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Because of previous reports of a possible correlation between echinocytogenesis and muscular dystrophies, we investigated the time-dependent development of echinocytes in the blood of normal subjects, patients, and healthy carriers of Duchenne dystrophy. There was a quantitatively significant increase of echinocytes in patients and carriers.

We investigated the number of echinocytes, the serum hemopexin level, and spectrin band II phosphorylation in the blood of normal subjects, patients, and carriers of Duchenne dystrophy. The patients and carriers exhibited quantitatively significant differences with respect to controls.

Control over the shape and morphology of single crystals is a theme of great interest in fundamental science and for technological application. Many synthetic strategies to achieve this goal are inspired by biomineralization processes. Indeed, organisms are able to produce crystals with high fidelity in shape and morphology utilizing macromolecules that act as modifiers. An alternative strategy can be the recovery of crystals from biomineralization products, in this case, seashells. In particular, waste mussel shells from aquaculture are considered. They are mainly built up of single crystals of calcite fibers and aragonite tablets forming an outer and an inner layer, respectively. A simple mechanochemical treatment has been developed to separate and recover these two typologies of single crystals. The characterization of these single crystals showed peculiar properties with respect to the calcium carbonate from quarry or synthesis. We exploited these biomaterials in the water remediation field using them as substrate adsorbing dyes. We found that these substrates show a high capability of adsorption for anionic dye, such as Eosin Y, but a low capability of adsorption for cationic dyes, such as Blue Methylene. The adsorption was reversible at pH 5.6. This application represents just an example of the potential use of these biogenic single crystals. We also envision potential applications as reinforcing fillers and optical devices. published

Basic information on the reproductive biology of many scleractinian corals species is limited or entirely lacking, particularly from temperate zones, though it is essential for a better understanding of their ecology. This study describes the morphological aspects and the annual cycle of gametogenesis and biometric parameterss of the papillose cup coral Paracyathus pulchellus collected at Palinuro (Italy, Southern Tyrrhenian Sea), filling a knowledge gap about the reproductive biology of a widespread Mediterranean and Northern Atlantic coral. Samples of P. pulchellus were collected by SCUBA diving between 5- and 10-meters depth during 18 monthly collections from June 2010 to December 2011. Sexually active polyps displayed either oocytes or spermaries, indicating that P. pulchellus was gonochoric. The sex ratio of sexually active polyps was 1:1. Gametogenesis began with undifferentiated germ cells arose in the gastrodermis that migrated towards the mesoglea of the mesentery where they completed the development. During spermatogenesis, spermary diameter increased from 25 to 83 µm. Oocyte diameter ranged from 9 to 146 µm and during oogenesis the nucleus/cytoplasm ratio decreased due to the accumulation of yolk. The nucleus migrated to the periphery of the oocyte adhering closely to the cell membrane. No embryo was observed in the coelenteric cavity of the polyps, suggesting an external development of planktotrophic larvae due to the small-sized mature oocytes and a possible broadcast spawning reproductive mode. Gonadal index of both females and males increased significantly from August until November and fertilization took place from November to January. Only sexually inactive individuals were observed from February to April, suggesting a quiescence period in both males and females. Seasonal variations in water temperature and photoperiod may have a key role in regulating gametogenesis. The analysis of the main biometric parameters (polyp width, height, dry skeletal mass, volume, surface/volume ratio and bulk skeletal density) showed a negative correlation between size and skeletal density, and no sexual dimorphism.

Tourism is one of the most important economic sectors worldwide, with significant overarching impact on the environment, including negative effects caused by tourist inappropriate behavior while on vacation. By providing informal educational activities, tourism also has an educative role that leads to positive learning outcomes and beneficial environmental effects. Here we present the short- and long-term outcomes of a project for environmental education (Glocal Education) carried out in three travel destinations, aimed at promoting sustainability variables (knowledge, attitude, and awareness) in participating tourists. Since psychological components can affect learning outcomes, we also considered tourist satisfaction in participating in the project and identification with its values, as well as the intention to travel with the hosting tour operator again in the future. Tourists were asked to complete evaluation questionnaires three times: before Glocal Education activities, right after activities (i.e., while still on vacation), and after at least one year from initial project participation. Short- and long-term learning outcomes were tested, and possible relations between these variables and psychological components (satisfaction, identification, and intention) of the learning experience were verified. Overall, knowledge, attitude and awareness increased in the short term, while in the long term, knowledge and attitude decreased, and awareness remained constant. In most cases, psychological components showed positive relation with sustainability variables, which suggested their important role in structuring and carrying out environmental education activities. This study suggests that informal environmental education activities can be advantageous for tourism stakeholders in terms of customer loyalty. Such activities can contribute to enhance environment literacy, by allowing tourists to observe the environmental impact caused by human activity, and understand how their day-to-day actions, even if small, might help address some of the current concerns for environmental conservation.

Atmospheric carbon dioxide enrichment alters seawater carbonate chemistry, thus threatening calcifying organisms such as corals. Coral populations at carbon dioxide vents are natural acidification experiments that mimic organism responses to seawater pH values projected for 2100. Even if demographic traits are paramount information to assess ecological relationships and habitat suitability, population dynamics studies on corals thriving under acidified conditions are lacking. Here, we investigate the demography and reproduction of populations of the solitary, symbiotic, temperate coral Balanophyllia europaea naturally living along a pH gradient at a Mediterranean CO2 vent. Gametogenesis and larval production were unaffected while recruitment efficiency collapsed at low and variable pH, contributing to coral abundance decline and suggesting that life stages between larval release and early polyp growth are hindered by acidification. Exploring these processes is crucial to assess coral fate in the forthcoming acidified oceans, to preserve coral ecosystems and the socioeconomic services they provide.

Ecotourism gives tourists the opportunity to improve knowledge and awareness of environmental issues while on vacation. Recreational environmental education has been proven an effective method to raise perception of human impact on ecosystems. “Glocal Education” is an education project aimed at developing environmental interest in tourists on vacation. The present study assessed the effectiveness of Glocal Education in improving tourist environmental interest. Using specific questionnaires, we evaluated project impact on tourists, tourist satisfaction regarding the project and customer loyalty towards the tour operator hosting the project. The study took place at three mass touristic facilities, where tourists were asked to fill a questionnaire before and after participating in educational activities (e.g., biology lessons, excursions). The average score of both questionnaires was then compared to evaluate possible improvement of tourist knowledge, attitude and awareness. Results showed that such activities had a significantly positive impact on tourist knowledge, attitude and awareness at all localities. High levels of satisfaction and loyalty towards the host tour operator were observed at all sites, which indicate that once a person is briefed about the correct approach to natural systems, they can become increasingly interested in taking action, developing an “advocate” role. This study shows how informal education activities can act as trigger for environmental awareness and behavior among tourists, providing them with the tools, knowledge, and motivation to critically discern what is and isn't environmentally friendly, not only in terms of products and services in their everyday life, but also when choosing their vacation spots.

Here we present a quantitative mechanism-based investigation aimed at comparing the cell uptake, intracellular trafficking, endosomal escape and final fate of lipoplexes and lipid-protamine/deoxyribonucleic acid (DNA) (LPD) nanoparticles (NPs) in living Chinese hamster ovary (CHO) cells. As a model, two lipid formulations were used for comparison. The first formulation is made of the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and the zwitterionic lipid dioleoylphosphocholine (DOPC), while the second mixture is made of the cationic 3β-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and the zwitterionic helper lipid dioleoylphosphatidylethanolamine (DOPE). Our findings indicate that lipoplexes are efficiently taken up through fluid-phase macropinocytosis, while a less efficient uptake of LPD NPs occurs through a combination of both macropinocytosis and clathrin-dependent pathways. Inside the cell, both lipoplexes and LPD NPs are actively transported towards the cell nucleus, as quantitatively addressed by spatio-temporal image correlation spectroscopy (STICS). For each lipid formulation, LPD NPs escape from endosomes more efficiently than lipoplexes. When cells were treated with DOTAP-DOPC-containing systems the majority of the DNA was trapped in the lysosome compartment, suggesting that extensive lysosomal degradation was the rate-limiting factors in DOTAP-DOPC-mediated transfection. On the other side, escape from endosomes is large for DC-Chol-DOPE-containing systems most likely due to DOPE and cholesterol-like molecules, which are able to destabilize the endosomal membrane. The lipid-dependent and structure-dependent enhancement of transfection activity suggests that DNA is delivered to the nucleus synergistically: the process requires both the membrane-fusogenic activity of the nanocarrier envelope and the employment of lipid species with intrinsic endosomal rupture ability. © 2013 Elsevier B.V.

While knowledge of the reproductive biology of tropical scleractinian corals is extensive, information from temperate zones is limited. The aim of this study is to describe the reproductive biology of Caryophyllia inornata, a temperate species, and to increase the understanding of the reproductive strategies of Mediterranean corals. Samples of C. inornata were collected during SCUBA surveys at Elba island. Sexually active individuals displayed either male or female germ cells, showing a gonochoric sexuality. C. inornata exhibited an unusual pattern of embryogenesis. Embryos appeared throughout the whole year in males and in sexually inactive individuals, and they did not show a seasonal pattern of development, as usually expected for sexual reproduction. This observation suggests the possibility of asexual origin. These embryogenetic sexually inactive individuals were larger in size than the embryogenetic sexually active ones, and they might be senile polyps that preserve the ability to produce embryos only by agamic reproduction.

The mechanisms underlying pain in Parkinson's disease (PD) are unclear. Although a few studies have reported that PD patients may have low pain threshold and tolerance, none could accurately assess whether there was a correlation between sensory thresholds and demographic/clinical features of PD patients. Thus, tactile threshold, pain threshold, and pain tolerance to electrical stimuli in the hands and feet were assessed in 106 parkinsonian patients (of whom 66 reported chronic pain) and 51 age- and sex-matched healthy subjects. Linear regression models determined relationships between psychophysical parameters and demographic/clinical features. Female gender, severity of disease, medical disease associated with painful symptoms, and dyskinesia were more frequently observed in PD patients experiencing pain, even though dyskinesia did not reach significance. Pain threshold and pain tolerance were significantly lower in PD patients than in control subjects, whereas the tactile threshold yielded comparable values in both groups. Multivariable linear regression analyses yielded significant inverse correlations of pain threshold and pain tolerance with motor symptom severity and Beck depression inventory. Pain threshold and pain tolerance did not differ between PD patients with and without pain. In the former group, there was no relationship between pain threshold and the intensity/type of pain, and number of painful body parts. These findings suggest that pain threshold and pain tolerance tend to decrease as PD progresses, which can predispose to pain development. Female gender, dyskinesia, medical conditions associated with painful symptoms, and postural abnormalities secondary to rigidity/bradikinesia may contribute to the appearance of spontaneous pain in predisposed subjects.

We packaged condensed DNA/protamine particles in multicomponent envelope-type nanoparticle systems (MENS) combining different molar fractions of the cationic lipids 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and 3β-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and the zwitterionic lipids dioleoylphosphocholine (DOPC) and dioleoylphosphatidylethanolamine (DOPE). Dynamic light scattering (DLS) and microelectrophoresis allowed us to identify the cationic lipid/DNA charge ratio at which MENS are small sized and positively charged, while synchrotron small-angle X-ray scattering (SAXS) and atomic force microscopy (AFM) revealed that MENS are well-shaped DNA/protamine particles covered by a lipid monobilayer. Transfection efficiency (TE) experiments indicate that a nanoparticle formulation, termed MENS-3, was not cytotoxic and highly efficient to transfect Chinese hamster ovary (CHO) cells. To rationalize TE, we performed a quantitative investigation of cell uptake, intracellular trafficking, endosomal escape, and final fate by laser scanning confocal microscopy (LSCM). We found that fluid-phase macropinocytosis is the only endocytosis pathway used by MENS-3. Once taken up by the cell, complexes that are actively transported by microtubules frequently fuse with lysosomes, while purely diffusing systems do not. Indeed, spatiotemporal image correlation spectroscopy (STICS) clarified that MENS-3 mostly exploit diffusion to move in the cytosol of CHO cells, thus explaining the high TE levels observed. Also, MENS-3 exhibited a marked endosomal rupture ability resulting in extraordinary DNA release. The lipid-dependent and structure-dependent TE boost suggests that efficient transfection requires both the membrane-fusogenic activity of the nanocarrier envelope and the employment of lipid species with intrinsic endosomal rupture ability. © 2013 American Chemical Society.

Mesophotic coral reef ecosystems remain largely unexplored with only limited information available on taxonomic composition, abundance and distribution. Yet, mesophotic reefs may serve as potential refugia for shallow-water species and thus understanding biodiversity, ecology and connectivity of deep reef communities is integral for resource management and conservation. The Caribbean coral, Montastraea cavernosa, is considered a depth generalist and is commonly found at mesophotic depths. We surveyed abundance and size-frequency of M. cavernosa populations at six shallow (10m) and six upper mesophotic (45m) sites in Bermuda and found population structure was depth dependent. The mean surface area of colonies at mesophotic sites was significantly smaller than at shallow sites, suggesting that growth rates and maximum colony surface area are limited on mesophotic reefs. Colony density was significantly higher at mesophotic sites, however, resulting in equal contributions to overall percent cover. Size-frequency distributions between shallow and mesophotic sites were also significantly different with populations at mesophotic reefs skewed towards smaller individuals. Overall, the results of this study provide valuable baseline data on population structure, which indicate that the mesophotic reefs of Bermuda support an established population of M. cavernosa.

The variety of reproductive processes and modes among coral species reflects their extraordinary regeneration ability. Scleractinians are an established example of clonal animals that can exhibit a mixed strategy of sexual and asexual reproduction to maintain their populations. This study provides the first description of the annual reproductive cycle and embryogenesis of the temperate species Caryophyllia inornata. Cytometric analyses were used to define the annual development of germ cells and embryogenesis. The species was gonochoric with three times more male polyps than female. Polyps were sexually mature from 6 to 8 mm length. Not only females, but also sexually inactive individuals (without germ cells) and males were found to brood their embryos. Spermaries required 12 months to reach maturity, while oogenesis seemed to occur more rapidly (5–6 months). Female polyps were found only during spring and summer. Furthermore, the rate of gamete development in both females and males increased significantly from March to May and fertilization was estimated to occur from April to July, when mature germ cells disappeared. Gametogenesis showed a strong seasonal influence, while embryos were found throughout the year in males and in sexually inactive individuals without a defined trend. This unusual embryogenesis suggests the possibility of agamic reproduction, which combined with sexual reproduction results in high fertility. This mechanism is uncommon and only four other scleractinians (Pocillopora damicornis, Tubastraea diaphana, T. coccinea and Oulastrea crispata) have been shown to generate their broods asexually. The precise nature of this process is still unknown.