health and well-being are compromised. OBJECTIVES: To determine the diagnostic accuracy of state (one time), minimally invasive clinical symptoms, signs and tests to be used as screening tests for detecting water-loss dehydration in older people by systematically reviewing studies that have measured a reference standard and at least one index test in people aged 65 years and over. Water-loss dehydration was defined primarily as including everyone with either impending or current water-loss dehydration (including all those with serum osmolality ≥ 295 mOsm/kg as being dehydrated). SEARCH METHODS: Structured search strategies were developed for MEDLINE (OvidSP), EMBASE (OvidSP), CINAHL, LILACS, DARE and HTA databases (The Cochrane Library), and the International Clinical Trials Registry Platform (ICTRP). Reference lists of included studies and identified relevant reviews were checked. Authors of included studies were contacted for details of further studies. SELECTION CRITERIA: Titles and abstracts were scanned and all potentially relevant studies obtained in full text. Inclusion of full text studies was assessed independently in duplicate, and disagreements resolved by a third author. We wrote to authors of all studies that appeared to have collected data on at least one reference standard and at least one index test, and in at least 10 people aged ≥ 65 years, even where no comparative analysis has been published, requesting original dataset so we could create 2 x 2 tables. DATA COLLECTION AND ANALYSIS: Diagnostic accuracy of each test was assessed against the best available reference standard for water-loss dehydration (serum or plasma osmolality cut-off ≥ 295 mOsm/kg, serum osmolarity or weight change) within each study. For each index test study data were presented in forest plots of sensitivity and specificity. The primary target condition was water-loss dehydration (including either impending or current water-loss dehydration). Secondary target conditions were intended as current (> 300 mOsm/kg) and impending (295 to 300 mOsm/kg) water-loss dehydration, but restricted to current dehydration in the final review.We conducted bivariate random-effects meta-analyses (Stata/IC, StataCorp) for index tests where there were at least four studies and study datasets could be pooled to construct sensitivity and specificity summary estimates. We assigned the same approach for index tests with continuous outcome data for each of three pre-specified cut-off points investigated.Pre-set minimum sensitivity of a useful test was 60%, minimum specificity 75%. As pre-specifying three cut-offs for each continuous test may have led to missing a cut-off with useful sensitivity and specificity, we conducted post-hoc exploratory analyses to create receiver operating characteristic (ROC) curves where there appeared some possibility of a useful cut-off missed by the original three. These analyses enabled assessment of which tests may be worth assessing in further research. A further exploratory analysis assessed the value of combining the best two index tests where each had some individual predictive ability. MAIN RESULTS: There were few published studies of the diagnostic accuracy of state (one time), minimally invasive clinical symptoms, signs or tests to be used as screening tests for detecting water-loss dehydration in older people. Therefore, to complete this review we sought, analysed and included raw datasets that included a reference standard and an index test in people aged ≥ 65 years.We included three studies with published diagnostic accuracy data and a further 21 studies provided datasets that we analysed. We assessed 67 tests (at three cut-offs for each continuous outcome) for diagnostic accuracy of water-loss dehydration (primary target condition) and of current dehydration (secondary target condition).Only three tests showed any ability to diagnose water-loss dehydration (including both impending and current water-loss dehydration) as stand-alone tests: expressing fatigue (sensitivity 0.71 (95% CI 0.29 to 0.96), specificity 0.75 (95% CI 0.63 to 0.85), in one study with 71 participants, but two additional studies had lower sensitivity); missing drinks between meals (sensitivity 1.00 (95% CI 0.59 to 1.00), specificity 0.77 (95% CI 0.64 to 0.86), in one study with 71 participants) and BIA resistance at 50 kHz (sensitivities 1.00 (95% CI 0.48 to 1.00) and 0.71 (95% CI 0.44 to 0.90) and specificities of 1.00 (95% CI 0.69 to 1.00) and 0.80 (95% CI 0.28 to 0.99) in 15 and 22 people respectively for two studies, but with sensitivities of 0.54 (95% CI 0.25 to 0.81) and 0.69 (95% CI 0.56 to 0.79) and specificities of 0.50 (95% CI 0.16 to 0.84) and 0.19 (95% CI 0.17 to 0.21) in 21 and 1947 people respectively in two other studies). In post-hoc ROC plots drinks intake, urine osmolality and axillial moisture also showed limited diagnostic accuracy. No test was consistently useful in more than one study.Combining two tests so that an individual both missed some drinks between meals and expressed fatigue was sensitive at 0.71 (95% CI 0.29 to 0.96) and specific at 0.92 (95% CI 0.83 to 0.97).There was sufficient evidence to suggest that several stand-alone tests often used to assess dehydration in older people (including fluid intake, urine specific gravity, urine colour, urine volume, heart rate, dry mouth, feeling thirsty and BIA assessment of intracellular water or extracellular water) are not useful, and should not be relied on individually as ways of assessing presence or absence of dehydration in older people.No tests were found consistently useful in diagnosing current water-loss dehydration. AUTHORS' CONCLUSIONS: There is limited evidence of the diagnostic utility of any individual clinical symptom, sign or test or combination of tests to indicate water-loss dehydration in older people. Individual tests should not be used in this population to indicate dehydration; they miss a high proportion of people with dehydration, and wrongly label those who are adequately hydrated.Promising tests identified by this review need to be further assessed, as do new methods in development. Combining several tests may improve diagnostic accuracy.

Robust RNAi enhancement via human Argonaute-2 overexpression from plasmids, viral vectors and cell lines.

As the only mammalian Argonaute protein capable of directly cleaving mRNAs in a small RNA-guided manner, Argonaute-2 (Ago2) is a keyplayer in RNA interference (RNAi) silencing via small interfering (si) or short hairpin (sh) RNAs. It is also a rate-limiting factor whose saturation by si/shRNAs limits RNAi efficiency and causes numerous adverse side effects. Here, we report a set of versatile tools and widely applicable strategies for transient or stable Ago2 co-expression, which overcome these concerns. Specifically, we engineered plasmids and viral vectors to co-encode a codon-optimized human Ago2 cDNA along with custom shRNAs. Furthermore, we stably integrated this Ago2 cDNA into a panel of standard human cell lines via plasmid transfection or lentiviral transduction. Using various endo- or exogenous targets, we demonstrate the potential of all three strategies to boost mRNA silencing efficiencies in cell culture by up to 10-fold, and to facilitate combinatorial knockdowns. Importantly, these robust improvements were reflected by augmented RNAi phenotypes and accompanied by reduced off-targeting effects. We moreover show that Ago2/shRNA-co-encoding vectors can enhance and prolong transgene silencing in livers of adult mice, while concurrently alleviating hepatotoxicity. Our customizable reagents and avenues should broadly improve future in vitro and in vivo RNAi experiments in mammalian systems.

The unforeseen costs of extraordinary experience.

People seek extraordinary experiences--from drinking rare wines and taking exotic vacations to jumping from airplanes and shaking hands with celebrities. But are such experiences worth having? We found that participants thoroughly enjoyed having experiences that were superior to those had by their peers, but that having had such experiences spoiled their subsequent social interactions and ultimately left them feeling worse than they would have felt if they had had an ordinary experience instead. Participants were able to predict the benefits of having an extraordinary experience but were unable to predict the costs. These studies suggest that people may pay a surprising price for the experiences they covet most.

Response of sphagnum peatland testate amoebae to a 1-year transplantation experiment along an artificial hydrological gradient.

Peatland testate amoebae (TA) are well-established bioindicators for depth to water table (DWT), but effects of hydrological changes on TA communities have never been tested experimentally. We tested this in a field experiment by placing Sphagnum carpets (15 cm diameter) collected in hummock, lawn and pool microsites (origin) at three local conditions (dry, moist and wet) using trenches dug in a peatland. One series of samples was seeded with microorganism extract from all microsites. TA community were analysed at T0: 8-2008, T1: 5-2009 and T2: 8-2009. We analysed the data using conditional inference trees, principal response curves (PRC) and DWT inferred from TA communities using a transfer function used for paleoecological reconstruction. Density declined from T0 to T1 and then increased sharply by T2. Species richness, Simpson diversity and Simpson evenness were lower at T2 than at T0 and T1. Seeded communities had higher species richness in pool samples at T0. Pool samples tended to have higher density, lower species richness, Simpson diversity and Simpson Evenness than hummock and/or lawn samples until T1. In the PRC, the effect of origin was significant at T0 and T1, but the effect faded away by T2. Seeding effect was strongest at T1 and lowest vanished by T2. Local condition effect was strong but not in line with the wetness gradient at T1 but started to reflect it by T2. Likewise, TA-inferred DWT started to match the experimental conditions by T2, but more so in hummock and lawn samples than in pool samples. This study confirmed that TA responds to hydrological changes over a 1-year period. However, sensitivity of TA to hydrological fluctuations, and thus the accuracy of inferred DWT changes, was habitat specific, pool TA communities being least responsive to environmental changes. Lawns and hummocks may be thus better suited than pools for paleoecological reconstructions. This, however, contrasts with the higher prediction error and species' tolerance for DWT with increasing dryness observed in transfer function models.

Ultrasound Stimulation of Piezoelectric Nanocomposite Hydrogels Boosts Chondrogenic Differentiation in Vitro, in Both a Normal and Inflammatory Milieu.

The use of piezoelectric nanomaterials combined with ultrasound stimulation is emerging as a promising approach for wirelessly triggering the regeneration of different tissue types. However, it has never been explored for boosting chondrogenesis. Furthermore, the ultrasound stimulation parameters used are often not adequately controlled. In this study, we show that adipose-tissue-derived mesenchymal stromal cells embedded in a nanocomposite hydrogel containing piezoelectric barium titanate nanoparticles and graphene oxide nanoflakes and stimulated with ultrasound waves with precisely controlled parameters (1 MHz and 250 mW/cm2, for 5 min once every 2 days for 10 days) dramatically boost chondrogenic cell commitment in vitro. Moreover, fibrotic and catabolic factors are strongly down-modulated: proteomic analyses reveal that such stimulation influences biological processes involved in cytoskeleton and extracellular matrix organization, collagen fibril organization, and metabolic processes. The optimal stimulation regimen also has a considerable anti-inflammatory effect and keeps its ability to boost chondrogenesis in vitro, even in an inflammatory milieu. An analytical model to predict the voltage generated by piezoelectric nanoparticles invested by ultrasound waves is proposed, together with a computational tool that takes into consideration nanoparticle clustering within the cell vacuoles and predicts the electric field streamline distribution in the cell cytoplasm. The proposed nanocomposite hydrogel shows good injectability and adhesion to the cartilage tissue ex vivo, as well as excellent biocompatibility in vivo, according to ISO 10993. Future perspectives will involve preclinical testing of this paradigm for cartilage regeneration.