Self-Supporting Hyaluronic Acid-Functionalized G-Quadruplex-Based Perfusable Multicomponent Hydrogels Embedded in Photo-Cross-Linkable Matrices for Bioapplications.

Dynamic G-quadruplex supramolecular hydrogels have aroused great interest in a broad range of bioapplications. However, neither the development of native extracellular matrix (ECM)-derived natural biopolymer-functionalized G-quadruplex hydrogels nor their use to create perfusable self-supporting hydrogels has been explored to date, despite their intrinsic potential as carrier vehicles of therapeutic agents, or even living cells in advanced regenerative therapies, or as platforms to enable the diffusion of nutrients and oxygen to sustain long-term cell survival. Herein, we developed a dynamic co-assembling multicomponent system that integrates guanosine (G), 3-aminophenylboronic acid functionalized hyaluronic acid (HA-PBA), and potassium chloride to bioengineer strong, homogeneous, and transparent HA-functionalized G-quadruplex hydrogels with injectable, thermo-reversible, conductive, and self-healing properties. The supramolecular polymeric hydrogels were developed by hydrogen bonding and π-π stacking interactions between G coupled via dynamic covalent boronate ester bonds to HA-PBA and stabilized by K+ ions, as demonstrated by a combination of experiments and molecular dynamics simulations. The intrinsic instability of the self-assembled G-quadruplex structures was used to bioengineer self-supporting perfusable multicomponent hydrogels with interconnected size and shape-tunable hollow microchannels when embedded in 3D methacrylated gelatin supporting matrices. The microchannel-embedded 3D constructs have shown enhanced cell viability when compared to the bulk hydrogels, holding great promise for being use as artificial vessels for enabling the diffusion of nutrients and oxygen essential for cell survival. The proposed approach opens new avenues on the use of ECM-derived natural biopolymer-functionalized dynamic G-quadruplex hydrogels to design next-generation smart systems for being used in tissue regeneration, drug screening, or organ-on-a-chip.

New nitroindazole-porphyrin conjugates: Synthesis, characterization and antibacterial properties.

The synthesis of new porphyrin-indazole hybrids by a Knoevenagel condensation of 2-formyl-5,10,15,20-tetraphenylporphyrin and N-methyl-nitroindazolylacetonitrile derivatives is reported. The target compounds were isolated in moderate to good yields (32-57%) and some of the isolated porphyrin-indazole conjugates showed good performance in the generation of singlet oxygen when irradiated with visible light. Their efficiency as photosensitizers in the photoinactivation of methicillin resistant Staphylococcus aureus-MRSA was evaluated. All derivatives showed to be able to photoinactivate the MRSA bacteria. Compound 3a appears to be the most promising photosensitiser (PS) in the photoinactivation of these bacteria, despite being the least efficient in singlet oxygen generation. The addition of potassium iodide (KI) significantly potentiated the antimicrobial Photodynamic Therapy (aPDT) process mediated by all the analysed porphyrin-indazole conjugates. The combined action of nitroindazole-porphyrins with potassium iodide (KI) action appears to be promising in the photoinactivation of MRSA.

Neuronal deletion of GSK3ß increases microtubule speed in the growth cone and enhances axon regeneration via CRMP-2 and independently of MAP1B and CLASP2.

BACKGROUND: In the adult central nervous system, axonal regeneration is abortive. Regulators of microtubule dynamics have emerged as attractive targets to promote axonal growth following injury as microtubule organization is pivotal for growth cone formation. In this study, we used conditioned neurons with high regenerative capacity to further dissect cytoskeletal mechanisms that might be involved in the gain of intrinsic axon growth capacity. RESULTS: Following a phospho-site broad signaling pathway screen, we found that in conditioned neurons with high regenerative capacity, decreased glycogen synthase kinase 3ß (GSK3ß) activity and increased microtubule growth speed in the growth cone were present. To investigate the importance of GSK3ß regulation during axonal regeneration in vivo, we used three genetic mouse models with high, intermediate or no GSK3ß activity in neurons. Following spinal cord injury, reduced GSK3ß levels or complete neuronal deletion of GSK3ß led to increased growth cone microtubule growth speed and promoted axon regeneration. While several microtubule-interacting proteins are GSK3ß substrates, phospho-mimetic collapsin response mediator protein 2 (T/D-CRMP-2) was sufficient to decrease microtubule growth speed and neurite outgrowth of conditioned neurons and of GSK3ß-depleted neurons, prevailing over the effect of decreased levels of phosphorylated microtubule-associated protein 1B (MAP1B) and through a mechanism unrelated to decreased levels of phosphorylated cytoplasmic linker associated protein 2 (CLASP2). In addition, phospho-resistant T/A-CRMP-2 counteracted the inhibitory myelin effect on neurite growth, further supporting the GSK3ß-CRMP-2 relevance during axon regeneration. CONCLUSIONS: Our work shows that increased microtubule growth speed in the growth cone is present in conditions of increased axonal growth, and is achieved following inactivation of the GSK3ß-CRMP-2 pathway, enhancing axon regeneration through the glial scar. In this context, our results support that a precise control of microtubule dynamics, specifically in the growth cone, is required to optimize axon regrowth.

Early axonal loss accompanied by impaired endocytosis, abnormal axonal transport, and decreased microtubule stability occur in the model of Krabbe's disease.

In Krabbe's disease (KD), a leukodystrophy caused by ß-galactosylceramidase deficiency, demyelination and a myelin-independent axonopathy contributes to the severe neuropathology. Beyond axonopathy, we show that in Twitcher mice, a model of KD, a decreased number of axons both in the PNS and in the CNS, and of neurons in dorsal root ganglia (DRG), occurred before the onset of demyelination. Despite the early axonal loss, and although in vitro Twitcher neurites degenerated over time, Twitcher DRG neurons displayed an initial neurite overgrowth and, following sciatic nerve injury, Twitcher axons were regeneration-competent, at a time point where axonopathy was already ongoing. Psychosine, the toxic substrate that accumulates in KD, induced lipid raft clustering. At the mechanistic level, TrkA recruitment to lipid rafts was dysregulated in Twitcher neurons, and defective activation of the ERK1/2 and AKT pathways was identified. Besides defective recruitment of signaling molecules to lipid rafts, the early steps of endocytosis and the transport of endocytic and synaptic vesicles were impaired in Twitcher DRG neurons. Defects in axonal transport, specifically in the retrograde component, correlated with decreased levels of dynein, abnormal levels of post-translational tubulin modifications and decreased microtubule stability. The identification of the axonal defects that precede demyelination in KD, together with the finding that Twitcher axons are regeneration-competent when axonopathy is already installed, opens new windows of action to effectively correct the neuropathology that characterizes this disorder.

The importance of ether-phospholipids: a view from the perspective of mouse models.

Ether-phospholipids represent an important group of phospholipids characterized by an alkyl or an alkenyl bond at the sn-1 position of the glycerol backbone. Plasmalogens are the most abundant form of alkenyl-glycerophospholipids, and their synthesis requires functional peroxisomes. Defects in the biosynthesis of plasmalogens are the biochemical hallmark of the human peroxisomal disorder Rhizomelic Chondrodysplasia Punctata (RCDP), which is characterized by defects in eye, bone and nervous tissue. The generation and characterization of mouse models with defects in plasmalogen levels have significantly advanced our understanding of the role and importance of plasmalogens as well as pathogenetic mechanisms underlying RCDP. A review of the current mouse models and the description of the combined knowledge gathered from the histopathological and biochemical studies is presented and discussed. Further characterization of the role and functions of plasmalogens will contribute to the elucidation of disease pathogenesis in peroxisomal and non-peroxisomal disorders. This article is part of a Special Issue entitled: Metabolic Functions and Biogenesis of Peroxisomes in Health and Disease.

Total fishmeal replacement by defatted Tenebrio molitor larvae meal induces alterations in intermediary metabolism of European sea bass (Dicentrarchus labrax).

The replacement of fishmeal (FM) by insect meal (IM) in aquafeed formulation has been thoroughly studied lately, but little is known about their impact on nutrient metabolism of fish. This study evaluated the impact not only of partial but also total FM replacement by IM on intermediary metabolism of European sea bass (Dicentrarchus labrax). A fishmeal-based diet was used as a control (CTRL) and two other diets were formulated to include 20% and 40% of defatted Tenebrio molitor larvae meal (dTM), replacing 50% (TM50) and 100% (TM100) of fishmeal (FM), respectively. After a 16-week feeding trial, a multidisciplinary approach including assessment of histological, biochemical, molecular, and enzymatic parameters was adopted to investigate hepatic and plasmatic responses to the different dietary formulations. The results obtained demonstrated that dTM can be successfully used to replace 50% of FM in diets for European sea bass, without adversely affecting liver health or intermediary metabolism of nutrients. As for TM100, although no signs of steatosis were observed in the liver, the activity of glycolytic and lipogenic genes and enzymes increased when compared to CTRL diet (P < 0.05), resulting in higher levels of plasmatic non-esterified fatty acids and triacylglycerides (P < 0.05), which in the long-term may compromise fish health, thus precluding such a high degree of substitution for use in practical diets for European sea bass.

Insect meal has been increasingly considered as novel protein source in diets for different animal models, including fish. European sea bass is one of the most important fish species in Europe, particularly in the Mediterranean. Fishmeal is considered a premium protein source in diets for aquaculture fish. Thus, this study evaluated the impact of partial and total fishmeal replacement by an insect meal (defatted Tenebrio molitor larvae meal) on intermediary metabolism of European sea bass. After 16 weeks of feeding, the results obtained demonstrated that selected insect meal can be successfully used to replace 50% of fish in diets for European sea bass, without adversely affecting liver health or intermediary metabolism of nutrients. On the other hand, total fishmeal replacement by insect meal may compromise fish health in the long-term.

Malignant peripheral nerve sheath tumor mimicking an adnexal mass: a radio-pathologic correlation.

We report the case of a pelvic malignant peripheral nerve sheath tumor mimicking an adnexal mass. A 59-year-old postmenopausal woman presented with a 3-month history of diffuse abdominal bloating and urinary frequency. Laboratory tests revealed an increased CA 125. Radiologic evaluation depicted a large, heterogeneous solid mass located right to the uterus, pushing it to the left. After a multidisciplinary board discussion, the diagnosis of a right adnexal lesion was assumed, and the patient was referred to surgery. The final diagnosis was only achieved after pathology examination, which prove to be a malignant peripheral nerve sheath tumor. This paper highlights some clinical, radiologic and pathological features of malignant peripheral nerve sheath tumors, a rare entity that should be considered as a differential in patients presenting with pelvic tumors of uncertain origin.

An in-depth characterisation of European seabass intestinal segments for assessing the impact of an algae-based functional diet on intestinal health.

Sustainable farming of fish species depends on emerging new feed ingredients, which can alter the features of the digestive tract and influence animals' overall health. Recent research has shown that functional feeds hold great potential for enhancing fish robustness by evoking appropriate responses at the intestine level. However, there is a lack of extensive and accurate descriptions of the morphology of the gastrointestinal tract of most farmed fish. We have characterised the intestine of European seabass thoroughly, by targeting four segments - anterior, mid, posterior and rectum. Results indicated that the anterior segment is mostly associated with absorption-related features; this segment has the largest absorptive area, the longest villi, and the highest number of neutral goblet cells (GC). The posterior segment and rectum have distinct histomorphometric features, but both seem to be important for immunity, displaying the highest count of acid GC and the highest expression of immune-related genes. The strongest proliferating cell nuclear antigen (PCNA) signal was observed in the anterior intestine and rectum, with PCNA+ cells appearing at the base of the villi and the corresponding villi branches. We have also evaluated the impact of a novel feed supplemented with a macro- and microalgae blend and found that there were no differences in terms of growth. However, the alterations observed in the mid intestine of fish fed the blend, such as thickening of the submucosa and lamina propria, an increased number of leucocytes, and higher expression of immune- and oxidative stress-related genes, suggest that algae may have an immunomodulatory effect. In the current article, we have described the morphology and expression patterns of the intestine segments of European seabass in detail and have presented a comprehensive report of the indices and methods used for the semi-quantitative and quantitative histomorphometric assessments, thereby providing useful information for future studies that aim to maintain intestinal health through dietary interventions.

Systemic delivery of bone marrow-derived mesenchymal stromal cells diminishes neuropathology in a mouse model of Krabbe's disease.

In Krabbe's disease, a demyelinating disorder, add-on strategies targeting the peripheral nervous system (PNS) are needed, as it is not corrected by bone-marrow (BM) transplantation. To circumvent this limitation of BM transplantation, we assessed whether i.v. delivery of immortalized EGFP(+) BM-derived murine mesenchymal stromal cells (BM-MSC(TERT-EGFP) ) targets the PNS of a Krabbe's disease model, the Twitcher mouse. In vitro, BM-MSC(TERT-EGFP) retained the phenotype of primary BM-MSC and did not originate tumors upon transplantation in nude mice. In vivo, undifferentiated EGFP(+) cells grafted the Twitcher sciatic nerve where an increase in Schwann cell precursors and axonal number was detected. The same effect was observed on BM-MSC(TERT-EGFP) i.v. delivery following sciatic nerve crush, a model of axonal regeneration. Reiterating the in vivo findings, in a coculture system, BM-MSC(TERT-EGFP) induced the proliferation of Twitcher-derived Schwann cells and the neurite outgrowth of both Twitcher-derived neurons and wild-type neurons grown in the presence of psychosine, the toxic substrate that accumulates in Krabbe's disease. In vitro, this neuritogenic effect was blocked by K252a, an antagonist of Trk receptors, and by antibody blockage of brain derived neurotrophic factor, a neurotrophin secreted by BM-MSC(TERT-EGFP) and induced in neighboring Schwann cells. In vivo, BM-MSC(TERT-EGFP) surmounted the effect of K252a, indicating their ability to act through a neurotrophin-independent mechanism. In summary, i.v. delivery of BM-MSC(TERT-EGFP) exerts a multilevel effect targeting neurons and Schwann cells, coordinately diminishing neuropathology. Therefore, to specifically target the PNS, MSC should be considered an add-on option to BM transplantation in Krabbe's disease and in other disorders where peripheral axonal loss occurs.

Processed by-Products from Soy Beverage (Okara) as Sustainable Ingredients for Nile Tilapia (O. Niloticus) Juveniles: Effects on Nutrient Utilization and Muscle Quality.

The apparent digestibility coefficients (ADCs) of differently processed okara meals were assessed in Nile tilapia diets: dried okara not autoclaved (FOK), dried okara autoclaved (AOK), okara hydrolyzed with Alcalase (ALOK) or Cynara cardunculus proteases (CYOK), and hydrolyzed okara fermented with lactic bacteria: Lactobacillus rhamnosus R11 (CYR11OK) or Bifidobacterium animalis ssp. lactis Bb12 (CYB12OK). Okara processing significantly affected nutrient digestibility: dry matter ADC was highest in CYR11OK (80%) and lowest in FOK (40%). The lowest protein digestibility was observed in CYR11OK (72%), and the highest in AOK (97%) and CYOK (91%), evidencing the effectiveness of the autoclave and the use of C. Cardunculus proteases to increase okara protein bioavailability. The inclusion of up to 20% of AOK or CYOK did not affect fish growth, nutrient utilization, or whole body composition of Nile tilapia. The flesh quality (color, pH, water activity, cohesiveness, elasticity and resilience) was not affected by the dietary incorporation of AOK or CYOK. Fish fed with AOK diets stand out for their high density of muscle fibers, particularly in AOK20, which can explain their high muscle firmness and may result in further hypertrophic growth. Altogether, results suggest that hydrolyzed or autoclaved okara are valuable ingredients for Nile tilapia diets.

Early-onset Alzheimer disease: the contribution of neuroimaging for the diagnosis.

The diagnosis of Alzheimer disease (AD) at an early age of onset may be a challenging task. The diagnosis of such a type of dementia may be even more difficult when concomitant depressive symptoms occur. We report the case of a 51-year-old woman who was admitted at a Psychiatric Day Hospital presenting with depressive symptoms, visuospatial deficits, apraxia, and minor memory loss. The patient underwent long-term antidepressant therapy, but despite the improvement of depressive symptoms, there was progressive cognitive deterioration. Otherwise, the prior clinical history was unremarkable, and there was no family history of dementia. The clinical examination revealed cognitive deficits in several domains. The patient scored 12 in the Mini-Mental State Examination. Routine laboratory tests were normal. Magnetic resonance (MR) imaging showed global brain volume loss more pronounced than would be expected for someone of the patient's age, especially in the precuneus-a pattern of posterior cortical atrophy consistent with the diagnosis of early-onset AD. Images obtained with 99mTc-HMPAO single-photon emission computed tomography (SPECT) also revealed marked brain hypoperfusion involving the left parietal lobe, far beyond the regions of brain volume loss. This clinical case report illustrates the relative contribution of both MR imaging and SPECT for the diagnosis of dementia in a patient with concomitant depressive symptoms. Apart from contributing to the diagnosis of dementia beyond the traditional exclusionary approach, neuroimaging is increasingly being used to classify its particular subtypes. The role of neuroimaging role in AD is also supported by a recent proposal of revised diagnostic criteria, which take into account abnormal biomarkers of disease.

Growth performance, bioavailability of toxic and essential elements and nutrients, and biofortification of iodine of rainbow trout (Onchorynchus mykiss) fed blends with sugar kelp (Saccharina latissima).

Aquaculture production is demanding novel feed ingredients that reflect natural marine nutrient levels, that are also essential to humans. In this regard, biofortification through addition of iodine-rich sugar kelp in feed formulations was assessed in a 12 week rainbow trout trial. Yttrium inclusion in feed allowed determinations of apparent absorption coefficients of essential and potentially toxic elements and apparent digestibility coefficient of nutrients. E.g. apparent absorption coefficients in trouts fortified feed with 1-4% dw kelp were 67-61% As, 32-40% Cd, <5% Fe; 80-83% I; 66-58% Se. Iodine concentrations in feed up to 239 mg/kg (~4% kelp) was proportional to iodine accumulation in trout fillets (R2 = 1.00) with 0.5% transfer ratio. Feed iodine concentrations up to 117 mg/kg (~2% kelp) did not affect growth performance negatively, but increased significantly protein efficiency ratio after eight weeks feeding. However, 4% kelp meal inclusion affected final growth and hepato somatic index, and caused histomorphological changes in the intestine. All fillets had low toxic element concentrations (As, Cd, Hg, Pb). The potential applicability of Saccharina latissima as feed ingredient to tailor iodine concentration in farmed fish is evident. Consuming of a 160 g fillet (2% kelp) contributes ~60% of recommended daily iodine intake for adults.

Dietary Creatine Supplementation in Gilthead Seabream (Sparus aurata) Increases Dorsal Muscle Area and the Expression of myod1 and capn1 Genes.

Creatine (Cr) is an amino acid derivative with an important role in the cell as energy buffer that has been largely used as dietary supplement to increase muscle strength and lean body mass in healthy individuals and athletes. However, studies in fish are scarce. The aim of this work is to determine whether dietary Cr supplementation affects muscle growth in gilthead seabream (Sparus aurata) juveniles. Fish were fed ad libitum for 69 days with diets containing three increasing levels of creatine monohydrate (2, 5, and 8%) that were compared with a non-supplemented control (CTRL) diet. At the end of the trial, the fast-twist skeletal muscle growth dynamics (muscle cellularity) and the expression of muscle-related genes were evaluated. There was a general trend for Cr-fed fish to be larger and longer than those fed the CTRL, but no significant differences in daily growth index (DGI) were registered among dietary treatments. The dorsal cross-sectional muscle area (DMA) of fish fed Cr 5 and Cr 8% was significantly larger than that of fish fed CTRL. The groups supplemented with Cr systematically had a higher relative number of both small-sized (≤20 µm) and large-sized fibers (≥120 µm). Dorsal total fibers number was highest in fish fed 5% Cr. In fish supplemented with 5% Cr, the relative expression of myogenic differentiation 1 (myod1) increased almost four times compared to those fed the CTRL diet. The relative expression of calpain 3 (capn3) was highest in fish fed diets with 2% Cr supplementation, but did not differ significantly from those fed the CTRL or Cr 5%. The myod1 gene expression had a positive and significant correlation with that of capn1, capns1a, and capn3 expression. These results suggest that the observed modulation of gene expression was not enough to produce a significant alteration in muscle phenotype under the tested conditions, as a non-significant increase in muscle fiber diameter and higher total number of fiber was observed, but still resulted in increased DMA. Additional studies may be required in order to better clarify the effect of dietary Cr supplementation in fish, possibly in conjunction with induced resistance training.

Photodynamic Inactivation of Candida albicans in Blood Plasma and Whole Blood.

The few approved disinfection techniques for blood derivatives promote damage in the blood components, representing risks for the transfusion receptor. Antimicrobial photodynamic therapy (aPDT) seems to be a promising approach for the photoinactivation of pathogens in blood, but only three photosensitizers (PSs) have been approved, methylene blue (MB) for plasma and riboflavin and amotosalen for plasma and platelets. In this study, the efficiency of the porphyrinic photosensitizer Tri-Py(+)-Me and of the porphyrinic formulation FORM was studied in the photoinactivation of Candida albicans in plasma and in whole blood and the results were compared to the ones obtained with the already approved PS MB. The results show that FORM and Tri-Py(+)-Me are promising PSs to inactivate C. albicans in plasma. Although in whole blood the inactivation rates obtained were higher than the ones obtained with MB, further improvements are required. None of these PSs had promoted hemolysis at the isotonic conditions when hemolysis was evaluated in whole blood and after the addition of treated plasma with these PSs to concentrates of red blood cells.

A ação educativa realizada pelo assistente social no grupo de primeiro atendimento em um hospital de alta complexidade / The educational action carried out by the social worker in the first care group in a high complexity hospital

INTRODUÇÃO: O presente estudo refere-se à dimensão da ação socioeducativa realizada pelos assistentes sociais no "grupo de atendimento aos pacientes de primeira consulta" em um hospital terciário especializado em cardiologia no Estado de São Paulo. Buscou-se analisar a contribuição da ação educativa do grupo de primeiro atendimento para assim aprimorar a ação educativa socializando a garantia de acessos aos direitos dos usuários do Sistema Único de Saúde (SUS). OBJETIVO: Analisar a contribuição da ação educativa do grupo de primeiro atendimento em um hospital de alta complexidade realizada pela equipe do Serviço Social. MÉTODOS: Trata-se de uma pesquisa de cunho qualitativo/quantitativa com total de 36 participantes: (23) pacientes (13) funcionários da instituição. Os critérios para a participação foram: pacientes que participaram do grupo e funcionários envolvidos no processo do grupo de primeiro atendimento. Trata-se de um estudo descritivo, exploratório e prospectivo que se valeu de uma abordagem individual. RESULTADOS: As respostas desta pesquisa foram divididas em quatro grupos (pacientes, médicos, assistentes sociais e equipe do Serviço de Arquivo Médico e Estatístico - SAME) todos envolvidos com o grupo de primeiro atendimento. Vale ressaltar que 83% dos pacientes entrevistados informaram estar muito satisfeitos ao participar do grupo de primeiro atendimento. O que se revelou também sobre a orientação a respeito da Lei N.10.241 sobre os direitos dos usuários dos serviços de saúde no Estado de São Paulo, foi o tema considerado de maior satisfação pelos pacientes em 73%, este tema em específico traz uma breve abordagem sobre cidadania, humanização e a lei que respalda os usuários do Sistema Único de Saúde, com o intuito de incentivar o paciente a ser o principal protagonista do seu tratamento de saúde e levar ao conhecimento de que o mesmo possui leis de proteção social. DISCUSSÃO: A pesquisa possibilitou identificar aspectos relevantes sobre a efetividade da ação educativa no que se refere ao acolhimento e acesso aos direitos. CONCLUSÃO: A referida ação educativa contribui para a viabilização do acesso aos direitos sociais, favorece o empoderamento no exercício da cidadania e fortalece a autonomia dos pacientes em tratamento da saúde cardiovascular. Palavras-Chaves: Serviço Social; Ação educativa; Acolhimento; Educação em Saúde.

Microencapsulation of Gulosibacter molinativorax ON4T cells by a spray-drying process using different biopolymers.

Molinate is a thiocarbamate herbicide used in rice crop protection. As other pesticides, molinate is a recognized environmental pollutant and bio-accumulated by some wildlife forms. Gulosibacter molinativorax ON4T is able to hydrolyse molinate into metabolites which are further degraded by other un-related bacteria. Hence, it can be used in molinate bioremediation processes. The aim of this work was to investigate the possibility of producing G. molinativorax ON4T microparticles, using different non-toxic biopolymers (arabic gum, modified chitosan, calcium alginate and sodium alginate) as encapsulating agents by a spray-drying process. Several formulations of microparticles were prepared, and their physicochemical structures were analyzed by scanning electron microscopy (SEM), laser granulometry analysis and zeta potential analysis. The obtained microparticles were evaluated considering their ability to degrade molinate, the metabolic activity (by colour development of the tetrazolium violet redox), and also the survival rate and shelf-life/storage stability of microparticles. Based on their molinate degrading activity, the biopolymers calcium alginate and modified chitosan cross-linked with tripolyphosphate appear to be the best options for the microencapsulation of the G. molinativorax ON4T. However, the microparticles produced with modified chitosan cross-linked with tripolyphosphate present the best combination of physical properties and activity degradation of molinate.

Effects of different rearing temperatures on muscle development and stress response in the early larval stages of Acipenser baerii.

The present study aims at investigating muscle development and stress response in early stages of Siberian sturgeon when subjected to different rearing temperatures, by analysing growth and development of the muscle and by assessing the stress response of yolk-sac larvae. Siberian sturgeon larvae were reared at 16°C, 19°C and 22°C until the yolk-sac was completely absorbed. Sampling timepoints were: hatching, schooling and complete yolk-sac absorption stage. Histometrical, histochemical and immunohistochemical analyses were performed in order to characterize muscle growth (total muscle area, TMA; slow muscle area, SMA; fast muscle area, FMA), development (anti-proliferating cell nuclear antigen -PCNA or anticaspase) as well as stress conditions by specific stress biomarkers (heat shock protein 70 or 90, HSP70 or HSP90). Larvae subjected to the highest water temperature showed a faster yolk-sac absorption. Histometry revealed that both TMA and FMA were larger in the schooling stage at 19°C while no differences were observed in the SMA at any of the tested rearing temperatures. PCNA quantification revealed a significantly higher number of proliferating cells in the yolk-sac absorption phase at 22°C than at 16°C. HSP90 immunopositivity seems to be particularly evident at 19°C. HPS70 immunopositivity was never observed in the developing lateral muscle.

The Actin-Binding Protein α-Adducin Is Required for Maintaining Axon Diameter.

The actin-binding protein adducin was recently identified as a component of the neuronal subcortical cytoskeleton. Here, we analyzed mice lacking adducin to uncover the function of this protein in actin rings. α-adducin knockout mice presented progressive axon enlargement in the spinal cord and optic and sciatic nerves, followed by axon degeneration and loss. Using stimulated emission depletion super-resolution microscopy, we show that a periodic subcortical actin cytoskeleton is assembled in every neuron type inspected including retinal ganglion cells and dorsal root ganglia neurons. In neurons devoid of adducin, the actin ring diameter increased, although the inter-ring periodicity was maintained. In vitro, the actin ring diameter adjusted as axons grew, suggesting the lattice is dynamic. Our data support a model in which adducin activity is not essential for actin ring assembly and periodicity but is necessary to control the diameter of both actin rings and axons and actin filament growth within rings.

Production of microparticles of molinate degrading biocatalysts using the spray drying technique.

Previous studies demonstrated the capability of mixed culture DC1 to mineralize the thiocarbamate herbicide molinate through the activity of molinate hydrolase (MolA). Because liquid suspensions are not compatible with long-term storage and are not easy to handle when bioremediation strategies are envisaged, in this study spray drying was evaluated as a cost-effective method to store and transport these molinate biocatalysts. Microparticles of mixed culture DC1 (DC1) and of cell free crude extracts containing MolA (MA) were obtained without any carrier polymer, and with calcium alginate (CA) or modified chitosan (MCt) as immobilizing agents. All the DC1 microparticles showed high molinate degrading activity upon storage for 6 months, or after 9 additions of ∼0.4 mM molinate over 1 month. The DC1-MCt microparticles were those with the highest survival rate and lowest heterogeneity. For MA microparticles, only MA-MCt degraded molinate. However, its Vmax was only 1.4% of that of the fresh cell free extract (non spray dried). The feasibility of using the DC1-MCt and MA-MCt microparticles in bioaugmentation processes was assessed in river water microcosms, using mass (g):volume (L) ratios of 1:13 and 1:0.25, respectively. Both type of microparticles removed ∼65-75% of the initial 1.5 mg L(-1) molinate, after 7 days of incubation. However, only DC1-MCt microparticles were able to degrade this environmental concentration of molinate without disturbing the native bacterial community. These results suggest that spray drying can be successfully used to produce DC1-MCt microparticles to remediate molinate polluted sites through a bioaugmentation strategy.

Myelin Lipids Inhibit Axon Regeneration Following Spinal Cord Injury: a Novel Perspective for Therapy.

Lack of axon regeneration following spinal cord injury has been mainly ascribed to the inhibitory environment of the injury site, i.e., to chondroitin sulfate proteoglycans (CSPGs) and myelin-associated inhibitors (MAIs). Here, we used shiverer (shi) mice to assess axon regeneration following spinal cord injury in the presence of MAIs and CSPG but in the absence of compact myelin. Although in vitro shi neurons displayed a similar intrinsic neurite outgrowth to wild-type neurons, in vivo, shi fibers had increased regenerative capacity, suggesting that the wild-type spinal cord contains additional inhibitors besides MAIs and CSPG. Our data show that besides myelin protein, myelin lipids are highly inhibitory for neurite outgrowth and suggest that this inhibitory effect is released in the shi spinal cord given its decreased lipid content. Specifically, we identified cholesterol and sphingomyelin as novel myelin-associated inhibitors that operate through a Rho-dependent mechanism and have inhibitory activity in multiple neuron types. We further demonstrated the inhibitory action of myelin lipids in vivo, by showing that delivery of 2-hydroxypropyl-ß-cyclodextrin, a drug that reduces the levels of lipids specifically in the injury site, leads to increased axon regeneration of wild-type (WT) dorsal column axons following spinal cord injury. In summary, our work shows that myelin lipids are important modulators of axon regeneration that should be considered together with protein MAIs as critical targets in strategies aiming at improving axonal growth following injury.