Biocompatible adipose extracellular matrix and reduced graphene oxide nanocomposite for tissue engineering applications.

Despite the immense need for effective treatment of spinal cord injury (SCI), no successful repair strategy has yet been clinically implemented. Multifunctional biomaterials, based on porcine adipose tissue-derived extracellular matrix (adECM) and reduced graphene oxide (rGO), were recently shown to stimulate in vitro neural stem cell growth and differentiation. Nevertheless, their functional performance in clinically more relevant in vivo conditions remains largely unknown. Before clinical application of these adECM-rGO nanocomposites can be considered, a rigorous assessment of the cytotoxicity and biocompatibility of these biomaterials is required. For instance, xenogeneic adECM scaffolds could still harbour potential immunogenicity following decellularization. In addition, the toxicity of rGO has been studied before, yet often in experimental settings that do not bear relevance to regenerative medicine. Therefore, the present study aimed to assess both the in vitro as well as in vivo safety of adECM and adECM-rGO scaffolds. First, pulmonary, renal and hepato-cytotoxicity as well as macrophage polarization studies showed that scaffolds were benign invitro. Then, a laminectomy was performed at the 10th thoracic vertebra, and scaffolds were implanted directly contacting the spinal cord. For a total duration of 6 weeks, animal welfare was not negatively affected. Histological analysis demonstrated the degradation of adECM scaffolds and subsequent tissue remodeling. Graphene-based scaffolds showed a very limited fibrous encapsulation, while rGO sheets were engulfed by foreign body giant cells. Furthermore, all scaffolds were infiltrated by macrophages, which were largely polarized towards a pro-regenerative phenotype. Lastly, organ-specific histopathology and biochemical analysis of blood did not reveal any adverse effects. In summary, both adECM and adECM-rGO implants were biocompatible upon laminectomy while establishing a pro-regenerative microenvironment, which justifies further research on their therapeutic potential for treatment of SCI.

Fabrication of Customizable and Reproducible 3D Chondrocyte-Laden Nanofibrous Architectures: Effect of Specific Fiber Alignments and Porosities on Chondrocyte Response under Cyclic Compression.

Electrospinning has been widely employed to fabricate complex extracellular matrix-like microenvironments for tissue engineering due to its ability to replicate structurally biomimetic micro- and nanotopographic cues. Nevertheless, these nanofibrous structures are typically either confined to bidimensional systems or confined to three-dimensional ones that are unable to provide controlled multiscale patterns. Thus, an electrospinning modality was used in this work to fabricate chondrocyte-laden nanofibrous scaffolds with highly customizable three-dimensional (3D) architectures in an automated manner, with the ultimate goal of recreating a suitable 3D scaffold for articular cartilage tissue engineering. Three distinct architectures were designed and fabricated by combining multiple nanofibrous and chondrocyte-laden hydrogel layers and tested in vitro in a compression bioreactor system. Results demonstrated that it was possible to precisely control the placement and alignment of electrospun polycaprolactone and gelatin nanofibers, generating three unique architectures with distinctive macroscale porosity, water absorption capacity, and mechanical properties. The architecture organized in a lattice-like fashion was highly porous with substantial pore interconnectivity, resulting in a high-water absorption capacity but a poor compression modulus and relatively weaker energy dissipation capacity. The donut-like 3D geometry was the densest, with lower swelling, but the highest compression modulus and improved energy dissipation ability. The third architecture combined a lattice and donut-like fibrous arrangement, exhibiting intermediary behavior in terms of porosity, water absorption, compression modulus, and energy dissipation capacity. The properties of the donut-like 3D architecture demonstrated great potential for articular cartilage tissue engineering, as it mimicked key topographic, chemical, and mechanical characteristics of chondrocytes' surrounding environment. In fact, the combination of these architectural features with a dynamically compressive mechanical stimulus triggered the best in vitro results in terms of viability and biosynthetic production.

Effects of graphene oxide and reduced graphene oxide nanomaterials on porcine endothelial progenitor cells.

Graphene oxide (GO) and reduced graphene oxide (rGO) have been widely used in the field of tissue regeneration and various biomedical applications. In order to use these nanomaterials in organisms, it is imperative to possess an understanding of their impact on different cell types. Due to the potential of these nanomaterials to enter the bloodstream, interact with the endothelium and accumulate within diverse tissues, it is highly relevant to probe them when in contact with the cellular components of the vascular system. Endothelial progenitor cells (EPCs), involved in blood vessel formation, have great potential for tissue engineering and offer great advantages to study the possible angiogenic effects of biomaterials. Vascular endothelial growth factor (VEGF) induces angiogenesis and regulates vascular permeability, mainly activating VEGFR2 on endothelial cells. The effects of GO and two types of reduced GO, obtained after vacuum-assisted thermal treatment for 15 min (rGO15) and 30 min (rGO30), on porcine endothelial progenitor cells (EPCs) functionality were assessed by analyzing the nanomaterial intracellular uptake, reactive oxygen species (ROS) production and VEGFR2 expression by EPCs. The results evidence that short annealing (15 and 30 minutes) at 200 °C of GO resulted in the mitigation of both the increased ROS production and decline in VEGFR2 expression of EPCs upon GO exposure. Interestingly, after 72 hours of exposure to rGO30, VEGFR2 was higher than in the control culture, suggesting an early angiogenic potential of rGO30. The present work reveals that discrete variations in the reduction of GO may significantly affect the response of porcine endothelial progenitor cells.

Effects of Graphene Oxide Nanosheets in Freshwater Biofilms.

Graphene oxide (GO) properties make it a promising material for graphene-based applications in areas such as biomedicine, agriculture, and the environment. Thus, its production is expected to increase, reaching hundreds of tons every year. One GO final destination is freshwater bodies, possibly affecting the communities of these systems. To clarify the effect that GO may impose in freshwater communities, a fluvial biofilm scraped from submerged river stones was exposed to a range (0.1 to 20 mg/L) of GO concentrations during 96 h. With this approach, we hypothesized that GO can: (1) cause mechanical damage and morphological changes in cell biofilms; (2) interfere with the absorption of light by biofilms; (3) and generate oxidative stress, causing oxidative damage and inducing biochemical and physiological alterations. Our results showed that GO did not inflict mechanical damage. Instead, a positive effect is proposed, linked to the ability of GO to bind cations and increase the micronutrient availability to biofilms. High concentrations of GO increased photosynthetic pigment (chlorophyll a, b, and c, and carotenoids) content as a strategy to capture the available light more effectively as a response to the shading effect. A significant increase in the enzymatic (SOD and GSTs activity) and low molecular weight (lipids and carotenoids) antioxidant response was observed, that efficiently reduced oxidative stress effects, reducing the level of peroxidation, and preserving membrane integrity. Being complex entities, biofilms are more similar to environmental communities and may provide more accurate information to evaluate the impact of GO in aquatic systems.

Customizing 3D Structures of Vertically Aligned Carbon Nanotubes to Direct Neural Stem Cell Differentiation.

Neural tissue-related illnesses have a high incidence and prevalence in society. Despite intensive research efforts to enhance the regeneration of neural cells into functional tissue, effective treatments are still unavailable. Here, a novel therapeutic approach based on vertically aligned carbon nanotube forests (VA-CNT forests) and periodic VA-CNT micropillars produced by thermal chemical vapor deposition is explored. In addition, honeycomb-like and flower-like morphologies are created. Initial viability testing reveals that NE-4C neural stem cells seeded on all morphologies survive and proliferate. In addition, free-standing VA-CNT forests and capillary-driven VA-CNT forests are created, with the latter demonstrating enhanced capacity to stimulate neuritogenesis and network formation under minimal differentiation medium conditions. This is attributed to the interaction between surface roughness and 3D-like morphology that mimics the native extracellular matrix, thus enhancing cellular attachment and communication. These findings provide a new avenue for the construction of electroresponsive scaffolds based on CNTs for neural tissue engineering.

Anisotropic 3D scaffolds for spinal cord guided repair: Current concepts.

A spinal cord injury (SCI) can be caused by unforeseen events such as a fall, a vehicle accident, a gunshot, or a malignant illness, which has a significant impact on the quality of life of the patient. Due to the limited regenerative potential of the central nervous system (CNS), SCI is one of the most daunting medical challenges of modern medicine. Great advances have been made in tissue engineering and regenerative medicine, which include the transition from two-dimensional (2D) to three-dimensional (3D) biomaterials. Combinatory treatments that use 3D scaffolds may significantly enhance the repair and regeneration of functional neural tissue. In an effort to mimic the chemical and physical properties of neural tissue, scientists are researching the development of the ideal scaffold made of synthetic and/or natural polymers. Moreover, in order to restore the architecture and function of neural networks, 3D scaffolds with anisotropic properties that replicate the native longitudinal orientation of spinal cord nerve fibres are being designed. In an effort to determine if scaffold anisotropy is a crucial property for neural tissue regeneration, this review focuses on the most current technological developments relevant to anisotropic scaffolds for SCI. Special consideration is given to the architectural characteristics of scaffolds containing axially oriented fibres, channels, and pores. By analysing neural cell behaviour in vitro and tissue integration and functional recovery in animal models of SCI, the therapeutic efficacy is evaluated for its successes and limitations.

Interfacing reduced graphene oxide with an adipose-derived extracellular matrix as a regulating milieu for neural tissue engineering.

Enthralling evidence of the potential of graphene-based materials for neural tissue engineering is motivating the development of scaffolds using various structures related to graphene such as graphene oxide (GO) or its reduced form. Here, we investigated a strategy based on reduced graphene oxide (rGO) combined with a decellularized extracellular matrix from adipose tissue (adECM), which is still unexplored for neural repair and regeneration. Scaffolds containing up to 50 wt% rGO relative to adECM were prepared by thermally induced phase separation assisted by carbodiimide (EDC) crosslinking. Using partially reduced GO enables fine-tuning of the structural interaction between rGO and adECM. As the concentration of rGO increased, non-covalent bonding gradually prevailed over EDC-induced covalent conjugation with the adECM. Edge-to-edge aggregation of rGO favours adECM to act as a biomolecular physical crosslinker to rGO, leading to the softening of the scaffolds. The unique biochemistry of adECM allows neural stem cells to adhere and grow. Importantly, high rGO concentrations directly control cell fate by inducing the differentiation of both NE-4C cells and embryonic neural progenitor cells into neurons. Furthermore, primary astrocyte fate is also modulated as increasing rGO boosts the expression of reactivity markers while unaltering the expression of scar-forming ones.

Three-dimensional nanofibrous and porous scaffolds of poly(ε-caprolactone)-chitosan blends for musculoskeletal tissue engineering.

One of the established tissue engineering strategies relies on the fabrication of appropriate materials architectures (scaffolds) that mimic the extracellular matrix (ECM) and assist the regeneration of living tissues. Fibrous structures produced by electrospinning have been widely used as reliable ECM templates but their two-dimensional structure restricts, in part, cell infiltration and proliferation. A recent technique called thermally-induced self-agglomeration (TISA) allowed to alleviate this drawback by rearranging the 2D electrospun membranes into highly functional 3D porous-fibrous systems. Following this trend, the present research focused on preparing polycaprolactone/chitosan blends by electrospinning, to then convert them into 3D structures by TISA. By adding different amounts of chitosan, it was possible to accurately modulate the physicochemical properties of the obtained 3D nanofibrous scaffolds, leading to highly porous constructs with distinct morphologic and mechanical features. Viability and proliferation studies using adult human chondrocytes also revealed that the biocompatibility of the scaffolds was not impaired after 28 days of cell culture, highlighting their potential to be included into musculoskeletal tissue engineering applications, particularly cartilage repair.

Effects of Graphene Oxide and Reduced Graphene Oxide Nanostructures on CD4+ Th2 Lymphocytes.

The activation of T helper (Th) lymphocytes is necessary for the adaptive immune response as they contribute to the stimulation of B cells (for the secretion of antibodies) and macrophages (for phagocytosis and destruction of pathogens) and are necessary for cytotoxic T-cell activation to kill infected target cells. For these issues, Th lymphocytes must be converted into Th effector cells after their stimulation through their surface receptors TCR/CD3 (by binding to peptide-major histocompatibility complex localized on antigen-presenting cells) and the CD4 co-receptor. After stimulation, Th cells proliferate and differentiate into subpopulations, like Th1, Th2 or Th17, with different functions during the adaptative immune response. Due to the central role of the activation of Th lymphocytes for an accurate adaptative immune response and considering recent preclinical advances in the use of nanomaterials to enhance T-cell therapy, we evaluated in vitro the effects of graphene oxide (GO) and two types of reduced GO (rGO15 and rGO30) nanostructures on the Th2 lymphocyte cell line SR.D10. This cell line offers the possibility of studying their activation threshold by employing soluble antibodies against TCR/CD3 and against CD4, as well as the simultaneous activation of these two receptors. In the present study, the effects of GO, rGO15 and rGO30 on the activation/proliferation rate of these Th2 lymphocytes have been analyzed by studying cell viability, cell cycle phases, intracellular content of reactive oxygen species (ROS) and cytokine secretion. High lymphocyte viability values were obtained after treatment with these nanostructures, as well as increased proliferation in the presence of rGOs. Moreover, rGO15 treatment decreased the intracellular ROS content of Th2 cells in all stimulated conditions. The analysis of these parameters showed that the presence of these GO and rGO nanostructures did not alter the response of Th2 lymphocytes.

Is Graphene Shortening the Path toward Spinal Cord Regeneration?

Along with the development of the next generation of biomedical platforms, the inclusion of graphene-based materials (GBMs) into therapeutics for spinal cord injury (SCI) has potential to nourish topmost neuroprotective and neuroregenerative strategies for enhancing neural structural and physiological recovery. In the context of SCI, contemplated as one of the most convoluted challenges of modern medicine, this review first provides an overview of its characteristics and pathophysiological features. Then, the most relevant ongoing clinical trials targeting SCI, including pharmaceutical, robotics/neuromodulation, and scaffolding approaches, are introduced and discussed in sequence with the most important insights brought by GBMs into each particular topic. The current role of these nanomaterials on restoring the spinal cord microenvironment after injury is critically contextualized, while proposing future concepts and desirable outputs for graphene-based technologies aiming to reach clinical significance for SCI.

Multiscale Sensing of Bone-Implant Loosening for Multifunctional Smart Bone Implants: Using Capacitive Technologies for Precision Controllability.

The world population growth and average life expectancy rise have increased the number of people suffering from non-communicable diseases, namely osteoarthritis, a disorder that causes a significant increase in the years lived with disability. Many people who suffer from osteoarthritis undergo replacement surgery. Despite the relatively high success rate, around 10% of patients require revision surgeries, mostly because existing implant technologies lack sensing devices capable of monitoring the bone-implant interface. Among the several monitoring methodologies already proposed as substitutes for traditional imaging methods, cosurface capacitive sensing systems hold the potential to monitor the bone-implant fixation states, a mandatory capability for long-term implant survival. A multifaceted study is offered here, which covers research on the following points: (1) the ability of a cosurface capacitor network to effectively monitor bone loosening in extended peri-implant regions and according to different stimulation frequencies; (2) the ability of these capacitive architectures to provide effective sensing in interfaces with hydroxyapatite-based layers; (3) the ability to control the operation of cosurface capacitive networks using extracorporeal informatic systems. In vitro tests were performed using a web-based network sensor composed of striped and interdigitated capacitive sensors. Hydroxyapatite-based layers have a minor effect on determining the fixation states; the effective operation of a sensor network-based solution communicating through a web server hosted on Raspberry Pi was shown. Previous studies highlight the inability of current bone-implant fixation monitoring methods to significantly reduce the number of revision surgeries, as well as promising results of capacitive sensing systems to monitor micro-scale and macro-scale bone-interface states. In this study, we found that extracorporeal informatic systems enable continuous patient monitoring using cosurface capacitive networks with or without hydroxyapatite-based layers. Findings presented here represent significant advancements toward the design of future multifunctional smart implants.

Effects of graphene oxide nanosheets in the polychaete Hediste diversicolor: Behavioural, physiological and biochemical responses.

Numerous applications exist for graphene-based materials, such as graphene oxide (GO) nanosheets. Increased concentrations of GO nanosheets in the environment have the potential to have a large negative effect on the aquatic environment, with consequences for benthic organisms, such as polychaetes. The polychaete Hediste diversicolor mobilises the sediments, hence altering the availability of contaminants and the nutrients biogeochemical cycle. As such, this study proposes to assess the effects of different GO nanosheet concentrations on the behaviour, feeding activity, mucus production, regenerative capacity, antioxidant status, biochemical damage and metabolism of H. diversicolor. This study evidenced that H. diversicolor exposed to GO nanosheets had a significantly lower ability to regenerate their bodies, took longer to feed and burrow into the sediment and produced more mucus. Membrane oxidative damage (lipid peroxidation) increased in exposed specimens. The increased metabolic rate (ETS) evidenced a higher energy expenditure in exposed organisms (high use of ready energy sources - soluble sugars) to fight the toxicity induced by GO nanosheets, such as SOD activity. The increase in SOD activity was enough to reduce reactive oxygen species (ROS) induced by GO on cytosol at the lowest concentrations, avoiding the damage on proteins (lower PC levels), but not on membranes (LPO increase). This study revealed that the presence of GO nanosheets, even at the lower levels tested, impaired behavioural, physiological, and biochemical traits in polychaetes, suggesting that the increase of this engineered nanomaterial in the environment can disturb these benthic organisms, affecting the H. diversicolor population. Moreover, given the important role of this group of organisms in coastal and estuarine food webs, the biogeochemical cycle of nutrients, and sediment oxygenation, there is a real possibility for repercussions into the estuarine community.

The influence of salinity on the toxicity of remediated seawater.

Mercury (Hg) is one of the most hazardous pollutants, due to its toxicity, biological magnification and worldwide persistence in aquatic systems. Thus, new efficient nanotechnologies (e.g. graphene oxide functionalized with polyethyleneimine (GO-PEI)) have been developed to remove this metal from the water. Aquatic environments, in particular transitional systems, are also subjected to disturbances resulting from climate change, such as salinity shifts. Salinity is one of the most relevant factors that influences the distribution and survival of aquatic species such as mussels. To our knowledge, no studies assessed the ecotoxicological impairments induced in marine organisms exposed to remediate seawater (RSW) under different salinity levels. For this, the focus of the present study was to evaluate the effects of seawater previously contaminated with Hg and remediated with GO-PEI, using the species Mytilus galloprovincialis, maintained at three different salinities (30, 20 and 40). The results obtained demonstrated similar histopathological and metabolic alterations, oxidative stress and neurotoxicity in mussels under RSW treatment at stressful salinity conditions (20 and 40) in comparison to control salinity (30). On the other hand, the present findings revealed toxicological effects including cellular damage and histopathological impairments in mussels exposed to Hg contaminated seawater in comparison to non-contaminated ones, at each salinity level. Overall, these results confirm the high efficiency of GO-PEI to sorb Hg from water with no noticeable toxic effects even under different salinities, leading to consider it a promising eco-friendly approach to remediate contaminated water.

Biomimetic Graphene/Spongin Scaffolds for Improved Osteoblasts Bioactivity via Dynamic Mechanical Stimulation.

Biomimetics offers excellent prospects for design a novel generation of improved biomaterials. Here the controlled integration of graphene oxide (GO) derivatives with a 3D marine spongin (MS) network is explored to nanoengineer novel smart bio-based constructs for bone tissue engineering. The results point out that 3D MS surfaces can be homogeneously coated by layer-by-layer (LbL) assembly of oppositely charged polyethyleneimine (PEI) and GO. Notably, the GOPEI@MS bionanocomposites present a high structural and mechanical stability under compression tests in wet conditions (shape memory). Dynamic mechanically (2 h of sinusoidal compression cyclic interval (0.5 Hz, 0-10% strain)/14 d) stimulates GOPEI@MS seeded with osteoblast (MC3T3-E1), shows a significant improvement in bioactivity, with cell proliferation being two times higher than under static conditions. Besides, the dynamic assays show that GOPEI@MS bionanocomposites are able to act as mechanical stimulus-responsive scaffolds able to resemble physiological bone extracellular matrix (ECM) requirements by strongly triggering mineralization of the bone matrix. These results prove that the environment created by the system cell-GOPEI@MS is suitable for controlling the mechanisms regulating mechanical stimulation-induced cell proliferation for potential in vivo experimentation.

Multi-layered electrospinning and electrospraying approach: Effect of polymeric supplements on chondrocyte suspension.

Articular cartilage was expected to be one of the first tissues to be successfully engineered, but replicating the complex fibril architecture and the cellular distribution of the native cartilage has proven difficult. While electrospinning has been widely used to reproduce the depth-dependent fibre architecture in 3D scaffolds, the chondrocyte-controlled distribution remains an unsolved problem. To incorporate cells homogeneously through the depth of scaffolds, a combination of polymer electrospinning and cell seeding is necessary. A multi-layer approach alternating between polymer electrospinning with chondrocyte electrospraying can be a solution. Still, the success of this process is related to the survival rate of the electrosprayed chondrocytes embedded within the electrospun mesh. In this regard, the present study investigated the impact of the multi-layered process and the supplementation of the electrospray chondrocyte suspension with different concentrations of Gelatin and Alginate on the viability of electrosprayed chondrocytes embedded within a Polycaprolactone/Gelatin electrospun mesh and on the mechanical properties of the resulting meshes. The addition of Gelatin in the chondrocyte suspension did not increase significantly (p > 0.05) the percentage of viable electrosprayed chondrocytes (25%), while 3 wt% Alginate addition led to a significant (p < 0.05) increase in chondrocyte viability (50%) relative to the case without polymer supplement (15%). Furthermore, the addition of both polymer supplements increased the mechanical properties of the multi-layer construct. These findings imply that this multi-layered approach can be applied to cartilage TE allowing for automated chondrocyte integration during scaffolds creation.

The Influence of Temperature Increase on the Toxicity of Mercury Remediated Seawater Using the Nanomaterial Graphene Oxide on the Mussel Mytilus galloprovincialis.

Mercury (Hg) has been increasing in waters, sediments, soils and air, as a result of natural events and anthropogenic activities. In aquatic environments, especially marine systems (estuaries and lagoons), Hg is easily bioavailable and accumulated by aquatic wildlife, namely bivalves, due to their lifestyle characteristics (sedentary and filter-feeding behavior). In recent years, different approaches have been developed with the objective of removing metal(loid)s from the water, including the employment of nanomaterials. However, coastal systems and marine organisms are not exclusively challenged by pollutants but also by climate changes such as progressive temperature increment. Therefore, the present study aimed to (i) evaluate the toxicity of remediated seawater, previously contaminated by Hg (50 mg/L) and decontaminated by the use of graphene-based nanomaterials (graphene oxide (GO) functionalized with polyethyleneimine, 10 mg/L), towards the mussel Mytilus galloprovincialis; (ii) assess the influence of temperature on the toxicity of decontaminated seawater. For this, alterations observed in mussels' metabolic capacity, oxidative and neurotoxic status, as well as histopathological injuries in gills and digestive tubules were measured. This study demonstrated that mussels exposed to Hg contaminated seawater presented higher impacts than organisms under remediated seawater. When comparing the impacts at 21 °C (present study) and 17 °C (previously published data), organisms exposed to remediated seawater at a higher temperature presented higher injuries than organisms at 17 °C. These results indicate that predicted warming conditions may negatively affect effective remediation processes, with the increasing of temperature being responsible for changes in organisms' sensitivity to pollutants or increasing pollutants toxicity.

Benefits in the Macrophage Response Due to Graphene Oxide Reduction by Thermal Treatment.

Graphene and its derivatives are very promising nanomaterials for biomedical applications and are proving to be very useful for the preparation of scaffolds for tissue repair. The response of immune cells to these graphene-based materials (GBM) appears to be critical in promoting regeneration, thus, the study of this response is essential before they are used to prepare any type of scaffold. Another relevant factor is the variability of the GBM surface chemistry, namely the type and quantity of oxygen functional groups, which may have an important effect on cell behavior. The response of RAW-264.7 macrophages to graphene oxide (GO) and two types of reduced GO, rGO15 and rGO30, obtained after vacuum-assisted thermal treatment of 15 and 30 min, respectively, was evaluated by analyzing the uptake of these nanostructures, the intracellular content of reactive oxygen species, and specific markers of the proinflammatory M1 phenotype, such as CD80 expression and secretion of inflammatory cytokines TNF-α and IL-6. Our results demonstrate that GO reduction resulted in a decrease of both oxidative stress and proinflammatory cytokine secretion, significantly improving its biocompatibility and potential for the preparation of 3D scaffolds able of triggering the appropriate immune response for tissue regeneration.

How efficient is graphene-based nanocomposite to adsorb Hg from seawater. A laboratory assay to assess the toxicological impacts induced by remediated water towards marine bivalves.

Advanced investigations on the use of graphene based nanomaterials have highlighted the capacity of these materials for wastewater treatment. Research on this topic revealed the efficiency of the nanocomposite synthetized by graphene oxide functionalized with polyethyleneimine (GO-PEI) to adsorb mercury (Hg) from contaminated seawater. However, information on the environmental risks associated with these approaches are still lacking. The focus of this study was to evaluate the effects of Hg in contaminated seawater and seawater remediated by GO-PEI, using the species Ruditapes philippinarum, maintained at two different warming scenarios: control (17 °C) and increased (22 °C) temperatures. The results obtained showed that organisms exposed to non-contaminated and remediated seawaters at control temperature presented similar biological patterns, with no considerable differences expressed in terms of biochemical and histopathological alterations. Moreover, the present findings revealed increased toxicological effects in clams under remediated seawater at 22 °C in comparison to those subjected to the equivalent treatment at 17 °C. These results confirm the capability of GO-PEI to adsorb Hg from water with no noticeable toxic effects, although temperature could alter the responses of mussels to remediated seawater. These materials seem to be a promise eco-friendly approach to remediate wastewater, with low toxicity evidenced by remediated seawater and high regenerative capacity of this nanomaterial, keeping its high removal performance after successive sorption-desorption cycles.

A Multifactorial Approach to Untangle Graphene Oxide (GO) Nanosheets Effects on Plants: Plant Growth-Promoting Bacteria Inoculation, Bacterial Survival, and Drought.

Drought is a limiting factor for agricultural productivity. Climate change threatens to expand the areas of the globe subjected to drought, as well as to increase the severity and duration of water shortage. Plant growth-promoting bacteria (PGPB) are widely studied and applied as biostimulants to increase plant production and to enhance tolerance to abiotic and biotic constraints. Besides PGPB, studies on the potential of nanoparticles to be used as biostimulants are also thriving. However, many studies report toxicity of tested nanoparticles in bacteria and plants in laboratory conditions, but few studies have reported effects of nanoparticles towards bacterial cells and communities in the soil. The combined application of nanoparticles and PGPB as biostimulant formulations are poorly explored and it is important to unravel the potentialities of their combined application as a way to potentiate food production. In this study, Rhizobium sp. E20-8 and graphene oxide (GO) nanosheets were applied on container-grown maize seedlings in watered and drought conditions. Bacterial survival, seedling growth (dry weight), and biochemical endpoints (photosynthetic pigments, soluble and insoluble carbohydrates, proline, lipid peroxidation, protein, electron transport system, and superoxide dismutase) were evaluated. Results showed that the simultaneous exposure to GO and Rhizobium sp. E20-8 was able to alleviate the stress induced by drought on maize seedlings through osmotic and antioxidant protection by GO and mitigation of GO effects on the plant's biochemistry by Rhizobium sp. E20-8. These results constitute a new lead on the development of biostimulant formulations to improve plant performance and increase food production in water-limited conditions.

Boosting in vitro cartilage tissue engineering through the fabrication of polycaprolactone-gelatin 3D scaffolds with specific depth-dependent fiber alignments and mechanical stimulation.

Due to the limited self-healing ability of natural cartilage, several tissue engineering strategies have been explored to develop functional replacements. Still, most of these approaches do not attempt to recreate in vitro the anisotropic organization of its extracellular matrix, which is essential for a suitable load-bearing function. In this work, different depth-dependent alignments of polycaprolactone-gelatin electrospun fibers were assembled into three-dimensional scaffold architectures to assess variations on chondrocyte response under static, unconfined compressed and perfused culture conditions. The in vitro results confirmed that not only the 3D scaffolds specific depth-dependent fiber alignments potentiated chondrocyte proliferation and migration towards the fibrous systems, but also the mechanical stimulation protocols applied were able to enhance significantly cell metabolic activity and extracellular matrix deposition, respectively.